BR112019017891A2 - PROCESS FOR THERMAL TREATMENT OF BIOLOGICAL TISSUE - Google Patents

Abstract

a process for thermal treatment of biological tissue includes repeatedly applying pulsed energy to a target tissue over a period of time in order to controllably raise a temperature of the target tissue to create a therapeutic effect on the target tissue without destroying or permanently damaging the target tissue. after the first treatment is completed, the application of pulsed energy to the target tissue is interrupted for a period of time. in a single treatment session, a second treatment is performed on the target tissue after the interval by repeatedly reapplying pulsed energy in the target tissue in order to raise the target tissue temperature in a controlled manner to therapeutically treat the target tissue without permanently destroying or damaging the target tissue

Description

TREATMENT PROCESS

BIOLOGICAL TISSUE THERMAL

DESCRIPTION

BACKGROUND OF THE INVENTION

[0001] The present invention generally relates to systems and processes for treating biological tissue, such as diseased biological tissue. More particularly, the present invention relates to a process for thermal treatment of biological tissue using pulsed energy that creates a therapeutic effect for a target tissue without destroying or permanently damaging the target tissue.

[0002] The inventors have discovered that there is a therapeutic effect on biological tissue and, particularly, on damaged or diseased biological tissue, controllably raising the temperature of the tissue to a predetermined temperature range, maintaining the average rise in tissue temperature during several minutes at a predetermined level or below it so as not to permanently damage the target tissue. More particularly, the inventors have found that electromagnetic radiation, such as in the form of several wavelengths of laser light, can be applied to retinal tissue in a way that does not destroy or damage retinal tissue while achieving beneficial effects in eye diseases . The inventors found that a beam of laser light can be generated and applied to cells of the retinal tissue in order to be therapeutic, but sublethal to the cells of the retinal tissue and thus preventing harmful photocoagulation in the retinal tissue that provides preventive and protective treatment. retinal tissue of the eye. Treatment typically involves application

Petition 870190083984, of 08/28/2019, p. 15/162

2/98 of a laser micropulse frequency to irradiate a portion of a diseased retina for a total duration of less than a second. Each micropulse is in the order of tens to hundreds of microseconds in length, with the microseconds separated by one to several milliseconds, which raises the temperature of the tissue in a controlled manner.

[0003] It is believed that increasing the temperature of the tissue in such a controlled manner selectively stimulates the activation of the heat shock protein and / or the production and facilitation of protein repair, which serves as a mechanism to treat the tissue therapeutically. It is believed that this micropulse frequency thermally activates heat shock proteins (HSPs) in the target tissue. In the case of retinal tissue, the process thermally activates HSPs in the retinal pigment epithelium (RPE) layer immediately behind the retinal layer containing visually sensitive cones and rods and that these activated HSPs restore the sick retina to its healthy condition by removing and repairing the damaged proteins. This results in improved RPE function, improved retinal function and self-regulation, acute restorative inflammation, reduced chronic inflammation and systematic immunodulation. These laser-triggered effects slow, stop or reverse retinal disease, improve visual function and reduce the risk of visual loss. It is believed that increasing tissue temperature in a controlled manner to selectively stimulate heat shock protein activation also has benefits in other tissues.

[0004] HSPs are a family of proteins that are produced by cells in response to exposure to stressful conditions. The production of high levels of protein from

Petition 870190083984, of 08/28/2019, p. 16/162

3/98 thermal shock can be triggered by exposure to different types of environmental stress conditions, such as infection, inflammation, exercise, cell exposure to toxins, oxidants, heavy metals, hunger, hypoxia, water deprivation and tissue trauma.

[0005] Heat shock proteins are known to play a role in responding to a large number of abnormal conditions in the body's tissues, including viral infection, inflammation, malignant transformations, exposure to oxidizing agents, cytotoxins and anoxia. Several heat shock proteins function as intracellular guides for other proteins and members of the HSP family are expressed or activated at low to moderate levels due to their essential role in maintaining the protein and simply monitoring the cell's proteins even under non-stressful conditions. These activities are part of the cell's own repair system, called the cell voltage response or thermal shock response.

[0006] Heat shock proteins are found in almost all cells and types of multicellular organisms, as well as in explanted tissues and cultured cells. HSPs typically comprise 3% to 10% of a cell's proteins, although under stress the percentage can rise to 15%. The protein density of mammalian cells was found to be in the range of (2-4) x 10 ¹⁸ CM ~ ³ . Thus, the percentages mentioned above mean that the density of HSPs is normally (1-4) x 10 CM, while under tension the density can rise to (3-6) x 10 CM.

[0007] Heat shock proteins are typically named according to their molecular weight and act in ways

Petition 870190083984, of 08/28/2019, p. 17/162

4/98 different. An especially ubiquitous heat shock protein is Hsp70, a protein with a molecular weight of 70 kilodaltons. It plays a particularly significant role in protecting proteins that are being formed and in rescuing damaged proteins. It contains a groove with an affinity for neutral and hydrophobic amino acid residues that can interact with peptides with a maximum of 7 residues. Hsp70 has peptide and ATPase binding domains that stabilize protein structures in unfolded and assembly-competent states. HSPs play a role in preventing the aggregation of abnormally folded proteins, many of which have exposed hydrophobic portions and facilitate the refolding of proteins into their appropriate conformations. Hsp70 does this by first binding to the abnormally folded or fragmented protein, a bond that is energetically possible by a site that binds ATP and hydrolyzes it into ADP.

[0008] Hsp70 heat shock proteins are a member of membrane-bound extracellular heat shock proteins that are involved in binding antigens and presenting them to the immune system. Hsp70 has been found to inhibit the activity of the influenza A virus ribonucleoprotein and blocks the replication of the virus. Tumor-derived heat shock proteins elicit specific protective immunity. Experimental and clinical observations have shown that heat shock proteins are involved in the regulation of autoimmune arthritis, type 1 diabetes mellitus, arterial sclerosis, multiple sclerosis and other autoimmune reactions.

[0009] Thus, it is believed that it is advantageous to be able to selectively and controlled increase a tissue temperature

Petition 870190083984, of 08/28/2019, p. 18/162

5/98 target to a predetermined temperature range over a short period of time, while maintaining the average rise in tissue temperature at a predetermined temperature over a longer period of time. This is believed to induce a response to heat shock in order to increase the number or activity of heat shock proteins in the body's tissue in response to infections or other abnormalities. However, this must be done in a controlled manner, in order not to damage or destroy the tissue or area of the body to be treated. It would also be desirable to maximize the amount of heat shock protein activation within the cells of a target tissue during a single treatment session. The present invention addresses these needs and provides other related advantages.

SUMMARY OF THE INVENTION

[00010] The present invention relates to a process for the thermal treatment of biological tissues by applying pulsed energy to a target tissue to therapeutically treat the target tissue. A first treatment for the target tissue is performed by generating pulsed energy and repeatedly applying the pulsed energy to the target tissue over a period of time in order to raise the temperature of the target tissue in a controlled manner to therapeutically treat the target tissue without destroying or permanently damaging the target tissue. The target tissue may comprise retinal tissue.

[00011] Pulsed energy has energy parameters, including wavelength or frequency, duty cycle and pulse frequency duration. Energy parameters are selected to raise tissue temperature

Petition 870190083984, of 08/28/2019, p. 19/162

6/98 target up to 11 ° C to achieve a therapeutic effect, in which the average rise in tissue temperature over several minutes is maintained at or below a predetermined level so as not to permanently damage the target tissue. The pulsed energy parameters can be selected so that the temperature of the target tissue is increased by approximately 6 ° C to 11 ° C at least during the application of the pulsed energy to the target tissue. The average rise in temperature of the target tissue over several minutes is maintained at 6 ° C or less, as well as at approximately 1 ° C or less over several minutes.

[00012] Pulsed energy can comprise a beam of light, a microwave, a radio frequency or an ultrasound. A device can be inserted into a body cavity to apply pulsed energy to the tissue. Pulsed energy can be applied to an external area of a body that is adjacent to the target tissue or has a blood supply close to the surface of the external area of the body.

[00013] The pulsed energy can comprise a radio frequency between approximately 3 to 6 megahertz (MHz). It can have a duty cycle of approximately 2.5% to 5%. It can have a pulsed frequency duration between approximately 0.2 and 0.4 seconds. The radio frequency can be generated with a device with coil radii between approximately 2 and 6 mm and approximately between 13 and 57 amps.

[00014] Pulsed energy can comprise a microwave frequency between 10 and 20 gigahertz (GHz). Microwaves can have a pulse rate of approximately 0.2 to 0.6 seconds. Microwaves can have a cycle of

Petition 870190083984, of 08/28/2019, p. 20/162

7/98 work between approximately 2% and 5%. Microwaves can have an average power between approximately 8 and 52 watts.

[00015] Pulsed energy may comprise a beam of pulsed light, such as one or more beams of laser light. The light beam can have a wavelength between approximately 530 nm and 1,300 nm and more preferably between 800 nm and 1,000 nm. The pulsed light beam can have a power between approximately 0.5 and 74 watts. The pulsed light beam has a duty cycle of less than 10% and, preferably, between 2.5% and 5%. The pulsed light beam can have a pulse frequency duration of approximately 0.1 and 0.6 seconds.

[00016] The pulsed energy can comprise a pulsed ultrasound, with a frequency between approximately 1 and 5 MHz. The ultrasound has a frequency duration of approximately 0.1 and 05 seconds. Ultrasound can have a duty cycle between approximately 2% and 10%. The ultrasound has a power between approximately 0.46 and 28.6 watts.

[00017] The first treatment comprises the application of pulsed energy to the target tissue for a period of less than ten seconds and, more typically, less than a second. The first treatment creates a level of activation of the heat shock protein in the target tissue.

[00018] The application of pulsed energy to the target tissue is interrupted for a period of time that preferably exceeds the time period of the first treatment. The time interval can comprise several seconds to several minutes, such as three seconds to three minutes or preferably between ten seconds and ninety seconds.

[00019] After the time interval and within a single treatment session, a second treatment is performed at

Petition 870190083984, of 08/28/2019, p. 21/162

8/98 target tissue, repeatedly reapplying pulsed energy in the target tissue in order to raise the temperature of the target tissue in a controlled manner to therapeutically treat the target tissue without destroying or permanently damaging the target tissue. The second treatment increases the level of heat shock protein activation in the target tissue, such that it is at a level that is higher than the level after the first treatment.

[00020] Other features and advantages of the present invention will be evident in the following more detailed description, taken in conjunction with the accompanying drawings that illustrate, by way of example, the principles of the invention.

BRIEF DESCRIPTION OF THE DRAWINGS

[00021] The accompanying drawings illustrate the invention. In such drawings:

FIGURES IA and 1B are graphs that illustrate the average power of a laser source compared to a source radius and duration of the laser pulse frequency;

FIGURES 2A and 2B are graphs that illustrate the time for the temperature to drop, depending on the radius of the laser source and the wavelength;

FIGURES 3 to 6 are graphs illustrating the peak ampere turns for various radiofrequencies, duty cycles and coil radii;

FIGURE 7 is a graph representing the time for the temperature rise to fall compared to the radius of the radio frequency coil;

FIGURES 8 and 9 are graphs that represent the average microwave power compared to the microwave frequency and pulse frequency durations;

Petition 870190083984, of 08/28/2019, p. 22/162

9/98

FIGURE 10 is a graph representing the time for the temperature to drop for various microwave frequencies;

FIGURE 11 is a graph that represents the power of the average ultrasound source compared to the frequency and duration of the pulse frequency;

FIGURES 12 and 13 are graphs that represent the time for the temperature drop for various ultrasound frequencies;

FIGURE 14 is a graph that represents the volume of the focal heated region compared to the frequency of ultrasound;

FIGURE 15 is a graph that compares temperature equations over pulse durations for an ultrasound energy source;

FIGURES 16 and 17 are graphs that illustrate the magnitude of the damage logarithm and the Arrhenius integrals of HSP activation as a function of temperature and pulse duration;

FIGURE 18 is a schematic view of a light generating unit that produces time series of pulses, having a light tube extending from it, according to the present invention;

FIGURE 19 is a cross-sectional view of a photostimulation delivery device that delivers electromagnetic energy to the target tissue, in accordance with the present invention;

FIGURE 20 is a schematic view illustrating a system used to generate a beam of laser light, in accordance with the present invention;

Petition 870190083984, of 08/28/2019, p. 23/162

10/98

FIGURE 21 is a schematic view of the optics used to generate a geometric pattern of laser light, in accordance with the present invention;

FIGURE 22 is a top plan view of an optical scanner used in accordance with the present invention;

FIGURE 23 is a partially exploded view of the optical scanner of FIG. 22, which illustrates the various parts of the components thereof;

FIGURE 24 illustrates the controlled deviation of the exposure of an exemplary geometric pattern grid of laser points to treat a target tissue, according to an embodiment of the present invention;

FIGURE 25 is a schematic view illustrating the use of a geometric object in the form of a controllably digitized line to treat an area of the target tissue;

Figure 26 is a schematic view similar to that of FIG. 25, but illustrating the geometric line or bar rotated to treat the target tissue;

FIGURE 27 is a schematic view illustrating an alternative embodiment of the system used to generate beams of laser light to treat tissue in accordance with the present invention;

FIGURE 28 is a schematic view illustrating yet another embodiment of a system used to generate laser light beams for treating tissue in accordance with the present invention;

FIGURE 29 is a schematic cross-sectional view of one end of an endoscope inserted into the

Petition 870190083984, of 08/28/2019, p. 24/162

11/98 nasal cavity and that treats its tissue, according to the present invention;

FIGURE 30 is a schematic and partially cross-sectional view of a bronchoscope that extends through the trachea and into the bronchus of a lung and provides treatment for it in accordance with the present invention;

FIGURE 31 is a schematic view of a colonoscope that provides photostimulation to an intestinal or colon area of the body, in accordance with the present invention;

FIGURE 32 is a schematic view of an endoscope inserted into a stomach and providing treatment to it, in accordance with the present invention;

FIGURE 33 is a partially sectioned perspective view of a capsule endoscope, used in accordance with the present invention;

FIGURE 34 is a schematic view of a pulsed high-intensity focused ultrasound to treat the body's internal tissue, in accordance with the present invention;

FIGURE 35 is a schematic view for delivering therapy to a patient's bloodstream through an earlobe, in accordance with the present invention;

FIGURE 36 is a cross-sectional view of a stimulation therapy device of the present invention used to deliver photostimulation to the blood through an ear lobe in accordance with the present invention;

FIGURES 37A to 37D are schematic views illustrated in the application of micropulsed energy to different treatment areas during a predetermined time interval, within a single treatment session and

Petition 870190083984, of 08/28/2019, p. 25/162

12/98 reapplying energy to previously treated areas, according to the present invention;

FIGURES 38 to 40 are graphs that represent the relationship between treatment power and time according to the modalities of the present invention;

FIGURE 41 is a graph representing the wavefront of two sources separated by a distance;

FIGURES 42A and 42B are graphs that represent the behavior of the components of the HSP cellular system over time, after a sudden increase in temperature;

FIGURES 43A to 43H are graphs showing the behavior of the components of the HSP cellular system in the first minute after a sudden increase in temperature;

FIGURES 44A and 44B are graphs that illustrate the variation in the activated concentrations of HSP and non-activated HSP in the cytoplasmic reservoir over an interval of one minute, according to the present invention; and

FIGURE 45 is a graph representing the reasons for improvement depending on the interval between treatments, according to the present invention.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[00022] As shown in the accompanying drawings, and as more fully described herein, the present invention relates to a system and method for providing pulsed energy, such as ultrasound, ultraviolet radio frequency, microwave radio frequency, one or more beams of light and the like, with energy parameters selected to cause a thermal time course in the tissue to raise the temperature of the tissue for a short time to a level sufficient to achieve a therapeutic effect while maintaining a temperature

Petition 870190083984, of 08/28/2019, p. 26/162

13/98 average tissue over an extended period below a predetermined level in order to avoid permanent tissue damage. It is believed that the creation of the thermal time course stimulates the activation or production of the heat shock protein and facilitates the repair of the protein without causing any damage.

[00023] The inventors have found that electromagnetic radiation can be applied to retinal tissue in a way that does not destroy or damage retinal tissue, while at the same time achieving beneficial effects in eye diseases. More particularly, a beam of laser light can be generated which is therapeutic but sublethal to the cells of the retinal tissue and thus prevents harmful photocoagulation in the retinal tissue which provides a preventive and protective treatment of the retinal tissue of the eye. This is believed to be due, at least in part, to the stimulation and activation of heat shock proteins and to the facilitation of protein repair in retinal tissue. This is disclosed in the United States patent applications, serial numbers 14 / 607,959, filed on January 28, 2015, 13 / 798,523, filed on March 13, 2013 and 13 / 481,124, filed on May 25, 2012, whose contents are incorporated by reference as if they were done in full.

[00024] Various parameters of the light beam must be taken into account and selected so that the combination of the selected parameters achieves the therapeutic effect without permanently damaging the tissue. These parameters include laser wavelength, laser source radius, average laser power, total pulse duration and pulse train duty cycle.

Petition 870190083984, of 08/28/2019, p. 27/162

14/98

[00025] The selection of these parameters can be determined by requiring that the Arrhenius integral for HSP activation is greater than 1 or unit. Arrhenius integrals are used to analyze the impacts of actions on biological tissue. See, for example, The CRC Handbook of Thermal Engineering, ed. Frank Kreith, Springer Science and Business Media (2000). At the same time, the selected parameters must not permanently damage the fabric. Thus, the Arrhenius integral for damage can also be used, where the resolved Arrhenius integral is less than 1 or unit.

[00026] Alternatively, the FDA / FCC restrictions on energy deposition per gram unit of tissue and temperature rise measured in minute periods are met in order to avoid permanent tissue damage. FDA / FCC requirements on energy deposition and temperature rise are widely used and can be referenced, for example, at www.fda.gov/medicaldevices/deviceregulationandguidance/guidanc edocuments / ucm073817.htm # attacha for electromagnetic sources and Anastosio and P. LaRivero, ed., Emerging Imaging Technologies. CRC Press (2012), for ultrasound sources. In general, increases in tissue temperature between 6 ° C and 11 ° C can create a therapeutic effect, activating heat shock proteins, maintaining the average tissue temperature over an extended period of time, such as for several minutes, such as six minutes, below a predetermined temperature, such as 6 ° C and even 1 ° C or less in certain circumstances, will not permanently damage the fabric.

[00027] The inventors found that the generation of a subliminal and sublethal laser beam of micropulse that has a

Petition 870190083984, of 08/28/2019, p. 28/162

15/98 wavelength greater than 532 nm and a duty cycle of less than 10% at a predetermined intensity or power and a predetermined pulse duration or exposure time creates a desirable photostimulation of the retina without any visible burn area or destruction of fabric. More particularly, a beam of laser light with a wavelength between 550 nm and 1,300 nm and in a particularly preferred embodiment between 810 nm and 1,000 nm, with a duty cycle of approximately 2.5% to 5% and an intensity or predetermined power (such as between 100 and 590 watts per square centimeter on the retina or approximately 1 watt per laser point for each treatment point on the retina) and a predetermined pulse length or exposure time (such as between 100 and 600 milliseconds or less ) creates a sublethal, true subliminal photostimulation of the retina in which all areas of the retinal pigment epithelium exposed to laser irradiation are preserved and available to contribute therapeutically. In other words, the inventors found that the increase in retinal tissue, at least to a therapeutic level, but below a cellular level or lethal tissue, recreates the benefit of the halo effect of prior art methods without destroying, burning or damaging the retinal tissue. This is referred to here as sub-threshold diode (SDM) micropulse laser treatment.

[00028] SDM does not produce laser-induced retinal damage (photocoagulation), and has no known adverse effects, and has been reported to be an effective treatment for various retinal disorders (including proliferative diabetic retinopathy (PDR) of macular edema) diabetic (DME), macular edema due to occlusion of the central vein of

Petition 870190083984, of 08/28/2019, p. 29/162

16/98 retina (BRVO), central serous chorioretinopathy (CSR), reversal of drug tolerance and prophylactic treatment of progressive degenerative retinopathies, such as age-related macular degeneration, Stargard's disease, conical dystrophies and pigmentous retinitis. The safety of SDM is such that it can be used transfoveally in the eyes with 20/20 visual acuity to reduce the risk of visual loss due to DME involving the early fovea.

[00029] One mechanism by which the SDM can work is the generation or activation of heat shock proteins (HSPs). Despite an almost infinite variety of possible cellular abnormalities, cells of all types share a common and highly conserved mechanism of repair: heat shock proteins (HSPs). HSPs are caused almost immediately, in seconds to minutes, by almost any type of stress or cell injury. In the absence of lethal cell damage, HSPs are extremely effective in repairing and returning the viable cell to a more normal functional state. Although HSPs are transient, usually peaking in hours and persisting for a few days, their effects can be long lasting. HSPs reduce inflammation, a common factor in many disorders.

[00030] Laser treatment can induce the production or activation of HSP and alter the expression of cytokines. The more sudden and severe the non-lethal cellular tension (such as laser irradiation), the faster and more robust the HSP activation. Thus, a breakdown of repetitive low-temperature thermal peaks at a very steep rate of change (~ 7 ° C with each 100 ps micropulse, or 70,000 ° C / s) produced by each SDM exposure is especially effective in stimulating

Petition 870190083984, of 08/28/2019, p. 30/162

17/98 activation of HSPs, particularly in comparison to non-lethal exposure for subliminal treatment with continuous wave lasers, which can only double the low average rise in tissue temperature.

[00031] Laser wavelengths below 550 nm produce increasingly cytotoxic photochemical effects. At 810 nm, SDM produces photothermal, rather than photochemical, cell voltage. Thus, SDM is able to affect the tissue without damaging it. The clinical benefits of SDM are, therefore, mainly produced by sub-morbid photothermal cell activation of HSP. In dysfunctional cells, the stimulation of HSP by SDM results in expression of normalized cytokines and, consequently, improves the structure and function of. The therapeutic effects of this low-intensity laser / tissue interaction are amplified by the application of high-density laser, recruiting all dysfunctional cells in the target tissue area by dense / confluent treatment of a large area of tissue, including all areas of pathology, thus maximizing the treatment effect. These principles define the SDM treatment strategy described here.

[00032] Because normally functioning cells do not need repair, stimulation of HSP in normal cells would tend to have no noticeable clinical effect. The patoselectivity of the effects of laser in the near infrared, such as SDM, affecting diseased cells, but not affecting normal cells in various types of cells, is consistent with the clinical observations of SDM. It has been reported that SDM has a clinically broad therapeutic range, unique among retinal laser modalities, consistent with the predictions of the National Maximum Standards Institute of

Petition 870190083984, of 08/28/2019, p. 31/162

18/98 maximum exposure allowed. While SDM can cause direct photothermal effects, such as entropy protein splitting and breakdown, SDM appears optimized for clinically safe and effective stimulation of HSP-mediated repair.

[00033] As noted earlier, while SDM stimulation of HSPs is nonspecific with respect to the disease process, the result of HSP-mediated repair is, by its nature, specific to the state of the dysfunction. HSPs tend to fix what's wrong, whatever. Thus, the observed effectiveness of SDM in retinal conditions is as different as BRVO, DME, PDR, CSR, age-related and genetic retinopathy and drug-tolerant NAMD. Conceptually, this feature can be considered a kind of way to Reset to Default of the SDM action. For the wide range of disorders in which cell function is critical, SDM normalizes cell function by triggering a reset (to factory default settings) through HSP-mediated cell repair.

[00034] The inventors have found that treating SDM in patients suffering from age-related macular degeneration (AMD) can slow progress or even stop the progression of AMD. Most patients showed significant improvement in the dynamic functional logMAR mesoptic visual acuity and in the mesoptic contrast visual acuity after SDM treatment. SDM is believed to work by directing, preserving and normalizing (going to normal) the function of the retinal pigment epithelium (RPE).

[00035] SDM has also been shown to stop or reverse manifestations of diabetic retinopathy disease status without

Petition 870190083984, of 08/28/2019, p. 32/162

19/98 treatment-related damage or adverse effects, despite the persistence of systemic diabetes mellitus. Based on this, there is a theory that SDM can work by inducing a return to more normal cellular function and expression of cytokines in RPE cells affected by diabetes, analogous to pressing the reset button on an electronic device to restore factory default settings. Based on previous information and studies, SDM treatment can directly affect cytokine expression by activating heat shock protein (HSP) in the target tissue.

[00036] As heat shock proteins play a role in responding to a large number of abnormal conditions in body tissue other than eye tissue, it is believed that similar systems and methodologies can be advantageously used in the treatment of such conditions abnormal, infections, etc. As such, the present invention relates to the controlled application of ultrasound or electromagnetic radiation to treat abnormal conditions, including inflammation, autoimmune conditions, and cancers that are accessible through optical fiber from endoscopes or surface probes, as well as electromagnetic / sound waves. For example, cancers on the surface of the prostate that have the greatest threat of metastasis can be accessed through optical fibers in a protoscope. Colon tumors can be accessed through a fiber optic system, such as those used in colonoscopy.

[00037] As indicated previously, micropulsed subliminal diode laser (SDM) photostimulation was effective in stimulating the direct repair of abnormally folded proteins in the ocular tissue. In addition to activating

Petition 870190083984, of 08/28/2019, p. 33/162

20/98

HSP, another way that can occur is because the temperature spikes caused by the micropulses in the form of a thermal time allow the diffusion of water inside the proteins and this allows the breaking of the peptide-peptide hydrogen bonds that prevent the protein from returning to its native state. The diffusion of water in proteins results in an increase in the number of restrictive hydrogen bonds by a factor of the order of a thousand. Thus, it is believed that this process can be applied to other diseases and tissues also advantageously.

[00038] As explained earlier, the energy source to be applied to the target tissue will have energetic and operational parameters that must be determined and selected in order to achieve the therapeutic effect while not permanently damaging the tissue. Using a light beam energy source, such as a laser beam, for example, the parameters of wavelength, duty cycle and duration of the total laser pulse train must be taken into account. Other parameters that can be considered include the radius of the laser source as well as the average laser power. Adjusting or selecting one of these parameters can have an effect on at least one other parameter.

[00039] FIGURES IA and 1B illustrate graphs showing the average power in watts compared to the radius of the laser source (between 0.1 cm and 0.4 cm) and the duration of the pulse frequency (between 0.1 and 0.6 seconds). FIG. IA shows a wavelength of 880 nm, while FIG. 1B has a wavelength of 1,000 nm. It can be seen in these figures that the required power decreases monotonically as the radius of the source decreases, as the total duration of the

Petition 870190083984, of 08/28/2019, p. 34/162

21/98 frequency increases and as the wavelength decreases. Preferred parameters for the radius of the laser source are 1 mm to 4 mm. For a wavelength of 880 nm, the minimum energy value is 0.55 watts, with a laser source radius of 1 mm and the total pulse frequency duration is 600 milliseconds. The maximum energy value for the 880 nm wavelength is 52.6 watts when the laser source radius is 4 mm and the total duration of the pulse frequency is 100 milliseconds. However, when selecting a laser with a wavelength of 1,000 nm, the minimum energy value is 0.77 watts with a laser source radius of 1 mm and a total pulse train duration of 600 milliseconds and a maximum value of 73.6 watt power when the radius is 4 mm and the total pulse duration is 100 milliseconds. The powers

peak corresponding during a pulse individual, are obtained of medium powers dividing by cycle in job. [00040] 0 volume of the region of fabric a be heated is determined by length wave, the length in absorption at the relevant fabric and the width of the beam. THE

total pulse duration and average laser power determine the total energy delivered to heat the tissue, and the pulse train duty cycle provides the associated peak or peak power associated with the average laser power. Preferably, the energy parameters of the pulsed energy source are selected so that approximately 20 to 40 joules of energy are absorbed by each cubic centimeter of the target tissue.

[00041] The absorption length is very small in the thin melanin layer in the pigmented epithelium of the retina. In others

Petition 870190083984, of 08/28/2019, p. 35/162

22/98 body parts, the absorption length is generally not that short. At wavelengths ranging from 400 nm to 2,000 nm, the depth of penetration and the skin are in the range of 0.5 mm to 3.5 mm. The depth of penetration into human mucous tissues in the range of 0.5 mm to 6.8 mm. Consequently, the heated volume will be limited to the outer or inner surface where the radiation source is placed, with a depth equal to the depth of penetration and a transversal dimension equal to the transversal dimension of the radiation source. As the light beam energy source is used to treat diseased tissues near external surfaces or near accessible internal surfaces, a radius of origin between 1 mm and 4 mm and operating at a wavelength of 880 nm produces a depth of penetration of approximately 2.5 mm and a wavelength of 1,000 nm produces a penetration depth of approximately 3.5 mm.

[00042] It has been determined that the target tissue can be heated to approximately 11 ° C for a short period of time, such as less than a second, to create the therapeutic effect of the invention while maintaining the average temperature of the target tissue at a range lower temperature, such as less than 6 ° C or up to 1 ° C or less over an extended period of time, such as several minutes. The duty cycle selection and the total duration of the pulse rate provide time intervals in which the heat can dissipate. A duty cycle of less than 10% and, preferably between 2.5% and 5%, with a total pulse duration between 100 milliseconds and 600 milliseconds, was considered effective. FIGS. 2A and 2B illustrate the fall time from 10 ° C to 1 ° C for a laser source with a radius between 0.1 cm and 0.4 cm, the length being

Petition 870190083984, of 08/28/2019, p. 36/162

23/98 880 nm wave in FIG. 2A and 1,000 nm in FIG. 2B. It can be seen that the drop time is shorter when using a wavelength of 880 nm, but the wavelength is within the requirements and acceptable operating parameters to achieve the benefits of the present invention without causing permanent tissue damage.

[00043] It has been found that the average increase in temperature of the desired target region, increasing at least 6 ° C and up to 11 ° C and preferably approximately 10 ° C, during the total irradiation period, results in the activation of HSP. The temperature control of the target tissue is determined by choosing the source and target parameters, so that the Arrhenius integral for HSP activation is greater than 1, while ensuring compliance with conservative FDA / FCC requirements to prevent damage or an Arrhenius integral of damage less than 1.

[00044] In order to comply with conservative FDA / FCC restrictions to prevent permanent damage to tissues, light beams and other sources of electromagnetic radiation, the average elevation of the target tissue temperature in any six minute period is 1 ° C or less. FIGS. 2A and 2B above illustrate the typical drop times required for the temperature in the heated target region to decrease by thermal diffusion from a temperature rise of approximately 10 ° C to 1 ° C, as seen in FIG. 2A when the wavelength is 880 nm and the source diameter is 1 millimeter, the temperature drop time is 16 seconds. The temperature drop time is 107 seconds when the source diameter is 4 mm. As shown in FIG. 2B, when the wavelength is 1,000 nm, the drop time of

Petition 870190083984, of 08/28/2019, p. 37/162

24/98 temperature is 18 seconds when the source diameter is 1 mm and 136 seconds when the source diameter is 4 mm. This is well within the time that the average temperature rise is maintained over several minutes, such as 6 minutes or less. While the temperature of the target tissue is increased, such as at approximately 10 ° C, very quickly, such as in a fraction of a second during the application of the energy source to the tissue, the relatively low duty cycle provides relatively long periods of time between the pulses of energy applied to the tissue and the duration of the relatively short pulse rate ensure sufficient diffusion and temperature drop within a relatively short period of time comprising several minutes, such as 6 minutes or less, that there is no permanent tissue damage.

[00045] The parameters differ for individual energy sources, including microwaves, infrared laser, radio frequency and ultrasound, because the absorbing properties of tissues differ for these different types of energy sources. The water content in the tissue can vary from one type of tissue to another, however, there is an observed uniformity of the properties of the tissues under normal or near normal conditions that allowed the publication of tissue parameters that are widely used by doctors in the design of treatments. The following are the tables that illustrate the properties of electromagnetic waves in biological media, with Table 1 referring to muscles, skin and tissues with high water content and Table 2 referring to fat, bones and tissues with low water content.

Petition 870190083984, of 08/28/2019, p. 38/162

25/98

Table 1. Properties of electromagnetic waves in a biological environment: muscle, skin and tissues with high water content

Reflection coefficient

Constant Length Frequency dielectric air wave (cm) (MHz) Depth wavelength Air-muscle interface Muscle-fat interface conductivity of r ₀ r 0

27.12

40.68

100

200

300

433

750

915

1,500

2,450

3,000 5,000 5,800 8,000 10,000

30,000 2,000 0.400 436 91.3 0.982 +179 3,000 160 0.625 118 21.6 0.956 +178 1.106 113 0.612 68.1 14.3 0.925 +177 0.651 -11.13 738 97.3 0.693 51.3 11.2 0.913 +176 0.652 -10.21 300 71.7 0.889 27 6.66 0.881 +175 0.650 -7.96 150 56.5 1.28 16.6 4.79 0.844 +175 0.612 -8.06 100 54 1.37 11.9 3.89 0.825 +175 0.592 -8.14 69.3 53 1.43 8.76 3.57 0.803 +175 0.562 -7.06 40 52 1.54 5.34 3.18 0.779 +176 0.532 -5.69 32.8 51 1.60 4.46 3.04 0.772 +177 0.519 -4.32 20 49 1.77 2.81 2.42 0.761 +177 0.506 -3.66 12.2 47 2.21 1.76 1.70 0.754 +177 0.500 -3.88 10 46 2.26 1.45 1.61 0.751 +178 0.495 -3.20 6 44 3.92 0.89 0.788 0.749 +177 0.502 -4.95 5.17 43.3 4.73 0.775 0.720 0.746 +177 0.502 -4.29 3.75 40 7.65 0.578 0.413 0.744 +176 0.513 -6.65 3 39.9 10.3 0.464 0.343 0.743 +176 0.518 -5.95

Table 2. Properties of electromagnetic waves in biological media: fat, bone and tissues with low water content

Reflection coefficient

Constant Length Conductivity Length P ^r ° f ^{ur |} air-fat interface fat-muscle interface

Dielectric wave frequency (M Hz) (cm) € L oL, wave (cm) r ₀ r 0

30,000

27.12 40.68

100

200

300

433 750 915 1,500 2,450 3,000 5,000 5,900 8,000 10,000

1,106 20 10.9 to 43.2 241 159 0.660 +174 0.651 +169 738 14.6 12.6 to 52.8 187 118 0.617 +173 0.652 +170 300 7.45 19.1 to 75.9 106 60.4 0.511 +168 0.650 +172 150 5.95 25.8 to 94.2 59.7 39.2 0.458 +168 0.612 +172 100 5.7 31.6 to 107 41 32.1 0.438 +169 0.592 +172 69.3 5.6 37.9 to 118 28.8 26.2 0.427 +170 0.562 +173 40 5.6 49.8 to 138 16.8 23 0.415 +173 0.532 +174 32.8 5.6 55.6 to 147 13.7 17.7 0.417 +173 0.519 +176 20 5.6 70.8 to 171 8.41 13.9 0.412 +174 0.506 +176 12.2 5.5 96.4 to 213 5.21 11.2 0.406 +176 0.500 +176 10 5.5 110 to 234 4.25 9.74 0.406 +176 0.495 +177 6 5.5 162 to 309 2.63 6.67 0.393 +176 0.502 +175 5.17 5.05 186 to 338 2.29 5.24 0.388 +176 0.502 +176 3.75 4.7 255 to 431 1.73 4.61 0.371 +176 0.513 +173 -M- 3 4.5 324 to 549 1.41 3.39 0.363 +175 0.518 + 174, -

[00046] The radiofrequency absorption lengths in the body tissue are long compared to the body dimensions. Consequently, the heated region is determined by the dimensions of the coil that is the source of the radio frequency energy and not by the absorption lengths. Long distances from a coil, the magnetic field (near) a

Petition 870190083984, of 08/28/2019, p. 39/162

26/98 coil drops as 1 / r ³ . At shorter distances, the electric and magnetic fields can be expressed in terms of the vector's magnetic potential, which in turn can be expressed in closed form in terms of elliptic integrals of the first and second types. Heating occurs only in a region that is comparable in size to the dimensions of the coil source itself. Therefore, if it is desired to preferably heat a region characterized by a radius, the original coil will be chosen to have a similar radius. The heating falls very quickly outside a hemispheric region of lightning due to the 1 / r ³ drop in the magnetic field. Since it is proposed to use radiofrequency in diseased tissue accessible only externally or from internal cavities, it is reasonable to consider a coil radius between approximately 2 and 6 mm.

[00047] The radius of the source coil (s) and the number of turns (NI) in the source coils provide the magnitude and spatial extension of the magnetic field and radio frequency is a factor that relates the magnitude of the electric field the magnitude of the magnetic field. Heating is proportional to the product of the conductivity and to the square of the electric field. For target tissues of interest that are close to external or internal surfaces, the conductivity is that of the skin and mucous tissue. The duty cycle of the pulse train, as well as the total duration of the train of a pulse train are factors that affect the total amount of energy that is supplied to the tissue.

[00048] Preferred parameters for a radio frequency energy source were determined as coil radii between 2 and 6 mm, radio frequencies in the range of 3 to 6 MHz, durations of

Petition 870190083984, of 08/28/2019, p. 40/162

27/98 total pulse train from 0.2 to 0.4 seconds and a duty cycle between 2.5% and 5%. FIGS. 3 to 6 show how the number of ampere turns varies as these parameters are varied, in order to provide a temperature increase that produces an Arrhenius integral of approximately one or unit for HSP activation. With reference to FIG. 3, for an RF frequency of 6 MHz, a pulse frequency duration between 0.2 and 0.4 seconds, a coil radius between 0.2 and 0.6 cm and a cycle of 5%, the peak of voltage (NI) is 13 in the 0.6 cm coil radius and 20 in the 0.2 cm coil radius. For a frequency of 3 MHz, as illustrated in FIG. 4, the peak ampere turns are 26 when the pulse frequency duration is 0.4 seconds and the coil radius is 0.6 cm and the duty cycle is 5%. However, with the same 5% duty cycle, the peak ampere turns are 40 when the coil radius is 0.2 cm and the pulse frequency lasts 0.2 seconds. A 2.5% duty cycle is used in FIGS. 5 and 6. This produces, as illustrated in FIG. 5, 18 amps turns to a 6 MHz radiofrequency with a 0.6 cm radius and a pulse frequency duration of 0.4 seconds and 29 amps turns when the coil radius is only 0.2 cm and the duration of the pulse rate is 0.2 seconds. With reference to FIG. 6, with a duty cycle of 2.5% and a radio frequency of 3 MHz, the peak ampere turns are 36 when the pulse frequency duration is 0.4 seconds and the coil radius is 0, 6 cm and 57 ampere turns when the pulse frequency lasts 0.2 seconds and the coil radius is 0.2 cm.

Petition 870190083984, of 08/28/2019, p. 41/162

28/98

[00049] The time, in seconds, for the temperature rise to drop from approximately 10 ° C to approximately 1 ° C for coil radii between 0.2 cm and 0.6 cm is illustrated for a radio frequency energy source in FIG . 7. The temperature drop time is approximately 37 seconds when the

coil radius radio frequency is 0 , 2 cm and about 233 seconds when the lightning gives coil in radio frequency is 0.5 cm. When the lightning gives coil in radio frequency is 0.6 cm, the fall is in about 336 seconds, what is still inside of acceptable range of time falling but in a track

top of it.

[00050] Microwaves are another source of electromagnetic energy that can be used in accordance with the present invention. The frequency of the microwaves determines the distance of penetration into the tissue. The gain of a conical microwave horn is large compared to the microwave wavelength, indicating under these circumstances that the energy is radiated mainly in a direct forward charge. Typically, a microwave source used in accordance with the present invention has a linear dimension in the order of one centimeter or less, so the source is less than the wavelength, in which case the microwave source can be approximated like a dipole antenna. These small microwave sources are easier to insert into the body's internal cavities and can also be used to radiate external surfaces. In this case, the heated region can be approached by a hemisphere with a radius equal to the microwave absorption length in the body tissue to be treated. How microwaves are used to treat nearby tissues

Petition 870190083984, of 08/28/2019, p. 42/162

29/98 of external surfaces or surfaces accessible from internal cavities, frequencies in the range of 10 to 20 GHz are used, in which the corresponding penetration distances are only between approximately 2 and 4 mm.

[00051] The rise in tissue temperature using a microwave energy source is determined by the average microwave power and the total pulse train duration. The duty cycle of the pulse train determines the peak power on a single pulse on a pulse train. Since the radius of the source is less than approximately 1 centimeter and frequencies between 10 and 20 GHz are typically used, a resulting pulse train duration of 0.2 and 0.6 seconds is preferred.

[00052] The required power monotonically decreases as the train's duration increases and the microwave frequency increases. For a frequency of 10 GHz, the average power is 18 watts when the pulse train lasts 0.6 seconds and 52 watts when the pulse train lasts 0.2 seconds. For a microwave frequency of 20 GHz, an average power of 8 watts is used when the pulse train is 0.6 seconds and can be 26 watts when the pulse train is only 0.2 seconds long . The corresponding peak power is obtained from the average power simply by dividing by the duty cycle.

[00053] With reference now to FIG. 8, a graph represents the average microwave power in watts of a microwave with a frequency of 10 GHz and a duration of the pulse frequency between 0.2 seconds and 0.6 seconds. FIG. 9 is a similar graph, but it shows the average microwave power for a microwave with a frequency of 20 GHz.

Petition 870190083984, of 08/28/2019, p. 43/162

30/98 since the average power of the microwave source varies according to the total duration of the frequency and the microwave frequency varies. The fixed condition, however, is that the Arrhenius integral for HSP activation in the heated region is approximately 1.

[00054] With reference to FIG. 10, a graph illustrates the time, in seconds, for the temperature to drop from approximately 10 ° C to 1 ° C compared to microwave frequencies between 58 MHz and 20,000 MHz. The minimum and maximum temperature drop for the range preferred microwave frequency is 8 seconds when the microwave frequency is 20 GHz and 16 seconds when the microwave frequency is 10 GHz.

[00055] The use of ultrasound as an energy source allows the heating of superficial tissues and tissues of different depths in the body, including deeper tissues. The absorption length of the ultrasound in the body is quite long, as evidenced by the wide use of images. Therefore, ultrasound can be focused on target regions deep within the body, with the heating of a focused ultrasound beam concentrated mainly on the approximately cylindrical focal region of the beam. The heated region has a volume determined by the focal waist of the ventilated disc and the length of the focal waist region, which is the confocal parameter. Multiple bundles of sources at different angles can also be used, with heating occurring in the overlapping focal regions.

[00056] For ultrasound, the relevant parameters for determining tissue temperature are the frequency of the ultrasound, the total duration of the train and the power of the transducer when the focal length and diameter of the transducer

Petition 870190083984, of 08/28/2019, p. 44/162

31/98 ultrasound is given. The frequency, focal length and diameter determine the volume of the focal region where the ultrasound energy is concentrated. It is the focal volume that comprises the target tissue volume for treatment. Transducers with a diameter of approximately 5 cm and a focal length of approximately 10 cm are readily available. Favorable focal dimensions are achieved when the ultrasound frequency is between 1 and 5 MHz and the total duration of the train is 0.1 to 0.5 seconds. For example, for a focal length of 10 cm and the transducer diameter of 5 cm, the focal volumes are 0.02 cc at 5 MHz and 2.36 cc at 1 MHz.

[00057] With reference now to FIG. 11, a graph illustrates the average power of the source in watts compared to the frequency (between 1 MHz and 5 MHz) and the duration of the pulse frequency (between 0.1 and 0.5 seconds). A transducer focal length of 10 cm and a source diameter of 5 cm were adopted. The power required to provide the Arrhenius integral for HSP activation of approximately 1 decreases monotonically as the frequency increases and the total duration of the train increases. Given the preferred parameters, the minimum power for a frequency of 1 GHz and the duration of 0.5 second is 5.72 watts, while for the frequency of 1 GHz and pulse train duration of 0.1 second, the power maximum is 28.6 watts. For a frequency of 5 GHz, 0.046 watts are needed for a pulse train duration of 0.5 seconds, and 0.23 watts are required for a pulse train duration of 0.1 second. The corresponding peak power during an individual pulse is obtained simply by dividing by the duty cycle.

Petition 870190083984, of 08/28/2019, p. 45/162

32/98

[00058] FIGURE 12 illustrates the time, in seconds, for the temperature to diffuse or fall from 10 ° C to 6 ° C when the ultrasound frequency is between 1 and 5 MHz. FIG. 13 illustrates the time, in seconds, to drop from approximately 10 ° C to approximately 1 ° C for ultrasound frequencies from 1 to 5 MHz. For the preferred focal length of 10 cm and the diameter of the transducer of 5 cm, the maximum time drop in temperature is 366 seconds when the ultrasound frequency is 1 MHz and the minimum temperature is 15 seconds when the microwave frequency is 5 MHz. As the FDA only requires that the temperature rise be lower than 6 ° C for test times of minutes, the drop time from 366 seconds to 1 MHz to reach a 1 ° C increase in the various minutes is allowed. As can be seen in FIGS. 12 and 13, the drop times for an increase of 6 ° C are much shorter, by a factor of approximately 70, than that of 1 ° C.

[00059] FIGURE 14 illustrates the volume of the focal heated region, in cubic centimeters, compared to ultrasound frequencies between 1 and 5 MHz. Considering ultrasound frequencies in the range of 1 to 5 MHz, the corresponding focal sizes for these frequencies vary from 3.7 mm to 0.6 mm and the length of the focal region varies from 5.6 cm to 1.2 cm. The corresponding treatment volumes vary between approximately 2.4 cc and 0.02 cc.

[00060] Examples of parameters that give a desired Arrhenius HSP activation greater than 1 and damage to the Arrhenius integral less than 1 are a total ultrasound power between 5.8 and 17 watts, pulse duration 0.5 seconds, 5 second pulse interval, with total number of pulses 10 within the total pulse current time of 50 seconds. The volume of

Petition 870190083984, of 08/28/2019, p. 46/162

33/98 target treatment would be approximately 1 mm on the side. Larger treatment volumes could be treated by an ultrasound system similar to a laser diffracted optical system, applying ultrasound to multiple adjacent columns, but separated and spaced simultaneously. The multiple focused ultrasound beams converge into a very small treatment target within the body, the convergence allowing for minimal warming, except for the overlapping beams on the target. This area would be heated and stimulate the activation of HSPs and facilitate the repair of proteins through transient high temperature peaks. However, given the pulsating aspect of the invention, as well as the relatively small area to be treated at any time, the treatment complies with the FDA / FCC requirements for long-term average temperature rise (minutes) <1K. An important distinction of the invention of existing therapeutic warming treatments for pain and muscle tension is that there are no high T spikes in existing techniques and these are necessary to effectively activate HSPs and facilitate the repair of proteins to provide healing at the cellular level.

[00061] The pulse rate mode of the power supply has a distinct advantage over a single pulse or gradual mode of power supply, with regard to the activation of the repairing HSPs and the facilitation of protein repair. There are two considerations that go into this advantage: [00062] First, a major advantage for activating HSP and repairing the protein in an SDM power supply mode comes from producing a peak temperature in the order of 10 ° C. This large increase in temperature has a major impact on

Petition 870190083984, of 08/28/2019, p. 47/162

34/98 Arrhenius integrals that quantitatively describe the number of HSPs that are activated and the rate of water diffusion in proteins, which facilitates protein repair. This is because the temperature goes into an exponential that has a great amplifying effect.

[00063] It is important that the temperature rise does not remain at the high value (10 ° C or more) for a long time, because then it would violate the FDA and FCC requirements that, in periods of minutes, the average temperature rise should be lower to ° C (or, in the case of ultrasound, 6 ° C).

[00064] An SDM mode of power supply satisfies these two considerations exclusively by carefully choosing the power, pulse time, pulse interval and the volume of the target region to be treated. The volume of the treatment region enters because the temperature must drop from its high value in the order of 10 ° C very quickly so that the long-term increase in average temperature does not exceed the FDA / FCC long-term limit of 6 ° C to ultrasound frequencies and 1 ° C or less for energy sources of electromagnetic radiation.

[00065] For a region of linear dimension L, the time it takes for the peak e-fold temperature in the fabric is approximately L ² / 16D, where D = 0, 00143 cm ² / s is the heat diffusion coefficient typical. For example, if L = 1 mm, the drop time will be approximately 0.4 seconds. Thus, for a region of 1 mm on a side, a frequency consisting of 10 pulses lasting 0.5 seconds, with an interval between pulses of 5 seconds, can achieve the desired momentary temperature rise without exceeding an increase

Petition 870190083984, of 08/28/2019, p. 48/162

35/98 long-term average temperature of 1 ° C. This is further demonstrated below.

[00066] The limitation of the heated volume is the reason why RF electromagnetic radiation is not such a good option for the SDM treatment of deep regions with the body as the ultrasound. The long depths of the skin (penetration distances) and ohmic heating along the depth of the skin result in a large heated volume whose thermal inertia does not allow obtaining a high peak temperature that activates the HSPs and facilitates the repair of proteins and the rapid temperature drop satisfies the FDA and FCC long-term limit on average temperature rise.

[00067] Ultrasound has already been used to heat the body regions therapeutically to relieve pain and muscle tension. However, the heating did not follow the SDM type protocol and does not have the temperature peaks that are responsible for the excitation of the HSPs.

[00068] Then consider a group of focused ultrasound beams that are directed to a target region within the body. To simplify the math, suppose that the beams are replaced by a single source with a spherical surface shape focused on the center of the sphere. The absorption lengths of the ultrasound can be reasonably long. Table 3 below shows the typical absorption coefficients for 1 MHz ultrasound. The absorption coefficients are approximately proportional to the frequency.

Petition 870190083984, of 08/28/2019, p. 49/162

36/98

Table 3. Typical absorption coefficients for ultrasound of

MHz in body tissue:

Body tissue Attenuation coefficient at 1 MHz (cm ^-1 )

Water 0.00046 Blood 0.0415 Fat 0. 145 Liver 0.115 to 0.217 Kidney 0.23 Muscle 0.3 to 0.76 Bone 1, 15

[00069] Assuming that the geometric variation of radiation received due to the focus dominates any variation due to attenuation, the intensity of the incoming ultrasound at a distance r from the focus can be written approximately as:

I (r) = P / (4nr ² ) [1] where P denotes the total power of ultrasound.

increase in temperature at the end of a short pulse of duration t _p in r is then dT (tp) = Patp / (4nC _v r ² ) [2] where α is the absorption coefficient and C _v is the specific volume of capacity thermal. This will be the case until _r is reached when the heat diffusion length in t _p becomes comparable to air or the diffraction limit of the focused beam is reached. For smaller r, the temperature rise is essentially independent of r. As an example, suppose that the diffraction limit is reached at a radial distance less than that determined by the heat diffusion. So r _dlf = (4Dt _p ) ^1/2 [3] where D is the heat diffusion coefficient and, for r <r _di f, the temperature increase in t _p is dT (r _dif , t _p ) = 3Pa / (8nC _v D) when r <r _dif [4]

Petition 870190083984, of 08/28/2019, p. 50/162

37/98

So, at the end of the pulse, we can write for the temperature increase:

dTp (r) = {Patp / (4nC _v } [(6 / r _dif ² ) U {rdif-r) + (1 / r ² ) U (r-rdif)] [5]

When applying the Green function to the heat diffusion equation,

G (r, t) = (4QDt) ~ ^3/2 exp [-r ² / (4Dt)] [6] to this initial temperature distribution, we found that the temperature dT (t) at the focal point r = 0 at the moment t is dT (t) = [dT _o / {(1/2) + (n ^1/2 / 6)}] [(1/2) (tp / t) ^3/2 + (n ^1/2 / 6 ) (tp / t)] [7] with dT _o = 3Pa / (8nC _v D) [8]

[00070] A good approximation for the eq. [7] is provided by: dT (t) «dT _o (tp / t) ^3/2 [9] as seen in FIG. 15, which is a comparison of the equations. [7] and [9] for dT (t) / dT _o in the target treatment zone. The lower curve is the approximate expression of eq [9].

The Arrhenius integral for an N-pulse train can now be evaluated with the temperature increase given by eq. [9]. In this expression, dT _N (t) = Σ dT (t-nt _T ) [11] where dT (t-nti) is the expression of eq. [9] with t replaced by t-nti 'and with t _T designating the interval between pulses.

[00071] The Arrhenius integral can be evaluated approximately by dividing the integration interval in the portion where the temperature peaks occur and the portion where the temperature peak is absent. The sum on the peak temperature contribution can be simplified by applying Laplace's end point formula to the integral on the peak of

Petition 870190083984, of 08/28/2019, p. 51/162

38/98 temperature. In addition, the integral over the portion when the peaks are absent can be simplified by noting that the increase in temperature without a peak reaches an asymptotic value very quickly, so that a good approximation is obtained by replacing the variable increase in time with its asymptotic value. When these approximations are made, eq. [10] becomes:

Ω = AN [{t _p (2k _B T _o ² / (3EdT _o )} exp [- (E / k _B ) 1 / (T _o + dT _o + dT _N (Nt _B ))] + exp [- ( E / k _B ) 1 / (T _o + dT _N (Nt _B ))]] [12] where dT _N (Nt _B ) «2.5 dT _o (t _p / 1 _τ ) ^3/2 [13] _ o / o (O 2,5 in eq. [13] arises from the sum over n of (Nn) and is the magnitude of the harmonic number (N, 3/2) for typical N of interest).

[00072] It is interesting to compare this expression to that used for SDM applied to the retina. The first term is very similar to the peak contribution in the case of the retina, except that the effective peak interval is reduced by a factor of 3 for this case of 3D converging radius. The second term, involving dT _N (Nti) is much smaller than in the case of the retina. There, the increase in the background temperature was comparable in magnitude to the increase in the peak temperature. But here, in the case of the convergent beam, the increase in the background temperature is much smaller for the ratio (tp / ti) ^3/2 . This points to the importance of the peak contribution to the activation or production of HSPs and to facilitate the repair of proteins, since the increase in the background temperature, which is similar to the increase in a continuous case of ultrasound heating, is insignificant compared to peak contribution. At the end of the pulse rate, even

Petition 870190083984, of 08/28/2019, p. 52/162

39/98 this increase in the low background temperature disappears quickly by the diffusion of heat.

[00073] FIGURES 16 and 17 show the magnitude of the Arrhenius integral logarithm for damage and for HSP activation or production as a function of dT _o for a pulse duration t _p = 0.5 s, pulse interval t _T = 10 if total number of pulses N = 10. Logarithm of Arrhenius integrals [eq. 12] for damage and for activation of HSP as a function of the temperature increase in degrees Kelvin of a single pulse dT _o , for a pulse duration t _p = 0.5 s, pulse interval t _T = 10 if a total number of ultrasound pulses N = 10. FIG. 16 shows the logarithm of the damage integral with the Arrhenius constants A = 8.71xl0 ³³ s ^_1 and E = 3.55x10 ~ ¹² ergs. FIG. 17 shows the logarithm of the HSP activation integral with Arrhenius constants A = 1.24x10 s and E = 2.66x10 ergs. The graphs in FIGS. 16 and 17 show that _damage does not exceed Q 1 to Td exceeds _the 11.3 K, while _hsp Q is greater than 1 throughout the range shown, the desired condition for cellular repair without damage.

[00074] Equation [8] shows that when α = 0.1 cnf ¹ , a dT _o of 11.5 K can be obtained with a total ultrasound power of 5.8 watts. This is easily achievable. If α is increased by a factor of 2 or 3, the resulting power is still easily attainable. The volume of the region where the temperature increase is constant (that is, the corresponding volume ar = r _d = (4Dt _p )) and 0, 00064 cc. This corresponds to a cube that is 0.86 mm on one side.

[00075] This simple example demonstrates that focused ultrasound should be used to stimulate HPSs deep in the body with easily accessible equipment:

Petition 870190083984, of 08/28/2019, p. 53/162

40/98

Total ultrasound power: 5.8 watts to 17 watts

Pulse time 0.5s

Pulse interval5 s

Total train duration (N = 10) 50 s

To expedite the handling of larger internal volumes, a SAPRA system can be used.

[00076] The pulsed energy source can be directed to an exterior of a body that is adjacent to the target tissue or has a blood supply close to the exterior surface of the body. Alternatively, a device can be inserted into a body cavity to apply the pulsed energy source to the target tissue. Whether the energy source is applied outside the body or inside the body and what type of device is used depends on the energy source selected and used to treat the target tissue.

[00077] Photostimulation according to the present invention can be effectively transmitted to an internal surface area or tissue of the body using an endoscope, such as a bronchoscope, proctoscope, colonoscope or the like. Each of these essentially consists of a flexible tube that itself contains one or more inner tubes. Typically, one of the inner tubes comprises a light tube or multimode optical fiber that conducts the light down the scope to illuminate the region of interest and allow the doctor to see what is on the illuminated side. Another inner tube could consist of wires that carry an electric current to allow the doctor to cauterize the illuminated tissue. Yet another inner tube may consist of a biopsy tool that would allow the doctor to cut and hold any illuminated tissue.

Petition 870190083984, of 08/28/2019, p. 54/162

41/98

[00078] In the present invention, one of these internal tubes is used as an electromagnetic radiation tube, such as a multimode optical fiber, to transmit the SDM or other pulses of electromagnetic radiation that are fed into the scope at the end that the doctor holds. Referring now to FIG. 18, a light generating unit 10, such as a laser having a desired wavelength and / or frequency used to generate electromagnetic radiation, such as laser light, in a controlled, pulsed manner to be delivered through a tube or tube. light 12 to a distal end of the nozzle 14, shown in FIG. 19, which is inserted into the body and laser light or other radiation 16 applied to the target tissue 18 to be treated.

[00079] With reference now to FIG. 20, a schematic diagram of a system for generating radiation from electromagnetic energy, such as laser light, including SDM, is shown. The system, generally referred to by reference number 20, includes a laser console 22, such as, for example, the micropulsed infrared diode laser near 810 nm in the preferred embodiment. The laser generates a beam of laser light that is passed through the optics, such as a lens or optical mask or a plurality of lenses and / or optical masks 24 as needed. The optics of the laser projector 24 passes the patterned light beam to a delivery device 26, such as an endoscope, to project the light from the laser beam onto the patient's target tissue. It will be understood that the box labeled 26 can represent both the laser beam projector and the distribution device, as well as a visualization / camera system, such as an endoscope or comprises two different components in use. The visualization / camera system 26 provides information for a

Petition 870190083984, of 08/28/2019, p. 55/162

42/98 display monitor 28, which can also include computerized hardware, data entry and control, etc. necessary to manipulate the laser 22, the optics 24 and / or the projection / visualization components 26.

[00080] With reference now to FIG. 21, in one embodiment, a plurality of light beams are generated, each of which has selected parameters so that a target tissue temperature can be controlled so as to treat the target tissue therapeutically without destroying or permanently damaging the target tissue. This can be done, for example, by passing the laser beam 30 through diffracting optics or otherwise generating a plurality of laser beam from the single laser beam 30 having the parameters selected. For example, the laser beam 30 can be passed through a collimating lens 32 and then through a mask 34. In a particularly preferred embodiment, mask 34 comprises a diffraction grating. The mask / diffraction grid 34 produces a geometric object or more typically a geometric pattern of multiple laser points produced simultaneously or other geometric objects. This is represented by the multiple laser light beams marked with reference number 36. Alternatively, the multiple laser points can be generated by a plurality of fiber optic waveguides. Any method of generating laser points allows the creation of a very large number of laser points simultaneously in a very wide treatment field. In fact, a very high number of laser points, perhaps numbering in the hundreds, even thousands or more, could be generated simultaneously to cover a certain area of the laser.

Petition 870190083984, of 08/28/2019, p. 56/162

43/98 target tissue, or possibly even the entire target tissue. A wide range of applications of small separate laser points applied simultaneously may be desirable, thus avoiding certain disadvantages and treatment risks that are known to be associated with large laser point applications.

[00081] Using optical resources with a resource size equivalent to the wavelength of the laser employed, for example, using a diffraction grid, it is possible to take advantage of the quantum mechanical effects that allow the simultaneous application of a very large number of laser points over a very large target area. The individual points produced by these diffraction grids are all of optical geometry similar to the input beam, with minimum power variation for each point. The result is a plurality of laser points with adequate irradiance to produce a harmless yet effective treatment application, simultaneously on a large target area. The present invention also contemplates the use of other objects and geometric patterns generated by other diffractive optical elements.

[00082] The laser light that passes through the diffracted mask 34, producing a periodic pattern at a distance from the mask 34, shown by the laser rays marked 36 in FIG. 21. The single laser beam 30 has thus been formed into hundreds or even thousands of individual laser rays 36 in order to create the desired pattern of dots or other geometric objects. These laser beams 36 can be passed through additional lenses, collimators, etc. 38 and 40 in order to transport the laser beams and form the desired pattern. These additional lenses, collimators, etc. 38 and 40 can further transform and redirect laser beams 36 as needed.

Petition 870190083984, of 08/28/2019, p. 57/162

44/98

[00083] Arbitrary patterns can be constructed by controlling the shape, spacing and pattern of the optical mask 34. The pattern and exposure points can be created and modified arbitrarily as desired, according to the application requirements by experts in the field of optical engineering. Photolithographic techniques, especially those developed in the field of semiconductor manufacturing, can be used to create the simultaneous geometric pattern of points or other objects.

[00084] The present invention can use a plurality of beams or points of therapeutic light generated simultaneously, such as numbering in the tens or even hundreds, as the parameters and methodology of the present invention create therapeutically effective treatment, but not destructive and not permanently harmful. Although hundreds or even thousands of simultaneous laser points can be generated and created and formed in patterns to be applied simultaneously to the fabric, due to the requirements of not overheating the fabric, there are restrictions on the number of treatment points or beams that can be simultaneously used in accordance with the present invention. Each individual beam or laser point requires minimum average power for the duration of the frequency to be effective. However, at the same time, the fabric cannot exceed certain temperature increases without being damaged. For example, using an 810 nm wavelength laser, the number of simultaneous points generated and used can vary from 1 and up to 100 when a duty cycle of 0.04 (4%) and a total frequency duration of 0 , 3 seconds (300 milliseconds) is used. Water absorption increases as the wavelength

Petition 870190083984, of 08/28/2019, p. 58/162

45/98 increases. For shorter wavelengths, for example, 577 nm, the laser power may be less. For example, at 577 nm the power can be decreased by a factor of 4 for the invention to be effective. Consequently, there can be only one laser point or up to approximately 400 laser points when using 577 nm laser light, without harming or damaging the tissue.

[00085] Typically, the system of the present invention incorporates a guidance system to ensure complete and total treatment of the retina with photostimulation of the retina. Fixing / tracking / recording systems consisting of a fixing target, tracking mechanism and linked to the operation of the system can be incorporated into the present invention. In a particularly preferred embodiment, the geometric pattern of the simultaneous laser points is shifted sequentially in order to achieve a complete and confluent surface treatment.

[00086] This can be done in a controlled manner using an optical scanner 50. FIGS. 22 and 23 illustrate an optical scanning mechanism 50 in the form of a MEMS mirror, with a base 52 with electronically operated controllers 54 and 56 which serve to tilt and move the mirror 58 as electricity is applied and removed from it. The application of electricity to the controller 54 and 56 causes the mirror 58 to move and thus the simultaneous pattern of laser points or other geometric objects reflected in it moves accordingly on the patient's retina. This can be done, for example, in an automated manner, using an electronic software program to adjust the optical scanner 50 until complete retinal coverage, or

Petition 870190083984, of 08/28/2019, p. 59/162

46/98 at least the desired portion of the retina to be treated, be exposed to phototherapy. The optical scanning mechanism can also be a small beam diameter scanning galvo mirror system, or similar system, such as that distributed by Thorlabs. Such a system is capable of scanning the lasers in the desired displacement pattern.

[00087] The stitch pattern is compensated for each exposure, in order to create space between the immediately previous exposure to allow heat dissipation and avoid the possibility of heat damage or tissue destruction. Thus, as illustrated in FIG. 24, the pattern, illustrated for exemplary purposes as a grid of sixteen points, is compensated for each exposure, in such a way that the laser points occupy a different space from the previous exhibitions. It will be understood that the diagrammatic use of circles or empty dots, as well as filled dots, serves only to illustrate previous and subsequent exposures of the dot pattern for the area, in accordance with the present invention. Spacing the laser points prevents overheating and tissue damage. It will be understood that this occurs until all the target tissue to be treated has received phototherapy or until the desired effect is achieved. This can be done, for example, by applying electrostatic torque to a micro-machined mirror, as illustrated in FIGS. 22 and 23. Combining the use of small laser points separated by free areas of exposure, avoiding the accumulation of heat and grids with a large number of points per side, it is possible to treat large target areas non-traumatically and invisibly with durations of short exposures much faster than is possible with current technologies.

Petition 870190083984, of 08/28/2019, p. 60/162

47/98

[00088] Repeating quickly and sequentially the redirection or displacement of any grid arrangement or geometric objects applied simultaneously, it is possible to obtain a complete coverage of the fabric quickly and without thermal tissue damage. This displacement can be determined by algorithms to guarantee the fastest treatment time and the lowest risk of damage due to thermal tissue, depending on the laser parameters and the desired application.

[00089] The next one was modeled using the Fraunhoffer approximation. With a mask with a nine by nine square matrix, with an aperture radius of 9 pm, an aperture spacing of 600 pm, using an 890 nm wavelength laser, with a 75 mm mask-lens separation and secondary mask of 2.5 mm by 2.5 mm, the following parameters will produce a grid with nineteen points per side separated by 133 pm with a point size radius of 6 pm. The number of exposures m needed to treat (cover confluently with applications of small dots) given the lateral length of the desired area A, given output pattern points per square side n, separation between points R, local radius r and the length of the desired square side to treat area A can be given by the following formula:

A / R \ ² m = —floor - nR \ 2r /

[00090] With the previous configuration, it is possible to calculate the number of operations required to treat different areas of the exposure field. For example, an area of 3 mm x 3 mm, which is useful for treatments, would require 98 displacement operations, requiring a treatment time of approximately thirty seconds. Another example would be an area

Petition 870190083984, of 08/28/2019, p. 61/162

48/98 of 3 cm x 3 cm, representing the entire surface of the human retina. For such a large treatment area, a much larger secondary mask size of 25 mm by 25 mm could be used, producing a treatment grid of 190 points per side separated by 133 pm with a spot size radius of 6 pm. Since the size of the secondary mask has been increased by the same factor as the desired treatment area, the number of displacement operations is approximately 98 and, therefore, the treatment time of approximately thirty seconds is constant.

[00091] Of course, the number and size of stitches produced in an arrangement of simultaneous patterns can be easy and highly varied, so that the number of sequential displacement operations required to complete treatment can be easily adjusted, depending on therapeutic requirements. the given application.

[00092] In addition, due to the small openings used in the grating or diffraction mask, a quantum mechanical behavior can be observed that allows the arbitrary distribution of the laser input energy. This would allow the generation of any arbitrary geometric shapes or patterns, such as a plurality of points in the grid pattern, lines or any other desired pattern. Other methods that generate geometric shapes or patterns, such as the use of multiple fibers or fiber optic microlenses, can also be used in the present invention. The time-saving use of simultaneous projection of geometric shapes or patterns allows treatment fields of new size, such as the 1.2 cm ^A 2 area to perform the treatment of the complete retina, in a single clinical setting or treatment session. .

Petition 870190083984, of 08/28/2019, p. 62/162

49/98

[00093] With reference now to FIG. 25, instead of a geometric pattern of small laser dots, the present invention contemplates the use of other objects or geometric patterns. For example, a single line 60 of laser light can be created, formed continuously or by means of a series of closely spaced points. An optical shift scanner can be used to sequentially scan the line over an area, illustrated by the down arrow in FIG. 25.

[00094] With reference now to FIG. 26, the same geometric object in a line 60 can be rotated, as illustrated by the arrows, in order to create a circular phototherapy field. The negative potential of this approach, however, is that the central area will be repeatedly exposed and could reach unacceptable temperatures. This can be overcome, however, by increasing the time between exposures or creating a gap in the line so that the central area is not exposed.

[00095] The field of photobiology reveals that different biological effects can be achieved by exposing target tissues to lasers of different wavelengths. The same can also be achieved by the consecutive application of multiple lasers of different or equal wavelength in sequence with varying periods of time of separation and / or with different radiating energies. The present invention provides for the use of multiple wavelengths of laser, light or radiant (or modes) applied simultaneously or in sequence to maximize or personalize the desired treatment effects. This method also minimizes the potential harmful effects. The optical methods and systems illustrated and

Petition 870190083984, of 08/28/2019, p. 63/162

50/98 described above provide the simultaneous or sequential application of multiple wavelengths.

[00096] FIGURE 27 schematically illustrates a system that couples multiple sources of treatment light to the optical subset that generates the patterns described previously. Specifically, this system 20 'is similar to the system 20 described in FIG. Previous 20. The main differences between the alternative system 20 'and the system previously described 20 are the inclusion of a plurality of laser consoles, the outputs of which are each fed to a fiber coupler 42. Each laser console can provide a beam of laser light with different parameters, like a different wavelength. The fiber coupler produces a single output that is passed through the optics of the laser projector 24 as described in the previous system. The coupling of the plurality of laser consoles 22 on a single optical fiber is achieved with a fiber coupler 42 as is known in the art. Other known mechanisms for combining multiple light sources are available and can be used to replace the fiber coupler described here.

[00097] In this system 20 ', the multiple light sources 22 follow a path similar to that described in the previous system 20, that is, collimated, diffracted, recollimated and directed towards the projector and / or fabric device. In this alternative system 20 ', the diffractive element must function in a different way than was previously described, depending on the wavelength of the light that passes through it, which results in a slightly variable pattern. The variation is linear with the wavelength of the light source being diffracted. Overall, the difference in the angles of

Petition 870190083984, of 08/28/2019, p. 64/162

51/98 diffraction is small enough that the different patterns of overlap can be directed along the same optical path through the projector device 26 to the fabric for treatment.

[00098] Since the resulting pattern varies slightly for each wavelength, a sequential shift to obtain complete coverage will be different for each wavelength. This sequential shift can be performed in two modes. In the first mode, all wavelengths of light are applied simultaneously without identical coverage. A travel direction pattern to obtain complete coverage for one of multiple wavelengths is used. Thus, while the light of the selected wavelength reaches a complete coverage of the tissue area, the application of the other wavelengths reaches an incomplete or overlapping coverage of the tissue. The second mode sequentially applies each light source of a variable wavelength with the appropriate direction pattern to achieve complete tissue coverage for that specific wavelength. This mode excludes the possibility of simultaneous treatment using multiple wavelengths, but allows the optical method to achieve identical coverage for each wavelength. This avoids incomplete or overlapping coverage for any of the optical wavelengths.

[00099] These modes can also be mixed and matched. For example, two wavelengths can be applied simultaneously with one wavelength reaching full coverage and the other reaching incomplete or overlapping coverage, followed by a third length

Petition 870190083984, of 08/28/2019, p. 65/162

52/98 wave applied sequentially and obtaining complete coverage.

[000100] FIGURE 28 diagrammatically illustrates yet another alternative embodiment of the inventive system 20 ''. This system 20 '' is generally configured the same as the system 20 shown in FIG. 20. The main difference lies in the inclusion of multiple channels of subsets that generate patterns tuned to a specific wavelength of the light source. Several laser consoles 22 are arranged in parallel, each taking directly its own optics from the laser projector 24. The optics of the laser projector of each channel 44a, 44b, 44c comprise a collimator 32, mask or diffraction net 34 and recolimizers 38, 40, as described in conjunction with FIG. Previous 21 - the entire set of lenses tuned to the specific wavelength generated by the corresponding laser console 22. The output of each set of optics 24 is then directed to a beam splitter 46 for combination with the other wavelengths. Those skilled in the art know that a beam splitter used in reverse can be used to combine multiple beams of light into a single outlet. The combined channel output of the final beam splitter 46c is then directed through the projector device 26.

[000101] In this 20 '' system, the optical elements for each channel are tuned to produce the exact pattern specified for the channel's wavelength. Consequently, when all channels are combined and properly aligned, a single direction pattern can be used to achieve complete fabric coverage for all wavelengths. The 20 '' system can use so many channels 44a, 44b, 44c, etc. and beam splitters 46a,

Petition 870190083984, of 08/28/2019, p. 66/162

53/98

6b, 4 6c, etc. as there are wavelengths of light being used in the treatment.

[000102] The implementation of the 20 '' system can take advantage of different symmetries to reduce the number of alignment restrictions. For example, the proposed grid patterns are periodic in two dimensions and targeted in two dimensions to obtain complete coverage. As a result, if the patterns of each channel are identical, as specified, the actual pattern of each channel would not need to be aligned to the same direction pattern to obtain complete coverage for all wavelengths. Each channel would only need to be aligned optically to achieve an efficient match.

[000103] In the 20 '' system, each channel starts with a light source 22, which can be of an optical fiber as in other modalities of the subset that generates patterns. This light source 22 is directed to the optical assembly 24 for collimation, diffraction, recolimation and directed to the beam divider that combines the channel with the main outlet.

[000104] It will be understood that the laser light generation systems illustrated in FIGS. 20 to 28 are exemplary. Other devices and systems can be used to generate an SDM laser light source that can be operably passed through a projection device, typically in the form of an endoscope that has a light tube or the like. Also, other forms of electromagnetic radiation can also be generated and used, including ultraviolet waves, microwaves, other radio frequency waves and laser light at predetermined wavelengths. In addition, ultrasound waves can also be generated and used to create a peak thermal temperature in the target tissue sufficient to activate or

Petition 870190083984, of 08/28/2019, p. 67/162

54/98 produce heat shock proteins in the cells of the target tissue without damaging the target tissue itself. To do this, typically, a pulsed source of ultrasonic or electromagnetic radiation energy is supplied and applied to the target tissue in a way that raises the temperature of the target tissue, such as between 6 ° C and 11 ° C, transiently, while only 6 ° C or 1 ° C or less in the long run, such as for several minutes.

[000105] It is believed that the stimulation of HSP production according to the present invention can be effectively used in the treatment of a wide range of tissue abnormalities, diseases and even infections. For example, viruses that cause colds mainly affect a small port of the respiratory epithelium in the nasal passages and nasopharynx. Similar to the retina, the respiratory epithelium is a thin, clear tissue. With reference to FIG. 29, a cross-sectional view of a human head 62 is shown with an endoscope 14 inserted in the nasal cavity 64 and the energy 16, such as laser light or the like, being directed to the tissue 18 to be treated within the nasal cavity 64. The tissue 18 to be treated can be inside the nasal cavity 64, including the nasal passages and nasopharynx.

[000106] To ensure the absorption of energy from the laser, or other energy source, the wavelength can be adjusted to an infrared (IR) absorption peak of the water or an adjuvant dye can be used to serve as a photosensitizer. In a case like this, the treatment would consist of drinking, or topically applying, the adjuvant, waiting a few minutes for the adjuvant to permeate the surface tissue and then administering laser light or another source of

Petition 870190083984, of 08/28/2019, p. 68/162

55/98 energy 16 to the target tissue 18 for a few seconds, such as through optical fibers in an endoscope 14, as illustrated in FIG. 29. To provide comfort to the patient, endoscope 14 can be inserted after applying a topical anesthetic. If necessary, the procedure can be repeated periodically, as in a day or more.

[000107] The treatment would stimulate the activation or production of heat shock proteins and facilitate the protein repair without damaging the cells and tissues to be treated. As discussed earlier, it was discovered that certain heat shock proteins play an important role in the immune response, as well as in the well-being of target cells and tissues. The energy source can be monochromatic laser light, such as 810 nm wavelength laser light, administered in a similar manner to that described in the previously referenced patent applications, but administered through an endoscope or the like, as illustrated in FIG. 29. The adjuvant dye would be selected in order to increase the absorption of the laser light. While this comprises a particularly preferred method and embodiment for carrying out the invention, it will be appreciated that other types of energy and distribution means can be used to achieve the same goals in accordance with the present invention.

[000108] With reference now to FIG. 30, there is a similar situation for influenza viruses, in which the primary target is the epithelium of the upper respiratory tree, in segments with diameters greater than 3.3 mm, that is, the six upper generations of the upper respiratory tree. A thin layer of mucus separates the target epithelial cells from the lumen of the pathways

Petition 870190083984, of 08/28/2019, p. 69/162

56/98 and it is in this layer that antigen-antibody interactions occur that result in virus inactivation.

[000109] With continuous reference to FIG. 30, the flexible light tube 12 of a bronchoscope 14 is inserted through the individual's mouth 66 through the throat and trachea 68 and into a bronchus 70 of the respiratory tree. All laser light or other energy source 16 is administered and delivered to the tissue in this area of the uppermost segments to treat the tissue and the area in the same manner as described above in relation to FIG. 29. It is envisaged that a laser wavelength or other energy would be selected to correspond to a peak of IR absorption from the water residing in the mucus to heat the tissue and stimulate the activation or production of HSP and facilitate protein repair , with its inherent benefits.

[000110] With reference now to FIG. 31, a colonoscope 14 may have a flexible optical tube 12 of the same inserted in the anus and rectum 72 and in the large intestine 74 or in the small intestine 76 in order to administer the selected laser light or other energy source 16 in the area and tissue to be treated , as illustrated. This could be used to help treat colon cancer, as well as other gastrointestinal problems.

[000111] Typically, the procedure could be performed similar to a colonoscopy in which all feces would be removed from the intestine and the patient would lie on his side and the doctor would insert the long, thin portion of the light tube 12 of the colonoscope 14 into the rectum and move it to the colon area, large intestine 74 or small intestine 76 to the area to be treated. The doctor can see through the monitor the path of the inserted flexible member 12 and even see the tissue at the tip of the colonoscope 14 inside the intestine, in order to see the area to be

Petition 870190083984, of 08/28/2019, p. 70/162

57/98 be treated. Using one of the other fiber optic or light tubes, the tip 78 of the telescope would be directed towards the tissue to be treated and the laser light source or other radiation 16 would be delivered through one of the colonoscope light tubes 14 to treat the area of the tissue to be treated, as previously described, in order to stimulate the activation or production of HSP in this tissue 18.

[000112] With reference now to FIG. 32, another example in which the present invention can be used advantageously in the tract is that it is often referred to as intestinal drip syndrome, a condition of the gastrointestinal tract (GI) marked by inflammation and another metabolic dysfunction. Since the GI tract is susceptible to metabolic dysfunction similar to that of the retina, it is expected that it will respond well to the treatment of the present invention. This can be done by means of treatment with diode subliminal micropulsed laser (SDM), as discussed above, or by other sources of energy and means as discussed herein and known in the art.

[000113] With continuous reference to FIG. 32, the flexible light tube 12 of an endoscope or the like inserted through the patient's mouth 66 through the throat and trachea area 68 and into the stomach 80, where the tip or end 7 8 of it is directed towards the tissue 18 to be treated and the laser light or other energy source 16 is directed to the tissue 18. Experts in the technique will realize that a colonoscope could also be used and inserted through the rectum 72 and into the stomach 80 or any tissue between the stomach and the rectum .

[000114] If necessary, a chromophore pigment can be administered to the gastrointestinal tissue orally to allow radiation absorption. If, for example, 810 nm radiation

Petition 870190083984, of 08/28/2019, p. 71/162

58/98 without focusing a laser diode or LED was used, the pigment would have an absorption peak at or near 810 nm. Alternatively, the wavelength of the energy source could be adjusted to a slightly longer wavelength at a peak of water absorption, so that no externally applied chromophore would be needed.

[000115] It is also contemplated by the present invention that a capsule endoscope 82, such as that illustrated in FIG. 33, can be used to administer the radiation and the energy source according to the present invention. Such capsules are relatively small in size, approximately one inch long, in order to be swallowed by the patient. When the capsule or pill 82 is swallowed and enters the stomach and passes through the gastrointestinal tract, when in the appropriate place, the capsule or pill 82 can receive energy and signals, such as through antenna 84, in order to activate the source of energy 86, such as a laser diode and related circuits, with an appropriate lens 88 focusing the laser light or radiation generated through a transparent cover for radiation 90 and on the tissue to be treated. It will be understood that the location of the endoscope capsule 82 can be determined by a variety of means, such as

Image external, tracking of signal or even through in an chamber miniature with lights through gives which doctor see at images of the tract GI through of which The pill or capsule 82

it was happening at that moment. The capsule or pill 82 can be supplied with its own energy source, such as by means of a battery or it can be fed externally through an antenna, in such a way that the laser diode 8 6 or another source that generates energy creates the wavelength

Petition 870190083984, of 08/28/2019, p. 72/162

59/98 desired and pulsed energy source to treat the tissue and the area to be treated.

[000116] As in the treatment of the retina in previous applications, the radiation would be pulsed to take advantage of the micropulse temperature peaks and the associated safety and the power could be adjusted so that the treatment was completely harmless to the tissue. This could involve adjusting the peak power, pulse times and repetition rate to raise the temperature by 10 ° C, keeping the long-term temperature rise below the FDA's 1 ° C limit. If pill form 82 is used, the device could be powered by a small rechargeable battery or by wireless inductive excitation or the like. The heated / stressed tissue would stimulate the activation or production of HSP and facilitate the protein repair and the benefits resulting from it.

[000117] From the previous examples, the technique of the present invention is limited to the treatment of conditions on surfaces close to the body or on internal surfaces easily accessible by means of optical fibers or other optical distribution means. The reason that the application of SDM to activate HSP activity is limited to regions close to the surface or optically accessible from the body is that the length of absorption of IR or visible radiation in the body is very short. However, there are deeper conditions in the tissue or body that can benefit from the present invention. Thus, the present invention contemplates the use of ultrasound and / or radio frequency (RF) as well as electromagnetic radiation (EM) of shorter wavelengths, such as microwaves, which have wavelengths of

Petition 870190083984, of 08/28/2019, p. 73/162

60/98 relatively long absorption into body tissues. The use of pulsed ultrasound is preferable to electromagnetic RF radiation to activate the corrective activity of HSP in abnormal tissue that is inaccessible to surface SDM or similar.

[000118] For deep tissues that are not close to an internal orifice, a light tube may not be an effective means of delivering pulsed energy. In that case, pulsed low frequency electromagnetic energy or, preferably, pulsed ultrasound can be used to cause a series of temperature spikes in the target tissue.

[000119] Thus, according to the present invention, a source of pulsed ultrasound or electromagnetic radiation is applied to the target tissue in order to stimulate the production or activation of HSP and facilitate protein repair in living animal tissue. In general, electromagnetic radiation can be ultraviolet waves, microwaves, other radio frequency waves, laser light at predetermined wavelengths, etc. On the other hand, if electromagnetic energy is used for target tissues deep away from natural orifices, the absorption lengths restrict the wavelengths to those of microwaves or radio frequency waves, depending on the depth of the target tissue. However, ultrasound should be preferred over long wavelength electromagnetic radiation to target tissues deep away from natural orifices.

[000120] Ultrasonic or electromagnetic radiation is pulsed in order to create a thermal course in time in the tissue that stimulates the production or activation of HSP and facilitates the repair of the protein without causing damage to the cells and tissue to be treated. The area and / or volume of the treated tissue is also controlled and minimized so that temperature spikes

Petition 870190083984, of 08/28/2019, p. 74/162

61/98 are of the order of several degrees, for example, approximately 10 ° C, while the long-term temperature rise is less than that required by the FDA limit, such as 1 ° C. It has been found that if a very large area or volume of tissue is treated, the rise in tissue temperature cannot be diffused fast enough to meet FDA requirements. However, limiting the area and / or volume of the treated tissue, as well as creating a pulsed energy source, accomplishes the objectives of the present invention to stimulate HSP activation or production by heating or otherwise stressing cells and tissues, allowing treated cells and tissues to dissipate any excess heat generated within acceptable limits.

[000121] With reference now to FIG. 34, with ultrasound, a specific region at the bottom of the body can be specifically targeted using one or more beams that are focused on the target location. The pulsating heating will, in large part, only be in the target region where the beams are focused and overlap. Pulsed ultrasound sources can also be

used for abnormalities on the surfaces as well how close from them. [000122] How illustrated in FIG. 34, a transducer in ultrasound 92 or similar generates a plurality of bundles in ultrasound 94 that are coupled to the skin through one gel of

acoustic-impedance correspondence and penetrate through the skin 96 and through undamaged tissue in front of the focus of the bundles 94 to a target organ 98, such as the illustrated liver and specifically to a target tissue 100 to be treated where the ultrasound bundles 94 are focused. As previously mentioned, the pulsating heating will then be only in the

Petition 870190083984, of 08/28/2019, p. 75/162

62/98 target region, focused 100, where focused bundles 94 overlap. The tissue in front of and behind the focused region 100 will not be heated or affected considerably.

[000123] The present invention contemplates not only the treatment of superficial or close tissues, such as the use of laser light or the like, deep tissues using, for example, focused or similar ultrasound beams, but also the treatment of blood diseases, such as sepsis. As previously indicated, the focused ultrasound treatment could be used both on the surface and on deep body tissue and could also be applied, in this case, in the treatment of blood. However, it is also contemplated that similar SDM treatment options that are typically limited to surface treatment or close to the surface of epithelial cells and the like, are used to treat blood disorders in areas where blood is accessible through a relatively thin layer. thin tissue, such as the earlobe.

[000124] With reference now to FIGS. 35 and 36, treatment of blood disorders simply requires the transmission of SDM or other electromagnetic radiation or ultrasound impulses to the ear lobe 102, where the SDM or other radiation energy source can pass through the ear lobe tissue to the blood that passes through the earlobe. It will be appreciated that this approach can also occur in other areas of the body where blood flow is relatively high and / or close to the surface of the tissue, such as the fingertips, inside the mouth or throat, etc.

[000125] With reference again to FIGS. 35 and 36, an ear lobe 102 is shown adjacent to a clamp device

Petition 870190083984, of 08/28/2019, p. 76/162

63/98

104 configured to transmit SDM radiation or the like. This could be, for example, by means of one or more laser diodes 106, which would transmit the desired frequency on the desired pulse and pulse train to the earlobe 102. Energy can be supplied, for example, by means of a drive of the lamp 108. Alternatively, the drive of the lamp 108 can be the actual source of laser light, which would be transmitted through the appropriate optics and electronics to the earlobe 102. The fixation device 104 would only be used to fix the lobe of the patient's ear and cause radiation to be limited to the patient's earlobe 102. This can be done using mirrors, reflectors, diffusers, etc. This could be controlled by a control computer 110, which would be operated by a keyboard 112 or the like. The system may also include a display and speakers 114, if necessary, for example, if the procedure is performed by an operator at a distance from the patient.

[000126] The treatment proposed with an electromagnetic pulse or ultrasound train has two major advantages over previous treatments that incorporate a single short or prolonged (long) pulse. First, the short individual pulses (preferably sub-seconds) in the frequency activate cellular reconfiguration mechanisms such as the activation of the HSP with higher reaction speed constants than those that operate in larger time scales (minute or hour). Second, the repeated pulses in the treatment provide large thermal spikes (in the order of 10,000) that allow the cell repair system to more quickly overcome the activation energy barrier that separates a cellular state.

Petition 870190083984, of 08/28/2019, p. 77/162

64/98 dysfunctional of the desired functional state. The net result is a low therapeutic threshold, in the sense that a lower average applied power and the total applied energy can be used to achieve the desired treatment goal.

[000127] Power limitations in today's micropulsed diode lasers require a reasonably long exposure duration. The greater the exposure, the more important is the ability to dissipate heat from the central point towards the unexposed tissue at the edges of the laser point. Thus, the micro-pulsed laser beam of an 810 nm diode laser should have an exposure envelope life of 500 milliseconds or less and, preferably, approximately 300 milliseconds. Of course, if the micro-pulsed diode lasers become more powerful, the duration of exposure should be shortened accordingly.

[000128] In addition to the power limitations, another parameter of the present invention is the duty cycle or the frequency of the micropulse train or the length of the thermal relaxation time between consecutive pulses. It has been found that the use of a duty cycle of 10% or higher adjusted to provide micropulsed laser to an irradiance at similar MPE levels significantly increases the risk of lethal cell injury. However, cycles of less than 10%, and preferably 5% or less, demonstrate adequate thermal elevation and treatment at the level of the MPE cell to stimulate a biological response, but remain below the level expected to produce lethal cell damage. The shorter the duty cycle, however, the longer the duration of the exposure envelope and, in some cases, can exceed 500 milliseconds.

Petition 870190083984, of 08/28/2019, p. 78/162

65/98

[000129] Each micropulse lasts a fraction of milliseconds, typically between 50 microseconds and 100 microseconds in duration. Thus, for the duration of the exposure envelope of 300 to 500 milliseconds and in a duty cycle of less than 5%, there is a significant amount of time spent between the micropulses to allow for the thermal relaxation time between consecutive pulses. Typically, a delay of between 1 and 3 milliseconds and, preferably, approximately milliseconds, of the thermal relaxation time between consecutive pulses is required. For proper treatment, cells are typically exposed or hit between 50 and 200 times, and preferably between 75 and 150 at each location and with 1 to milliseconds of relaxation time or interval, the total time according to the described modalities previously to treat a particular area being exposed to laser points is generally less than a second, such as between 100 milliseconds and 600 milliseconds on average. The thermal relaxation time is necessary in order not to overheat the cells within that location or point and to prevent the cells from being damaged or destroyed. Although periods of time from 100 to 600 milliseconds may not seem long, given the small size of the laser points and the need to treat a relatively large area of the target tissue, treatment of the entire target tissue takes a significant amount of time, particularly for a patient being treated.

[000130] Other pulsed energy sources, including microwaves, radiofrequencies and ultrasound are also preferably pulsed in nature and have duty cycles and / or pulse frequencies and thus delay

Petition 870190083984, of 08/28/2019, p. 79/162

66/98 time or intervals between applications of micropulse energy to the target tissue. In addition, the target tissue previously treated with the energy micropulse must naturally dissipate the heat created by the application of energy so as not to exceed a predetermined upper temperature level that could permanently damage or even destroy the cells of the target tissue. Typically, the area or volume of the target tissue to be treated is much larger than the area or volume of the target tissue that is treated at any time by the energy sources, even if multiple bundles of energy are created and applied to the target tissue.

[000131] Consequently, the present invention can use the interval between consecutive applications at the same location to apply energy to a second treatment area, or additional areas, of the target tissue that is spaced from the first treatment area. The pulsed energy is returned to the first treatment site, or previous treatment sites, within the predetermined time interval, in order to provide sufficient thermal relaxation time between consecutive pulses, but also to sufficiently treat cells in those locations or areas properly, increasing sufficiently the temperature of these cells over time by repeatedly applying energy to that location in order to achieve the desired therapeutic benefits of the invention.

[000132] It is important to return to a previously treated location within a predetermined period of time to allow the area to cool sufficiently during that period, but also to treat it within the required time window. In the case of pulsed laser light applications, the laser light is returned to the previously treated location within one to

Petition 870190083984, of 08/28/2019, p. 80/162

67/98 three milliseconds and, preferably, approximately two milliseconds, as you cannot wait a second or two and return to a previously treated area that has not yet received the necessary complete treatment, as the treatment will not be as effective or perhaps not even effective. However, during that time, typically about 2 milliseconds, at least one other area and typically multiple areas, can be treated with a laser light application, as the laser light pulses are typically 50 seconds at 100 microseconds in duration. This is referred to here as microdisplacement. The number of additional areas that can be treated is limited only by the duration of the mycopulse and the ability to move light beams from one area to another in a controlled manner.

[000133] Currently, about four additional areas that are sufficiently spaced from each other can be treated during thermal relaxation intervals starting with a first treatment area when using laser light. Thus, multiple areas can be treated, at least partially, during the 200 to 500 millisecond exposure envelope for the first area. Thus, in a single time interval, instead of just 100 simultaneous light points being applied to a treatment area, approximately 500 light points can be applied during that time in different treatment areas. This would be the case, for example, with a beam of laser light with a wavelength of 810 nm. For shorter wavelengths, such as 572 nm, even a larger number of individual locations can be exposed to laser beams to create points of light. So, instead of a maximum of approximately

Petition 870190083984, of 08/28/2019, p. 81/162

68/98

400 simultaneous points, approximately 2,000 points could be covered during the interval between micropulse treatments for a given area or location. Typically, each site has between 50 and 200 and, more typically, between 75 and 150, light applications applied over the duration of the exposure envelope (typically 200 to 500 milliseconds) to achieve the desired treatment. In accordance with an embodiment of the present invention, the laser light would be reapplied to previously treated areas in sequence during the rewash time intervals for each area or location. This would occur repeatedly until a predetermined number of laser light applications in each area to be treated were achieved.

[000134] Likewise, one or more microwave, radiofrequency and / or ultrasound beams could be applied to additional second treatment areas of the target tissue, separated from the first treatment area, and after a predetermined return interval , if necessary, for the first treatment area of the target tissue to reapply the pulsed energy to it. The pulsed energy can be reapplied to a previously treated area in sequence, during the relaxation time intervals for each area or location, until a desired number of applications have been reached for each treatment area. The treatment areas must be separated by at least a predetermined minimum distance to allow for thermal relaxation and dissipation and to avoid thermal damage to tissues. Pulsed energy parameters, including wavelength or frequency, duty cycle and pulse frequency duration are selected to raise the temperature of the target tissue to

Petition 870190083984, of 08/28/2019, p. 82/162

69/98 ° C, as approximately between 6 ° to 11 ° C, during the application of the pulsed energy source to the target tissue to achieve a therapeutic effect, such as stimulating the production of HSP within the cells. However, the cells of the target tissue must have a period of time to dissipate heat so that the average rise in tissue temperature over several minutes is maintained at or below a predetermined level, such as 6 ° C or less, or even 1 ° C or less, for several minutes, to not permanently damage the target tissue.

[000135] This is illustrated schematically in FIGS. 37A to 37D. FIG. 37A illustrates with solid circles a first area having beams of energy, such as beams of laser light, applied to it as a first application. The beams are moved in a controlled way or microscoupled to a second exposure area, followed by a third exposure area and a fourth exposure area, as illustrated in FIG. 37B, until the sites in the first exposure area need to be treated again by means of vehicles applied again within the thermal relaxation time interval. The locations within the first exposure area would then have energy bundles reapplied to them, as illustrated in FIG. 37C. Secondary or subsequent exposures would occur in each exposure area, as illustrated in FIG. 37D by the increasingly shaded points or circles until the desired number of exposures or blows or applications of energy in the target tissue area has been reached to therapeutically treat these areas, illustrated diagrammatically by the black circles in the exposure area 1 in FIG. 37D. When a first exhibition area or

Petition 870190083984, of 08/28/2019, p. 83/162

70/98 is completed, this allows the system to add an additional exposure area, a process that is repeated until the entire area to be treated has been fully treated. It should be understood that the use of solid circles, broken line circles, partially shaded circles and fully shaded circles are for explanatory purposes only, as in fact the exposure of energy or laser light according to the present invention is invisible and not detectable either the human eye and by known detection devices and techniques.

[000136] Adjacent exposure areas must be separated by at least a predetermined minimum distance to avoid thermal damage to the fabric. Such a distance is at least 0.5 in diameter from the immediately preceding location or treated area and more preferably between 1 and 2 distances in diameter. Such spacing refers to the places actually treated in a previous exhibition area. It is contemplated by the present invention that a relatively large area can actually include multiple areas of exposure that are shifted in a different way than illustrated in FIG. 37. For example, the display areas may comprise the fine lines illustrated in FIGS. 25 and 26, which would be repeatedly exposed in sequence until all necessary areas were fully exposed and treated. According to the present invention, the time needed to treat the area to be treated is significantly reduced, such as by a factor of 4 or 5 times, so that a single treatment session takes much less time for the doctor and the patient does not need to be uncomfortable for such a long period of time.

Petition 870190083984, of 08/28/2019, p. 84/162

71/98

[000137] According to this modality of the invention of applying one or more treatment bundles at once, and moving the treatment bundles to a series of new locations, then returning the bundles to depict the same location or area repeatedly also requires less energy compared to the methodology of keeping the beams in the same locations or area for the duration of the exposure envelope. With reference to FIGS. 38 to 40, there is a linear relationship between the pulse length and the required power, but there is a logarithmic relationship between the heat generated.

[000138] With reference to FIG. 38, a graph is shown in which the x-axis represents the Log of the average power in watts of a laser and the y-axis represents the treatment time, in seconds. The lower curve is for panmacular treatment and the upper curve is for panretinal treatment. This would be for a beam of laser light with a micropulse time of 50 microseconds, a period of 2 milliseconds between

the wrists and the duration gives frequency in a dot in 300 milliseconds. The areas in each Score gives retina are in 100 microns, and The power of laser for these points in 100 microns in retina is 0, 74 watts. The area panmacular is 0, 55 ² ,

requiring 7,000 total panmacular points and the panretinal area o and 3.30, requiring 42,000 laser points for full coverage. Each point of RPE requires minimum energy for its replacement mechanism to be properly activated, according to the present invention, namely, 38.85 joules for panmacular and 233.1 joules for panretinal. As expected, the shorter the treatment time, the greater the average power required. However, there is an upper limit on the power

Petition 870190083984, of 08/28/2019, p. 85/162

72/98 average allowed, which limits how short the treatment time can be.

[000139] As mentioned earlier, there are not only energy restrictions in relation to the available and used laser light, but also in relation to the amount of energy that can be applied to the eye without damaging the eye tissue. For example, the temperature rise in the lens of the eye is limited, such as between 4 ° C, so as not to overheat and damage the lens, such as causing cataracts. Thus, an average power of 7.52 watts could raise the temperature of the lens to approximately 4 ° C. This power limitation increases the minimum treatment time.

[000140] However, with reference to FIG. 39, the total pulse power required is lower in the case of the microdisplacement of moving the laser points repeatedly and sequentially and returning to previously treated locations, so that the total energy distributed and the total average power during the treatment time are the same . FIGS. 39 and 40 show how the total potency depends on the treatment time. This is shown in FIG. 39 for panmacular treatment and in FIG. 40 for panretinal treatment. The top and solid line or curve represents the mode where there are no microdisplacements taking advantage of the thermal relaxation time interval, as described and illustrated in FIG. 24, while the lower dashed line represents the situation for such micro displacements, as described and illustrated in FIG.

37. FIGS. 39 and 40 show that, for a given treatment time, the total peak power is lower with microdisplacements than without microdisplacements. This means that less energy is needed for a given

Petition 870190083984, of 08/28/2019, p. 86/162

73/98 treatment, using the microdisplacement modality of the present invention. Alternatively, the permissible peak power can be used to advantage, reducing the total treatment time.

[000141] Thus, according to FIGS. 38 to 40, a log power of 1.0 (10 watts) would require a total treatment time of 20 seconds, using the microdisplacement modality of the present invention, as described herein. This would take more than 2 minutes without the microdisplaces and, instead, leave the micropulsed light beams in the same location or area for the entire duration of the treatment envelope. There is a minimum treatment time according to the power. However, this treatment time with microdisplacement is much less than without microdisplacement. Since the laser power required is much less with microdisplacement, it is possible to increase the power in some cases, in order to reduce the treatment time for a given retinal treatment area. The product of the treatment time and the average power are fixed for a given treatment area, in order to achieve the therapeutic treatment according to the present invention. This can be implemented, for example, by applying a larger number of beams or therapeutic laser light points simultaneously at reduced power. Of course, since the laser light parameters are selected to be therapeutically effective, but not destructive or permanently harmful to cells, no guidance or tracking beams are needed, only treatment beams, since all areas can be treated in accordance with the present invention.

Petition 870190083984, of 08/28/2019, p. 87/162

74/98

[000142] Although the present invention is described for use in conjunction with a micropulse laser, theoretically a continuous wave laser could potentially be used instead of a micropulse laser. However, with the continuous wave laser, there is concern about overheating, as the laser is moved from one location to another, as the laser does not stop and there may be heat leakage and overheating between the treatment areas. Thus, although it is theoretically possible to use a continuous wave laser, in practice it is not ideal and the micropulsed laser is preferred.

[000143] While the information provided in conjunction with charts 38 to 40 is derived from observations and calculations of laser light beams as a source of energy applied to the retinal tissue of the eye, it is believed that the application of pulsed laser light to another tissue will achieve similar results in that the movement of the treatment bundles to a series of new locations, then bringing the bundles back to repeatedly treat the same location or area repeatedly will not only save time, but will also require less energy compared to the keep the bundles in the same location or area for the entire duration of the exposure envelope. Likewise, it is believed that this energy conservation will also be achieved with other sources of pulsed energy, including microwave, radiofrequency and ultrasound energy sources.

[000144] According to the microdisplacement technique described above, the displacement or direction of the light beam pattern can be done using an optical scanning mechanism, such as that illustrated and described together with FIGS. 22 and 23. For situations where the length of

Petition 870190083984, of 08/28/2019, p. 88/162

75/98 wave of illumination or energy is much less than the distance to the volume to be illuminated or exposed, the targeting can be performed using phased arrangements. The lighting or energy in this case is said to be the distant field. Phased arrangements can be used for the application of microwave and ultrasound lighting or even for the laser beam source.

[000145] The targeting for microwaves, ultrasound and even for laser energy sources can be done through the use of multiple sources that provide an arrangement. The basic idea for directing the lighting radiation pattern of an array is the constructive (and destructive) interference between the radiation of the individual members of the array of sources. With reference to FIG. 41, to illustrate this, it is only necessary to consider two adjacent members of the arrangement. FIG. 41 represents the wavefront originating from two adjacent sources.

[000146] It is evident that for a wavefront that is represented at an angle Θ with respect to the distance between the two sources, the amplitude of the wave from the source on the left is proportional to exp [icot] while the amplitude of the wave from the source to right is proportional to exp [icot - kasinQ -φ], where ω is the angular frequency of radiation and k = 2π / λ.

[000147] For constructive interference, these two waves must be in phase, that is, çconstrutivo = kasinQ + 2nn [1]

[000148] For destructive interference, these two waves must be out of phase, that is, çdestructive = kasinQ + (2n +1) π [2]

Petition 870190083984, of 08/28/2019, p. 89/162

76/98

[000149] Thus, the illumination will be great in the directions Θ given by sinQ = (l / ka) [çconstrutivo-2nn] [3]

[000150] In other words, the radiation can be directed in different desired directions Θ simply by choosing different intervals φ.

[000151] The intervals can be introduced electronically in the circuits to excite the sources of radiation. The means to do this have also been well discussed in the published literature: analog range circuits are available, as well as digital range circuits.

[000152] The radiation standards for microwaves, ultrasound and laser sources are well targeted. If we estimate the divergence of the radiation beam from a source of transversal dimension 2b by the expression of the Airy disk

01/2 = 0.6 À / b [4]

[000153] So, at a target distance D from the source, the half width w of the illuminated region is approximately w = 0.6 ÀD / b [5]

[000154] If we need the separation of the illuminated regions to be 2w, then the separation of the source s is approximately 3w: a = 1.8 ÀD / b [6]

[000155] This can be a small separation if the font size is chosen to be much larger than the radiation's wavelength.

[000156] For example, for ultrasound, suppose we have a 5 MHz source with a transversal dimension of 1 cm and suppose that the desired distance is 10 cm. Then the separation distance is ~ 0.5 cm.

Petition 870190083984, of 08/28/2019, p. 90/162

77/98

[000157] As another example, a commercially available standard microwave gain horn source operating at 140 to 220 GHz has cross-sectional dimensions of 13.9 mm by 10.8 mm and a depth dimension of 32.2 mm . For 200 GHz, the wavelength is 0.15 cm and for a target distance of 10 cm, the target width given by equation [5] is 1.2 x 0.15 x 10 / 0.6 = 3 cm. For the spacing of the horns, eq. [6] then gives 9 cm.

[000158] Then apply the eqs. [4] to [6] to obtain approximate estimates of a directional array of 810 nm laser radiation. Suppose b = 2x810 nm and suppose D = 1 mm. Then the eqs. [4] to [6] provide Θ1 / 2 = 0.3, w = 0.3 mm and a = 0.9 mm.

[000159] For the application of radio frequency, however, the wavelength of radio frequency radiation is typically much larger than the dimensions of a human body. In this case, it is said that the treatment volume is in the field close to the radio frequency source. Phased arrangements are not useful in the near field and a different method of targeting is needed.

[000160] For radiofrequency treatment, the wavelength of the radiation is much greater than the dimensions of the body. Thus, for 3 to 6 MHz, the wavelengths vary from 10,000 cm to 5,000 cm. Thus, the target region in the body is in the field close to the source, that is, the distance and dimensions of the target are much smaller than the wavelength of the RF radiation. This means that the relevant treatment fields are not radiation fields (as they were in the case of microwave, ultrasound and laser treatments), but induction fields.

Petition 870190083984, of 08/28/2019, p. 91/162

78/98

[000161] The induction field of an RF coil is only large over dimensions comparable to the dimension of the coil. The induction magnetic fields drop rapidly as l / r3 for distances greater than that. Thus, for a coil on the body surface, we can imagine the treatment volume as approximately a hemisphere with a radius equal to that of the coil.

[000162] For coils with radii between 2 and 6 mm, the treatment volumes for these coils are very close to the surface (distances comparable to the dimensions of the coil). Larger coils can be used for deeper targets. According to the spacing criteria discussed earlier, the spacing between the coils in a surface matrix would be chosen to be comparable to the individual dimensions of the coil.

[000163] As mentioned earlier, the controlled way of applying energy to the target tissue is designed to increase the temperature of the target tissue to therapeutically treat the target tissue without destroying or permanently damaging the target tissue. Such heating is believed to activate HSPs and that thermally activated HSPs work to redefine diseased tissue to a healthy condition, for example, by removing and / or repairing damaged proteins. The inventors believe that maximizing this activation of HSP improves the therapeutic effect on the target tissue. As such, understanding the behavior and activation of HSP and HSP system species, their generation and activation, temperature ranges for HSP activation and time limits for HSP activation or generation and deactivation can be used to optimize the thermal treatment of biological target tissue .

Petition 870190083984, of 08/28/2019, p. 92/162

79/98

[000164] As mentioned earlier, the target tissue is heated by the pulsed energy for a short period of time, such as ten seconds or less, and typically less than one second, such as between 100 milliseconds and 600 milliseconds. The time that energy is effectively applied to the target tissue is typically much less than this, in order to provide time intervals for the relaxation of heat, so that the target tissue does not overheat and is damaged or destroyed. For example, as mentioned earlier, pulses of light from the laser can last in the order of microseconds with several milliseconds of time laps.

[000165] Thus, understanding the HSP sub-second behaviors may be important for the present invention. The thermal activation of HSPs in the SDM is typically described by an associated Arrhenius integral,

Ω = Jdt A exp [-E / k _B T (t)] [1] where the integral is in the treatment time and

A is the Arrhenius rate constant for HSP activation E is the activation energy

T (t) is the temperature of the thin layer of RPE, including the laser-induced temperature rise

[000166] The temperature increase induced by the laser - and therefore the activation of the Arrhenius integral - depends on the treatment parameters (for example, laser power, duty cycle, total train duration) and the properties of the RPE ( for example, absorption coefficients, density of HSPs). It has been clinically verified that effective SDM treatment is obtained when the Arrhenius integrals are of the order of the unit.

[000167] Arrhenius' integral formalism takes into account only a direct reaction, that is, only the activation reaction

Petition 870190083984, of 08/28/2019, p. 93/162

80/98 of the HSP): does not take into account any reverse reaction in which the activated HSP returns to their inactivated states. For typical sub-second durations of SDM treatments, this appears to be quite adequate. However, for long periods of time (for example, a minute or more), this formalism is not a good approximation: in these longer times, a whole series of reactions occur resulting in much lower rates of HSP activation. This is the case during the proposed minute or intervals between SDM applications in the disclosure of the present invention.

[000168] In the published literature, the production and destruction of heat shock proteins (HSPs) in cells of longer duration is generally described by a set of 9 to 13 simultaneous differential mass equilibrium equations that describe the behavior of the various molecular species involved in the life cycle of an HSP molecule. These simultaneous equations are usually solved by computer to show the time behavior of HSPs and other species after the temperature has been suddenly increased.

[000169] These equations are all conservation equations based on the reactions of the various molecular species involved in the activity of HSPs.

To describe the behavior of HSfs in. approximately one minute between repeated applications of SDM, we will use the equations described in M. Rybinski, Z.Szymanska, S. Lasota, A. Gambin (2013) Modeling the efficacy of hyperthermia treatment. Journal of the Royal Society Interface 10, No. 88, 20130527 (Rybinski et al (2013)). The species considered in Rybinski et al (2013) are shown in Table 1.

Petition 870190083984, of 08/28/2019, p. 94/162

81/98

Table 1. Species of the HSP system in the description by Rybinski et al (2013):

HSP ubiquitous heat shock protein of molecular weight 70 Da (in activated free state)

HSF 'thermal shock factor (transcription) that lacks DNA binding capacity

HSF ₃ heat shock factor HSF3 (trimer) capable of binding to DNA, formed from HSF

HSE> * HSE heat shock element, a DNA site that initiates HSP transcription when bound to the HSF ₃ mRNA -μ molecule, from messenger RNA to produce HSP

A substrate for HSP binding: a damaged protein.

P protein properly folded

HSP.HSF -> i is a HSP complex bound to HSF (HSPs not activated)

HSFj.HSE -> i um. HSE-linked HSF ₃ complex, which induces the transcription and creation of a new HSP mRNA molecule

HSP.S -x an HSP complex linked to damaged proteins (HSP that actively repairs the. Protein)

[000170] The coupled simultaneous mass conservation equations for these 10 species are summarized below as eqs. [2] to [11]:

d [HSP] / dt = (li + kio) [HSPS] + 1 ₂ [HSPHSF] + k ₄ [mRNA]

- ki [S] [HSP] -k ₂ [HSP] [HSF] -1 ₃ [HSP] [HSF ₃ ] - k ₉ [HSP] [2] d {HSF] / dt = 1 ₂ [HSPHSF] + 21 ₃ [HSP] [HSF ₃ ] + k ₆ [HSPHSF] [S]

Petition 870190083984, of 08/28/2019, p. 95/162

82/98

-k ₂ [HSP] [HSF] - 3k ₃ [HSF] ³ - 1 ₆ [HSPS] [HSF] [3] d [S] / dt = ku {[P] + li [HSPS] + 1 ₆ [SPS ] [HSF] - ki [S] [HSP] k ₆ [HSPHSF] [S] [4] d [HSPHSF] / dt = k ₂ [HSP] [HSF] + 1 ₆ [HSPS] [HSF] + 1 ₃ [HSP] [HSF ₃ ] - 1 ₂ [HSPHSF] - k ₆ [HSPHSF] [S] [5]

d [HSPS] / dt = k ₂ [S] [HSP] + kg [HSPHSF] [S] - (li + ki ₀ ) [HSPS] - lg [HSPS] [HSF] [6] d [HSF ₃ ] / dt = k ₃ [HSF] ³ + 1 ₇ [HSF ₃ ] [HSE] - 1 ₃ [HSP] [HSF ₃ ] - k ₇ [HSF ₃ ] [HSE] [7]

d [HSE] / dt = 1 ₇ [HSF ₃ ] [HSE] - k ₇ [HSF ₃ ] [HSE] [8 d [HSF ₃ HSE] / dt = k ₇ [HSF ₃ ] [HSE] - 1 ₇ [ HSF ₃ ] [HSE] [9] d [mRNA] / dt = k ₈ [HSF ₃ HSE] - k ₅ [mRNA] [10] d [P] / dt = k ₁₀ [HSPS] - k _n [P] [11]

[000171] In these expressions, [] represents the concentration

the amount inside the holder. For Rybinski et al (2013), the initial concentrations at the equilibrium temperature of 310K are given in Table 2.0

Table 2. Initial values of species in 310K for a typical cell in arbitrary units [Rybinski et al (2013)]. The arbitrary units are chosen by Rybinski et al for computational convenience: to make the quantities of interest in the range of 0.01 to 10.

[HSP (0)] 0.308649 [HSF (0)] 0. 150836 [S (0)] 0. 113457 [HSPHSF (0)] 2.58799 [HSPS (0)] 1.12631 [HSF ₃ (0)] 0.0444747 [HSE (0)] 0.957419 [HSF ₃ HSE (0)] 0.0425809

Petition 870190083984, of 08/28/2019, p. 96/162

83/98

[mRNA (O) ] 0.114641 [P (0)] 8.76023 [000172] The rate constants of Rybinski et al (2013) are shown in Table 3. Table 3. Rybinski et al (2013) calculate constants giving

min ¹ rates for arbitrary concentration units in the previous table.

li = = 0.0175 ki = 1.47 1 ₂ = 0.0175 k ₂ = 1.47 13 = = 0.020125 k ₃ = 0.0805 k ₄ = = 0.1225 k ₅ ^: = 0.0455 ke ^: = 0.0805 16 = = 0.00126 k ₇ = = 0.1225 I7 ^: = 0.1225 kg ^: = 0.1225 kg ^: = 0.0455 kio = 0.049 kn = 0.00563271 [000173] The initial concentration values in Table 2 and the

speed constants in Table 3 were determined by

Rybinski et al (2013) to correspond to experimental data about 0 cc > general behavior of the HSP system when the temperature increased by 5 ° C by several (for example, example 350) minutes.

Petition 870190083984, of 08/28/2019, p. 97/162

84/98

[000174] Note that the initial concentration of HSPs is 100 x 0.308649 / (8.76023 +0.113457 +1.1231)} = 3.09% of the total number of proteins present in the cell.

[000175] Although the speed constants in Table 3 are used by Rybinski et al. for T = 310 +5 + 315K it is likely that there are velocity constants very similar to other temperatures. In this context, the qualitative behavior of the simulations is similar for a large variety of parameters. For convenience, we will assume that the values of the velocity constants in Table 3 are a good approximation for the values at the equilibrium temperature of T === 310K.

[000176] The behavior of the different components in the cell of Rybinski et al is shown in FIG. 42 for 350 minutes for the situation where the temperature is suddenly increased by 5K at t = 0 in an 310K environment.

[000177] With continuous reference to FIG. 42, the behavior of the components of the HSP cellular system is shown for 350 minutes after a sudden temperature rise from 37 ° C to

42 ° C.

[000178] Here, the concentrations of the components are presented in arbitrarily computationally convenient units. S represents denatured or damaged proteins that have not yet been affected by HSPs; HSP represents free (activated) heat shock proteins; HSP: S represents activated HSPs that are bound to the damaged proteins and perform repairs; HSP: HSF represents HSPs (inactive) that are linked to thermal shock factor monomers; HSF represents a thermal shock factor monomer; HSF3 represents a thermal shock factor trimer

Petition 870190083984, of 08/28/2019, p. 98/162

85/98 that can penetrate the nuclear membrane to interact with a thermal shock element in the DNA molecule; HSE: HSF3 represents a trimester of thermal shock factor linked to a thermal shock element in the DNA molecule that initiates the transcription of a new mRNA molecule; mRNA represents the messenger RNA molecule that results from HSE: HSF3 and that leads to the production of a new HSP molecule (activated) in the cell's cytoplasm.

[000179] FIGURE 42 shows that initially the concentration of activated HSPs is the result of the release of HSPs sequestered in the HSPHSF molecules in the cytoplasm, with the creation of new HSPs from the cell nucleus by means of mRNA not occurring until 60 minutes after the temperature rise occurs. FIG. 42 also shows that activated HSP are very quickly bound to damaged proteins to begin their repair work. For the cell represented, the sudden increase in temperature also results in a temporary increase in the concentration of damaged protein, with the peak in the concentration of damaged protein occurring about 30 minutes after the temperature increase.

[000180] FIGURE 42 shows what the equations of Rybinski et al predict the variation of 10 different species over a period of 350 minutes. However, the present invention relates to the application of SDM in species variation over the much shorter O (minute) interval between two applications of SDM at any single location on the retina. It will be understood that the preferred modality of SDM in the form of laser light treatment is analyzed and described, but it is also applicable to other energy sources.

Petition 870190083984, of 08/28/2019, p. 99/162

86/98

[000181] With reference now to FIGS. 43A-43H, ο behavior of the components of the HSP cellular system during the first minute after a sudden temperature rise from 37 ° C to 42 ° C using the equations of Rybinski et al. (2013) with the initial values and speed constants in Tables 2 and 3 is shown. The abscissa represents the time in minutes and the ordinate shows the concentration in the same arbitrary units as in FIG. 43.

[000182] FIGURE 43 shows that the nuclear source of HSPs practically plays no role over a period of 1 minute and that the main source of new HSPs in the cytoplasm arises from the release of sequestered HSPs from the HSPHSF molecule reservoir. It also shows that a good fraction of the newly activated HSPs bind to damaged proteins to initiate the repair process.

[000183] The initial concentrations in Table 2 are not the equilibrium values of the species, that is, they do not provide d [. . . ] / dt = 0, as evidenced by the curves in FIGS. 42 and 43. The equilibrium values that provide d [...] / dt = 0 corresponding to the speed constants in Table 3 are those listed in Table 4.

Table 4. Species equilibrium values in arbitrary units [Rybinski et al (2013)] corresponding to the velocity constants in Table 3. The arbitrary units are those chosen by Rybinski et al for computational convenience: to make the quantities of interest in the range from 0.01 to 10.

[HSP (equilibrium)] 0.315343

[HSF (equil)] 0.255545

[S (equilibrium)] 0.542375

Petition 870190083984, of 08/28/2019, p. 100/162

87/98

[HSPHSF (equil)] 1.982248

[HSPS (equilibrium)] 5.05777

[HSF ₃ (equilibrium)] 0.210688

[HSE (equilibrium)] 0.206488

[HSF ₃ HSE (equilibrium)], 643504

[mRNA (equilibrium)] 0.1171274

[P (equilibrium)] 4.39986

[000184] Note that the equilibrium concentration of HSPs is 100 x {0.315343 / (4.39986 + 5.05777 + 0.542375)} = 3.15% of the total number of proteins present in the cell. This is comparable, but less than expected, from 5% to 10% of the total proteins found by other researchers. However, we do not try to adjust the percentage upwards, hoping that the general behavior will not be significantly altered, as indicated by other researchers.

[000185] The inventors have found that a first treatment for the target tissue can be carried out by repeatedly applying pulsed energy (e.g., SDM) to the target tissue over a period of time in order to raise the temperature of the target tissue in a controlled manner to therapeutically treat the target tissue without destroying or permanently damaging the target tissue. A treatment comprises the total number of applications of pulsed energy to the target tissue over a given period of time, such as tens or even hundreds of applications of light or other energies to the target tissue over a short period of time, such as a period of less than ten seconds and, more typically, a period of less than one second, such as 100 milliseconds to 600 milliseconds. This treatment increases the temperature of the target tissue to activate heat shock proteins and related components.

Petition 870190083984, of 08/28/2019, p. 101/162

88/98

[000186] What has been found, however, is that if the application of pulsating energy to the target tissue is interrupted for a period of time, such as a period of time that exceeds the first period of time that comprises the first treatment, which can include several seconds to several minutes, such as three seconds to three minutes or more, preferably ten seconds to 90 seconds, and then a second treatment is performed on the target tissue after the time interval within a session or medical consultation single treatment, in which the second treatment also involves repeatedly reapplying the pulsating energy in the target tissue in order to controllably raise the temperature of the target tissue to therapeutically treat the target tissue without destroying or permanently damaging the target tissue, the amount of activated HSPs and related components in the target tissue cells is increased, resulting in a more effective and comprehensive treatment of biological tissue. In other words, the first treatment creates a level of activation of the heat shock protein in the target tissue and the second treatment increases the level of activation of the heat shock protein in the target tissue above the level due to the first treatment. Thus, conducting multiple treatments on the patient's target tissue within a single treatment session or office visit improves the overall treatment of the biological tissue, provided that the second treatment or additional treatments are carried out after an interval of time that does not exceed several minutes, but that is long enough to allow the temperature to relax so as not to damage or destroy the target tissue.

Petition 870190083984, of 08/28/2019, p. 102/162

89/98

[000187] This technique can be referred to here as a step, as the levels of activated HSP production increase with the treatment or subsequent treatments within the same treatment session as the office visit. This step technique can be described by combining Arrhenius' integral approach to subsecond phenomena with the treatment of intervals by Rybinski et al. (2013) between repeated subsecond applications of SDM or other pulsed energy.

[000188] For the proposed step SDM (repetitive SDM applications) in this disclosure of the invention there are some important differences from the situation represented in Figure 42:

• SDM can be applied prophylactically to a healthy cell, but often SDM will be applied to a diseased cell. In this case, the initial concentration of damaged proteins [S (0)] may be higher than that given in Table 4. We will not try to explain this, assuming that the qualitative behavior will not be changed.

• The duration of a single SDM application is only sub-seconds, instead of the minutes shown in Figure 42. The Rybinski and other velocity constants are much smaller than the Arrhenius constants: the latter provide integrals of

Arrhenius of the order of unity for, subsecond durations, while the speed constants of Rybinski and others are too small to do this. This is an example of the different effective rate constants that exist when the time scales of interest are different:

Petition 870190083984, of 08/28/2019, p. 103/162

90/98 Rybinski et al rates apply to phenomena that occur in minutes, while Arrhenius rate constants apply to subsecond phenomena.

[000189] Thus, to analyze what happens in the proposed stepwise SDM technique to improve the effectiveness of SDM, the treatment of the Arrhenius integral appropriate for the subsecond phenomena must be combined with the treatment of Rybinski et al (2013 ) appropriate for the phenomena that occurred during the order of an interval of minutes between repeated SDM applications:

• Sub-second application of SDM described by the integral formalism of Arrhenius • O interval (minute) between SDM applications described by equations by Rybinski et al (2013)

[000190] Specifically, we consider two successive applications of SDM, each SDM micropulse frequency having a subsecond duration.

• For the short subsecond time scale, it is assumed that the non-activated HSPs that are the source of the activated (free) HSP are all contained in the HSPHSF molecules in the cytoplasm. Thus, the first application of SDM is taken to reduce the cytoplasmic reservoir of non-activated HSPs in the initial population of HSPHSF molecules from

[HSPHSF (equil)] to [HSPHSF (equil)] exp [-Ω], • and increase the initial molecular population of HSP from

[HSP (equil)] to [HSP (equilibr)] + [HSPHSF (equilibr)] (1exp [-Ω])

Petition 870190083984, of 08/28/2019, p. 104/162

91/98 • as well as increasing the initial molecular population of HSF from

[HSF (equil)] to [HSF (equilibr)] +

[HSPHSF (equilibr)] (l-exp [-Q]) • It is assumed that the equilibrium concentrations of all other species remain the same after the first application of the SDM • Rybinski et al's equations are then used to calculate what happens to [HSP] and [HSPHSF] in the interval Àt = 0 (minute) between the first SDM application and the second SDM application, with the initial values of HSP, HSF and HSPHSF after the first SDM application considered as

[HSP (SDMl)] = [HSP (equil)] + [HSPHSF (equilibr)] (1-exp [Ω])

[HSF (SDMl)] = [HSF (equil)] + [HSPHSF (equilibr)] (1-exp [Ω]) and

[HSPHSF (SDM1)] = [HSPHSF (equil)] exp [-Ω] • For the second application of the SDM after the Àt interval, the values of [HSP], [HSF] and {HSPHSF] after the SDM will be considered as being

[HSP (SDM2)] = [HSP (Àt)] + [HSPHSF (Àt)] (1-exp [Ω])

[HSF (SDM2)] = [HSF (Àt)] + [HSPHSF (Àt)] (1-exp [Ω]) and

[HSPHSF (SDM2)] = [HSPHSF (Àt)] exp [-Ω]

Petition 870190083984, of 08/28/2019, p. 105/162

92/98 where [HSP (Xt)], [HSF (Xt)] and [HSPHSF (Xt)] are the values determined by the equations of Rybinski et al (2013) at time Xt.

Our current interest is to compare [HSP [SDM2]] repeated with of [HSP [SDM1]], for to see if the application SDM in a range Xt after first application of SDM resulted in HSPs more activated (free) at the cytoplasm. THE β ratio (Xt, Ω) =

[HSP (SDM2)] / [HSP (SDM1)] {[{[HSP (Xt)] + [HSPHSF (Xt)] (1-exp [-Ω])} / {

[HSP (O)] + [HSPHSF (0)] (1-exp [-Ω])} provides a direct measure of the improvement in the degree of HSP activation for a repeated application of the SDM after an Xt interval of the first application of the SDM .

[000191] The concentrations of HSP and HSPHSF can vary widely in the Xt interval between SDM applications.

[000192] FIGURES 44A and 44B illustrate the variation in activated concentrations [HSP] and HSP not activated in the cytoplasmic reservoir [HSPHSF] during an interval Xt = 1 minute between SDM applications when the Arrhenius integral of SDM Ω = 1 and equilibrium concentrations are given in Table 4.

[000193] Although only a single repetition (one step) is treated here, it is evident that the procedure could be repeated to provide multiple step events as a means of improving the effectiveness of SDM or another therapeutic method involving activation of tissue HSPs.

Petition 870190083984, of 08/28/2019, p. 106/162

93/98

[000194] The effects of the variation in the magnitude of the Arrhenius integral Ω and the Àt interval between two different treatments separated by a time interval are shown by the following examples and results.

[000195] Nine examples generated with the procedure described above are presented below. All examples are of a treatment that consists of two SDM treatments, with the second occurring in an Àt time following the first and they explore:

• The effect of different Arrhenius integrals of magnitude Ω in the SDM treatments [Three different Ω are considered: Ω = 0.2, 0.5 and 1.0] • The impact of varying the Àt interval between the two SDM treatments [Three different Àt are considered: Àt = 15 s, 30 s and 60 s.

[000196] As previously indicated, the activation of the Arrhenius Integral depends both on the treatment parameters (for example, laser power, duty cycle, total train duration) and on the properties of the RPE (for example, absorption coefficients, density HSPs).

[000197] Table 5 below shows the effect of different Ω (Ω = 0.2, 0.5, 1) on the HSP content of a cell when the interval between the two SDM treatments is Xt = 1 minute. Here, the cell is considered to have the equilibrium concentrations of Rybinski et al (2013) for the ten species involved, given in Table 4.

[000198] Table 5 shows four concentrations of HSP (in the arbitrary units of Rybinski et al.) Each corresponding to four different times:

Petition 870190083984, of 08/28/2019, p. 107/162

94/98 • Before the first SDM treatment: [HSP (equil)] • Immediately after the first application of SDM: [HSP (SDMI)] • At the end of the Àt interval after the first SDM treatment: [HSP (Àt)] • Immediately after the second SDM treatment in Àt: [HSP (SDM2)] • The improvement factor in a single treatment is also shown: β = [HSP (SDM2)] / [HSP (SDMI)]

Table 5. HSP concentrations in the four times described in the text: Effect of the variation of SDM Ω for two SDM applications in a cell when the treatments are separated by Àt = 0.25 minutes = 15 seconds.

[HSP (equilibrium)] [HSP (SDMl)] [HSP (Àt)] [HSP (SDM2)] β

Ω = 0.2 0.315 0.95 1.27 0.67 0.54 Ω = 0.5 0.315 1, 10 0.77 1.34 1.22 Ω = 1.0 0.315 1.57 0.93 1.71 1, 09 [000199] The Table 6 is the same as Table 5, except that

for an interval between SDM treatments of Àt = 0.5 minutes = seconds.

Petition 870190083984, of 08/28/2019, p. 108/162

95/98

Table 6. HSP concentrations in the four times described in the text: Effect of the variation of SDM Ω for the two treatments of SDM in a cell when the treatments are separated by Àt = 0.5 minutes = 30 seconds.

[HSP (equilibrium)] [HSP (SDMl)] [HSP (Àt)] [HSP (SDM2)] β Ω = 0.2 0.315 0.67 0.44 0.77 1, 14 Ω = 0.5 0.315 1, 10 0.58 1, 18 1, 08 Ω = 1.0 0.315 1.57 0.67 1.59 1.01 [000200] The Table 7 is the same as Tables 5 and 6, except that

treatments are separated by one minute or sixty seconds.

Table 7. HSP concentrations at the four times described in

text: Effect of variation of SDM Ω for two treatments SDM in a cell normal (healthy) When the treatments are separated by Àt = 1 minute = 60 seconds. [HSP (equilibrium) ] [HSP (SDMl)] [HSP (Àt)] [HSP (SDM2)] β Ω = 0.2 0.315 0.67 0.30 0.64 0.95 Ω = 0.5 0.315 1, 10 0.37 1, 06 0.96 Ω = 1.0 0.315 1.57 0.48 1.51 0.96 [000201] Tables 5 to 7 show what:

• The first treatment of SDM increases [HSP] by a large factor for all three Q's, although the

Petition 870190083984, of 08/28/2019, p. 109/162

96/98 increase is greater the greater Ω. Although not shown explicitly in the tables, the increase in [HSP] occurs at the expense of the cytoplasmic reservoir of hijacked (not activated) HSPs: [HSPHSF (SDM1)] is much smaller than [HSPHSF (equil)] • [HSP] in the Àt interval between the two SDM treatments, with the decrease being greater the greater the Àt. (The decrease in [HSP] is accompanied by an increase in both [HSPHSF], as shown in Figure 44, and in [HSPS] during the Àt interval, indicating a rapid replacement of the cytoplasmic reservoir of inactivated HSPs and a rapid onset of HSPs to damaged proteins.) • For Àt less than 60 seconds, there is an improvement in the number of activated (free) HSP in the cytoplasm for two treatments of SDM, instead of a single treatment.

• The improvement increases as Àt becomes smaller.

• For Àt to become as large as 60 seconds, however, the ratio β = [HSP (SDM2)] / [HSP (SDMI)] becomes smaller than the unit, indicating that there is no improvement in the two treatments of SDM in comparison with a single treatment of SDM, although this result may vary depending on the parameters of the energy source and the type of tissue that is treated.

• The improvement for Àt <60 seconds is greater the smaller the Arrhenius integral of SDM Ω.

Petition 870190083984, of 08/28/2019, p. 110/162

97/98

[000202] The results for the improvement rate β = [HSP (SDM2)] / [HSP (SDMI)] are summarized in Figure 45, where the improvement ratio β = [HSP (SDM2)] / [HSP (SDMI )] as a function of the interval between treatments of SDM Xt (in seconds) for three

integral values in Arrhenius of SDM Ω e for the three range values Xt = 15 s, 30 s and 60 s. The most high is for Ω = 0.2; The curve of middle is for Ω = 0 , 5; and The curve bottom is for Ω = 1 , 0. Est es results are to the

rate constants of Rybinski et al (2013) in Table 3 and the concentrations of species in equilibrium in Table 4.

[000203] It should be noted that the results of Tables 5 to 7 and FIG. 45 are for the constants by Rybinski et al. (2013) in Table 3 and the equilibrium concentrations in Table 4. The actual and velocity constant concentrations in a cell can differ from these values and, thus, the number results in Tables 5 to 7 and FIG. 45 should be considered representative and not absolute. However, they should not be significantly different. Thus, performing multiple intra-session treatments in a single location or target tissue area, such as a single retinal site, with the second and subsequent treatments after the first after an interval between three seconds to three minutes and preferably between ten seconds and ninety seconds, it should increase the activation of HSPs and related components and thus the effectiveness of the overall treatment of the target tissue. The resulting stepping effect achieves incremental increases in the number of heat shock proteins that are activated, increasing the therapeutic effect of the treatment. However, if the time interval between the first and the subsequent treatment is very

Petition 870190083984, of 08/28/2019, p. 111/162

98/98 large, the step effect will be decreased or not achieved.

[000204] The technique of the present invention is especially useful when the treatment parameters or tissue characteristics are such that the Arrhenius integral associated with activation is low and when the interval between repeated applications is short, such as less than ninety seconds and preferably less than a minute. Consequently, such multiple treatments must be performed within the same treatment session, as in a single office visit, in which different treatments may have a window of time between them, in order to achieve the benefits of the technique of the present invention. .

[000205] Although several modalities have been described in detail for purposes of illustration, several modifications can be made without departing from the scope and spirit of the invention. Thus, the invention should not be limited, except in accordance with the appended claims.

Claims (11)

1. SYSTEM FOR THERMAL TREATMENT OF BIOLOGICAL TISSUE, comprising: an intermittent pulsed energy source having energy parameters including wavelength or frequency, duty cycle and pulse train duration, in order to raise the temperature of a target tissue to eleven degrees Celsius at least during application of the pulsed energy source in the target tissue over a first period of time to create a first level of heat shock protein activation in the target tissue; characterized by understanding: a timer controlling activation gives source in energy pulsated to stop the application gives source in energy pulsed to tissue target by one interval in time what exceed the first period of time; is for to apply the source power pulsed at target tissue after the time interval within a single treatment session to increase the level of heat shock protein activation in the target tissue. System according to claim 1, characterized in that the timer activates the pulsed energy source during the first period of time of less than ten seconds and, preferably, less than one second. System according to claims 1 and 2, characterized in that the timer deactivates the pulsed energy source to interrupt the application of the pulsed energy source to the target tissue for a period of time ranging from several seconds to several minutes, and preferably three Petition 870190083984, of 08/28/2019, p. 159/162 2/3 seconds to three minutes and, more preferably, ten seconds to ninety seconds. System according to any of claims 1 to 3, characterized in that the parameters of the pulsed energy source are selected in such a way that the temperature of the target tissue is increased between six degrees Celsius to eleven degrees Celsius. System according to any of claims 1 to 4, characterized in that the timer activates the pulsed energy source to maintain the average temperature rise of the target tissue for several minutes at six degrees Celsius or less or, preferably, one degree Celsius or less for several minutes. 6. System according to any of claims 1 to 5, characterized in that the pulsed energy source comprises a beam of light, a microwave, a radio frequency or an ultrasound. 7. System according to claim 6, characterized in that a radio frequency is between approximately three and six megahertz, has a duty cycle between approximately 2.5% and 5% and a pulse train duration between approximately 0.2 and 0, 4 seconds. System according to claim 7, characterized in that it includes a device for generating the radio frequency having a coil radius between approximately 2 and 6 mm and between approximately 13 and turns of 57 ampere turns. 9. System according to claim 6, characterized in that the microwave frequency is between approximately 10 and 20 GHz, has a pulse train duration Petition 870190083984, of 08/28/2019, p. 160/162 3/3 between approximately 0.2 and 0.6 seconds, a duty cycle between approximately 2% and 5% and an average power between approximately 8 and 52 watts. 10. System according to claim 6 characterized by a wavelength between approximately 530 nm and 1,300 nm, a working time of less than a pulse train duration between approximately 0.1 and 0.6 seconds, and preferably where the pulsed light beam has a wavelength between 800 nm and 1,000 nm and a power between approximately 0.5 and 74 watts. 11. System according to claim 6, characterized in that the ultrasound has a frequency between approximately 1 MHz and 5 MHz, a train duration between approximately 0.1 and 0.5 seconds and a duty cycle between approximately 2% and 10% and a power between approximately 0.46 and 28.6 watts.