Compositions containing pafolacyanin for identifying malignant lesions

Abstract

The composition is configured to be administered to a subject undergoing a malignant lesion resection procedure. The composition comprises a pharma- ceutically effective amount of pafolacyanin or a pharma- ceutically acceptable salt thereof. The pafolacyanin or a pharma- ceutically acceptable salt thereof is configured to bind one or more malignant lesions and emit an optical signal. The compositions, methods, and techniques described herein allow visualization of cancerous tumors, cells, and / or lesions that conventional compositions and techniques would not otherwise identify.

Description

[Technical field]

[0001] Related Applications This patent application claims the benefit of priority to U.S. Patent Application No. 18 / 111,704, filed February 20, 2023, and U.S. Provisional Patent Application No. 63 / 312,965, filed February 23, 2022, the contents of each of which are incorporated by reference in their entirety into this disclosure. [Background technology]

[0002] background Many treatment regimens for cancer, such as ovarian and lung cancer, include surgery. For example, a patient with cancer may undergo surgery with the goal of removing cancer cells. In some aspects, a method or system may be used to assist the surgeon in visualizing the malignant lesion intended to be removed via surgery. In some such cases, an imaging agent may be provided to the patient, and a light source or imaging technology may be used to capture the imaging agent within the patient's body. For example, a patient may receive the imaging agent via ingestion, intravenous injection, topically, or via another method. Such visualization techniques may result in better surgical outcomes (e.g., a higher percentage of cancer cells removed) in comparison to surgery without visualization techniques.

[0003] Although conventional visualization compositions, methods, and techniques may provide better visualization of cancer lesions within a patient's body than surgery without any visualization composition, method, or technique, such conventional compositions, methods, and techniques may not be completely effective in identifying cancerous areas of the body. Thus, there is a need for compositions and methods that are more effective in identifying cancerous lesions in a patient's body.

[0004] Embodiments of the present invention address these and other challenges, both individually and collectively.

[0005] Further limitations and drawbacks of conventional and legacy approaches will become apparent to those skilled in the art through comparison of such systems with certain aspects of the present disclosure as set forth in the remainder of this application with reference to the drawings. Summary of the Invention [Means for solving the problem]

[0006] Abstract The present inventors have recognized the need for improved visualization techniques and compositions for identifying cancerous lesions in a patient's body. Thus, the compositions, methods, and techniques described herein allow for the visualization of cancerous tumors, cells, and / or lesions that conventional compositions and techniques would not otherwise identify.

[0007] Current technology is believed to target folate receptor (FR), which may be overexpressed in certain cancer cells (e.g., ovarian cancer cells and lung cancer cells). The compositions, methods, and technology of the present disclosure include pafolacyanin, which is believed to bind to FR-expressing cancer cells and fluoresce when exposed to light to allow visualization of FR-expressing cancer cells bound to pafolacyanin. This technology can improve the detection of cancer cells and the specificity of said detection.

[0008] In one aspect, a composition configured to be administered to a subject undergoing a malignant lesion resection procedure, the composition comprising a pharma- tically effective amount of pafolacyanin, wherein the pafolacyanin is configured to fluoresce to identify one or more malignant lesions.

[0009] In some aspects, one or more of the malignancies include ovarian cancer. In other aspects, one or more of the malignancies include lung cancer. In yet other aspects, the pharmacologic effective amount of pafolacyanin comprises at least about 0.25 mg of pafolacyanin per about 1 kg body weight of the subject. In further aspects, the composition is configured to be administered intravenously to the subject.

[0010] In some aspects, the composition comprises an injectable solution. In other aspects, the solution is blue-green. In other aspects, the solution is clear to translucent. In still other aspects, the composition comprises between about 1 mg and about 20 mg of pafolacyanin per about 1.6 mL volume of solution. In further aspects, the composition comprises about 3.2 mg of pafolacyanin per about 1.6 mL volume of solution.

[0011] In some aspects, about 3.2 mg of perforacyanine per about 1.6 mL volume of solution is equivalent to about 3.4 mg of perforacyanine sodium per about 1.6 mL volume of solution. In other aspects, the perforacyanine sodium has the following chemical structure: [ka] has.

[0012] In some aspects, the composition includes at least one of sodium chloride, potassium phosphate monobasic, or sodium phosphate dibasic heptahydrate. In other aspects, the composition includes between about 1 mg and about 50 mg of sodium chloride per about 1.6 mL volume of solution. In yet other aspects, the composition includes about 14.4 mg of sodium chloride per about 1.6 mL volume of solution. In further aspects, the composition includes between about 0.01 mg and about 5 mg of potassium phosphate monobasic per about 1.6 mL volume of solution.

[0013] In some aspects, the composition comprises about 0.23 mg potassium phosphate monobasic per about 1.6 mL volume of solution. In other aspects, the composition comprises between about 0.1 mg and about 10 mg sodium phosphate dibasic heptahydrate per about 1.6 mL volume of solution. In yet other aspects, the composition comprises about 1.27 mg sodium phosphate dibasic heptahydrate per about 1.6 mL volume of solution. In further aspects, the composition comprises about 3.2 mg pafolacyanin, about 14.4 mg sodium chloride, about 0.23 mg potassium phosphate monobasic, and about 1.27 mg sodium phosphate dibasic heptahydrate per about 1.6 mL volume of solution.

[0014] In some aspects, the pH of the composition is between about 6 and about 8. In other aspects, the composition includes at least one of sodium hydroxide or hydrochloric acid. In yet other aspects, at least one of sodium hydroxide or hydrochloric acid is used to adjust the pH of the composition. In further aspects, a solution containing the composition is diluted prior to administering the composition to the subject.

[0015] In some aspects, the solution is diluted with a 5% dextrose solution. In other aspects, the solution is diluted with about 220 mL to about 250 mL of a 5% dextrose solution. In yet other aspects, the composition is configured to be administered to the subject about 1 to about 24 hours prior to the malignancy resection procedure. In further aspects, the composition is configured to be administered to the subject over a period of time.

[0016] In some aspects, the period of time comprises about 1 to about 3 hours. In other aspects, the perforacyanin fluoresces upon exposure to light in the near infrared range. In still other aspects, the perforacyanin fluoresces upon exposure to light having a wavelength between about 760 nm and about 785 nm. In further aspects, the perforacyanin fluoresces between about 790 nm and about 815 nm.

[0017] In some aspects, the composition is contained in a vial. In other aspects, the vial is a single dose vial. In still other aspects, a carton contains at least 10 vials. In further aspects, the at least 10 vials are individually packaged.

[0018] In some aspects, the subject is an animal. In other aspects, the subject is a mammal. In still other aspects, the subject is a human. In a further aspect, the subject is an adult.

[0019] In one aspect, a method of treating a subject undergoing a malignant lesion resection procedure includes administering to the subject a pharma- tically effective amount of a composition comprising pafolacyanin; and causing the pafolacyanin to fluoresce.

[0020] In some aspects, the method includes identifying one or more malignant lesions based on the fluorescence of the pafolacyanin. In other aspects, the method includes resecting one or more of the malignant lesions. In yet other aspects, one or more of the malignant lesions include ovarian cancer. In a further aspect, one or more of the malignant lesions include lung cancer.

[0021] In some aspects, the pharma- ceutically effective amount comprises at least about 0.25 mg of pafolacyanin per about 1 kg body weight of the subject. In other aspects, the composition is administered intravenously to the subject. In still other aspects, the composition comprises an injectable solution. In a further aspect, the solution is blue-green in color.

[0022] In some aspects, the solution is clear to translucent. In other aspects, the solution contains between about 1 mg and about 20 mg of perforacyanin per about 1.6 mL volume of solution. In yet other aspects, the composition contains about 3.2 mg perforacyanin per about 1.6 mL volume of solution. In further aspects, about 3.2 mg perforacyanin per about 1.6 mL volume of solution is equivalent to about 3.4 mg perforacyanin sodium per about 1.6 mL volume of solution.

[0023] In some aspects, the sodium pafolacyanine has the following chemical structure: [ka] has.

[0024] In some aspects, the solution further comprises at least one of sodium chloride, potassium phosphate monobasic, or sodium phosphate dibasic heptahydrate. In other aspects, the solution comprises between about 1 mg and about 50 mg of sodium chloride per about 1.6 mL volume of solution. In yet other aspects, the solution comprises about 14.4 mg of sodium chloride per about 1.6 mL volume of solution. In further aspects, the solution comprises between about 0.01 mg and about 5 mg of potassium phosphate monobasic per about 1.6 mL volume of solution.

[0025] In some aspects, the solution contains about 0.23 mg potassium phosphate monobasic per about 1.6 mL volume of solution. In other aspects, the solution contains between about 0.1 mg and about 10 mg sodium phosphate dibasic heptahydrate per about 1.6 mL volume of solution. In yet other aspects, the solution contains about 1.27 mg sodium phosphate dibasic heptahydrate per about 1.6 mL volume of solution. In further aspects, the solution contains about 3.2 mg pafolacyanin, about 14.4 mg sodium chloride, about 0.23 mg potassium phosphate monobasic, and about 1.27 mg sodium phosphate dibasic heptahydrate per about 1.6 mL volume of solution.

[0026] In some aspects, the pH of the solution is between about 6 and about 8. In other aspects, the solution further comprises at least one of sodium hydroxide or hydrochloric acid. In yet other aspects, the at least one of sodium hydroxide or hydrochloric acid is used to adjust the pH of the solution. In a further aspect, the method includes diluting the solution prior to administering the composition to the subject.

[0027] In some aspects, diluting the solution comprises diluting the solution with a 5% dextrose solution. In other aspects, diluting the solution comprises diluting the solution with about 220 mL to about 250 mL of a 5% dextrose solution. In yet other aspects, administering a pharmacologic effective amount of the composition comprises administering a pharmacologic effective amount of the composition about 1 to about 24 hours prior to the malignant lesion resection procedure. In further aspects, administering a pharmacologic effective amount of the composition comprises administering a pharmacologic effective amount of the composition over a period of time.

[0028] In some aspects, the period of time includes about 1 to about 3 hours. In other aspects, the step of causing the perforation to fluoresce includes exposing the perforation to light in the near infrared range. In yet other aspects, the step of causing the perforation to fluoresce includes exposing the perforation to light having a wavelength between about 760 nm and about 785 nm. In a further aspect, the perforation fluoresces between about 790 nm and about 815 nm.

[0029] In some aspects, the composition is contained in a vial. In other aspects, the vial is a single dose vial. In yet other aspects, a carton contains 10 vials. In further aspects, the 10 vials are individually packaged.

[0030] In some aspects, the subject is an animal. In other aspects, the subject is a mammal. In still other aspects, the subject is a human. In further examples, the subject is an adult.

[0031] In some aspects, the fluorescence of the above-mentioned Pafolacyanin is detected using imaging system or imaging software.In another aspect, the imaging system or imaging software is selected from the group consisting of imaging system FAST (fiber optic array scanning technology), flow cytometry, confocal microscopy, two-photon microscopy, epifluorescence microscopy, fluorescence microscopy, fluorescent goggles, and innovative wearable equipment.In one aspect, the fluorescence of the above-mentioned Pafolacyanin is detected in vivo.

[0032] In some aspects, the method further comprises guiding a flexible probe to the malignant lesion after administration of the composition comprising the pafolacyanin. In some aspects, the flexible probe is a flexible endoscope, a fluorescence endoscopic imaging probe, a fiberscope, a videoscope, a gastroscope, a colonoscope, a bronchoscope, a laryngoscope, a cystoscope, a duodenoscope, an intestinal endoscope, a ureteroscope, a sigmoidoscope, an intestinal endoscope, a biliary endoscope, a nasal laryngoscope, an angioscope, or a hysteroscope. In other aspects, the flexible probe is equipped to detect wavelengths having absorption and emission maxima between about 400 nm and about 900 nm.

[0033] In some aspects, the resection procedure is performed using an iBiopsy, an iKnife, an iLaser, an iBurner, an electric cutting loop, a rotating blade, a curved blade, an expandable blade, a dissector with a cutting blade, a blunt dissector, forceps, an electrolyzable element, a biopsy needle, a microwave ablation probe, a radiofrequency ablation probe, a cryoablation probe, or a laser.

[0034] In some aspects, the ablation procedure is non-invasive.

[0035] In some aspects, the resection procedure is performed manually or using robotic-assisted techniques. [Brief description of the drawings]

[0036] BRIEF DESCRIPTION OF THE DRAWINGS [Figure 1] FIG. 1 is a graph illustrating the relative binding of pafolacyanin, the L-isomer of pafolacyanin (OTL0039), and folic acid to FRs.

[0037] [Figure 2-1] Figure 2A is a whole-body fluorescence image of a mouse bearing an FR-positive KB tumor injected with pafolacyanin. Figure 2B is a whole-body fluorescence image of a mouse bearing an FR-positive KB tumor injected with the L-isomer of pafolacyanin (OTL0039). Figure 2C is a fluorescence image of tissue distribution 2.5 hours after injection of pafolacyanin. Figure 2D is a fluorescence image of tissue distribution 2.5 hours after injection of the L-isomer of pafolacyanin (OTL0039). [Figure 2-2] Same as above.

[0038] [Diagram 3] Figure 3A is a whole body fluorescence image of a mouse bearing a FR-negative A549 tumor injected with Pafolacyanin. Figure 3B is a fluorescence image of the tissue distribution of the mouse bearing a FR-negative A549 tumor injected with Pafolacyanin of Figure 3A.

[0039] [Figure 4] FIG. 4 is a graph illustrating the mean concentrations of pafolacyanin in male and female rats and dogs following intravenous administration of pafolacyanin. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0040] Detailed Description The present technology provides compositions that include pafolacyanin and methods of using such compositions (eg, methods of identifying or visualizing cancer cells in a patient's body).

[0041] As used herein, the following conventional unit abbreviations and terms are used as follows: "pg" refers to picogram. "ng" refers to nanogram. "μg" refers to microgram. "mg" refers to milligram. "g" refers to gram. "kg" refers to kilogram. "mL" refers to milliliter. "h" refers to hour. "t" refers to time.

[0042] As used herein, "patient" and / or "subject" refer to a human or an animal. In some aspects, the human is an adult. In other aspects, the human may be a child. In some aspects, the animal is a mammal. In other aspects, the animal is a non-mammalian animal. "Patient" and "subject" are used interchangeably throughout this disclosure.

[0043] As used herein, "pharmacologically effective amount" refers to an effective dose or concentration that produces a biological response.

[0044] As used herein, "and / or" means any one or more of the items in the list connected by "and / or." As an example, "x and / or y" means any element of the set of these elements {(x),(y),(x,y)}. In other words, "x and / or y" means "one or both of x and y." As another example, "x, y, and / or z" means any element of the seven-element set {(x),(y),(z),(x,y),(x,z),(y,z),(x,y,z)}. In other words, "x, y, and / or z" means "one or more of x, y, and z."

[0045] The term "exemplary" means serving as a non-limiting example, instance, or illustration. As used herein, the terms "eg," and "for example" refer to a list of one or more non-limiting examples, instances, or illustrations.

[0046] As used herein, the terms "about" and "approximately," when used to modify or describe a value (or range of values), mean reasonably close to that value or range of values ​​(e.g., ±10% of that value or range of values). Thus, the aspects described herein are not limited to only the values ​​and ranges of values ​​described, but rather should include deviations that operate reasonably.

[0047] As used herein, the use of the singular includes the plural unless specifically stated otherwise.

[0048] As mentioned above, malignant lesion resection procedures, or surgical removal of cancer cells or other malignant diseases, are part of many treatment plans for such cancers or malignant diseases. In some cases, surgical removal can be one of the most effective ways to treat cancer. For example, removal of all detectable malignant lesions may result in approximately 50% of all cancer patients not experiencing detectable recurrence of disease. In the event that the disease recurs, the malignant lesion resection procedures may still extend life expectancy or reduce the morbidity of the patient. Therefore, for malignant lesion resection procedures to have the best possible outcome, it is important that as many cancer cells as possible are removed during surgery.

[0049] In many cases, visualization methods are used to identify malignant lesions. Traditional visualization methods include identification based on color, texture, and / or morphology of the tumor. Additionally or alternatively, tumors may be identified based on differences in plasticity, elasticity, or solidity from adjacent healthy tissue. In some aspects, these such methods may be performed by the surgeon without the use of additional tools or compositions, or with tools and compositions typically used during surgical procedures. For example, surgeons may use their own senses to identify malignant lesions based on color, texture, morphology, plasticity, elasticity, and / or solidity. Another example of a traditional visualization technique is the use of fluorescent dyes that may passively flow from the tumor to the draining lymph nodes. Such traditional dyes may fluoresce in the visible light range. The fluorescent lymph nodes (e.g., due to the dye) may be visually identified and resected to determine whether cancer cells have metastasized to the lymph nodes. However, in many cases, at least some of the malignant lesions may not be identified and therefore may not be resected using these traditional visualization methods. Furthermore, some conventional dyes may autofluoresce in the visible light range, may have a relatively short shelf life, may be relatively unstable, and may have poor tissue penetration.

[0050] Thus, there is a need for improved visualization techniques and compositions to better identify malignant lesions in the patient's body. Disclosed are compositions, formulations, and methods for identifying cancer cells. The disclosed compositions, formulations, and methods include Pafolacyanin. Without being bound by theory, Pafolacyanin is believed to have increased binding affinity to folate receptor (FR)-positive cancers. The above compositions and formulations that include Pafolacyanin bind to FRs of malignant lesions that express FRs, which can then be used to identify malignant lesions. Such compositions, formulations, and methods can result in increased visualization and identification of malignant lesions that express FRs, compared to the conventional visualization methods discussed above. In this way, the use of the compositions, formulations, and methods disclosed herein can result in improved outcomes for cancer patients (e.g., resection of a higher percentage of malignant lesions, increased life expectancy, better chance of cancer remission, and / or reduced mortality). Additionally, the disclosed techniques may have reduced autofluorescence, increased shelf life, increased stability, and / or more efficient tissue penetration compared to conventional dyes.

[0051] As discussed above, the compositions, formulations, and methods include pafolacyanin, which is believed to have increased binding affinity to FR. In this way, the disclosed compositions and methods can result in improved visualization of cancer lesions in cancers that express or overexpress FR. Such cancers include, for example, ovarian cancer, renal cancer, lung cancer, endometrial cancer, breast cancer, and colon cancer. Other cancers that are not specifically disclosed herein may also express or overexpress FR, FR. In such cases, the compositions, formulations, and methods disclosed herein can result in improved identification of those cancers as well. The use of the compositions, formulations, and methods disclosed herein is contemplated for any type of cancer or other malignant disease.

[0052] Thus, the present disclosure allows the identification of biological tissues expressing FRs by contacting the tissue with a composition comprising the present disclosure of pafolacyanin and allowing time for the composition to distribute in the tissue and interact with the site of FRs. After a sufficient time for such interaction has elapsed, the tissue is irradiated with excitation light so that the composition fluoresces. The fluorescence is then observed to visualize malignant lesions. In a similar manner, the composition of the present disclosure is used to identify target cell types in biological samples by contacting the biological sample with such a composition for a period of time and under conditions that allow the binding of the composition to at least one cell of the target cell type. The bound composition is then made to fluoresce to allow the identification of the tissue to which the composition is bound.

[0053] The compositions disclosed herein include an optical imaging agent. In some aspects, the optical imaging agent is pafolacyanin. In some aspects, the compositions include pafolacyanin as a pharma- ceutically acceptable salt. In some aspects, the compositions include pafolacyanin as a tetrasodium salt. For example, the compositions may include pafolacyanin sodium. Pafolacyanine sodium has the chemical formula (S)-2-(4-(((2-amino-4-oxo-3,4-dihydropteridin-6-yl)methyl)amino)benzamido)-3-(4-(((E)-2-((E)-2-(3,3-dimethyl-5-sulfonato-1-(4-sulfonatobutyl)-3H-indol-1-ium-2-yl)vinyl)-6-((E)-2-(3,3-dimethyl-5-sulfonato-1-(4-sulfonatobutyl)indolin-2-ylidene)ethylidene)cyclohex-1-en-1-l)oxy)phenyl)propanoate hydrate tetrasodium and a molar mass of 1414.42 g / mol. The general structure of the pafolacyanine sodium of the present technology is as follows: [ka] It is.

[0054] Without intending to be bound by theory, it is believed that Pafolacyanin binds to FR-expressing cancer cells.In some aspects, Pafolacyanin can bind to FR-expressing cancer cells with at least about 1 nanomolar (nM) affinity, at least about 0.75 nM affinity, at least about 0.5 nanomolar (nM) affinity, at least about 0.25 nanomolar (nM) affinity, at least about 0.1 nanomolar (nM) affinity, at least about 0.05 nanomolar (nM) affinity, at least about 0.01 nanomolar (nM) affinity, or at least about 0.001 nanomolar (nM) affinity.Pafolacyanin is then internalized into cancer cells via receptor-mediated endocytosis, and thus concentrates in FR-positive ocular tissues.

[0055] Once absorbed in tissue, the compositions of the present disclosure can be used to identify cancer cells by causing pafolacyanin to fluoresce. Pafolacyanin is excited using light having at least one excitation wavelength in the near infrared range (e.g., between about 600 nm and about 1000 nm). In some aspects, pafolacyanin absorbs light in the near infrared range of about 600 nm to about 900 nm, about 700 nm to about 900 nm, about 700 nm to about 800 nm, about 750 nm to about 800 nm, or about 760 nm to about 785 nm. In some aspects, peak absorption by pafolacyanin can occur at about 776 nm. In some aspects, pafolacyanin emits fluorescence between about 600 nm to about 900 nm, about 700 nm to about 900 nm, about 700 nm to about 850 nm, about 750 nm to about 850 nm, or about 790 nm to about 815 nm. In some aspects, the peak emission by pafolacyanin can occur at about 796 nm. Fluorescence by pafolacyanin bound to the FR of cancer cells can allow visualization of said cancer cells. In some aspects, visualization using pafolacyanin can be improved over conventional visualization methods (e.g., more cancer cells can be identified). Furthermore, due to the use of near-infrared light instead of light in the visual range, the compositions and formulations discussed herein can be more stable, have a longer shelf life, produce less autofluorescence, and penetrate tissues better than conventional dyes.

[0056] In some aspects, an operating room for a surgical procedure may be equipped with overhead lights that produce wavelengths of light in the range discussed above. For example, an operating room may include at least one lamp that produces light of an appropriate wavelength. In this manner, at least one lamp that produces light of an appropriate wavelength may be directed toward a body cavity, surgical opening, etc., to cause the perforacyanin to bind to and fluoresce in cancer cells. In some aspects, a surgeon (or other professional) may directly observe the fluorescence without needing additional tools, accessories, or special equipment to identify malignant lesions. In other examples, additional tools, accessories, or special equipment may additionally or alternatively be used to visualize the fluorescence of perforacyanin and identify malignant lesions. For example, an endoscopic device may be used to deliver excitation light to the site, receive the fluorescence emanating from the site, and / or aid in forming an image of the fluorescence from the malignant lesion. Additionally or alternatively, in some aspects, an image processing device (such as, for example, a CCD camera, a display, a photon collection device, etc.) may be used to aid in visualization of the cancer cells.

[0057] The compositions and formulations disclosed herein can be administered by any route known to those of skill in the art, such as, for example, intravenously, topically, intraarticularly, intracisternally, intraocularly, intracerebroventricularly, intrathecally, intramuscularly, intraperitoneally, intradermally, intratracheally, or intracavitary. In some aspects, the most appropriate administration route can vary depending on the disease state to be treated, or the location of the suspected condition or tumor to be identified.

[0058] In some aspects, the composition of the present disclosure can be administered intravenously.For example, Pafolacyanin can be administered intravenously as a solution.The solution can contain Pafolacyanin and any other composition, ingredient, drug, solvent, buffer, preservative, antioxidant, diluent, etc. in an effective amount (e.g., the amount of Pafolacyanin that allows identification of malignant lesions in the patient's body).

[0059] The composition may be administered before surgery begins, before an incision is made during a surgical procedure, after surgery begins, after the surgical cavity or tumor site is revealed by surgery, or at other suitable times. In some aspects, the composition may be administered intravenously to the patient at least about 1 hour before the start of fluorescence imaging. For example, the pafolacyanin may be administered between about 1 hour and about 24 hours before surgery, between about 1 hour and about 12 hours before surgery, between about 1 hour and about 9 hours before surgery, between about 1 hour and about 7 hours before surgery, between about 1 hour and about 5 hours before surgery, between about 1 hour and about 3 hours before surgery, or between about 1 hour and about 2 hours before surgery. In some such instances, allowing at least 1 hour before surgery may allow the composition to sufficiently bind to FR-expressing cancer cells to fluoresce and thus allow visualization of the cancer cells. In other aspects, the composition may be provided to the patient less than about 1 hour before surgery.

[0060] In some aspects, the Pafolacyanin composition may be provided (intravenously) to the patient over a period of time. For example, in some aspects, the Pafolacyanin may be administered to the patient over a period of about 1 hour or longer. In some such aspects, the composition may be administered to the patient using an infusion line over a period of about 1 hour, about 1.5 hours, about 2 hours, or about 3 hours. For example, the composition may be administered to the patient over a period of between about 1 hour and about 3 hours. In other aspects, the composition may be administered over a different period of time.

[0061] In some aspects, the compositions disclosed herein contain about 0.5 mg or more, about 2.5 mg or more, about 5.0 mg or more, about 7.5 mg or more, about 10 mg or more, about 20 mg or more, about 30 mg or more, about 40 mg or more, about 50 mg or more, about 60 mg or more, about 70 mg or more, The formulations may contain amounts of about 80 mg or more, about 90 mg or more, or about 100 mg or more of pafolacyanin, including any further increments thereof (e.g., 0.1 mg, 0.2, mg, 0.25 mg, 0.3 mg, 0.4 mg, 0.5 mg, 0.6 mg, 0.7 mg, 0.75 mg, 0.8 mg, 0.9 mg, or 1.0 mg) and multiplication factors thereof (e.g., x1, x2, x2.5, x5, x10, x100, etc.). In some aspects, the formulations include between about 1 mg and about 20 mg, between about 1 mg and about 10 mg, between about 1 mg and about 5 mg, between about 1 mg and about 4 mg, or between about 2 mg and about 4 mg. For example, some formulations may contain about 3.2 mg of pafolacyanin. In aspects where the formulation comprises pafolacyanine sodium, the formulation may contain between about 1 mg and about 20 mg, between about 1 mg and about 10 mg, between about 1 mg and about 5 mg, between about 1 mg and about 4 mg, between about 2 mg and about 4 mg, or about 3.4 mg of pafolacyanine sodium.

[0062] Pafolacyanin may be administered to a patient in any suitable dose. In some aspects, suitable doses of Pafolacyanin include, but are not limited to, between about 0.01 mg / kg and about 1 mg / kg of patient body weight, between about 0.01 mg / kg and about 0.5 mg / kg of patient body weight, between about 0.01 mg / kg and about 0.25 mg / kg of patient body weight, between about 0.01 mg / kg and about 0.1 mg / kg of patient body weight, between about 0.01 mg / kg and about 0.09 mg / kg of patient body weight, between about 0.01 mg / kg and about 0.07 mg / kg of patient body weight, between about 0.25 mg / kg and about 0.07 mg / kg of patient body weight, or between about 0.25 mg / kg and about 0.05 mg / kg of patient body weight. In some aspects, about 0.25 mg of Pafolacyanin is administered per kg of patient body weight. In other aspects, different amounts of Pafolacyanin may be administered per kg of patient body weight. A similar dose of a composition containing pafolacyanine sodium may be administered to the patient.

[0063] In some aspects, the present technology provides a combination formulation comprising Pafolacyanin and at least one other composition, ingredient, drug, solvent, buffer, preservative, antioxidant, diluent, etc. For example, the at least one other composition, ingredient, drug, solvent, buffer, preservative, antioxidant, or diluent can be a salt. In some such aspects, the salt can include sodium chloride, potassium phosphate monobasic, sodium phosphate dibasic heptahydrate, sodium hydrogen sulfate, calcium chloride, potassium iodide, or another salt. As a non-limiting aspect, the combination formulation can include Pafolacyanin (and / or Pafolacyanin sodium), sodium chloride, potassium phosphate monobasic, and sodium phosphate dibasic heptahydrate. In some aspects, the combination formulation can further or instead include an acid, a base, and / or a buffer. For example, in some such aspects, the combination formulation may include sodium hydroxide, hydrochloric acid, perchloric acid, hydroiodic acid, nitric acid, sulfuric acid, potassium hydroxide, and / or calcium hydroxide. As a non-limiting aspect, the combination formulation may include sodium hydroxide and / or hydrochloric acid.

[0064] As another non-limiting example, the combination formulation may contain pafolacyanine (and / or pafolacyanine sodium), sodium chloride, potassium phosphate monobasic, sodium phosphate dibasic heptahydrate, sodium hydroxide, and / or hydrochloric acid. In some aspects, the combination formulation may contain between about 1 mg and about 50 mg, between about 1 mg and about 25 mg, between about 1 mg and about 20 mg, between about 1 mg and about 15 mg, between about 5 mg and about 50 mg, between about 5 mg and about 25 mg, between about 5 mg and about 20 mg, between about 5 mg and about 15 mg, between about 10 mg and about 15 mg, or about 14.4 mg of sodium chloride. In some aspects, the combination formulation may contain between about 0.01 mg and about 5 mg, between about 0.01 mg and about 1 mg, between about 0.01 mg and about 0.5 mg, between about 0.01 mg and about 0.25 mg, between about 0.05 mg and about 5 mg, between about 0.05 mg and about 1 mg, between about 0.05 mg and about 0.5 mg, between about 0.05 mg and about 0.25 mg, between about 0.1 mg and about 0.25 mg, or about 0.23 mg of potassium phosphate monobasic. In some aspects, the combination formulation may contain between 0.1 mg and about 10 mg, between about 0.1 mg and about 5 mg, between about 0.1 mg and about 2.5 mg, between about 0.5 mg and about 10 mg, between about 0.5 mg and about 5 mg, between about 0.5 mg and about 2.5 mg, between about 1 mg and about 10 mg, between about 1 mg and about 5 mg, between about 1 mg and about 2.5 mg, or about 1.27 mg of potassium phosphate monobasic. In some such aspects, the combination formulation may contain about 3.2 mg pafolacyanine (equivalent to about 3.4 mg of pafolacyanine sodium), 14.4 mg sodium chloride, 0.23 potassium phosphate monobasic, 1.27 mg sodium phosphate dibasic heptahydrate per about 1.6 mL volume of solution. Thus, in some aspects, 3.2 mg of pafolacyanine (equivalent to about 3.4 mg of pafolacyanine sodium) per about 1.6 mL volume of solution of the combination formulation may be administered to the patient. In other aspects, the combination formulation may include additional or alternative compositions, ingredients, agents, solvents, buffers, preservatives, antioxidants, diluents, and the like.Additionally, in other aspects, the combined preparations may contain different amounts of any of the compositions, ingredients, agents, solvents, buffers, preservatives, antioxidants, diluents, and the like.

[0065] In some aspects, the pH of the composition can be between about 6 and about 9, between about 6 and about 8, between about 7 and about 8, or between about 7.1 and about 7.8. In other aspects, the pH can be different. In some aspects, a buffer or another composition can be used to adjust the pH of the formulation to between about 6 and about 9, between about 6 and about 8, between about 7 and about 8, or between about 7.1 and about 7.8. For example, the pH of the composition formulation can be adjusted using sodium hydroxide and / or hydrochloric acid. In other aspects, the pH can be adjusted using a different buffer, composition, substance, or solvent (e.g., perchloric acid, hydroiodic acid, nitric acid, sulfuric acid, potassium hydroxide, and / or calcium hydroxide).

[0066] In some aspects, the dosages discussed above may be provided in single-dose vials. For example, a single-dose vial may contain 3.2 mg of pafolacyanine (or 3.4 mg of pafolacyanine sodium) in about 1.6 mL of the combined formulation solution. In other aspects, the formulation may contain between about 1 mg and about 20 mg, between about 1 mg and about 10 mg, between about 1 mg and about 5 mg, between about 1 mg and about 4 mg, or between about 2 mg and about 4 mg of pafolacyanine, or between about 1 mg and about 10 mg, between about 1 mg and about 5 mg, between about 1 mg and about 4 mg, between about 2 mg and about 4 mg, or about 3.4 mg of pafolacyanine sodium. Such combined formulations may be a clear aqueous solution of dark blue-green color. In other aspects, the dosages may not be provided in single-dose vials. In such aspects, the vial (or other packaging) can contain any suitable number of doses (such as, for example, about 20 doses, about 10 doses, about 5 doses, or about 2 doses). In some aspects, the vial can be a sealed amber glass vial. In some aspects, the vials can be individually packaged and supplied in cartons containing at least about 5 vials, at least about 10 vials, at least about 20 vials, at least about 50 vials, or at least about 100 vials. In other aspects, the compositions can be in alternative forms, have different quantities, be packaged differently, and / or have different appearances.

[0067] In some aspects, the compositions disclosed herein may be diluted before administration to a patient. For example, the perforacyanin may be diluted in a dextrose solution (such as a 5% dextrose solution). In some such aspects, about 0.25 mg per kg of patient weight may be diluted in about 220 mL to about 250 mL of a 5% dextrose solution. In other examples, additional or alternative solutions and / or concentrates of solutions may be used to dilute the compositions of the present disclosure.

[0068] The technology described in the present invention and its advantages can be better understood by referring to the following examples. These examples are provided to describe certain aspects of the technology. It is not intended to limit the scope and spirit of the technology by providing these specific examples. It is understood by those skilled in the art that the full scope of the technology described in the present invention encompasses the subject matter defined by the claims appended hereto, and any changes, modifications or equivalents of these claims. EXAMPLES

[0069] Working Example Example 1: Detection of malignant lesions with a composition containing pafolacyanin The safety and efficacy of the compositions disclosed herein were evaluated in a randomized, multicenter, open-label study. The study enrolled 178 women who were diagnosed with or had a high clinical suspicion of ovarian cancer and were scheduled to undergo primary surgical cytoreduction, intermediate cytoreduction, or recurrent ovarian cancer surgery. 150 women with high suspicion or confirmed ovarian cancer received a composition containing pafolacyanin (administered at 0.25 mg / kg at least 1 hour before the start of fluorescent imaging). Among them, 134 women with a mean age of 60 years (range 33-81 years) underwent both regular optical imaging and fluorescent imaging evaluation (the Intent-to-Image set).

[0070] The study evaluated a population of patients with at least one evaluable ovarian cancer lesion confirmed by central pathology review that was detected with a composition containing pafolacyanin under fluorescent light, but not detected under normal light or palpation, and not otherwise identified for resection prior to surgery. Detection rates were estimated in women who underwent both normal light and fluorescence (Intent-to-Image set). See Table 1. Table 1 [Table 1-1] [Table 1-2]

[0071] The patient-level false positive rate for the composition containing pafolacyanin in near-infrared fluorescence for the detection of ovarian cancer lesions confirmed by central pathology was 20.2% (95% confidence interval (13.7%, 28.0%)).

[0072] Example 2: Pharmacodynamics The tumor-to-background ratio was examined at various mass doses of pafolacyanin. A high tumor-to-background ratio was observed at a dose of 0.25 mg pafolacyanin per kg of patient weight.

[0073] Example 3: In vitro pharmacodynamics KB cells (a derivative of HeLa cells) overexpressing FR were combined with [3H]-folate in the presence of increasing concentrations (0.1 nM-1 μM) of pafolacyanin (L-isomer), OTL0039 (D-isomer) or folate. Pafolacyanin clearly exhibited high affinity (KD=10.4 nM) for FR. This compared well with the binding affinity of folate (KD=7.4 nM) as shown in Figure 1. In contrast, the D-isomer of pafolacyanin (OTL0039) exhibited a relatively low affinity (KD=81.8 nM) for the receptor. Pafolacyanin competed well with [3H]-folate for the receptor.

[0074] Example 4: In vivo Pharmacodynamics Whole-body imaging and tissue distribution of pafolacyanin and OTL0039 were performed in female nu / nu mice bearing FR-positive KB cell tumor xenografts. Results showed fluorescence primarily in the tumor 2.5 hours after intravenous (IV) administration of 10 nmol pafolacyanin, and fluorescent signals were also observed in the kidney (Figures 2A and 2C). Uptake of pafolacyanin in the kidney was expected, since the apical membrane of the renal proximal tubules has been clearly shown to express high levels of FR. All other normal tissues showed little or no fluorescent signal, resulting in an excellent tumor-to-normal tissue fluorescence ratio. The tissue distribution pattern of OTL0039 (Figure 2D) was similar to that of pafolacyanin (Figure 2C), but had a weaker fluorescent signal intensity than pafolacyanin in both tumor (Figure 2B) and kidney. This observation was consistent with the lower binding affinity of the D-isomer to FR compared to the L-isomer. Tumor fluorescence was brighter in mice receiving pafolacyanin compared to other folate-conjugated near-infrared dyes (eg, LS288, IR800 and ZW800).

[0075] The optimal dose range of pafolacyanin in terms of tumor-to-background ratio in mice bearing FR-positive tumors was 1-30 nmol / mouse, with lower fluorescence intensity observed in tumors at lower doses (0.3 nmol) and higher fluorescence intensity observed in other tissues at doses above 30 nmol. In a second experiment, pafolacyanin whole-body imaging and tissue distribution was again investigated; however, this experiment was performed in female nu / nu mice bearing FR-negative A549 cell tumor xenografts. In this study, fluorescence was mainly noted in the kidneys, and no fluorescence was observed in FR-negative tumors (Figure 3). Thus, pafolacyanin did not target FR-negative tumors in vivo, clearly demonstrating the specificity of the imaging agent for FR-positive tumors.

[0076] Example 5: Secondary Pharmacodynamics Pafolacyanin (5 μM) was evaluated in an in vitro ligand binding screen (ExpresSProfile; Cerep 100010238), which assessed the potential for pafolacyanin "off-target" binding to a broad panel of 55 pharmacologically relevant receptors, ion channels and transporters. Results from that binding experiment showed that positive values ​​for % specific binding of pafolacyanin ranged between 0.2-29.7% and negative values ​​ranged between -0.3%-16.8%. These data suggest no significant affinity of pafolacyanin for a comprehensive set of off-target proteins and support the specificity of pafolacyanin for FR binding.

[0077] Example 6: Pharmacokinetics Mean maximum plasma concentration of pafolacyanin (C max ) was 59.1 ± 5.94 ng / mL, and the area under the plasma concentration-time curve to infinity (AUC inf ) was 63.6 ± 12.6 ng·hr / mL.

[0078] discharge The elimination half-life of pafolacyanin is 0.44±0.23 hours, with a mean plasma clearance of 28.6±4.97 L / hour.

[0079] metabolism Pafolacyanine sodium is not metabolized by cytochrome P450 (CYP) enzymes.

[0080] excretion After a single IV infusion of radiolabeled pafolacyanine sodium, approximately 35% of the dose was recovered in the urine (19.1%) and feces (15.8%) after approximately 3-5 weeks.

[0081] Specific populations No clinically significant differences in the pharmacokinetics of pafolacyanine were identified based on age 18–89 years, weight 41.6–133.6 kg, mild to moderate renal impairment (creatine clearance (CLcr) 30–89 mL / min), or mild to moderate hepatic impairment (total bilirubin <3 × upper limit of normal (ULN) and aspartate transaminase (AST) >ULN).

[0082] absorption Following a single IV administration of pafolacyanin to male and female rats, systemic exposure to pafolacyanin (AUClast) appeared similar between males and females at the 46.3 mg / kg dose level (Table 2). There was a slight trend toward higher AUClast values ​​in females vs. males at the two lower dose levels; however, due to sparse blood sampling, the significance of this slight trend could not be assessed. AUClast values ​​were calculated, but the time associated with the last measurable concentration was variable (Tlast; 5 min to 24 h), which confounded the ability to assess dose proportionality. Terminal data were generally sparse and / or the terminal phase was not well defined (Figure 4). Thus, parameters dependent on accurate determination of the terminal phase (AUCinf, T 1 / 2 , CL, Vz) were not reported.

[0083] Following a single IV administration of pafolacyanin to male and female dogs, systemic exposure to pafolacyanin (C0, AUClast) appeared similar between males and females at all dose levels (Table 3). As with rats, AUClast values ​​in dogs were calculated from a range of concentration-time data with Tlast values ​​ranging from 5 min to 4 h. Furthermore, dogs with reportable TK at 0.9 mg / kg were limited (N=1-2). In conclusion, dose proportionality was not evaluated for dogs. Terminal data were generally sparse (i.e., 2 or fewer measurable concentrations in the terminal phase (Figure 4)). As with rats, parameters dependent on accurate determination of the terminal phase (AUCinf, T 1 / 2 , CL, Vz) were not reported. Table 2 [Table 2] Table 3 [Table 3]

[0084] Repeated administration Male and female rats received weekly IV doses of pafolacyanin (0.9 mg / kg, 9.3 mg / kg, and 13.9 mg / kg) for 14 days. TK parameters are summarized in Table 4. On both days 1 and 13, the increase in pafolacyanin exposure (AUCinf) tended to be slightly greater than dose proportional, although it is noted that sparse blood sampling was used so variability in AUCs could not be assessed. After a 14.9-fold increase in dose, AUCinf increased approximately 18-fold on day 1 and approximately 20-fold on day 13 (for both sexes combined). There were no obvious gender-related differences, and no changes in accumulation or TK parameters were evident after repeated weekly dosing for 3 weeks. CL was slow (relative to hepatic blood flow) and Vss was moderate. Mean T 1 / 2 ranged from approximately 6 to 11 hours. Table 4 [Table 4]

[0085] In addition, male and female dogs also received weekly IV doses of pafolacyanin (3.1 mg / kg, 30.8 mg / kg, and 46.3 mg / kg) over a 14-day period. Overall, drug concentrations increased as dose was increased, although overlapping mean profiles at the two highest dose levels was noted. Systemic exposure to pafolacyanin (AUClast) was approximately dose-proportional over the range of doses administered (0.9-13.9 mg / kg) and within the observed variability (Table 5). No obvious or consistent gender differences were present, and no changes in accumulation or TK were evident after repeated weekly dosing over a 3-week period. On both days 0 and 13, CL was slow (relative to hepatic blood flow) and Vss was moderate. Mean T 1 / 2 ranged from approximately 13 to 18 hours. Table 5 [Table 5]

[0086] Overall, in both species, the mean CL was found to be slow relative to hepatic blood flow, and the mean Vss values ​​were higher than the total body water volume, 668 mL / kg and 693 mL / kg for rats and monkeys, respectively, suggesting distribution of perforacyanin to tissues. There were no observable changes in TK with repeated dosing, and PK for both sexes was similar.

[0087] distribution Average distribution volume (V z ) was 17.1 ± 5.99 L, indicating distribution to tissues.

[0088] The plasma protein binding of pafolacyanin is 93.7%. No significant partitioning to erythrocytes was observed.

[0089] Tissue distribution was evaluated in male and female rats after receiving a single 2 mg / kg (approximately 25 μCi / kg) IV dose of [14C]paforacyanine. Radioactivity levels (paforacyanine equivalents) in tissues were determined over time using QWBA. Radioactivity was rapidly (within 15 minutes) and widely distributed to most tissues. Tissue-to-plasma ratios for radioactivity were >1.0 for all tissues in male and female animals, except bone (femur), brain, eye, and fat. This indicates the distribution of radioactivity from [14C]paforacyanine to tissues. The overall highest concentration of radioactivity at 24 hours post-dose was measured in the kidneys of male and female rats. Approximately 11% of the dose remained in the kidneys at 96 hours post-dose. The lowest concentrations of radioactivity were found in the brain and eye. The concentrations of [14C]pafolacyanine-derived radioactivity in the eye, uvea, and skin of male pigmented (Long Evans) rats were generally similar to those found in albino Sprague Dawley rats, suggesting that there was no affinity for [14C]pafolacyanine-derived radioactivity for melanin-containing tissues.

[0090] Plasma protein binding The plasma protein binding of pafolacyanin was studied in vitro in fresh plasma from humans, Sprague Dawley rats, and beagle dogs using equilibrium dialysis. Pafolacyanin was introduced into plasma samples in duplicate resulting in a final concentration of 5 μM. The mean (±SD) percentage of plasma protein bound pafolacyanin was 99.1±0.1%, 98.6±0.1%, and 93.7±1.5% for rats, dogs, and humans, respectively. Pafolacyanin was highly bound to plasma proteins in all species examined.

[0091] PK parameters in humans A total of 73 potentially eligible healthy volunteers provided written informed consent to participate in a Phase 1a study to evaluate the safety and pharmacokinetics of pafolacyanine versus placebo. Fifty-four were found suitable to participate, of which 30 were enrolled in the study. Twenty-four subjects were Caucasian, one was Black / African American, one was Asian, one was mixed race, and three were of another race. Table 6 shows the characteristics of the study population. Table 6 [Table 6]

[0092] The pharmacokinetic (PK) analysis population was defined as all subjects who were randomized and received study treatment. Of note, drug infusion was stopped prematurely in one subject and discontinued in another. The pharmacokinetics of pafolacyanine following a single IV dose of 0.25 mg / kg, 0.05 mg / kg, 0.1 mg / kg, or 0.2 mg / kg were determined using noncompartmental analysis; key PK parameters are summarized in Table 7. Table 7 [Table 7] a OTL38 dissolved in 20 mL 0.9% NaCl over a 10-60 min infusion b OTL38 dissolved in 220 mL of 5% dextrose solution over a 60-minute infusion c OTL38 with pretreatment with antihistamine (clemastine)

[0093] Maximum concentrations (Cmax) for each dose were obtained near the end of the infusion. After completion of the infusion, plasma concentrations declined rapidly. Drug concentration-time profiles obtained at the higher doses suggest that the decline in plasma concentrations was (at least) biphasic. Exposure (as measured by Cmax and AUClast) increased with increasing dose of pafolacyanine. When the dose was increased 8-fold (from 0.25 to 0.2 mg / kg), Cmax values ​​increased approximately 12-fold, whereas AUClast values ​​increased approximately 22-fold. This suggested that the increase in exposure with dose was more than dose-proportional. Clearance values ​​(CL) decreased with dose. This indicates that clearance may be saturable at higher doses, consistent with a more than dose-proportional increase in exposure. Overall, clearance was moderate to low relative to hepatic blood flow (approximately 84 L / hr for a 70 kg subject) and varied from approximately 29 to 10 L / hr over the dose range. Drug half-life values ​​were short, ranging from approximately 0.4 to 2.7 hours.

[0094] Example 7: Safety evaluation of a composition containing pafolacyanin The safety of the composition of the present disclosure was evaluated in three open-label clinical trials, two of which (N=150 and N=44) were in patients with ovarian cancer, and one of which (N=100) was in patients with lung cancer.A total of 294 patients received 0.25mg / kg of the composition of the present disclosure via intravenous administration.The mean age of the patients was 63.5 years; 51% were 65 years or older; 89% of the patients were female; and 84% of the patients were white.

[0095] Adverse reactions occurring in >1% of patients were: nausea (15%), vomiting (5.8%), abdominal pain (2.7%), flushing (1.7%), dyspepsia (1%), chest discomfort (1%), pruritus (1%) and hypersensitivity (1%). In 2.4% of patients, these adverse reactions occurred during administration of the composition. Reactions typically occurred within 15 minutes of beginning the infusion.

[0096] Example 8: Ex vivo evaluation of camera sensitivity and specificity using pafolacyanin Sodium pafolacyanine (2 mg / mL, 1414.2 g / mol (or Da), λex=774-776 nm, λem=794-796 nm, in 1.6 mL PBS) was serially diluted 100-fold from 100 μM to 1 pM using PBS or 5% dextrose solution. The dilutions were used as negative controls. ICG (10 μM in the same dilutions) was used as a positive control. The serially diluted samples were transferred to a 96-well black plate and imaged with the camera under evaluation to determine the sensitivity of the camera.

[0097] penetration depth Fresh pork was purchased from Butcher Block and cut into 1 cm thick pieces. 100 μL of 10 μM sodium perforacyanine solution was dispensed into a glass tube and placed under the pork. Images were taken using the camera under evaluation. Increasing numbers of pork pieces were imaged to determine the penetration depth of the camera.

[0098] Preliminary phantom testing was performed using an agarose-based gel phantom to construct an agar phantom with the geometry of the desired organ. Glass tubes ranging in size from 3 mm to 10 mm with 10 μM sodium perforacyanine (100 μL) were placed in the agar phantom gel at various measurement distances and imaged to determine the penetration depth of the camera.

[0099] field of view A 100 μL solution of 10 μM sodium pafolacyanine was dispensed into nine glass tubes and positioned on the center and eight arrows as shown in Figure 5. Images were taken using the camera under evaluation to determine uniform field of view.

[0100] Example 9: In vivo evaluation of camera sensitivity and specificity using pafolacyanin KB cells (approx. 2 x 10 6HeLa cells (a derivative of a human cervical carcinoma cell line expressing the folate receptor / cell) and A549 cells (an alveolar basal carcinoma cell line that does not express the folate receptor) were obtained from ATCC (Rockville, MD) and grown as monolayers for at least six passages at 37°C in a 5% carbon dioxide:95% air-humidified atmosphere using folate-free or regular RPMI-1640 medium (Gibco, NY) containing 10% heat-inactivated fetal bovine serum (Atlanta Biological, GA) and 1% penicillin-streptomycin (Gibco, NY) before use in the study.

[0101] Athymic female nude (nu / nu) mice (5-6 weeks old, 18-20 g) were purchased from Envigo (Indianapolis, IN) and maintained on gamma-irradiated, folate-deficient diet (Envigo, IN) for 2 weeks before the start of the study. Animals were housed 5 / cage on barrier, pathogen-free covered racks. Autoclaved tap water and food were provided when needed. Animals were housed in a pathogen-free environment with a standard 12-h light / dark cycle for the duration of the study.

[0102] Seven-week-old female nu / nu mice were cultured at 1.0 × 10 6 KB or A549 cells / mouse were inoculated subcutaneously in the shoulder or neck. The A549 xenograft model was used as a negative control to determine the receptor specificity of sodium pafolacyanine. Tumor growth was measured vertically every 2 days using a caliper (body weight was also monitored on the same schedule) and tumor volume was calculated as 0.5×L×W. 2 (L = longest axis and W = axis perpendicular to L in millimeters). 3After reaching a median 100 mg / kg (volume unit), animals (5 mice / group) were intravenously injected with an appropriate dose of pafolacyanine sodium (2-4 nmol, i.e., 0.0125-0.25 mg / kg HDE) in PBS or 5% dextrose solution. For whole-body imaging and biodistribution studies, animals were euthanized by CO2 asphyxiation 2-3 h after administration of pafolacyanine sodium. For time-dependent studies, animals were imaged under anesthesia using isoflurane. After whole-body imaging, animals were dissected and selected tissues were analyzed for fluorescent activity using an objective camera, and tissue ROIs were calculated using appropriate software. The tumors were further broken down into smaller pieces (e.g., 1 / 2, 1 / 4, 1 / 8, 1 / 16, etc.) and placed either directly or surgically into organs of interest (e.g., lungs, liver, kidneys, etc.) of larger animals (e.g., pigs, dogs, human cadavers, or other animals) and imaged to determine the sensitivity of the camera (i.e., to examine the camera's ability to determine the smallest tumors in an appropriate environment).

[0103] Example 10: Embryo-Fetal Development (EFD) Animal Study Pafolacyanin was administered intravenously during the period of organogenesis, i.e., from gestational day (GD) 6 to GD 17 in rats at doses of 0.015 mg / kg / day, 0.15 mg / kg / day, and 1.5 mg / kg / day (human equivalent doses (HEDs) were 0.002 mg / kg / day, 0.24 mg / kg / day, and 0.242 mg / kg / day), and from GD 7 to GD 20 in rabbits at doses of 0.3 mg / kg / day, 1 mg / kg / day, and 3 mg / kg / day (HEDs were 0.97 mg / kg / day, 0.323 mg / kg / day, and 0.968 mg / kg / day). No drug-related maternal or embryo-fetal developmental toxicity was observed. No observed side effects levels were 1.5 mg / kg / day in rats and 3 mg / kg / day in rabbits. The predicted systemic exposure was 158-fold (rats) and 570-fold (rabbits) the human exposure at a human dose of 0.25 mg pafolacyanin per kg patient body weight, based on a comparison of the areas under the plasma concentration-time curve (AUC).

[0104] Although the present disclosure has been described with reference to certain specific embodiments, it will be understood by those skilled in the art that various modifications may be made and equivalents may be substituted without departing from the scope of the present disclosure and the appended claims. In addition, many modifications may be made to adapt a particular situation or material to the teachings of the present disclosure without departing from its scope. Thus, the present disclosure is not limited to the exact embodiments disclosed, but is intended to include all aspects that fall within the scope of the appended claims.

[0105] All patents, patent applications, publications, and descriptions set forth above are incorporated herein by reference in their entirety.

Claims

1. A composition configured for administration to a subject seeking visualization of cancerous tumors, cells, and / or lesions, the composition comprising between about 2 mg and about 4 mg of pafolacyanin or a pharmaceutically acceptable salt thereof per about 1.6 mL volume of solution, wherein the pafolacyanin or a pharmaceutically acceptable salt thereof is configured to bind to a folate receptor in tissue, and the composition allows visualization of the tissue under excitation light of a wavelength absorbable by the pafolacyanin or a pharmaceutically acceptable salt thereof.

2. The composition described in claim 1, wherein the tissue comprises ovarian cancer, a malignant lung lesion or a non-malignant lung lesion.

3. The composition described in claim 1, characterized in that the pafolacyanin or a pharmaceutically acceptable salt thereof is administered at a dose of between about 0.01 mg and about 0.05 mg per kg body weight of the subject.

4. The composition described in claim 1, wherein the composition is configured to be administered intravenously to the subject.

5. The pharmaceutically acceptable salt of pafolacyanine, having the following chemical structure: 【Chemistry 4】 10. The composition of claim 1, wherein 6. The composition of claim 1, further comprising at least one of a chloride salt or a phosphate salt.

7. The composition of claim 6, wherein the chloride salt is sodium chloride or calcium chloride.

8. The composition described in claim 7, wherein the composition contains between about 1 mg and about 50 mg of the chloride salt per about 1.6 mL volume of solution.

9. The composition of claim 6, wherein the phosphate is potassium phosphate or sodium phosphate.

10. The composition of claim 9, wherein the composition comprises between about 0.010 mg and about 5 mg of potassium phosphate per about 1.6 mL volume of solution.

11. The composition of claim 9, wherein the composition contains between about 0.1 mg and about 10 mg of sodium phosphate per about 1.6 mL volume of solution.

12. The composition described in claim 1, wherein the pH of the composition is between about 6 and about 8.

13. The composition described in claim 1, wherein the composition is configured to be administered to the subject about 1 to about 24 hours before the visualization.

14. The composition of claim 1, wherein the pafolacyanine or a pharmaceutically acceptable salt thereof fluoresces upon exposure to light in the near-infrared range.

15. The composition described in claim 14, wherein the excitation light is between about 750 nm and about 790 nm.

16. The composition described in claim 1, characterized in that the pafolacyanin or its pharmaceutically acceptable salt is administered at a dose of between about 0.01 mg and about 0.05 mg per kg body weight of the subject.

17. The composition described in claim 1, wherein the composition is diluted prior to administration to the subject.

18. The composition of claim 17, wherein the composition is diluted with a dextrose solution.

19. The composition described in claim 18, wherein the composition is diluted with about 220 mL to about 250 mL of the dextrose solution.

20. The composition of claim 1, wherein the composition is used to identify tissue during surgery.

21. The composition described in claim 20, wherein the surgery is for ovarian cancer.

22. The composition described in claim 20, wherein the surgery is for lung cancer.

23. The composition of claim 1, wherein the composition comprises an injectable solution.

24. The composition of claim 23, wherein the solution is blue-green.

25. The composition of claim 23, wherein the solution is transparent or translucent.

26. The composition described in claim 1, wherein the composition is configured to be administered to the subject over a period of time.

27. ​​The composition described in claim 26, wherein the certain period of time includes about 1 to about 3 hours.

28. The composition of claim 1, wherein the composition is contained in a vial.

29. The composition of claim 28, wherein the vial is a single-dose vial.

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