Uses of Bipolar Trans Carotenoids in the Treatment of Cancer

[0001] This application claims priority to U.S. Provisional Application No. 63/356,947 filed June 29, 2022, and U.S. Provisional Application No. 63/369,355 filed July 25, 2022, the contents of each of which are hereby incorporated by reference in their entireties.

Field

[0002] Provided are methods for the treatment of solid tumors, including glioblastoma multiforme (GBM). For instance, provided are diffusion enhancing compounds and their use in the treatment of solid tumors in conjunction with certain imaging modalities that measure the level of hypoxia within a tumor, radiation therapy (RT), chemotherapy, other anti-cancer therapies, or any combination of the foregoing.

Background

Cellular Respiration

[0003] The cells that make up our body’s tissues require a consistent supply of oxygen to generate sufficient energy to meet our body’s metabolic demands. Through a process known as cellular respiration, our cells combine oxygen from the air we breathe with glucose from the food we eat to generate this energy.

[0004] When oxygen in the air is inhaled through our lungs, it is transported through our body via the circulatory system, carried by the hemoglobin molecules in our red blood cells. Upon reaching the body’s tissues (e.g., skeletal muscle or organs) where there is demand for the oxygen, the oxygen is released by the hemoglobin molecules in the red blood cells and moves through the blood and blood plasma through a process known as diffusion. In this diffusion process, oxygen molecules move passively from the area of high oxygen concentration in the blood to the area of lower oxygen concentration in the surrounding tissue.

[0005] Our ability to deliver a sufficient supply of oxygen throughout our body in this manner depends on several factors - the oxygen concentration in the air we’re breathing; the efficiency of oxygen uptake into our blood from the lungs; our cardiac output (i.e., how effectively our heart pumps blood to circulate the oxygen); the concentration of red blood cells, which carry the oxygen-transporting hemoglobin, in our blood; the hemoglobin’s oxygen saturation level; and the efficiency of oxygen diffusion from our red blood cells, through the water molecules in the blood plasma, and into the surrounding tissue cells.

Hypoxia

[0006] Hypoxia is a condition in which, due to a breakdown or deficiency in this oxygen transport process, there is an insufficient supply of oxygen to meet the energy demands of the tissue cells. Hypoxia is associated with the pathophysiology of many of medicine’s most intractable and difficult-to-treat conditions, including cancers, cardiovascular diseases, cerebrovascular diseases, respiratory diseases, skin and soft tissue diseases, and neurodegenerative diseases, as tissues can only survive for brief periods of time, sometimes just minutes, under hypoxic conditions.

[0007] All currently approved treatments for hypoxia work by augmenting or supplementing the systemic availability of oxygen. For example, respirators use pressure to push oxygen across the lungs into the blood to increase oxygen concentration, certain medications and medical devices are designed to improve cardiac output, and blood transfusions are used to increase the concentration of red blood cells and oxygen-carrying hemoglobin molecules. While these treatments are effective in certain circumstances, there are significant associated risks, such as lung damage resulting from the use of respirators, toxic side effects and drug-to-drug interactions of medications that increase cardiac output, infections related to blood transfusions, and the risk of creating excessive oxygen-related tissue toxicity, or hyperoxia, by providing a patient with excessive amounts of oxygen. In addition, in certain indications complicated by localized hypoxia (such as cancerous tumors), these currently approved therapies focused on increasing the systemic availability of oxygen are ineffective in treating the localized hypoxia associated with the condition.

[0008] As a result, hypoxia remains a critical, complicating factor in many intractable and difficult-to-treat conditions affecting people of all ages - including hypoxic solid tumors and other cancers - and there continues to be significant unmet needs in the treatment of hypoxia, without causing hyperoxia, and conditions complicated hypoxia.

Hypoxic Solid Tumors

[0009] The American Cancer Society estimated that 1.9 million new cancer cases would be diagnosed in the U.S. during 2021 and, historically, solid tumors comprise approximately 90% of all adult cancer diagnoses. Cancerous tumors are specifically susceptible to developing hypoxia due to a combination of rapid growth, structural abnormalities of the tumor microvessels, and disturbed circulation within the tumor. Hypoxic conditions within the tumor microenvironment have negative consequences for patient outcomes and treatment, including:

• increased resistance to ionizing radiation, chemotherapy, immunotherapy, and other treatment methods;

• a more clinically aggressive phenotype;

• increased potential for invasive growth and tumor progression; and

• increased regional and distal tumor metastasis.

Accordingly, many of the challenges faced by practitioners in treating some of the most fatal and difficult-to-treat solid tumors, including GBM, are a product of the tumors’ highly hypoxic nature.

[0010] As a result, hypoxia remains a critical, complicating factor in many intractable and difficult-to-treat conditions affecting people of all ages - including hypoxic solid tumors and other cancers - and new methods for addressing hypoxia, without causing hyperoxia, are needed.

Summary

[0011] Provided is a method (Method 1) of treating cancer in a patient (e.g., a human) in need thereof, wherein the method comprises administering a therapeutically effective amount of a diffusion enhancing compound to the patient, and administering radiation therapy and/or chemotherapy.

[0012] Further provided is a method (Method 2) of treating glioma in a patient (e.g., a human) in need thereof, wherein the method comprises administering a therapeutically effective amount of a diffusion enhancing compound to the patient and administering radiation therapy and/or chemotherapy.

[0013] Further provided is a method (Method 3) of treating glioblastoma multiforme in a patient (e.g., a human) in need thereof, wherein the method comprises administering a therapeutically effective amount of a diffusion enhancing compound to the patient and administering radiation therapy and/or chemotherapy.

[0014] Further provided is a method (Method 4) of treating a solid tumor in a patient (e.g., a human) in need thereof, wherein the method comprises: (a) administering to the patient a first diffusion enhancing compound and optionally a positron emission tomography radiotracer;

(b) acquiring a measure of the solid tumor hypoxia (e.g., by one or more of polarographic electrode, a fiber-optic probe, positron emission tomography, magnetic resonance imaging, electron paramagnetic resonance oximetry, and/or endogenous contrast magnetic resonance);

(c) developing a radiation therapy protocol for the patient; and

(d) administering to the patient radiation therapy according to the protocol, wherein optionally a second diffusion enhancing compound, which may be the same or different from the first diffusion enhancing compound, is administered before the radiation therapy.

[0015] Further provided is a method (Method 5) of treating a solid tumor in a patient (e g., a human) in need thereof, wherein the method comprises:

(a) acquiring a first measure of the solid tumor hypoxia (e.g., by one or more of polarographic electrode, a fiber-optic probe, positron emission tomography, magnetic resonance imaging, electron paramagnetic resonance oximetry, and/or endogenous contrast magnetic resonance);

(b) administering to the patient a first diffusion enhancing compound and optionally a positron emission tomography radiotracer,

(c) acquiring a second measure of the solid tumor hypoxia (e.g., by one or more of polarographic electrode, a fiber-optic probe, positron emission tomography, magnetic resonance imaging, electron paramagnetic resonance oximetry, and/or endogenous contrast magnetic resonance) following administration of the diffusion enhancing compound and optionally the positron emission tomography radiotracer;

(d) developing a radiation therapy protocol for the patient; and

(e) administering to the patient radiation therapy according to the protocol, wherein optionally a second diffusion enhancing compound, which may be the same or different from the first diffusion enhancing compound, is administered before the radiation therapy.

Detailed Description [0016] The subject disclosure relates to diffusion enhancing compounds and novel methods of their use in the treatment of solid tumors, alone or in conjunction with certain imaging modalities that measure the level of hypoxia within a tumor, radiation therapy (RT), chemotherapy, other anti-cancer therapies, or any combination of the foregoing.

[0017] As used herein, patient includes human and non-human. Preferably, a patient is human.

[0018] Cancerous tumors are susceptible to developing hypoxia due to a combination of rapid growth, structural abnormalities of the tumor micro-vessels, and disturbed circulation within the tumor.

[0019] Since current standard of care for GBM was established, consisting of surgical resection or biopsy, followed by radiotherapy with concomitant temozolomide chemotherapy, followed by maintenance temozolomide for 6 to 12 months, little progress has been made in the treatment of GBM. Currently, therapeutic modalities are insufficient to control high-grade glioma progression overall due to its aggressive nature. Surgical resection is not always possible, and regrowth of tumors as well as acute morbidity is likely. Radiotherapy has high toxicity, and re-treating recurrences carries a higher risk due to loss in neurogeneration potential. Although chemotherapy has been shown to improve prognosis and time of progression, there are important drawbacks to this treatment modality, including bypassing the blood brain barrier, interactions with anti-seizure medications, resistance to therapy, recurrent tumors, and intrinsic or acquired resistance.

[0020] Tumor oxygenation may be assessed by fiber optic probes, magnetic resonance imaging, electron paramagnetic resonance (EPR) oximetry, endogenous contrast magnetic resonance, polarographic electrodes, and positron emission tomography (PET).

[0021] Positron emission tomography requires injection of a radiotracer (e.g., nitroimidazole) that diffuses into cells and is then reduced intracellularly. This intracellular process is reversible under normoxic conditions but, under hypoxic conditions, the radiolabeled molecules are trapped and react with cellular macromolecules such as nucleic acids and proteins, as the reduction requires the activity of reductases that are only present in viable hypoxic cells. Accordingly, the accumulation and detection of radiotracers is enhanced in hypoxic regions, whereas the necrotic cells will not be visible to PET imaging. The quantification of the tracer uptake is generally expressed as the tumor-to-background (TBR) ratio at a given time after the tracer injection. 2-Nitroimidazoles have been developed as radiotracers because they have a nitro (NO2) group linked to the imidazole structure and, as a result, can undergo up to six electron reductions, eventually resulting in an amino group (NH ₂ ). For PET imaging, these tracers are labeled with radioisotopes - fluorine-18 ( ¹⁸ F) or carbon-11 ( ⁿ C). 18F-Fluoromisonidazole (F- MISO) and 18F-Fluoroazomycin- Arabinoside (F-FAZA) are examples of radiotracers. With F- MISO, a TBR ratio of 1.2 may delineate regions of hypoxia after a minimum delay of 2 hours with the best signal -to-noise ratio generally observed approximately 4 hours after tracer injection.

[0022] Because tumor hypoxia is a factor that impacts the efficacy of radiotherapy and chemotherapy, oxygen mapping of a tumor, for instance, with PET, may be particularly useful with intensity modulated radiation therapy (IMRT) since the hypoxic regions of a tumor are well delineated, thus making it possible to ensure those regions receive higher radiation doses and the most oxygenated regions lower radiation doses.

[0023] Administering a diffusion enhancing compound, such as trans sodium crocetinate (TSC), in combination with oxygen mapping of a tumor may allow for even more accurate and detailed information about the tumor microenvironment, for instance, regions most in need of higher radiation doses or potentially most vulnerable to radiation doses, thereby allowing for the development of a personalized, directed, and ultimately effective treatment regimen that spares healthy tissue. Further, a diffusion enhancing compound would contribute to increased oxygenation of the hypoxic cells, thereby enhancing potency of the radiation.

[0024] Previous work with trans sodium crocetinate (TSC) reported that it exhibited a U- shaped dosage curve. Specifically, Manabe et al., J Neurosurgery, 2010, 113 (4), 802-809, in testing the effect of TSC on cerebral infarct volume in a model of permanent (24-hour) focal ischemia at dosages ranging from 0.023 to 4.580 mg/kg, found maximal protective effect at an intermediate dosage of 0.092 mg/kg. Mohler et al., Vase Med., 2011, 16 (5), 346-353, in administering TSC ranging from 0.25 mg/kg to 2.0 mg/kg to patients with peripheral artery disease, found a peak increase in peak walking time after 5 days of treatment in the 1.50 mg/kg dosing arm. Mohler reported the greatest increase in claudication onset time was observed at the lowest dose of TSC of 0.25 mg/kg. U.S. Patent No. 8,030,350 reported that for HCT116 human colon carcinoma tumors on mice, TSC tested at doses 0.07 mg/kg, 0.14 mg/kg, 0.18 mg/kg, 0.28 mg/kg, 0.54 mg/kg, and 1 .35 mg/kg with radiotherapy, optimal doses were 0.07 mg/kg, 0.14 mg/kg, and 0.18 mg/kg. U.S. Patent No. 11,185,523 discusses a “low” dose range of 0.15-0.35 mg/kg and a “high” dose range of 0.75-2.0 mg/kg for humans.

[0025] Applicant has now found that administration of trans sodium crocetinate unexpectedly enhances delivery of oxygen to hypoxic solid tumors in humans more effectively at higher dosages without causing hyperoxia.

[0026] The methods described herein include administration of a therapeutically effective amount of a diffusion enhancing compound such as TSC.

[0027] Diffusion enhancing compounds for use in the methods described herein include bipolar trans carotenoid salts having the formula:

YZ-TCRO-ZY, where:

Y = a cation which can be the same or different,

Z = a polar group which can the same or different and which is associated with the cation,

TCRO = a linear trans carotenoid skeleton with conjugated carbon-carbon double bonds and single bonds, and having pendant groups X, wherein the pendant groups X, which can be the same or different, are a linear or branched hydrocarbon group having 10 or less carbon atoms, or a halogen.

[0028] Advantageously, the bipolar trans carotenoid is the all trans form of crocetin (trans-crocetin), which may be in acid or pharmaceutically acceptable salt form. Trans sodium crocetinate (TSC) (e.g., synthetic TSC) is shown as Formula I below.

Formula I

[0029] In one embodiment, the absorbency (e.g., in an aqueous solution) of the bipolar trans carotenoid salt (e.g., trans sodium crocetinate) at the highest peak which occurs in the visible wavelength range divided by the absorbency of a peak occurring in the ultraviolet wavelength range is greater than 7 (e.g., 7 to 8.5), e.g., greater than 7.5 (e.g., 7.5 to 9, e.g., 7.5 to 8.5), e.g., greater than 8 (e.g., 8 to 8.8), e.g., greater than 8.5. In another embodiment, the absorbency (e.g., in an aqueous solution) of the TSC at the highest peak which occurs in the visible wavelength range divided by the absorbency of a peak occurring in the ultraviolet wavelength range is greater than 7 (e.g., 7 to 8.5), e.g., greater than 7.5 (e.g., 7.5 to 9, e.g., 7.5 to 8.5), e.g., greater than 8 (e.g., 8 to 8.8), e.g., greater than 8.5.

[0030] The bipolar trans carotenoid salt (e.g., trans sodium crocetinate) is at least 90% pure as measured by high performance liquid chromatography (HPLC), e.g., > 95% pure as measured by HPLC, e.g., > 96% pure as measured by HPLC. In an advantageous embodiment, the TSC is at least 90% pure as measured by high performance liquid chromatography (HPLC), e.g., > 95% pure as measured by HPLC, e.g., > 96% pure as measured by HPLC.

[0031] In an advantageous embodiment, the bipolar trans carotenoid salt is in a composition also comprising a cyclodextrin. For instance, wherein TSC is in a composition also comprising a cyclodextrin (e.g., wherein the TSC is in a lyophilized composition with a cyclodextrin).

[0032] Advantageously, the cyclodextrin is gamma-cyclodextrin. For instance, the bipolar trans carotenoid salt is TSC which is in a composition also comprising gamma-cyclodextrin (e.g., wherein the TSC is in a lyophilized composition with gamma-cyclodextrin).

[0033] In an embodiment of the invention, the composition further comprises mannitol.

[0034] The diffusion enhancing compound is administered intravenously or intramuscularly (e.g., as an intravenous injection or infusion or intramuscular injection).

[0035] Advantageously, the diffusion enhancing compound is admixed with sterile water for injection to form an injection. TSC is administered intravenously or intramuscularly (e.g., as an intravenous injection or infusion or intramuscular injection). For instance, wherein TSC is admixed with sterile water for injection to form an injection.

[0036] Advantageously, the diffusion enhancing compound is TSC and is administered at a dose of 2-2.5 mg/kg, or 2.5-5 mg/kg, e.g., 3-5 mg/kg.

[0037] In another embodiment, provided is a diffusion enhancing compound (e g., a bipolar trans carotenoid salt (e.g., TSC)) for use in any of the methods described herein. [0038] In another embodiment, provided is use of a diffusion enhancing compound (e.g., a bipolar trans carotenoid salt (e.g., TSC)) in the manufacture of a medicament for any of the methods described herein.

[0039] In another embodiment, provided is a pharmaceutical composition comprising an effective amount of a diffusion enhancing compound (e.g., a bipolar trans carotenoid salt (e.g., TSC)) for use in any of the methods described herein.

Compounds and Compositions of the Invention

Diffusion Enhancing Compounds

[0040] Diffusion enhancing compounds include those compounds described in U.S. Pat. 7,759,506, U.S. Pat. 8,030,350, U.S. Pat. 8,901,174, U.S. Pat 8,206,751, and U.S. Pat.

11,185,523, each of which is hereby incorporated by reference in its entirety.

[0041] Included are bipolar trans carotenoid compounds having the formula: YZ-TCRO-ZY where:

• Y=a cation

• Z=a polar group which is associated with the cation, and

• TCRO=trans carotenoid skeleton, such as TSC.

[0042] More specifically, the subject invention relates to trans carotenoids including trans carotenoid diesters, dialcohols, diketones and diacids, bipolar trans carotenoids (BTC), and bipolar trans carotenoid salts (BTCS) and uses of such compounds having the structure: YZ-TCRO-ZY where:

• Y (which can be the same or different at the two ends)=H or a cation other than H, preferably Na ⁺ or K ⁺ or Li ⁺ . Y is advantageously a monovalent metal ion. Y can also be an organic cation, e.g., IUN ⁺ , RsS ⁺ , where R is H, or CtTUn+i where n is 1-10, advantageously 1- 6. For example, R can be methyl, ethyl, propyl or butyl.

• Z (which can be the same or different at the two ends)=polar group which is associated with H or the cation. Optionally including the terminal carbon on the carotenoid (or carotenoid related compound), this group can be a carboxyl (COO-) group or a CO group (e.g. ester, aldehyde or ketone group), or a hydroxyl group. This group can also be a sulfate group (OSO3 ) or a monophosphate group (OPO3 ), (OP(OH)O ₂ ~), a diphosphate group, triphosphate or combinations thereof. This group can also be an ester group of COOR where the R is C _n H _2n +i.

• TCRO=trans carotenoid or carotenoid related skeleton (advantageously less than 100 carbons) which is linear, has pendant groups (defined below), and typically comprises “conjugated” or alternating carbon-carbon double and single bonds (in one embodiment, the TCRO is not fully conjugated as in a lycopene). The pendant groups (X) are typically methyl groups but can be other groups as discussed below. In an advantageous embodiment, the units of the skeleton are joined in such a manner that their arrangement is reversed at the center of the molecule. The 4 single bonds that surround a carbon-carbon double bond all lie in the same plane. If the pendant groups are on the same side of the carbon-carbon double bond, the groups are designated as cis (also known as “Z”); if they are on the opposite side of the carbon-carbon bond, they are designated as trans (also known as “E”). Throughout this case, the isomers will be referred to as cis and trans.

[0043] The compounds of the subject invention are trans. The cis isomer typically is a detriment — and results in the diffusivity not being increased. In one embodiment, a cis isomer can be utilized where the skeleton remains linear. The placement of the pendant groups can be symmetric relative to the central point of the molecule or can be asymmetric so that the left side of the molecule does not look the same as the right side of the molecule either in terms of the type of pendant group or their spatial relationship with respect to the center carbon.

[0044] The pendant groups X (which can be the same or different) are hydrogen (H) atoms, or a linear or branched hydrocarbon group having 10 or less carbons, advantageously 4 or less, (optionally containing a halogen), or a halogen. X could also be an ester group (COO — ) or an ethoxy/methoxy group. Examples of X are a methyl group (CH3), an ethyl group (C2H5), a phenyl or single aromatic ring structure with or without pendant groups from the ring, a halogencontaining alkyl group (Cl -CIO) such as CH ₂ C1, or a halogen such as Cl or Br or a methoxy (OCH3) or ethoxy (OCH ₂ CH ₃ ). The pendant groups can be the same or different but the pendant groups utilized must maintain the skeleton as linear.

[0045] Although many carotenoids exist in nature, carotenoid salts do not. Commonly- owned U.S. Pat. No. 6,060,511 hereby incorporated by reference in its entirety, relates to trans sodium crocetinate (TSC). The TSC was made by reacting naturally occurring saffron with sodium hydroxide followed by extractions that selected primarily for the trans isomer.

[0046] Naturally occurring carotenoid compounds are composed of both cis and trans isomers. The presence of the cis and trans isomers of a carotenoid or carotenoid salt can be determined by looking at the ultraviolet-visible spectrum for the carotenoid sample dissolved in an aqueous solution. Given the spectrum, the value of the absorbence of the highest peak which occurs in the visible wave length range of 380 to 470 nm (the number depending on the solvent used and the chain length of the BTC or BTCS. The addition of pendant groups or differing chain lengths will change this peak absorbance but someone skilled in the art will recognize the existence of an absorbance peak in the visible range corresponding to the conjugated backbone structure of these molecules.) is divided by the absorbency of the peak which occurs in the UV wave length range of 220 to 300 nm can be used to determine the purity level of the trans isomer. When the trans carotenoid diester (TCD) or BTCS is dissolved in water, the highest visible wave length range peak will be at between 380 nm to 470 nm (depending on the exact chemical structure, backbone length and pendant groups) and the UV wave length range peak will be between 220 to 300 nm. According to M. Craw and C. Lambert, Photochemistry and Photobiology, Vol. 38 (2), 241-243 (1983) hereby incorporated by reference in its entirety, the result of the calculation (in that case crocetin was analyzed) was 3.1, which increased to 6.6 after purification.

[0047] Performing the Craw and Lambert analysis, using a cuvette designed for UV and visible wavelength ranges, on the trans sodium salt of crocetin of commonly owned U.S. Pat. No. 6,060,511 (TSC made by reacting naturally occurring saffron with sodium hydroxide followed by extractions which selected primarily for the trans isomer), the value obtained averages about 6.8. Performing that test on the synthetic TSC of the subject invention, that ratio is greater than 7.0 (e g. 7.0 to 8.5), advantageously greater than 7.5 (e.g. 7.5-8.5), most advantageously greater than 8. The synthesized material is a “purer” or highly purified trans isomer.

Formulation and Administration

[0048] A detailed description of formulation and administration of diffusing enhancing compounds can be found in commonly owned U.S. Patent 8,293,804, U.S. application 12/801,726, and U.S. Patent 8,206,751, each of which is hereby incorporated by reference in its entirety. A detailed description of formulation and administration of diffusing enhancing compounds can also be found in commonly owned U.S. Patent No. 8,030,350, which is hereby incorporated by reference in its entirety.

[0049] A diffusion enhancing compound such as TSC can be administered by various routes for rapid delivery to the hypoxic tissue. For example, the compound, which can be formulated with other compounds including excipients, can be administered at the proper dosage as an intravenous injection (IV) or infusion, or an intramuscular injection (IM).

[0050] The IV injection route is an advantageous route for giving TSC for many of the uses of the subject application.

[0051] In addition to intravenous injection, routes of administration for specially formulated trans carotenoid molecules include intramuscular injection, delivery by inhalation, oral administration and transdermal administration.

Cyclodextrins

[0052] Naturally occurring carotenoid compounds present a variety of additional obstacles in terms of large scale applications as a pharmaceutical for enhancing blood diffusivity as well. One problem is the preparation of the active agent in sufficiently pure and large amounts. Another, more serious problem is the fact that crocetin is nearly insoluble in aqueous solutions, making preparation of a drug for intravenous administration particularly difficult. Finally, reaching and maintaining adequate levels of the trans isomer to improve oxygen diffusivity over a length of time proves difficult.

[0053] In order to administer some pharmaceuticals, it is necessary to add another compound which will aid in increasing the absorption/solubility/concentration of the active pharmaceutical ingredient (API). Such compounds are called excipients, and cyclodextrins are examples of excipients. Cyclodextrins are cyclic carbohydrate chains derived from starch. They differ from one another by the number of glucopyranose units in their structure. The parent cyclodextrins contain six, seven and eight glucopyranose units, and are referred to as alpha, beta and gamma cyclodextrins respectively. Cyclodextrins were first discovered in 1891, and have been used as part of pharmaceutical preparations for several years.

[0054] Cyclodextrins are cyclic (alpha- l,4)-linked oligosaccharides of alpha-D-gluco- pyranose containing a relatively hydrophobic central cavity and hydrophilic outer surface. In the pharmaceutical industry, cyclodextrins have mainly been used as complexing agents to increase the aqueous solubility of poorly water-soluble drugs, and to increase their bioavailability and stability. In addition, cyclodextrins are used to reduce or prevent gastrointestinal or ocular irritation, reduce or eliminate unpleasant smells or tastes, prevent drug-drug or drug-additive interactions, or even to convert oils and liquid drugs into microcrystalline or amorphous powders.

[0055] There are a number of cyclodextrins that can be used with the diffusion enhancing compounds dislcosed herein. See for example, U.S. Pat. No. 4,727,064, hereby incorporated by reference in its entirety. Advantageous cyclodextrins are y-cyclodextrin, 2-hydroxylpropyl-y- cyclodextrin and 2-hydroxylpropyl-P-cyclodextrin, or other cyclodextrins which enhance the solubility of the BTC.

[0056] The use of gamma-cyclodextrin with TSC increases the solubility of TSC in water by 3-7 times. Although this is not as large a factor as seen in some other cases for increasing the solubility of an active agent with a cyclodextrin, it is important in allowing for the parenteral administration of TSC in smaller volume dosages to humans (or animals). Dosages of TSC and gamma-cyclodextrin have resulted in aqueous solutions containing as much as 44 milligrams of TSC per ml of solution, with an advantageous range of 20-30 mg/ml of solution. The solutions need not be equal-molar. The incorporation of the gamma cyclodextrin also allows for TSC to be absorbed into the blood stream when injected intramuscularly. Absorption is quick, and efficacious blood levels of TSC are reached quickly (as shown in rats).

[0057] The cyclodextrin formulation can be used with other trans carotenoids and carotenoid salts. The subject invention also includes novel compositions of carotenoids which are not salts (e.g. acid forms such as crocetin, crocin or the intermediate compounds noted above) and a cyclodextrin. In other words, trans carotenoids which are not salts can be formulated with a cyclodextrin. Mannitol can be added for osmolality, or the cyclodextrin BTC mixture can be added to isotonic saline (see below).

[0058] The amount of the cyclodextrin used is that amount which will contain the trans carotenoid but not so much that it will not release the trans carotenoid. Advantageously, the ratio of cyclodextrin to BTC, e.g., TSC, is 4 to 1 or 5 to 1. See also U.S. Patent No. 8,974,822, the content of which is hereby incorporated by reference in its entirety.

Cyclodextrin-Mannitol

[0059] A trans carotenoid such as TSC can be formulated with a cyclodextrin as noted above and a non -metabolized sugar such as mannitol (e.g. d-mannitol to adjust the osmotic pressure to be the same as that of blood). Solutions containing over 20 mg TSC/ml of solution can be made this way. This solution can be added to isotonic saline or to other isotonic solutions in order to dilute it and still maintain the proper osmolality.

Mannitol/Acetic Acid

[0060] A BTCS such as TSC can be formulated with mannitol such as d-mannitol, and a mild buffering agent such as acetic acid or citric acid to adjust the pH. The pH of the solution should be around 8 to 8.5. It should be close to being an isotonic solution, and, as such, can be injected directly into the blood stream.

Water+Saline

[0061] A BTCS such as TSC can be dissolved in water (advantageously injectable water). This solution can then be diluted with water, normal saline, Ringer’s lactate or phosphate buffer, and the resulting mixture either infused or injected.

Buffers

[0062] A buffer such as glycine, bicarbonate, or sodium carbonate can be added to the formulation at a level of about 50 mM for stability of the BCT such as TSC.

TSC and Gamma-Cyclodextrin

[0063] The ratio of TSC to cyclodextrin is based on TSC:cyclodextrin solubility data. For example, 20 mg/ml TSC, 8% gamma cyclodextrin, 50 mM glycine, 2.33% mannitol with pH 8.2+/-0.5, or 10 mg/ml TSC and 4% cyclodextrin, or 5 mg/ml and 2% cyclodextrin. The ratios of these ingredients can be altered somewhat, as is obvious to one skilled in this art.

[0064] Mannitol can be used to adjust osmolality and its concentration varies depending on the concentration of other ingredients. The glycine is held constant. TSC is more stable at higher pHs. pH of around 8.2+/-0.5 is required for stability and physiological compatibility. The use of glycine is compatible with lyophilization. Alternatively, the TSC and cyclodextrin is formulated using a 50 mM bicarbonate buffer in place of the glycine.

Endotoxin Removal of Gamma-Cyclodextrin

[0065] Commercially available pharmaceutical grade cyclodextrin has endotoxin levels that are incompatible with intravenous injection. The endotoxin levels must be reduced in order to use the cyclodextrin in a BTC formulation intended for intravenous injection.

Lyophilization

[0066] Lyophilization can be used to produce an easily reconstituted injectable solution. Kits and Dual Chamber Delivery Systems

[0067] The diffusion enhancing compound such as TSC can be lyophilized and put in a vial which can be part of a vial kit system which also includes a vial with diluent such as water for injection, and a syringe for administration.

[0068] Dual-chamber delivery systems allow reconstitution of the lyophilized diffusion enhancing compound directly inside the system be it a syringe or a cartridge. The lyophilized diffusion enhancing compound such as TSC is located in one chamber and the diluent (e.g. water for injection) in the other. The drug is reconstituted j ust before administration. It is a simple and controllable process completed in a few easy steps.

[0069] In one embodiment, the diffusion enhancing compound such as TSC is loaded in an auto-injector. An auto-injector (or auto-injector) is a medical device designed to deliver a dose of a particular drug. Most auto-injectors are spring-loaded syringes. By design, autoinjectors are easy to use and are intended for self-administration by patients, or administration by untrained personnel. The site of injection is typically the thigh or the buttocks. The auto-injector typically keeps the needle tip shielded prior to injection and also has a passive safety mechanism to prevent accidental firing (injection). Injection depth can be adjustable or fixed and a function for needle shield removal can be incorporated. Just by pressing a button, the syringe needle is automatically inserted and the drug is delivered.

Methods of Treatment

GBM and Other Cancers

[0070] The subject disclosure relates to the treatment of various tumors and/or cancers, in particular GBM. Tumors are hypoxic with many tumor types being highly hypoxic. Hypoxic tumors are more resistant to radiotherapy and chemotherapy and, in GBM in particular, tumoral hypoxia correlates with worst outcomes. Through HIF1 alpha up-regulation, hypoxia is associated with multiple negative effects that lead to aggressive tumor phenotypes. These effects include increased angiogenesis, increased metastasis, as well as increased resistance to chemotherapy and radiation therapy. Hypoxia via HIFla affects many genes involved in cancer progression. Bipolar trans carotenoids such as TSC alter expression of HIF1 targeted genes in hypoxic conditions. For example, studies have shown that the VEGF A gene, which is upregulated with hypoxia and shows increased expression in GBM tumors, is down regulated with TSC. [0071] Given the resulting increased tumor aggressiveness, metastatic spread, resistance to therapy, rate of recurrence, and decreased local control and survival, hypoxia measurements may improve treatment planning and early assessment of efficacy in GBM. An imaging method (e.g., PET tracers, e.g., F-MISO or F-FAZA) is a desirable approach for measuring hypoxia because it is noninvasive, can provide high spatial resolution, has reasonable cost, and is easy to use in clinical trials. Imaging modalities may also provide additional information regarding one or more of parameters related to reliable biomarkers for grading tumors, predicting malignant transformation, planning treatments, and monitoring responses.

[0072] In addition, administration of a bipolar trans carotenoid such as TSC can enhance the cytotoxicity of chemotherapeutic agents in a tumor, as well as reduce or treat the neurotoxicity or neuropathy that the chemotherapy agents can cause.

[0073] The methods of the subject disclosure are directed to administering a dose of a bipolar trans carotenoid (such as TSC) at a dose and at the proper time prior to administration of radiation therapy, chemotherapy, and/or a PET imaging tracer such as F-MISO or F-FAZA (in each case, as discussed above) to assist in mapping the hypoxic regions of the tumor to provide more personalized and effective treatment to the patient. The administration of the bipolar trans carotenoid, due to its hypoxia reducing ability, may also decrease angiogenesis, decrease metastasis, and down regulate HIFla production in the tumor.

GBM Radiation Therapy

[0074] Target volume for radiation therapy may be based upon postoperative-enhanced MRI. Preoperative imaging may be used for correlation and improvied identification. Planning target volumes for radiation may extend beyond bony margins and the skin surface. Planning target volume (PTV) may be determined by adding a margin to clinical target volume (CTV) to account for variation in beam alignment, patient position, organ motion, and other movement. [0075] Initial gross tumor volume (GTV1) may be determined by either the T2 or FLAIR abnormality on a post-operative MRI scane. GTV1 may include all postoperative-enhanced MRI enhancement, and the surgical cavity. Areas of vascular infarction or compromise of normal brain should be excluded if they are deemed to not be part of the original pre-operative tumor volume. Comparing pre- and post-operative scans will assist in that process.

[0076] Initial clinical target volume (CTV1) may be GTV1 plus a margin of 1 cm or 2 cm (e g , 2 cm), which may be reduced around natural barriers to tumor growth such as the skull, ventricle, falx, etc. to as low as 0 mm to “fixed” barriers, i.e., bone, and falx and as low as 3-5 mm to “non-rigid” barriers such as brain stem, ventricles, etc. If no surrounding edema is present (e.g., the FLAIR and/or T2 MR images do not demonstrate any significant abnormality), the CTV 1 in those instances may include the post-operative MRI enhancement and the surgical resection cavity, plus a 2 cm margin, with possible reduction as described above. If no surrounding edema is present, the CTV1 may include the contrast-enhancing lesion plus a 2.5 cm margin.

[0077] Initial planning target volume (PTV1) may be CTV1 plus an additional margin of 1 to 5 mm (e.g., 3 to 5 mm, e.g., 3 mm), depending upon localization method and reproducibility at each center. PTV margins account for variations in set-up and reproducibility.

[0078] The boost gross tumor volume (GTV2) may be the contrast enhanced T1 abnormality on the post-operative MRI scan. This can also include the surgical cavity margins. Areas of vascular infarction or compromise of normal brain should be excluded if they are deemed to not be part of the original pre-operative tumor volume, and a comparison of the pre- and post-operative scans will assist in this process. The exception to this is the polar tumor, e.g., temporal lobe, frontal lobe, or occipital lobe tip, where a gross total resection is achieved, and no post-operative “cavity” remains, consequential to the partial or total lobectomy.

[0079] The boost clinical target volume (CTV2) may be the GTV2 plus a margin of 1 or 2 cm (e.g., 2 cm). The CTV2 margin may be reduced around natural barriers to tumor growth such as the skull, ventricles, falx, etc. to as low as 0 mm to “fixed” barriers, i.e., bone, and falx and as low as 3-5 mm to “non-rigid” barriers such as brain stem, ventricles, etc.

[0080] The boost or conedown planning target volume (PTV2) may be CTV2 plus an additional margin of 1 to 5 mm (e.g., 3 to 5 mm, e.g., 3 mm), depending upon localization method and reproducibility at each center. PTV margins account for variations in set-up and reproducibility.

[0081] Reducing PTV margins to modify organ at risk (OAR) dose(s) is not generally permissible. However, OAR may be defined, along with a planning risk volume (PRV) for each OAR. Each PRV may be its OAR plus 3 mm. If an OAR is in immediate proximity to a PTV such that dose to the OAR cannot be constrained within protocol limits, a second PTV (PTV overlap) may be defined as the overlap between the planned target volume (PTV1 or PTV2) and the particular PRV of concern. The overlap may be the intersection between PTV1 or PTV2 and the PRV. Dose to the PTV overlap should be as close as permissible to target dose for PTV1 (e.g., 46 Gy) or PTV2 (e.g., 14 Gy ) while not exceeding the OAR dose limit.

Uses of the Compounds and Compositions of the Invention

[0082] The diffusion enhancing compound can be administered by various routes. For example, the compound (which can be formulated with other compounds), can be administered at the proper dosage as an intravenous injection or infusion, an intramuscular injection, or in an oral form. The IV injection route is an advantageous route for giving a diffusion enhancing compound such as TSC.

[0083] Provided is a method (Method 1) of treating cancer in a patient (e.g., a human) in need thereof, wherein the method comprises administering a therapeutically effective amount of a diffusion enhancing compound to the patient, and administering radiation therapy and/or chemotherapy.

[0084] Further provided is a method (Method 2) of treating glioma in a patient (e.g., a human) in need thereof, wherein the method comprises administering a therapeutically effective amount of a diffusion enhancing compound to the patient and administering radiation therapy and/or chemotherapy.

[0085] Further provided is a method (Method 3) of treating glioblastoma multiforme in a patient (e.g., a human) in need thereof, wherein the method comprises administering a therapeutically effective amount of a diffusion enhancing compound to the patient and administering radiation therapy and/or chemotherapy.

[0086] Further provided is a method (Method 4) of treating a solid tumor in a patient (e.g., a human) in need thereof, wherein the method comprises:

(a) administering to the patient a first diffusion enhancing compound and optionally a positron emission tomography radiotracer;

(b) acquiring a measure of the solid tumor hypoxia (e.g., by one or more of polarographic electrode, a fiber-optic probe, positron emission tomography, magnetic resonance imaging, electron paramagnetic resonance oximetry, and/or endogenous contrast magnetic resonance);

(c) developing a radiation therapy protocol for the patient; and

(d) administering to the patient radiation therapy according to the protocol, wherein optionally a second diffusion enhancing compound, which may be the same or different from the first diffusion enhancing compound, is administered before the radiation therapy.

[0087] Further provided is a method (Method 5) of treating a solid tumor in a patient (e.g., a human) in need thereof, wherein the method comprises:

(a) acquiring a first measure of the solid tumor hypoxia (e.g., by one or more of polarographic electrode, a fiber-optic probe, positron emission tomography, magnetic resonance imaging, electron paramagnetic resonance oximetry, and/or endogenous contrast magnetic resonance),

(b) administering to the patient a first diffusion enhancing compound and optionally a positron emission tomography radiotracer;

(c) acquiring a second measure of the solid tumor hypoxia (e.g., by one or more of polarographic electrode, a fiber-optic probe, positron emission tomography, magnetic resonance imaging, electron paramagnetic resonance oximetry, and/or endogenous contrast magnetic resonance) following administration of the diffusion enhancing compound and optionally the positron emission tomography radiotracer;

(d) developing a radiation therapy protocol for the patient; and

(e) administering to the patient radiation therapy according to the protocol, wherein optionally a second diffusion enhancing compound, which may be the same or different from the first diffusion enhancing compound, is administered before the radiation therapy.

[0088] Further provided are any one of Methods 1-5 as follows:

1.1. Any one of Methods 1-5, wherein the diffusion enhancing compound (e.g., first diffusion enhancing compound and/or second diffusion enhancing compound) is a bipolar trans carotenoid salt having the formula:

YZ-TCRO-ZY, where:

Y = a cation which can be the same or different,

Z = a polar group which can the same or different and which is associated with the cation,

TCRO = a linear trans carotenoid skeleton with conjugated carbon-carbon double bonds and single bonds, and having pendant groups X, wherein the pendant groups X, which can be the same or different, are a linear or branched hydrocarbon group having 10 or less carbon atoms, or a halogen.

1.2. Any one of Methods 1-5 or 1.1, wherein the diffusion enhancing compound (e.g., first diffusion enhancing compound and/or second diffusion enhancing compound) is trans- crocetin, in acid or pharmaceutically acceptable salt form. For instance, any one of Methods 1-5 or 1.1, wherein the diffusion enhancing compound (e.g., first diffusion enhancing compound and/or second diffusion enhancing compound) is trans sodium crocetinate (TSC) (e g., synthetic TSC). For instance, any one of Methods 1-5 or 1.1, wherein the diffusion enhancing compound is trans sodium crocetinate (TSC) (e.g., synthetic TSC) (e.g., the first and second diffusion enhancing compounds are the same and are TSC (e.g., synthetic TSC)).

1.3. Any one of Methods 1-5, 1.1, or 1.2, wherein the absorbency (e.g., in an aqueous solution) of the bipolar trans carotenoid salt (e g., trans sodium crocetinate) at the highest peak which occurs in the visible wavelength range divided by the absorbency of a peak occurring in the ultraviolet wavelength range is equal to or greater than 7 (e.g., 7 to 8.5), e.g., equal to or greater than 7.5 (e.g., 7.5 to 9, e.g., 7.5 to 8.5), e.g., equal to or greater than 8

(e g., 8 to 8.8), e g., greater than 8.5. For instance, wherein the absorbency (e.g., in aqueous solution) of the highest peak which occurs in the visible wavelength range of 380 to 470 nm divided by the absorbency of the peak which occurs in the UV wavelength range of 220 to 300 nm is equal to or greater than 7 (e.g., 7 to 8.5), e.g., equal to or greater than 7.5 (e.g., 7.5 to 9, e.g., 7.5 to 8.5), e.g., equal to or greater than 8 (e.g., 8 to 8.8), e.g., greater than 8.5.

1.4. Any one of Methods 1-5 or 1.1-1.3, wherein the absorbency (e.g., in an aqueous solution) of the TSC at the highest peak which occurs in the visible wavelength range divided by the absorbency of a peak occurring in the ultraviolet wavelength range is equal to or greater than 7 (e.g., 7 to 8.5), e.g., equal to or greater than 7.5 (e.g., 7.5 to 9, e.g., 7.5 to 8.5), e.g., equal to or greater than 8 (e.g., 8 to 8.8), e.g., greater than 8.5. For instance, wherein the absorbency (e.g., in aqueous solution) of the TSC at the highest peak which occurs in the visible wavelength range of 380 to 470 nm divided by the absorbency of the peak which occurs in the UV wavelength range of 220 to 300 nm is equal to or greater than 7 (e.g., 7 to 8.5), e.g., equal to or greater than 7.5 (e.g., 7.5 to 9, e.g., 7.5 to 8.5), e.g., equal to or greater than 8 (e.g., 8 to 8.8), e.g., greater than 8.5.

1 .5. Any one of Methods 1 -5 or 1 .1-1 .4, wherein the bipolar trans carotenoid salt (e.g., trans sodium crocetinate) is at least 90% pure as measured by high performance liquid chromatography (HPLC), e.g., > 95% pure as measured by HPLC, e.g., > 96% pure as measured by HPLC.

1.6. Any one of Methods 1-5 or 1.1-1.5, wherein the TSC is at least 90% pure as measured by high performance liquid chromatography (HPLC), e.g., > 95% pure as measured by HPLC, e.g., > 96% pure as measured by HPLC.

1.7. Any one of Methods 1-5 or 1.1-1.6, wherein the bipolar trans carotenoid salt is in a composition also comprising a cyclodextrin. For instance, wherein TSC is in a composition also comprising a cyclodextrin (e.g., wherein the TSC is in a lyophilized composition with a cyclodextrin).

1.8. Method 1.7, wherein the cyclodextrin is selected from the group consisting of alpha cyclodextrin, beta cyclodextrin, 2-hydroxypropyl-beta-cyclodextrin, 2-hydroxypropyl- gamma-cyclodextrin, and gamma cyclodextrin.

1.9. Method 1.8, wherein the cyclodextrin is selected from the group consisting of alpha cyclodextrin, 2-hydroxypropyl-beta-cyclodextrin, and gamma cyclodextrin.

1.10. Method 1.9, wherein the cyclodextrin is gamma-cyclodextrin. For instance, wherein the bipolar trans carotenoid salt is TSC which is in a composition also comprising gamma-cyclodextrin (e.g., wherein the TSC is in a lyophilized composition with gamma- cyclodextrin).

1.11. Any one ofMethods 1-5 or 1.1-1.10, wherein the composition further comprises mannitol.

1.12. Any one ofMethods 1-5 or 1.1-1.11, wherein the diffusion enhancing compound (e.g., first diffusion enhancing compound and/or second diffusion enhancing compound) is administered intravenously or intramuscularly (e.g., as an intravenous bolus injection or intravenous infusion or intramuscular injection). For instance, any one ofMethods 1-5 or 1.1- 1.11, wherein the diffusion enhancing compound (e.g., first diffusion enhancing compound and/or second diffusion enhancing compound) is admixed with sterile water for injection to form an injection. Any one ofMethods 1-5 or 1.1-1.11, wherein TSC is administered intravenously or intramuscularly (e.g., as an intravenous bolus injection or intravenous infusion or intramuscular injection). For instance, any one ofMethods 1-5 or 1.1-1.11, wherein TSC is admixed with sterile water for injection to form an injection. 1.13. Any one of Methods 1-5 or 1.1-1.12, wherein the diffusion enhancing compound (e.g., first diffusion enhancing compound and/or second diffusion enhancing compound) is administered intravenously (e.g., as an intravenous bolus injection).

1.14. Any one of Methods 4, 5, or 1.1-1.13, wherein the second diffusion enhancing compound (e.g., as described above) is administered before the radiation therapy.

1.15. Any one ofMethods 1-5 or 1.1-1.14, wherein the patient is administered a positron emission tomography radiotracer (e.g., 18F-Fluoromisonidazole (F-MISO) or 18F- Fluoroazomycin-Arabinoside (F-FAZA)).

1.16. Any one ofMethods 1-5 or 1.1-1.15, wherein the patient is administered the first diffusion enhancing compound < 30 minutes prior to administration of the positron emission tomography radiotracer (e.g., 18F-FMISO or 18F-FAZA).

1.17. Any one of Method 5 or 1.1-1.16, wherein the patient is administered the first diffusion enhancing compound < 30 minutes prior to second measure of the solid tumor hypoxia.

1.18. Any one ofMethods 1-5 or 1.1-1.17, wherein the diffusion enhancing compound (e.g., first diffusion enhancing compound and/or second diffusion enhancing compound) is administered at a dose of 0.05-5 mg/kg, e.g., 0.05-2.5 mg/kg, e.g., 0.1-2 mg/kg, or 2.5-5 mg/kg, e.g., 3-5 mg/kg. For instance, any one ofMethods 1-5 or 1.1-1.17, wherein the diffusion enhancing compound (e.g., the first diffusion enhancing compound and the second diffusion enhancing compound) is trans-crocetin, in free or pharmaceutically acceptable salt form (e.g., trans sodium crocetinate (TSC)), and is administered at a dose of 0.05-5 mg/kg, e.g., 0.05-2.5 mg/kg (e.g., 1-2.5 mg/kg or 1.5-2.5 mg/kg or 0.5 mg/kg or 1 mg/kg or 1.5 mg/kg or 2 mg/kg or 2.5 mg/kg), e.g., 2-2.5 mg/kg, or 2.5-5 mg/kg, e.g., 3-5 mg/kg. For instance, any one ofMethods 1-5 or 1.1-1.17, wherein the diffusion enhancing compound

(e g., the first diffusion enhancing compound and the second diffusion enhancing compound) is trans sodium crocetinate (TSC), and is administered at a dose of 0.05-5 mg/kg, e.g., 0.05- 2.5 mg/kg (e.g., 1-2.5 mg/kg or 1.5-2.5 mg/kg or 0.5 mg/kg or 1 mg/kg or 1.5 mg/kg or 2 mg/kg or 2.5 mg/kg), e.g., 2-2.5 mg/kg, or 2.5-5 mg/kg, e.g., 3-5 mg/kg.

1.19. Any one ofMethods 1-5 or 1.1-1.18, wherein the cancer is a solid tumor. Any one ofMethods 1-5 or 1.1-1.18, wherein the solid tumor is glioblastoma multiforme.

1 .20. Any one ofMethods 1 -5 or 1 .1-1 .19, wherein a diffusion enhancing compound is administered before radiation therapy (e.g., the second diffusion enhancing compound is administered before radiation therapy). For instance, any one of Methods 1-5 or 1.1-1.19, wherein a diffusion enhancing compound (e.g., the second diffusion enhancing compound) (e.g., TSC) is administered 30 minutes to 90 minutes before radiation therapy. Or, for instance, any one of Methods 1-5 or 1.1-1.19, wherein a diffusion enhancing compound (e.g., the second diffusion enhancing compound) (e.g., TSC) is administered 15 minutes to 120 minutes before radiation therapy, e.g., 30 minutes to 60 minutes before radiation therapy, e.g., 45 minutes to 60 minutes before radiation therapy.

1.21. Any one of Methods 1-5 or 1.1-1.20, wherein the radiation therapy is external beam radiation therapy.

1.22. Any one of Methods 1-5 or 1.1-1.21, wherein the radiation therapy is three- dimensional conformal radiation therapy, intensity modulated radiation therapy, proton beam therapy, or stereotactic radiation therapy. For instance, any one of Methods 1-5 or 1.1-1.21, wherein the radiation therapy is intensity modulated radiation therapy.

1.23. Any one of Methods 1-5 or 1.1-1.22, wherein the radiation therapy is focal [or stereotactic] radiation therapy.

1.24. Any one of Methods 1-5 or 1.1-1.23, wherein the radiation therapy is internal beam radiation therapy.

1.25. Any one of Methods 1-5 or 1.1-1.24, wherein the radiation therapy is directed to a localized area within the tumor, in each case, based on upon the level of tumoral hypoxia demonstrated by the measure of the solid tumor hypoxia (e.g., by the patient’s PET imaging).

1.26. Any one of Methods 1-5 or 1.1-1.25, wherein the radiation therapy is administered in an amount of 0.1 Gy and 5 Gy per radiation therapy session.

1.27. Any one of Methods 1-5 or 1.1-1.26, wherein the radiation therapy is administered in an amount of 2 Gy per radiation therapy session.

1.28. Any one ofMethods 1-5 or 1.1-1.27, wherein the patient is administered chemotherapy.

1.29. Method 1.28, wherein the chemotherapy is one or more compounds selected from the group consisting of alkylating agents, antimetabolites, antitumor antibiotics, topoisomerase inhibitors, and anti -microtubule agents.

1 .30. Method 1 .29, wherein said chemotherapy is one or more compounds selected from the group consisting of temozolomide, gemcitabine, 5 -fluorouracil (5-FU), irinotecan, oxaliplatin, nab-paclitaxel (albumin-bound paclitaxel), capecitabine, cisplatin, elotinib, paclitaxel, docetaxel, and irinotecan liposome.

1.31. Any one ofMethods 1-5 or 1.1-1.30, wherein the solid tumor is glioblastoma multiforme (GBM) and the patient is administered temozolomide (e.g., 75 mg/m ² ) orally once daily (7 days a week) on an empty stomach over a period of 42 calendar days with a maximum of 49 days and radiation therapy. On radiation therapy days, the patient is administered the temozolomide within 1 to 3 hours prior to radiation therapy. For instance, any one of Methods 1-5 or 1.1-1.30, wherein the solid tumor is glioblastoma multiforme (GBM) and the patient is administered temozolomide (e.g., 75 mg/m ² ) orally once daily (7 days a week) on an empty stomach over a period of 6 weeks and radiation therapy. On radiation therapy days, the patient is administered the temozolomide within 1 to 3 hours prior to radiation therapy.

1.32. Any one ofMethods 1-5 or 1.1-1.31, wherein the solid tumor is glioblastoma multiforme (GBM) and the patient is administered 2Gy/day of radiation therapy. For instance, any one of Methods 1-5 or 1.1-1.31, wherein the solid tumor is glioblastoma multiforme (GBM) and the patient is administered fractionated focal irradiation in daily fractions of 2 Gy.

1.33. Any one ofMethods 1-5 or 1.1-1.32, wherein the solid tumor is glioblastoma multiforme (GBM) and the radiation therapy is administered for 5 days each week for 6 weeks. For instance, any one of Methods 1-5 or 1.1-1.32, wherein the total radiation therapy for 6 weeks is 60Gy (e.g., 2 Gy given 5 days per week for 6 weeks).

1.34. Any one of Methods 1-5 or 1.1-1.33, wherein the radiation therapy is focal radiation therapy. Any one of Methods 1-5 or 1.1-1.33, wherein the radiation therapy is fractionated focal radiation therapy.

1.35. Any one ofMethods 1-5 or 1.1-1.34, wherein the solid tumor is glioblastoma multiforme (GBM) and the diffusion enhancing compound (e.g., the second diffusion enhancing compound) (e.g., TSC) is administered between 30 to 60 minutes prior to a (e.g., prior to each) radiation therapy session (e.g., the diffusion enhancing compound (e.g., the second diffusion enhancing compound) is administered 3 days or 5 days each week for 6 weeks 30 to 60 minutes prior to radiation therapy). 1.36. Any one of Methods 1-5 or 1.1.35, wherein the solid tumor is glioblastoma multiforme (GBM) and after the combination radiation therapy and chemotherapy (e.g., and after 28 days of rest), the patient is administered temozolomide orally. For instance, the patient is administered 150 mg/m ² to 200 mg/m ² for Days 1 through 5 of a 28-day cycle (e.g., up to 6 cycles). For instance, the patient is administered 150 mg/m ² first cycle and 200 mg/m ² all subsequent cycles, as tolerated) administered on Days 1 through 5 of each 28-day cycle (e.g., up to 6 cycles).

1.37. Any one of Methods 1-5 or 1.1-1.36, wherein a diffusion enhancing compound (e g., as described above, e.g., TSC) is administered 30 minutes to 120 minutes (e.g., 45 minutes to 60 minutes or 1 hour to 2 hours) prior to administration of chemotherapy.

1.38. Any one ofMethods 1-5 or 1.1-1.37, wherein the patient is human.

1.39. Any one ofMethods 1-5 or 1.1-1.38, wherein the diffusion enhancing compound (e.g., first diffusion enhancing compound and second diffusion enhancing compound) is TSC and is administered intravenously, e.g., by a continuous intravenous infusion.

1.40. Any one ofMethods 1-5 or 1.1-1.39, wherein the diffusion enhancing compound (e.g., the first diffusion enhancing compound and the second diffusion enhancing compound) is TSC and is administered by a bolus intravenous injection.

1.41. Any one ofMethods 1-5 or 1.1-1.40, wherein the total dose (e.g., total daily dose) of the TSC does not result in a visual disturbance (e.g., a yellow visual disturbance).

1.42. Any one ofMethods 1-5 or 1.1-1.41, wherein the solid tumor is glioblastoma multiforme (GBM) and the patient is biopsy only.

1.43. Any one ofMethods 1-5 or 1.1-1.42, wherein the solid tumor is glioblastoma multiforme (GBM) and the patient has a subtotal resection.

1.44. Any on ofMethods 1-5 or 1.1-1.43, wherein the solid tumor is glioblastoma multiforme (GBM) and the patient has not had a gross total resection.

1.45. Any one ofMethods 1-5 or 1.1-1.44, wherein the solid tumor is glioblastoma multiforme (GBM) and the patient is > 70 years old at start of treatment.

1.46. Any one ofMethods 1-5 or 1.1-1.45, wherein the solid tumor is glioblastoma multiforme (GBM) and the patient is < 70 years old at start of treatment.

1.47. Any one ofMethods 1-5 or 1.1-1.46, wherein the solid tumor is glioblastoma multiforme (GBM) and the methyl guanine methyltransferase (MGMT) status of the patient is methylated.

1.48. Any one ofMethods 1-5 or 1.1-1.47, wherein the solid tumor is glioblastoma multiforme (GBM) and the methylguanine methyltransferase (MGMT) status of the patient is unmethylated.

1.49. Any one ofMethods 1-5 or 1.1-1.48, wherein the solid tumor is glioblastoma multiforme (GBM) and the patient also wears a portable treatment that creates electric fields that disrupt cancer cell division (e.g., an Optune).

1.50. Any one ofMethods 1-5 or 1.1-1.49, wherein the solid tumor is glioblastoma multiforme (GBM) and the overall survival of patients at 12 months is improved compared to patients not administered a diffusion enhancing compound.

1.51. Any one ofMethods 1-5 or 1.1-1.50, wherein the solid tumor is glioblastoma multiforme (GBM) and the overall survival of patients at 24 months is improved compared to patients not administered a diffusion enhancing compound.

1.52. Any one ofMethods 1-5 or 1.1-1.51, wherein the solid tumor is glioblastoma multiforme (GBM) and the health-related quality of life of patients is improved compared to patients not administered a diffusion enhancing compound (e.g., as measured using European Organization for Research Treatment of Cancer Quality of Life Questionnaire).

1.53. Any one ofMethods 1-5 or 1.1-1.52, wherein the solid tumor is glioblastoma multiforme (GBM) and the Kamofsky Performance Status is improved compared to patients not administered a diffusion enhancing compound.

1.54. Any one ofMethods 1-5 or 1.1-1.53, wherein the solid tumor is glioblastoma multiforme (GBM) and progression-free survival at 6 months (PFS6) by magnetic resonance imaging (MRI) assessment using Response Assessment in Neuro-Oncology (RANG) criteria is improved compared to patients not administered a diffusion enhancing compound.

1.55. Any one of Methods 1-5 or 1. 1-1.54, wherein the solid tumor is glioblastoma multiforme (GBM) and the radiation therapy is delivered to the gross tumor volume with a 2- to-3-cm margin for the clinical target volume.

1.56. Any one ofMethods 1-5 or 1.1-1.55, wherein the patient has histologically confirmed newly diagnosed high-grade IDH-wildtype GBM.

1.57. Any one ofMethods 1-5 or 1.1-1.56, wherein the tumor is a hypoxic solid tumor.

1 .58. Any one ofMethods 1 -5 or 1 .1-1 .57, wherein the radiation therapy is intensity modulated radiotherapy (IMRT) and/or 3D-cathode ray tube radiotherapy (CRT).

1.59. Any one of Methods 1-5 or 1.1-1.58, wherein the radiation therapy is fixed gantry IMRT, helical tomotherapy, and/or volumetric arc therapy (VMAT).

1.60. Any one ofMethods 1-5 or 1.1-1.59, wherein the solid tumor is gliobastoma multiforme and with radiation therapy the initial planning target volume (PTV1) is treated to 46 Gy in 23 fractions, then after 46 Gy, the conedown or boost planning target volume (PTV2) is treated to a total of 60 Gy with 7 additional fractions of 2 Gy each (14 Gy boost dose). Doses may be specified such that at least 95% of the planning target volume (PTV) shall receive 100% of the prescribed dose. PTV1 and/or PTV2 may be measured as described herein, for instance, above or in Example 1.

1.61. Method 1.60, wherein the prescription isodose (46 Gy for PTV1 and 60 Gy for PTV2) covers > 95% of the planning target volume (PTV). PTV, PTV1, and/or PTV2 may be measured as described herein, for instance, above or in Example 1.

1.62. Method 1.60 or 1.61, wherein the allowable dose within PTV1 (target dose 46 Gy) is between 41.4 Gy (90% of 46 Gy) and 50.6 Gy (110% of 46 Gy). PTV1 may be measured as described herein, for instance, above or in Example 1.

1.63. Any one of Methods 1.60-1.62, wherein the allowable dose within PTV2 (target dose 60 Gy) is between 54 Gy (90% of 60 Gy) and 66 Gy (110% of 60 Gy). PTV2 may be measured as described herein, for instance, above or in Example 1.

1.64. Any one ofMethods 1-5 or 1.1-1.63, wherein the measure of the solid tumor hypoxia (e.g., the first and/or second measure of the solid tumor hypoxia) is by positron emission tomography.

[0089] Further provided is a diffusion enhancing compound (e.g., a bipolar trans carotenoid salt (e.g., TSC)), e.g., as described in any one ofMethods 1-5 or 1.1-1.64, for use in any one of Methods 1-5 or 1.1-1.64.

[0090] Further provided is use of a diffusion enhancing compound (e.g., a bipolar trans carotenoid salt (e.g., TSC)), e.g., as described in any one of Methods 1-5 or 1.1-1.64, in the manufacture of a medicament for any one ofMethods 1-5 or 1.1-1.64.

[0091] Further provided is a pharmaceutical composition comprising an effective amount of a diffusion enhancing compound (e.g., a bipolar trans carotenoid salt (e.g., TSC)), e.g., as described in any one ofMethods 1-5 or 1.1-1.64, for use in any one ofMethods 1-5 or 1.1-1.64. Examples

[0092] The following is illustrative and not limiting of the compounds, compositions, and methods of the present disclosure. Other suitable modifications and adaptations of a variety of conditions and parameters normally encountered which are obvious to those skilled in the art are within the spirit and scope of this disclosure.

Example 1 - Study of Trans Sodium Crocetinate (TSC) When Administered with Standard of Care in Glioblastoma Patients

[0093] Obj ective s of the study are :

• Progression-free survival at 6 months (PFS6) by magnetic resonance imaging (MRI) assessment using Response Assessment in Neuro-Oncology (RANG) criteria

• Overall survival (OS) at 12 months

• Before and after hypoxia-related changes in the brain using fluorine- 18- fluoromisonidazole (18F-FMISO) or fluorine- 18-fluoroazomycin arabinoside (18F- FAZA) positron emission tomography (PET) tracer scans after a single dose of TSC (for Part A only)

• Health-related quality of life of patients treated with TSC and SOC

• Performance status of patients treated with TSC and SOC

• OS at 24 months

• Hypoxia changes (e.g., volumetric hypoxia changes) in PET tracer scans using 18F- FMISO or 18F-FAZA after a single dose of TSC

• Change in health-related quality of life using European Organization for Research Treatment of Cancer Quality of Life Questionnaire (EORTC QLQ-C30) at screening, Week 13, Week 25, End of Treatment (EOT), and Week 52 visits

• Change in Karnofsky Performance Status (KPS) from screening to Week 4, Week 8, Week 11, Week 15, Week 19, Week 23, Week 27, Week 31, EOT, and Week 52 visits

Methodology/Study Design

[0094] Adult patients who have confirmed histopathological diagnosis of glioblastoma (GBM) with status post biopsy or subtotal resection of the tumor, and who meet all eligibility criteria, will be enrolled into the study. Gross total resection in patients with GBM are excluded. [0095] Patients will be screened within 4 weeks of surgery to assess their study eligibility. [0096] After the screening period, the study (including both the Dose Escalation Run-In and Main Study) will be conducted in 2 parts; Part A to evaluate safety and pharmacodynamics (PD) and Part B to evaluate safety and efficacy of TSC. Patients will enter Part B no more than 14 days after completion of the repeat hypoxia PET tracer scan (with TSC) in Part A. Baseline for each assessment will be defined as the last measurement obtained during screening or Week 0 prior to the start of treatment in Week 1 (TSC plus SOC).

[0097] Safety, tolerability, pharmacokinetics, and pharmacodynamics of TSC at doses of 1.5 mg/kg, 2.0 mg/kg, and up to 2.5 mg/kg in combination with concomitant standard of care (radiation therapy (RT) + temozolomide) will be assessed via a Dose Escalation Run-In prior to initiating the Main Study.

[0098] The Dose Escalation Run-In will evaluate three cohorts of patients each at ascending TSC doses of 1.5, 2.0, and 2.5 mg/kg administered via intravenous injection. The patients that are to be part of the Dose Escalation Run-In will receive the same TSC dose (1.5, 2.0, 2.5 mg/kg) during Parts A and B of the protocol.

Part A:

[0099] In Part A, each patient who meets eligibility criteria will have a baseline and repeat 18F-FMISO hypoxia PET scan or baseline and repeat 18F-FAZA hypoxia PET scan. The baseline hypoxia PET tracer scan will be conducted without TSC. Within 48 to 72 hours of the baseline scan, the same tracer repeat hypoxia PET scan will be conducted. Prior to the repeat hypoxia PET tracer scan, a single IV dose of TSC given at 1.5, 2.0, or 2.5 mg/kg will be administered < 30 minutes prior to the 18F-FMISO or 18F-FAZA tracer administration. The PET scan then will commence following respective local practice for 18F-FMISO or 18F-FAZA hypoxia measurements. Pharmacodynamic and safety data will be collected.

Part B:

[00100] During Part B, all patients will receive TSC and SOC treatments for newly diagnosed GBM. All patients will be followed up to 12 months after the start of TSC treatment for efficacy and safety assessments and up to 24 months for overall survival. PK samples will be collected.

[00101] Patients will have laboratory parameters tested weekly and must maintain minimum values to continue the treatment period. [00102] The SOC treatment for GBM will follow the Stupp protocol which consists of temozolomide plus RT for 6 weeks, followed by 28 days of rest, followed by up to 6 cycles (each cycle of 28 days) of adjuvant temozolomide treatment.

[00103] The Stupp protocol should be followed; however, minor modifications to a patient’s radiotherapy plan may be needed (and thus will be allowed) for optimization of patient care if determined appropriate by the PI, study committee, and/or institutional standard of practice. Details of any such modification to the radiotherapy plan will be captured.

Dosing Schedule During 6-Week Treatment Period

[00104] During the 6-week treatment period, patients will receive:

• Temozolomide 75 mg/m ² orally once daily (7 days a week) on an empty stomach over a period of 42 calendar days with a maximum of 49 days; on RT session days, given at the same time relative to each RT session (within 1 to 3 hours prior to RT).

• Focal RT delivered as 60Gy/30 fractions scheduled at 2Gy/day for 5 days each week for 6 weeks.

• TSC 1.5, 2.0, or 2.5 mg/kg IV (IV push) for 5 days each week administered between 30 to 60 minutes prior to each RT session.

[00105] Radiotherapy will be delivered to the gross tumor volume with a 2-to-3-cm margin for the clinical target volume. Radiotherapy is planned with dedicated computed tomography (CT) and three-dimensional planning systems. Conformal radiotherapy will be delivered with linear accelerators with nominal energy of 6 MV or more.

[00106] Pneumocystis carinii pneumonia (PCP) prophylaxis will be given per local practice guidelines during temozolomide + RT administration.

[00107] During the 6-week treatment period, TSC should only be administered if the patient will receive a RT session (beam on) on that day. TSC should be interrupted when radiation is interrupted; TSC dosing will continue if radiation continues, even if temozolomide treatment is stopped for up to two weeks.

Dosing Schedule During Temozolomide Maintenance Period

[00108] All patients will receive oral temozolomide (150 mg/m ² first cycle and 200 mg/m ² all subsequent cycles, as tolerated) administered on Days 1 through 5 of each 28-day cycle (up to 6 cycles).

Radiation Therapy (RT) Information RT Technical Factors

[00109] This protocol requires photon treatment. Proton therapy is excluded. Intensity modulated radiotherapy (IMRT) and 3D-cathode ray tube (CRT) are allowed. Any of the methods of IMRT, including fixed gantry IMRT, helical tomotherapy, or volumetric arc therapy (VMAT) may be used, subject to protocol localization and dosimetry constraints. Computerized tomography (CT)-based treatment planning is necessary to assure accuracy in the selection of field arrangements. MRI-fusion for accurate target delineation is strongly recommended. Treatment shall be delivered with megavoltage (MV) machines of a minimum energy of 6 MV photons. Selection of the appropriate photon energy(ies) should be based on optimizing the radiation dose distribution within the target volume and minimizing dose to non-target normal tissue. Source skin distance for SSD techniques or source axis distance for SAD techniques must be at least 80 cm. Electron, particle, or implant boost is not permissible. IMRT delivery will require megavoltage radiation therapy machines of energy > 6 MV.

Localization and Simulation

[00110] Simulation may include a virtual simulation using a treatment planning computed tomography (CT). A planning CT scan of the cranial contents with 3.0 mm slice thickness or less may be fused with the pre- and post-operative MRI scans. Rigid registration is permitted, but deformable fusion is not permitted. Post-operative MRI with T2/fluid attenuation inversion recovery (FLAIR) and contrast enhanced T1 sequences is required.

RT Treatment Planning/Target Volumes

[00111] Treatment plans may include multiple fixed beams (3D conformal) or IMRT techniques (subject to protocol localization and dosimetry constraints). CT-based treatment planning is necessary to assure accuracy in the selection of field arrangements. MRI-fusion for accurate target delineation is recommended.

RT Initial Target Volume

[00112] Target volumes will be based upon postoperative-enhanced MRI. Preoperative imaging should be used for correlation and improved identification. Two planning target volumes (PTVs) will be defined, as outlined below. PTV may extend beyond bony margins and the skin surface.

[00113] The initial gross tumor volume (GTV1) will be defined by either the T2 or the FLAIR abnormality on the post-operative MRI scan. This must also include all postoperative- enhanced MRI enhancement, and the surgical cavity. Areas of vascular infarction or compromise of normal brain should be excluded if they are deemed to not be part of the original pre-operative tumor volume, and a comparison of the pre- and post-operative scans will assist in this process. [00114] The initial clinical target volume (CTV1) defined as the GTV1 plus a margin of 2 cm, which may be reduced around natural barriers to tumor growth such as the skull, ventricles, falx, etc. to as low as 0 mm to “fixed” barriers, i.e., bone, and falx and as low as 3 - 5 mm to “non-rigid” barriers such as brain stem, ventricles, etc. If no surrounding edema is present (i.e., the FLAIR and/or T2 MR images do not demonstrate any significant abnormality), the CTV1 in those instances should include the post-operative MRI enhancement and the surgical resection cavity plus a 2 cm margin, with reduction permitted as described above.

[00115] Reducing PTV margins to modify organ at risk (OAR) dose(s) is not generally permissible. However, OAR must be defined, along with a planning risk volume (PRV) for each OAR. Each PRV will be its OAR plus 3 mm. If an OAR is in immediate proximity to a PTV such that dose to the OAR cannot be constrained within protocol limits, a second PTV (PTV overlap), defined as the overlap between the initial planned target volume (PTV1) and the particular PRV of concern, may be created. Dose to the PTV overlap must be as close as permissible to 46 Gy while not exceeding the OAR dose limit.

RT Boost Target Volume

[00116] The boost gross tumor volume (GTV2) will be defined by the contrast enhanced T1 abnormality on the post-operative MRI scan. This must also include the surgical cavity margins. Areas of vascular infarction or compromise of normal brain should be excluded if they are deemed to not be part of the original pre-operative tumor volume, and a comparison of the pre- and post-operative scans will assist in this process. The exception to this is the polar tumor, e.g., temporal lobe, frontal lobe, or occipital lobe tip, where a gross total resection is achieved, and no post-operative “cavity” remains, consequential to the partial or total lobectomy.

[00117] The boost clinical target volume (CTV2) will be the GTV plus a margin of 2.0 cm. The CTV2 margin may be reduced around natural barriers to tumor growth such as the skull, ventricles, falx, etc. to as low as 0 mm to “fixed” barriers, i.e., bone, and falx and as low as 3 - 5 mm to “non-rigid” barriers such as brain stem, ventricles, etc.

[00118] The boost or conedown planning target volume (PTV2) is an additional margin of 3 to 5 mm, depending upon localization method and reproducibility, at each center. PTV margins account for variations in set-up and reproducibility. Reducing PTV margins to modify OAR dose(s) is not generally permissible. However, OAR must be defined, along with a PRV for each OAR. Each PRV will be its OAR plus 3 mm. If an OAR is in immediate proximity to a PTV such that dose to the OAR cannot be constrained within protocol limits, a second PTV (PTV overlap), defined as the overlap between the PTV2 and the particular PRV of concern, may be created (the overlap is the intersection between the PTV1 and the PRV). Dose to the PTV overlap must be as close as permissible to 14 Gy while not exceeding the OAR dose limit.

RT Dose Guidelines

[00119] For both IMRT and 3D-CRT plans, 1 treatment of 2 Gy will be given daily 5 days per week for a total of 30 sessions with 60 Gy over 6 weeks. The PTV1 will be treated to 46 Gy in 23 fractions. After 46 Gy, the PTV2 will be treated to a total of 60 Gy, with 7 additional fractions of 2 Gy each (14 Gy boost dose). All portals shall be treated during each treatment session. Doses are specified such that at least 95% of the planning target volume (PTV) shall receive 100% of the prescribed dose; Dose-volume histograms (DVH) are necessary to make this selection.

Critical Structures

[00120] All structures should be contoured on the planning CT, using the post-operative MRI for guidance. Due to variance in eye position between the CT and MRI, if possible, the lenses, retina, and optic nerves should be contoured using the CT dataset only. Special consideration should be given to avoid doses greater than the prescription dose within the scalp as well as limiting exit dose through the oral cavity and mucosa. The maximum point (defined as a volume greater than 0.03 cc) doses permissible to the structures are listed in the table below.

Table 1.

RT Compliance Criteria [00121] For all patients, as mentioned above, 2 PTV prescriptions, PTV1 and PTV2 will be used and the prescription isodose (46 Gy for PTV1 and 60 Gy for PTV2) must cover > 95% of the PTV volume; therefore, the total dose in the PTV2 volume will be 60 Gy. The allowable dose within PTV1 (target dose 46 Gy) must be between 41.4 Gy (90% of 46 Gy) and 50.6 Gy (110% of 46 Gy). The allowable dose within PTV2 (target dose 60 Gy) must be between 54 Gy (90% of 60 Gy) and 66 Gy (110% of 60 Gy). If the minimum dose falls below and/or the maximum dose falls above these parameters, a deviation will be recorded.

Eligibility Criteria for Inclusion and Exclusion:

[00122] Patients must meet the following inclusion criteria to be eligible for the study:

1. Male or female adult patients aged > 18 years at the date of signing informed consent

2. Histologically confirmed newly diagnosed high-grade IDH-wildtype GBM with confirmed pathology and prior subtotal tumor resection or biopsy up to 4 weeks prior to screening

3. Patient is willing and able to provide informed consent to participate in the study

4. No contraindications to MRI with gadolinium contrast, 18F-FAZA PET, or 18F-FMISO PET

5. Karnofsky Performance Status (KPS) score > 70%

6. Stable or decreasing dose of corticosteroids over 14 days prior to screening

7. No prior treatment with TSC

8. No investigational agent within 4 weeks prior to screening

9. Adequate hematological, renal, and hepatic function: a. Absolute neutrophil count > 1500/mm ³ , platelets > 100,000/mm ³ , hemoglobin > 9.0 g/dL b. Creatinine Clearance CrCl > 30 ml/min as measured by Cockcroft-Gault equation c. Blood urea nitrogen (BUN) within 2 times the upper limit of normal (ULN) d. Total bilirubin < 1.5 x ULN e. Transaminases < 4 x ULN

10. Must be without seizures or controlled on stable doses of anti-epileptic drugs for at least 14 days prior to screening

11. Absence of human immunodeficiency virus (HIV) infection, chronic hepatitis B, or hepatitis C infection; absence of any other serious medical condition which could interfere with medication intake

12. Female patients who do not have a surgical procedure precluding pregnancy (e g., hysterectomy) must have either a negative urine or serum pregnancy test at screening

13. Each patient must agree to take contraceptive measures for the duration of treatments and for at least 1 month after last dose of study treatment

14. Willing and able to fully comply with study procedures, restrictions, and other protocol requirements

[00123] Patients who meet any of the following exclusion criteria are not eligible to participate in the study:

1. Gross-total tumor resection (i.e., removal of all visible tumor at the time of surgery and confirmed by post-operative MRI)

2. Pathologically confirmed IDH1/2 mutation

3. Detection of metastases below the tentorium or beyond the cranial vault or leptomeningeal involvement

4. History of thrombotic or hemorrhagic stroke or myocardial infarction within 6 months of screening

5. Other chemotherapy or anti -turn or treatment for brain tumor (other than therapies required by the inclusion criteria of this protocol)

6. Pregnant or breastfeeding women

7. Uncontrolled intercurrent illness that would limit compliance with study requirements, or disorders associated with significant immunocompromised state

8. Any comorbid condition that confounds the ability to interpret data from the study as judged by the Investigator or medical monitor

9. Infratentorial tumor

10. 18F-FAZA PET or 18F-FMISO PET scan within 1 week of screening

11. Body weight of > 400 pounds.

12. QTcB > 470 milliseconds

Investigational Product, Dose, and Mode of Administration:

Investigational Product:

[00124] TSC will be supplied in 10 mL vials containing 100 mg of sterilized lyophilized powder and will be reconstituted with 5 mL Sterile Water for Injection, United States Pharmacopeia (USP) yielding 20 mg/mL.

TSC Dose Level (Parts A and B): 1 5, 2.0, or 2.5 mg/kg TSC Administration Route:

[00125] Part A: Single bolus IV injection < 30 minutes prior to 18F-FMISO or 18F-FAZA repeat PET tracer scan only.

[00126] Part B: Single bolus IV injection 30 to 60 minutes prior to RT sessions (beam on), 5 doses per week, for 6 weeks of RT.

Standard of Care:

[00127] RT (fractionated focal irradiation in daily fractions of 2 Gy given 5 days per week (Monday through Friday) for 6 weeks, for a total of 60Gy) plus continuous orally once daily temozolomide capsule (75 mg/m ² of body-surface area, 7 days per week from the first to the last day of radiotherapy), 4 weeks of rest, followed by up to 6 cycles of adjuvant temozolomide (150 to 200 mg per square meter for Days 1 through 5 of each 28-day cycle).

Treatments Administered

Table 2.

Subgroup Analysis

[00128] The following subgroups will be evaluated for the efficacy outcomes:

• Biopsy only versus subtotal resection

• Optune during adjuvant temozolomide versus no Optune during adjuvant temozolomide

• Age > 70 years versus Age < 70 years

• Methylguanine methyltransferase (MGMT) gene promoter status (methylated, unmethylated, or undetermined)

• Time from histological diagnosis to start of Part A (< median time versus > median time) [00129] Optune is an approved concomitant therapy for this study.

[00130] Progression-free survival is defined as the time interval from the date of histological diagnosis to the date of first occurrence of disease progression or death from any cause, whichever occurs first. The PFS6 will be assessed using the RANG criteria.

[00131] Progression-free survival by MRI using RANG criteria will be calculated from the date of histological diagnosis to the date of first occurrence of disease progression or death (from any cause), whichever occurs first. [00132] Tumor response will be performed according to the Imaging Acquisition Guidelines (e.g., same imaging method, type of MRI, slice of thickness, etc.) to ensure the standardization of the technique, limit of potential bias, and ensure quality of the data.

[00133] The RANG criteria define measurable disease as bidimensional contrast-enhancing lesions with clearly defined margins, with 2 perpendicular diameters of at least 10 mm, visible on > 2 axial slices. The non-measurable disease is defined as either unidimensional measurable lesions, masses with margins not clearly defined as frequently noted in the surgical margins, or lesions with maximal perpendicular diameters of < 10 mm. See Leao et al., “Response assessment in neuro-oncology criteria for gliomas: practical approach using conventional and advanced techniques,” Am J Neuroradiol. 2020;41(l): 10-20.

[00134] Overall survival will be defined as the time interval from the date of histological diagnosis to the date of death due to any cause. Overall survival at 12 months will be calculated from the date of histological diagnosis to the time of death (from any cause) and will be analyzed as a binary outcome (percentage of patients alive).

[00135] Overall survival at 24 months will be calculated from the date of histological diagnosis to the time of death (from any cause) and will be analyzed as a binary outcome (percentage of patients alive).

[00136] Quality of life will be assessed using EORTC QLQ-C30. EORTC QLQ-C30 is designed to measure cancer patients’ physical, psychological, and social functions. The questionnaire is composed of multi-item scales and single items.

[00137] Kamofsky Performance Status is a standard way of measuring the ability of cancer patients to perform ordinary tasks. The KPS scores range from 0 to 100. A higher score means the patient is better able to carry out daily activities. Karnofsky Performance Status may be used to determine a patient’s prognosis, to measure changes in a patient’s ability to function, or to decide if a patient could be included in a clinical study. See Karnofsky et al., “The clinical evaluation of chemotherapeutic agents in cancer,” in: MacLeod CM, ed. Evaluation of chemotherapeutic agents. Columbia University Press. 1949: 191-205.

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It will be readily apparent to those skilled in the art that the numerous modifications and additions can be made to both the present compounds and compositions, and the related methods without departing from the disclosed methods and compositions.