Compositions, systems, and methods for treating cancer using tumor therapeutic sites and chimeric antigen receptor (CAR) immune cells.

Abstract

This invention discloses compositions, systems, and methods for reducing the viability of cancer cells, treating cancer, and preventing an increase in the volume of tumors present in the body. The systems and methods include administering at least one composition comprising at least one chimeric antigen receptor (CAR) immune cell and simultaneously applying an alternating electric field.

Description

Technical Field

[0001] The present invention is a composition comprising at least one chimeric antigen receptor (CAR) immune cell for use in a method of treating cancer in a subject, the method comprising: (1) applying an alternating electric field to a target region of the subject for a certain period of time; (2) administering the composition to the subject, relates to a composition.

[0002] This application claims the benefit of U.S. Provisional Application No. 63 / 493,131, filed Mar. 30, 2023. The entire contents of the above patent application(s) are hereby expressly incorporated herein by reference. Statement Regarding Federally Sponsored Research None.

Background Art

[0003] A tumor therapeutic field (TT field) is a low-intensity (e.g., 1–3 V / cm) alternating electric field in a medium-frequency range (e.g., 100–500 kHz) that targets solid tumors by inhibiting mitosis. This non-invasive treatment targets solid tumors and is described, for example, in U.S. Patents 7,016,725, 7,089,054, 7,333,852, 7,565,205, 8,244,345, 8,715,203, 8,764,675, 10,188,851, and 10,441,776. The TT field is typically transmitted through two pairs of transducer arrays that generate a field perpendicular to the tumor being treated, with the electrode arrays constituting each of these pairs of transducer arrays located opposite the part of the body being treated. Specifically, in the OPTUNE® system, one pair of electrodes is positioned on the left and right (LR) sides of the tumor, and the other pair of electrodes is positioned on the front and back (AP) sides of the tumor. The TT field is approved for the treatment of glioblastoma multiforme (GBM) and can be delivered, for example, via the OPTUNE® system (Novocure Limited, St. Helier, Jersey), which includes a transducer array placed on the patient's shaved scalp.

[0004] In the OPTUNE® device, each transducer array used for TT field delivery consists of a series of ceramic disc electrodes bonded to the patient's skin (e.g., the patient's shaved scalp for GBM treatment) via a layer of conductive medical gel. The purpose of the medical gel is to conform to the body's contours and provide good electrical contact between the array and the skin, thereby allowing the gel interface to crosslink the skin and reduce interference. The device is intended to be worn continuously by the patient for 2-4 days, after which it is removed for hygienic care and reshaving (if necessary), and its new array set is reattached. Thus, the medical gel remains in substantially continuous contact with the area of ​​the patient's skin for 2-4 days at a time. In addition, the array can be shifted a few centimeters in either direction, allowing the skin to heal with each treatment period. Therefore, the area of ​​skin that was covered with electrodes / gel for 2-4 days is exposed for 2-4 days with the replaced electrodes slightly shifted, after which the device can be reapplied to the original area of ​​skin for the next 2-4 days. [Overview of the project] [Problems that the invention aims to solve]

[0005] Chimeric antigen receptor T cells (CAR-T cells) are currently used as a conventional cancer treatment in certain hematological malignancies, such as B-cell acute lymphoblastic leukemia (B-ALL) and diffuse large B-cell lymphoma (DLBCL). However, the use of CAR-T cells to treat solid tumors remains hindered by the immunosuppressive tumor microenvironment (TME) (Kozani et al. (2022) Front.Immunol., 13:795164).

[0006] Numerous studies have been conducted to understand which factors lead to better outcomes for CAR-T cells in order to utilize CAR-T therapy in solid tumors. CXCL9 and CXCL10 from TME recruit T cells to tumors via CXCR3 expression on T cells. Other chemokines such as CXCL16, CXCL1, and CCL19 also contribute to T cell recruitment (Tantalo et al. (2021) J Immunother Cancer, 9:e002555, Seitz et al. (2022) British J of Cancer, 126:1470-1480). In addition, it has been previously shown that intratumoral expression of both RANTES (CCL5) and IL-15 attracts CAR T cells and increases the survival of mice carrying neuroblastoma tumors (Nishio et al. (2014) Cancer Res, 74(18):5195-205). [Brief explanation of the drawing]

[0007] [Figure 1] The graph shows the analysis of serum from mice treated in a TT field compared to serum from mice treated with pseudoheat (heat sham) after cytokine array incubation. Multiple mouse cancer models were used. [Modes for carrying out the invention]

[0008] Before describing in detail, in illustrative language and results, at least one embodiment of the concept(s) of the present invention, it should be understood that the application of the concept(s) of the present invention is not limited to the details of configurations and component arrangements described below. The concept(s) of the present invention can be carried out in other embodiments or in various ways. Therefore, the language used herein is intended to give as broad a scope and meaning as possible. Furthermore, the embodiments are intended to be illustrative, not exhaustive. It should also be understood that the expressions and terms used herein are for illustrative purposes only and should not be considered limiting.

[0009] Unless otherwise defined herein, scientific and technical terms used in connection with the concepts(s) of the present invention shall have the meaning generally understood by those skilled in the art. Furthermore, unless otherwise required by context, singular terms include plurals, and plural terms include singular terms. The techniques and procedures described herein are generally carried out in accordance with conventional methods well known in the art, as described in the various general and more specific references cited and explained throughout this specification. The nomenclature used in connection with analytical chemistry, organic synthesis chemistry, medicinal chemistry, and pharmaceutical chemistry described herein, as well as their experimental procedures and techniques, are well known and commonly used in the art. Standard techniques are used for chemical synthesis and chemical analysis.

[0010] All patents, published patent applications, and non-patent publications described herein represent the skill level of experts in the art relating to the currently disclosed inventive concepts(s). All patents, published patent applications, and non-patent publications referenced in any part of this Application are expressly incorporated by reference in whole to the same extent that individual patents or publications are specifically and individually indicated to be incorporated by reference.

[0011] All compositions, assemblies, systems, kits, and / or methods disclosed herein can be prepared and performed without excessive experimentation in light of this disclosure. While the compositions, assemblies, systems, kits, and methods of the concept(s) of the present invention are described in relation to specific embodiments, it will be apparent to those skilled in the art that variations can be applied to the compositions and / or methods, and the steps or order of steps of the methods, described herein without departing from the concept(s), spirit, and scope of the present invention. These similar substitutions and modifications, apparent to those skilled in the art, are considered to be within the spirit, scope, and concept of the concept(s) of the invention as defined in the appended claims.

[0012] The following terms used in accordance with this disclosure shall be understood to have the following meanings unless otherwise specified:

[0013] The use of the terms “a” or “an” in a claim and / or specification in combination with the term “including” may mean “one,” but also coincides with the meanings of “one or more,” “at least one,” or “one or two or more.” Therefore, unless the context explicitly indicates otherwise, the terms “a,” “an,” and “the” include multiple references. Thus, for example, “compound” may refer to one or more compounds, two or more compounds, three or more compounds, four or more compounds, or more compounds. The term “multiple” refers to “two or more.”

[0014] The use of the term "at least one" is understood to include quantities greater than one, including but not limited to one, such as two, three, four, five, ten, fifteen, twenty, thirty, forty, fifty, and one hundred. The term "at least one" may extend to 100 or even more, depending on the term it is attached to, and the quantity 100 / 1000 is not considered limiting, as satisfactory results may be obtained with higher restrictions. In addition, the use of the term "at least one of X, Y, and Z" is understood to include X only, Y only, Z only, and any combination of X, Y, and Z. The use of ordinal terms (such as "first," "second," "third," and "fourth") is intended solely to distinguish two or more items and does not imply any order or importance of one item being superior to another, or any additional order.

[0015] The use of the term “or” in a claim is used to mean an inclusive “and / or” unless it is explicitly indicated that it refers only to the alternatives, or unless the alternatives are mutually exclusive. For example, the condition “A or B” is satisfied by any of the following: A is not true (or exists) and B is not false (or does not exist); A is not true (or exists) and B is true (or exists); and both A and B are true (or exist).

[0016] In this specification, any reference to “one embodiment,” “embodiment,” “several embodiments,” “example,” “for example,” or “example” means that a particular element, feature, structure, or characteristic described in relation to an embodiment is encompassed in at least one embodiment. For example, even if the phrases “in some embodiments” or “in one example” appear in different parts of the specification, they do not necessarily all refer to the same embodiment. Furthermore, all references to one or more embodiments or examples shall be construed as not limiting the claims.

[0017] Throughout this application, the term “approximately” is used to indicate that a value includes variations in inherent errors of the composition / apparatus / device, the method used to determine the value, or variations that exist between the subjects of study. For example, but not limited to, where the term “approximately” is used, a given value may vary from a given value by plus or minus 20 percent, or 15 percent, or 12 percent, or 11 percent, or 10 percent, or 9 percent, or 8 percent, or 7 percent, or 6 percent, or 5 percent, or 4 percent, or 3 percent, or 2 percent, or 1 percent, as is appropriate for performing the disclosed method and as understood by those skilled in the art.

[0018] As used herein and in the claims, the terms “comprising” (and any form of “comprise,” such as “comprises,” etc.), “having” (and any form of “have,” such as “has,” etc.), “including” (and any form of “includes,” such as “include,” etc.), and “containing” (and any form of “contains,” such as “contain,” etc.) are inclusive or unrestricted and do not exclude any additional elements or method steps not mentioned herein.

[0019] As used herein, the term “or any combination thereof” refers to all permutations and combinations of the enumerated items preceding the term. For example, “A, B, C, or any combination thereof” is intended to include at least one of A, B, C, AB, AC, BC, or ABC, and also at least one of BA, CA, CB, CBA, BCA, ACB, BAC, or CAB, where the order is important in a particular context. Continuing this example, combinations that explicitly include repetitions of one or more items or terms are explicitly included, such as BB, AAA, AAB, BBC, AAABCCCC, CBBAAA, CABABB, etc. A person skilled in the art will understand that, unless specifically evident from the context, there is generally no limit to the number of items or terms that can be combined.

[0020] As used herein, the term "substantially" means that the event or situation described thereafter occurs completely, or that the event or situation described thereafter occurs within a wide range or degree. For example, when related to a particular event or situation, the term "substantially" means that the event or situation described thereafter occurs at least 80% of the time, at least 85% of the time, at least 90% of the time, or at least 95% of the time. For example, the term "substantially adjacent" can mean that two items are 100% adjacent to each other, that two items are very close to each other but not 100% adjacent, or that a portion of one of the two items is not 100% adjacent to the other item but is very close to the other item.

[0021] The term "pharmaceutically acceptable" refers to compounds and compositions suitable for administration to humans and / or animals without excessive side effects such as toxicity, irritation, and / or allergic reactions, with a reasonable benefit / risk ratio.

[0022] As used herein, the terms "patient" or "subject" include human and veterinary subjects. "Mammal" for the purposes of treatment refers to any animal classified as a mammal, including, but not limited to, humans, domestic animals, farm animals, non-human primates, and other animals with mammary tissue.

[0023] The term "treatment" refers to both therapeutic treatments and prophylactic measures. Persons who require treatment include not only those who already suffer from a particular symptom / disease / infection, but also those who are at risk of developing a particular symptom / disease / infection (such as those who require prophylactic measures), but are not limited thereto. The term "treating" refers to administering a drug / element / method to a patient for the purpose of treatment and / or prevention.

[0024] As used herein, the terms "therapeutic composition" or "pharmaceutical composition" refer to agents that can be administered in vivo to provide a therapeutic and / or prophylactic effect.

[0025] Administering a therapeutically effective amount or a prophylactically effective amount is intended to provide a therapeutic benefit in the treatment, prevention, and / or management of a disease, condition, and / or infection. The specific amount that is therapeutically effective can be readily determined by a skilled medical practitioner and can vary depending on factors known in the art such as the type of the condition / disease / infection, the patient's medical history and age, the stage of the condition / disease / infection, and the co-administration of other agents (but not limited thereto).

[0026] As used in accordance with the concepts (s) of the present invention, the term "effective amount" refers to an amount of a biologically active molecule or its conjugate or derivative, or an amount of a treatment protocol (e.g., an alternating electric field), sufficient to exert a detectable therapeutic effect without undue side effects (such as toxicity, irritation, allergic reaction, etc. (but not limited thereto)) commensurate with a reasonable benefit / risk ratio. The therapeutic effect can include, but is not limited to, preventing, inhibiting, or alleviating the occurrence of at least one condition, disease, and / or infection. The effective amount for a subject varies depending on the type of the subject, the physical constitution and health status of the subject, the nature and severity of the condition / disease / infection to be treated, the administration method, the treatment period, the nature of the combination therapy (if any), the specific formulation used, etc. Therefore, the exact effective amount cannot be specified in advance. However, the effective amount in a particular situation can be determined by a person skilled in the art by performing routine experiments based on the information provided herein.

[0027] As used herein, the term “concurrent therapy” is to be used interchangeably with the terms “combination therapy” and “adjunctive therapy” and is understood to mean that a patient in need of treatment is treated or administered with another drug for a medical condition / disease / infection in conjunction with the treatment disclosed herein. This concurrent therapy may be sequential therapy in which the patient is first treated with one treatment protocol / pharmaceutical composition, then with another treatment protocol / pharmaceutical composition, or two treatment protocols / pharmaceutical compositions are administered simultaneously. Furthermore, it will be understood that one administration step (e.g., administration in a TT field) may occur over a longer period than other administration steps (i.e., oral or injectable administration of a substance). In these examples of various administration periods, the term “concurrently” will be understood to mean that the shorter administration step completely overlaps with the longer administration step. However, the term “concurrently” includes performing a shorter administration step at any point between longer administration steps (e.g., at the beginning, middle, or end of a longer administration step, or any other period between them), as well as performing a shorter administration step completely one or more times within the duration of a longer administration step. Therefore, the term "simultaneously" does not require that the two administration steps be performed over the exact same duration.

[0028] As used herein, the terms “administration” and “administering” are understood to encompass all known routes of administration in the art, including, but not limited to, oral, topical, transdermal, parenteral, subcutaneous, intranasal, mucosal, intramuscular, intraperitoneal, intravitreous, and intravenous routes, and to encompass both topical and systemic applications. In addition, the compositions of this disclosure (and / or methods of administration thereof) may be designed to provide delayed release, controlled release, or sustained release using formulation techniques well known in the art.

[0029] As used herein, the term “target region” refers to a region containing all or part of the cancer, cancer cells, and / or tumor being treated.

[0030] Turning to the concept(s) of the present invention, a combination therapy for cancer is disclosed herein. The combination therapy involves using chimeric antigen receptor (CAR) immune cells and an alternating electric field (e.g., a TT field) simultaneously. The addition of an alternating electric field (e.g., a TT field) and CAR immune cells provides a synergistic effect of improvement in the treatment of cancer.

[0031] Certain non-limiting embodiments of this disclosure relate to a method for reducing the viability of cancer cells. The method comprises (1) applying an alternating electric field to cancer cells for a certain period of time, and (2) administering at least one composition to the cancer cells, wherein the at least one composition comprises at least one chimeric antigen receptor (CAR) immune cell. Such a method may be carried out in vitro or in vivo.

[0032] Certain additional non-limiting embodiments of this disclosure relate to a method for treating cancer in a subject. The method includes (1) applying an alternating electric field to a target area of ​​the subject for a certain period of time, and (2) administering to the subject at least one composition, wherein the at least one composition comprises at least one CAR immune cell.

[0033] Certain additional non-limiting embodiments of the present disclosure relate to a method for reducing the volume of a tumor present in a living organism, wherein the tumor comprises a plurality of cancer cells. The method comprises (1) applying an alternating electric field to a target region of a subject for a certain period of time, and (2) administering to the subject at least one composition, wherein the at least one composition comprises at least one CAR immune cell.

[0034] Certain additional non-limiting embodiments of the present disclosure relate to a method for preventing an increase in the volume of a tumor, where the tumor is present in the body of a living organism and comprises a plurality of cancer cells. The method comprises (1) applying an alternating electric field to a target area of ​​a subject for a certain period of time, and (2) administering to the subject at least one composition, wherein the at least one composition comprises at least one CAR immune cell.

[0035] Certain additional non-limiting embodiments of the present disclosure relate to a method comprising the steps of (1) applying an alternating electric field to a target region of a subject for a certain period of time, and (2) administering to the subject at least one composition comprising at least one chimeric antigen receptor (CAR) immune cell.

[0036] Certain additional non-limiting embodiments of the present disclosure relate to a composition comprising at least one chimeric antigen receptor (CAR) immune cell for use in a method of treating cancer in a subject, the method comprising: 1) applying an alternating electric field to a target region of the subject for a certain period of time; and (2) administering the composition to the subject.

[0037] Certain additional non-limiting embodiments of the present disclosure relate to chemotaxis for use in a method of treating cancer in a subject, the method comprising: (1) applying an alternating electric field to a target region of the subject containing cancer cells to cause the chemotaxis to be expressed by the cancer cells of the subject; and (2) administering to the subject at least one composition, wherein the at least one composition comprises at least one chimeric antigen receptor (CAR) immune cell.

[0038] Certain additional, non-limiting embodiments of the present disclosure relate to a kit for reducing the viability of cancer cells, the kit comprising at least one composition comprising at least one chimeric antigen receptor (CAR) immune cell, and an electric field generator configured to apply an alternating electric field to the cancer cells for a certain period of time.

[0039] Certain additional, non-limiting embodiments of the present disclosure relate to a kit for reducing the viability of cancer cells, the kit comprising components for preparing a composition containing CAR immune cells by isolating at least one immune cell or its precursor from a subject or allogeneic donor, modifying at least one immune cell to express a chimeric antigen receptor, and culturing at least one chimeric antigen receptor (CAR) immune cell, the kit optionally further comprising an electric field generator configured to apply an alternating electric field to cancer cells for a certain period of time.

[0040] In certain (but not limiting) embodiments of the present invention, cancer or cancer cells are in the form of a solid tumor.

[0041] In certain (but not limiting) embodiments of any of the methods described above or otherwise discussed herein, the administration of an alternating electric field increases the cytotoxicity of at least one chimeric antigen receptor (CAR) immune cell against cancer cells in a subject compared to the administration of at least one chimeric antigen receptor (CAR) immune cell to a subject in the absence of an applied alternating electric field.

[0042] Any step (1) and (2) of the method of the present disclosure may be carried out simultaneously, sequentially, and in particular substantially simultaneously, or sequentially in whole or in part. If the steps are carried out sequentially in whole or in part, at least one composition comprising at least one CAR immune cell may be administered before or after the application of the alternating electric field is initiated.

[0043] The methods of this disclosure may be used to treat any type of cancer cell / cancer / tumor that responds to treatment with an alternating electric field (e.g., a TT field) and / or CAR immune cells. Non-limiting examples of cancer cells / cancer / tumors that can be treated according to this disclosure include hepatocellular carcinoma / cancer cells, glioblastoma / glioblastoma cells, pleural mesothelioma / mesothelioma cells, differentiated thyroid cancer / cancer cells, advanced renal cell carcinoma / cancer cells, ovarian cancer / cancer cells, pancreatic cancer / cancer cells, lung cancer / cancer cells, breast cancer / cancer cells, and any combination thereof.

[0044] Any type of conductive or nonconductive electrode(s) and / or transducer array(s) available for generating an alternating electric field, known in the art or otherwise discussed herein, may be used to generate an alternating electric field in accordance with the methods of this disclosure. Non-limiting examples of electrodes and transducer arrays that can be used to generate an alternating electric field in accordance with this disclosure include, but are not limited to, those that function as part of the TT field systems described in U.S. Patents Nos. 7,016,725, 7,089,054, 7,333,852, 7,565,205, 8,244,345, 8,715,203, 8,764,675, 10,188,851, and 10,441,776, as well as those that function as part of the TT field systems described in U.S. Patents Nos. 2018 / 0160933, 2019 / 0117956, 2019 / 0307781, and 2019 / 0308016.

[0045] An alternating current electric field can be generated at any frequency in accordance with this disclosure. For example (but not limited to), the alternating current electric field may be generated at 50kHz, 75kHz, 100kHz, 125kHz, 150kHz, 175kHz, 200kHz, 225kHz, 250kHz, 275kHz, 300kHz, 325kHz, 350kHz, 375kHz, 400kHz, 425kHz, 450kHz, 475kHz, 500kHz, 550kHz, 600kHz, 650kHz, 700kHz, 750kHz, 800kHz, 850kHz, 900kHz, 950kHz, 1MHz, 2MHz, 3MHz, 4MHz, The frequencies may include 5MHz, 6MHz, 7MHz, 8MHz, 9MHz, 10MHz, etc., as well as ranges formed from any of the above values ​​(e.g., the range of approximately 50kHz to approximately 10MHz, approximately 50kHz to approximately 1MHz, approximately 50kHz to approximately 500kHz, approximately 100kHz to approximately 500kHz, approximately 150kHz to approximately 300kHz, etc.), and ranges formed by combining two integers between the above two values ​​(e.g., the range of approximately 122kHz to approximately 313kHz, approximately 78kHz to approximately 298kHz, etc.).

[0046] In certain (but not limited) embodiments, the alternating electric field may be applied at two or more different frequencies. If two or more frequencies exist, each frequency is selected from any of the above values, a range formed from any of the above values, or a range of two integers that fall between the two above values.

[0047] In certain (but non-limiting) embodiments, the following frequencies may be used for specific cancers: approximately 200 kHz for GBM, approximately 150 kHz for NCLC, approximately 200 kHz for breast cancer, approximately 150 kHz for pancreatic cancer, approximately 150 kHz for brain metastases from NCLC, and approximately 150 kHz for liver cancer.

[0048] The alternating electric field may have any electric field intensity within the target region / subject / cancer cell, as long as it functions in accordance with this disclosure. For example (but not limited to these), an AC electric field has a voltage of at least approximately 1 V / cm, approximately 1.5 V / cm, approximately 2 V / cm, approximately 2.5 V / cm, approximately 3 V / cm, approximately 3.5 V / cm, approximately 4 V / cm, approximately 4.5 V / cm, approximately 5 V / cm, approximately 5.5 V / cm, approximately 6 V / cm, approximately 6.5 V / cm, approximately 7 V / cm, approximately 7.5 V / cm, approximately 8 V / cm, approximately 9 V / cm, approximately 9.5 V / cm, approximately 10 V / cm, approximately 10.5 V / cm, approximately 11 V / cm, approximately 11.5 V / cm, approximately 12 V / cm, approximately 12.5 V / cm, approximately 13 V / cm, approximately 13.5 V / cm, approximately 14 V / cm, approximately 14.5 V / cm, approximately 15 V / cm, approximately 15.5 V / cm, approximately 16 V / cm, and approximately 16. The electric field strength in the target region / subject / cancer cells may be in the following ranges: 5V / cm, approximately 17V / cm, approximately 17.5V / cm, approximately 18V / cm, approximately 18.5V / cm, approximately 19V / cm, approximately 19.5V / cm, approximately 20V / cm, etc., as well as ranges formed from any of the above values ​​(e.g., approximately 1V / cm to approximately 20V / cm, approximately 1V / cm to approximately 10V / cm, approximately 1V / cm to approximately 4V / cm, etc.), and ranges formed by combining two integers that fall between two of the above reference values ​​(e.g., approximately 1.1V / cm to approximately 18.6V / cm, approximately 1.2V / cm to approximately 9.8V / cm, approximately 1.3V / cm to approximately 4.7V / cm, etc.). Generally, it is desirable to utilize the highest possible electric field strength without causing overheating, given that the electric field strength is typically limited by temperature measurements.

[0049] In some cases, the electric field in at least a portion of the target region / subject / cancer cells is induced by an applied voltage determined by computer simulations of the target region / subject / cancer cells. In some cases, the electric field in at least a portion of the target region / subject / cancer cells is induced by an applied voltage of at least 50 V RMS (root mean square) or at least 50 V p2p (peak to peak), and optionally, the applied voltage is at least 100 V RMS or at least 100 V p2p. In some embodiments, an applied voltage of at least 50 V induces an electric field having an electric field strength of at least 1 V / cm (e.g., at least 5 V / cm) in at least a portion of the target region / subject / cancer cells.

[0050] An alternating current electric field may be applied in a unidirectional direction between a pair of arrays, or alternately applied in two or more directions / channels (e.g., front-to-back and left-to-right) between two or more pairs of arrays. For example, certain TT field devices (such as the OPTUNE® system (Novocure Limited, St. Helier, Jerse)) operate in two directions to increase the likelihood that dividing cells will be aligned with the electric field, allowing the electric field to exert the desired anti-mitotic effect. However, it is understood that the scope of the present invention also includes the application of an alternating current electric field in a unidirectional direction. The term “alternating current electric field” as used herein is understood to include not only application in a single direction / channel but also application in two or more directions / channels, and further, the term “alternating current electric field” as used herein is understood to include not only the application of a single alternating current electric field but also the application of multiple alternating current electric fields continuously over a period of time.

[0051] An alternating electric field may be applied for any duration or cumulative period sufficient to achieve a reduction in the viability of cancer cells and / or a decrease in tumor volume (and / or prevention of an increase in tumor volume). The period during which the alternating electric field is applied includes both continuous and cumulative periods. That is, the period during which the alternating electric field is applied includes not only a single session (i.e., continuous application) but also multiple sessions with short breaks in between (i.e., continuous application over a cumulative period). For example, a subject may be allowed to take breaks during treatment with an alternating electric field device, and is expected to keep the device in place and operating on their body for at least about 60%, at least about 70%, or at least about 80% of the entire treatment period (e.g., one day, one week, two weeks, one month, two months, three months, four months, five months, etc.).

[0052] For example, though not limited to, an alternating electric field can last for at least 1 hour, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 7 hours, 8 hours, 9 hours, 10 hours, 11 hours, 12 hours, 15 hours, 18 hours, 21 hours, 24 hours, 27 hours, 30 hours, 33 hours, 36 hours, 39 hours, 42 hours, 45 hours, 48 ​​hours, 51 hours, 54 hours, 57 hours, 60 hours, 63 hours, 66 hours, 69 hours, 72 hours, 75 hours, 78 hours, 81 hours, 84 hours, 87 hours, 90 hours, and 9 It can be applied over a continuous or cumulative period of time such as 3 hours, approximately 96 hours, approximately 5 days, approximately 6 days, approximately 7 days, approximately 8 days, approximately 9 days, approximately 10 days, approximately 11 days, approximately 12 days, approximately 13 days, approximately 14 days, approximately 21 days, approximately 1 month, approximately 2 months, approximately 3 months, approximately 4 months, approximately 5 months, approximately 6 months, etc., as well as a range formed from any of the above values ​​(e.g., a range from approximately 1 hour to approximately 6 months, a range from approximately 24 hours to approximately 72 hours, etc.), and a range formed by combining two integers between the two above values ​​(e.g., a range from approximately 14 hours to approximately 68 hours, etc.).

[0053] In certain (but not limited) embodiments, the period during which an alternating electric field is applied is at least about 24 hours, with the device positioned on the body and operating for at least about 80% of that period.

[0054] In certain (but non-limiting) embodiments, an alternating electric field is applied to a subject's cancer cell / target region for a period sufficient to increase the expression of at least one chemoattractant in the cancer cells compared to control cancer cells not exposed to the alternating electric field. Non-limiting examples of chemoattractants include CXCL9, CXCL10, CCL5, IL-15, and any combination thereof.

[0055] Any CAR immune cells known in the art or otherwise discussed herein may be used in accordance with this disclosure, insofar as the CAR immune cells can function as described herein. Non-limiting examples include CAR-T cells, CAR-NK cells, CAR-macrophages, and combinations thereof. The CAR immune cells may be autologous (and thus obtained from the subject being treated) or allogeneic (and thus obtained from another subject). Autologous or allogeneic CAR immune cells can be cultured directly from the subject and then transplanted / injected into cancer patients. Alternatively (and / or in addition), pluripotent stem cells may be obtained from the subject and subsequently differentiated in vitro into CAR immune cells that are subsequently transplanted / injected into cancer patients.

[0056] Various other types of CAR immune cells that may be used in accordance with this disclosure are known in the art, commercially available from multiple sources, and therefore do not require further description.

[0057] A composition containing at least one CAR immune cell may be provided in any formulation known in the art or otherwise discussed herein. In certain (but not limited) embodiments, the composition containing at least one CAR immune cell comprises one or more pharmaceutically acceptable carriers (the composition is therefore also referred to as the “pharmaceutical composition”). Not limited examples of suitable pharmaceutically acceptable carriers include water, saline, dextrose solution, fructose or mannitol, calcium carbonate, cellulose, ethanol, oils of animal, plant or synthetic origin, carbohydrates such as glucose, sucrose, or dextran, antioxidants such as ascorbic acid or glutathione, chelating agents, low molecular weight proteins, detergents, liposome carriers, buffer solutions such as sodium chloride, saline, or phosphate-buffered saline, and / or other substances, diluents, excipients such as polyethylene glycol (PEG), or any combination thereof that are physiologically acceptable and / or can be safely used. For pharmaceutically acceptable carriers suitable for pharmaceutical formulations, see, for example, Remington: The Science and Practice of Pharmacy, 23rd ed (2020).

[0058] In certain (but not limited) embodiments, a composition comprising at least one CAR immune cell may further comprise one or more additional activators. A variety of active drugs that can be used in combination with alternating electric fields or CAR immune cells are known in the art, and certain combination therapies have been approved by the FDA or are currently in clinical trials. Non-limiting examples of therapeutic agents that can be used concurrently with CAR immune cells in accordance with this disclosure include immune checkpoint inhibitors, steroids (e.g., dexamethasone), tocilizumab, interleukin-1 (IL-1), IL-1R, IL-2, IL-11, IL-15, IL-17, IL-18, IL-21, and docetaxel; anti-PD-1 therapies such as (but not limited to) pembrolizumab (KEYTRUDA®, Merck & Co., Inc., Rahway, NJ), tislerizumab, nivolumab, and semiprimab; anti-PD-L1 therapies such as atezolizumab, avelumab, and durvalumab; lenvatinib, paclitaxel, docetaxel, ifosfamide, and etoposide (VEPESID®, Bristol-Myers Squibb Co., New Zealand). This includes, but is not limited to, chemotherapeutic agents such as gemcitabine, lomustine, nab-paclitaxel, temozolomide, and carboplatin (York, NY); TKI inhibitors such as, but is not limited to, everolimus; mTOR inhibitors; Akt inhibitors; PI3K inhibitors; PARP inhibitors; FGF inhibitors; anti-LAB3 agents; anti-CTLA-4 agents; aromatase inhibitors such as letrozole (but is not limited to these); biological agents such as monoclonal antibodies (e.g., denosumab, pembrolizumab, lirirumab, monalizumab); and any combination thereof.

[0059] In addition, any composition of this disclosure may contain other drugs that enable the administration of the composition via a particular route of administration. For example, but not limited to, compositions may be formulated for administration via oral, topical, transdermal, parenteral, subcutaneous, intranasal, mucosal, intramuscular, intraperitoneal, intravitreous, and / or intravenous routes. Based on the route of administration, a composition may also contain one or more additional components (e.g., CAR immune cells and / or additional therapeutic agents) in addition to the activator. Examples of additional secondary compounds that may be present include, but are not limited to, fillers, salts, buffers, preservatives, stabilizers, solubilizers, wetting agents, emulsifiers, dispersants, and other substances well known in the art.

[0060] In certain (but not limited) embodiments, at least one composition comprising at least one CAR immune cell is administered to a subject by injection or transplantation. For example (but not limited to), in some cases it may be desirable to administer CAR immune cells at a local / topical level to ensure targeting of CAR immune cells to specific locations within the subject's body and to inhibit nonspecific interactions in other parts of the body, while in other cases a more systemic administration may be desired.

[0061] At least one composition comprising at least one CAR immune cell may be administered before or after the application of an alternating electric field is initiated. In certain (but not limited) embodiments, at least one composition comprising at least one CAR immune cell may be administered after the application of an alternating electric field has been initiated. In particular (but not limited to), at least one composition comprising at least one CAR immune cell may be administered during the application of the alternating electric field (e.g., before the period during which the alternating electric field is applied has elapsed) and / or after the application of the alternating electric field has elapsed.

[0062] For example (but not limited to these), at least one composition comprising at least one CAR immune cell is subjected to an alternating electric field for at least about 3 hours, about 6 hours, about 9 hours, about 12 hours, about 15 hours, about 18 hours, about 21 hours, about 24 hours, about 27 hours, about 30 hours, about 33 hours, about 36 hours, about 39 hours, about 42 hours, about 45 hours, about 48 hours, about 51 hours, about 54 hours, about 57 hours, about 60 hours, and about 63 hours from the start of application of the alternating electric field. The administration may occur after a certain number of hours, approximately 66 hours, 69 hours, 72 hours, 75 hours, 78 hours, 81 hours, 84 hours, 87 hours, 90 hours, 93 hours, 96 hours, approximately 5 days, 6 days, 7 days, etc., as well as a range formed from any of the above values ​​(e.g., a range of approximately 24 hours to approximately 96 hours), and a range formed by combining two integers between the two above values ​​(e.g., a range of approximately 14 hours to approximately 94 hours). In certain (but not limited) embodiments, at least one composition comprising at least one CAR immune cell is administered at least approximately 24 hours after the start of application of the alternating electric field.

[0063] In other non-limiting examples, at least one composition comprising at least one CAR immune cell may be administered after the period in which the alternating electric field has been applied has elapsed, or at least one composition comprising at least one CAR immune cell may be introduced within approximately 3 hours, approximately 6 hours, approximately 9 hours, approximately 12 hours, approximately 15 hours, approximately 18 hours, approximately 21 hours, approximately 24 hours, approximately 27 hours, approximately 30 hours, approximately 33 hours, approximately 36 hours, approximately 39 hours, approximately 42 hours, approximately 45 hours, approximately 48 hours, approximately 51 hours, approximately 54 hours, approximately 57 hours, approximately 60 hours, approximately 63 hours, approximately 66 hours, approximately 69 hours, approximately 72 hours, approximately 75 hours, approximately 78 hours, approximately 81 hours, approximately 84 hours, approximately 87 hours, approximately 90 hours, approximately 93 hours, approximately 96 hours, approximately 5 days, approximately 6 days, approximately 7 days, etc.

[0064] In certain (but not limited) embodiments, at least one composition comprising at least one CAR immune cell is administered within approximately 96 hours after the period has elapsed.

[0065] A composition comprising at least one CAR immune cell may be administered to cancer cells / subjects at any concentration that provides a therapeutically effective concentration of CAR immune cells. In certain non-limiting embodiments, the application of an alternating electric field reduces the amount of CAR immune cells required to be therapeutically effective compared to the usual therapeutically effective dose administered in the absence of the alternating electric field. For example, but not limited to, the therapeutically effective concentration of CAR immune cells may be reduced by at least about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, or more than about 75% compared to the dose of CAR immune cells known to be therapeutically effective in the absence of the alternating electric field. In certain (but non-limiting) embodiments, the therapeutically effective concentration of CAR immune cells may be reduced by at least about 50% compared to the dose of CAR immune cells known to be therapeutically effective in the absence of the alternating electric field.

[0066] CAR immune cell-containing compositions may be administered to subjects at any concentration capable of inducing an inflammatory response against tumor or cancer cells. For example, but not limited to, CAR immune cells may be administered at concentrations of approximately 10 cells / kg body weight, approximately 100 cells / kg body weight, approximately 1000 cells / kg body weight, approximately 104 cells / kg body weight, approximately 105 cells / kg body weight, approximately 106 cells / kg body weight, approximately 107 cells / kg body weight, approximately 108 cells / kg body weight, approximately 109 cells / kg body weight, approximately 1010 cells / kg body weight, approximately 1011 cells / kg body weight, approximately 1012 cells / kg body weight, approximately 1013 cells / kg body weight, approximately 1014 cells / kg body weight, approximately 1015 cells / kg body weight, or higher, as well as in a range formed from any of the above values ​​(e.g., the range of approximately 104 to approximately 109 cells / kg body weight).

[0067] In certain (but not limited to) embodiments, the method includes one or more additional steps. For example (but not limited to), the method may further include step (3) discontinuing the application of an alternating electric field (for example, for purposes such as allowing cells / tissues to recover, but not limited to). Furthermore, either step (1) and / or (2) may be repeated one or more times.

[0068] In certain (but not limited) embodiments, the CAR immune cell-containing composition may be administered by any dosing regimen known in the art. For example, the CAR immune cell-containing composition may be administered as a single dose or in multiple doses over a defined therapeutic period, but is not limited thereto. For example (but is not limited thereto), a therapeutically effective concentration of the CAR immune cell-containing composition may be administered at intervals such as once every 4 hours, once every 8 hours, once every 12 hours, once every 1 day, once every 2 days, once every 3 days, once every 1 week, twice every week, three times every week, once every 2 weeks, once every 3 weeks, once every 1 month, etc., as well as in ranges formed from any of the above values ​​(e.g., from once every 4 to 8 hours, from once every week to once every 1 month, etc.).

[0069] In certain (but not limited to) embodiments, the method involves simultaneous treatment with two or more compositions. Thus, the method may include an additional step of (4) administering at least a second composition to cancer cells / subjects. In certain (but not limited to) embodiments, at least a second composition may contain one or more active substances disclosed herein or separately considered for use with CAR immune cell-containing compositions.

[0070] Various substances and therapies used in combination with CAR immune cells are known in the field, and certain combination therapies have been approved by the FDA or are currently in clinical trials. Non-limiting examples of therapeutic agents that can be used in contemporaneous (sequential) therapy with CAR immune cells in accordance with this disclosure include immune checkpoint inhibitors, steroids (e.g., dexamethasone), tocilizumab, interleukin-1 (IL-1), IL-1R, IL-2, IL-11, IL-15, IL-17, IL-18, IL-21, and docetaxel; anti-PD-1 agents such as (but not limited to) pembrolizumab (KEYTRUDA®, Merck & Co., Inc., Rahway, NJ), tislerizumab, nivolumab, and semiprimab; anti-PD-L1 agents such as atezolizumab, avelumab, and durvalumab; lenvatinib, paclitaxel, docetaxel, ifosfamide, and etoposide (VEPESID®, Bristol-Myers Squibb Co., New Zealand). This includes, but is not limited to, chemotherapeutic agents such as gemcitabine, lomustine, nab-paclitaxel, temozolomide, and carboplatin (York, NY); TKI inhibitors such as, but is not limited to, everolimus; mTOR inhibitors; Akt inhibitors; PI3K inhibitors; PARP inhibitors; FGF inhibitors; anti-LAB3 agents; anti-CTLA-4 agents; aromatase inhibitors such as letrozole (but is not limited to these); biological agents such as monoclonal antibodies (e.g., denosumab, pembrolizumab, lirirumab, monalizumab); and any combination thereof.

[0071] Alternatively, additional therapies may include one or more mechanisms to promote the production of CAR immune cells within the subject. Non-limited examples of additional therapies that function in this way include the use of gene editing technologies (e.g., TALON and CRISPR) to induce immune cells provided to produce CARs, as well as nanotechnology and mRNA-based approaches that enable the purification of CAR immune cells in the body.

[0072] If present, step (4) may be carried out substantially simultaneously with, or in whole or in part, the administration of the first composition in step (2), thereby administering the two separate compositions simultaneously or in whole or in part sequentially. In addition, the two compositions administered in steps (2) and (4) may be administered via the same route (e.g., both intravenously), or the two compositions may be administered via different routes (e.g., one composition orally and the other intravenously).

[0073] If present, any additional dosing step (4) may be performed before or after the application of the alternating electric field is initiated, during the application of the alternating electric field, and / or after the application of the alternating electric field has elapsed, in the same manner and time frame as described above for the first composition.

[0074] That is, for example (but not limited to these), the second composition is subjected to at least about 3 hours, about 6 hours, about 9 hours, about 12 hours, about 15 hours, about 18 hours, about 21 hours, about 24 hours, about 27 hours, about 30 hours, about 33 hours, about 36 hours, about 39 hours, about 42 hours, about 45 hours, about 48 hours, about 51 hours, about 54 hours, about 57 hours, about 60 hours, about 63 hours, about 66 hours, It may be administered after approximately 69 hours, approximately 72 hours, approximately 75 hours, approximately 78 hours, approximately 81 hours, approximately 84 hours, approximately 87 hours, approximately 90 hours, approximately 93 hours, approximately 96 hours, approximately 5 days, approximately 6 days, approximately 7 days, and similarly, after a range formed from any of the above values ​​(e.g., a range of approximately 24 hours to approximately 96 hours), and after a range formed by combining two integers that fall between two of the above values ​​(e.g., a range of approximately 14 hours to approximately 94 hours). In certain (but not limited) embodiments, the second composition is administered at least approximately 24 hours after the application of the alternating electric field begins.

[0075] In other non-limiting examples, the second composition may be administered after the period of application of the alternating electric field has elapsed, in which case the second composition is administered within approximately 3 hours, approximately 6 hours, approximately 9 hours, approximately 12 hours, approximately 15 hours, approximately 18 hours, approximately 21 hours, approximately 24 hours, approximately 27 hours, approximately 30 hours, approximately 33 hours, approximately 36 hours, approximately 39 hours, approximately 42 hours, approximately 45 hours, approximately 48 hours, approximately 51 hours, approximately 54 hours, approximately 57 hours, approximately 60 hours, approximately 63 hours, approximately 66 hours, approximately 69 hours, approximately 72 hours, approximately 75 hours, approximately 78 hours, approximately 81 hours, approximately 84 hours, approximately 87 hours, approximately 90 hours, approximately 93 hours, approximately 96 hours, approximately 5 days, approximately 6 days, approximately 7 days, etc. In certain (but non-limiting) embodiments, the second composition is administered within approximately 96 hours after the period has elapsed.

[0076] In addition, for example (but not limited to these), the second composition may be administered for at least about 1 minute, about 5 minutes, about 10 minutes, about 15 minutes, about 30 minutes, about 45 minutes, about 1 hour, about 2 hours, about 3 hours, about 4 hours, about 5 hours, about 6 hours, about 7 hours, about 8 hours, about 9 hours, about 10 hours, about 11 hours, about 12 hours, about 15 hours, about 18 hours, about 21 hours, about 24 hours, about 27 hours, about 30 hours, about 33 hours, about 36 hours, about 39 hours, about 42 hours, about 45 hours, about 48 hours, It may be administered after approximately 51 hours, approximately 54 hours, approximately 57 hours, approximately 60 hours, approximately 63 hours, approximately 66 hours, approximately 69 hours, approximately 72 hours, approximately 75 hours, approximately 78 hours, approximately 81 hours, approximately 84 hours, approximately 87 hours, approximately 90 hours, approximately 93 hours, approximately 96 hours, approximately 5 days, approximately 6 days, approximately 7 days, etc., as well as a range formed from any of the above values ​​(e.g., a range of approximately 24 hours to approximately 96 hours), and a range formed by combining two integers between the two above values ​​(e.g., a range of approximately 14 hours to approximately 94 hours). In certain (but not limited) embodiments, the second composition is administered at least approximately 12 hours after the administration of the first substance.

[0077] In certain (but not limited) embodiments, the method may further include the step of (5) administering at least one additional therapy to cells / subjects. Any therapy known in the art or considered herein for use in combination with alternating electric fields (e.g., TT fields) and / or CAR immunotherapy may be utilized in accordance with the method of the present disclosure. Non-limited examples of additional therapies that may be utilized include radiotherapy, photodynamic therapy, transarterial chemoembolization (TACE), or combinations thereof.

[0078] Steps (1) and (2), and any of the optional steps (3), (4), and (5), may be repeated one or more times. Each step may be repeated any number of times as needed. When step (1) is repeated, the transducer array may be positioned in a slightly different location from the subject's original position, and by rearranging the array in this way, the treatment of the tumor / cancer may be further enhanced. In addition, step (2) and any of the optional steps (4) and (5) (if any) of administering the composition / additional therapy may be repeated at varying numbers and intervals to follow known and / or generally accepted dosage / treatment plans for the composition / therapy(s) / therapy(s).

[0079] The use of ordinal numbers in any step is illustrative only, and any one or more of steps (3), (4), and (5) may be included individually or in combination with each other. That is, the method of the disclosure includes performing step (3) if step (4) or (5) is not present, performing step (4) if step (3) or (5) is not present, and performing step (5) if steps (3) and (4) are not present. In other words, the scope of the methods disclosed herein is to carry out steps (1) to (2) (repeating each step as many times as necessary), to carry out steps (1) to (3) (repeating one or more of steps (1) to (3) as many times as necessary), to carry out steps (1) to (2) and (4) (repeating one or more of steps (1) to (2) and (4) as many times as necessary), to carry out steps (1) to (2) and (5) (repeating one or more of steps (1) to (2) and (5) as many times as necessary), step This includes performing (1) to (4) (repeating one or more of steps (1) to (4) the required number of times), performing steps (1) to (3) and (5) (repeating one or more of steps (1) to (3) and (5) the required number of times), performing steps (1) to (2) and (4) to (5) (repeating one or more of steps (1) to (2) and (4) to (5) the required number of times), and performing all of steps (1) to (5) (and also repeating one or more of steps (1) to (5) the required number of times).

[0080] While the use of combination therapy of two substances is explicitly described above, it is understood that the scope of this disclosure also includes combination therapy of three or more compositions. Accordingly, this method may include one or more additional steps (similar to steps (1) and (4)) in which an additional composition is administered to a subject. The additional substance administered in this method may be selected from any of the substances disclosed herein or (as disclosed herein above with respect to any step (4)) that are considered for use in combination with at least one CAR immune cell. Furthermore, the administration of the additional substance may be carried out substantially simultaneously with, or entirely or partially sequentially with, the administration of the first and second compositions / substances, in the same manner(s) and timeframe(s) as described above for the first and second compositions / substances.

[0081] Certain non-limiting embodiments of this disclosure are any of the components of AC electric field (e.g., TT field) generating systems disclosed or otherwise considered herein (e.g., U.S. Patents No. 7,016,725, 7,089,054, 7,333,852, 7,565,205, 8,244,345, 8,715,203, 8,764,675, 10,188,851, and 10,441,776, and U.S. Patent Applications US2018 / 0160933, US2019 / 0117956). The present invention relates to a kit comprising one or more transducer arrays and / or one or more hydrogel compositions (such as those disclosed in US2019 / 0307781 and US2019 / 0308016) in combination with at least one of the compositions comprising at least one CAR immune cell disclosed herein or separately considered, and / or a component for isolating, culturing and / or genetically modifying at least one immune cell or its precursor from a subject or allogeneic donor. The kit may optionally further comprise one or more of any compositions disclosed herein or separately considered (e.g., one or more of any compositions containing at least one additional activator, and / or one or more of any compositions for culturing / differentiating / genetically modifying at least one immune cell or its precursor). The kit may optionally further comprise one or more devices (or one or more components of a device) used in one or more additional therapeutic steps.

[0082] In certain (non-limiting) embodiments, the kit may further include instructions for performing any of the methods disclosed herein or otherwise considered. For example (but not limited to), the kit may include instructions for applying one or more components of an alternating electric field (e.g., TT field) generator to a patient's skin; instructions for applying an alternating electric field to a patient; instructions for isolating immune cells or their precursors from a subject or allogeneic donor; instructions for culturing and / or differentiating immune cells or their precursors in vitro; instructions for genetically modifying immune cells; instructions for formulating compositions containing CAR immune cells; instructions for the timing and method of administering a composition containing at least one CAR immune cell, and optionally for administering one or more additional compositions; and / or instructions for when to start and stop the alternating electric field in connection with the administration of a composition containing at least one CAR immune cell, and / or the administration of one or more additional compositions, and / or a therapeutic step.

[0083] In addition to the components described in detail above, the kit may further contain other components / reagents for carrying out any of the specific methods described herein or otherwise considered. For example (but not limited to), the kit may further include (i) components for preparing the skin before discarding the hydrogel composition and / or transducer array on the skin (e.g., razor, cleansing composition, wipe / towel, etc.); (ii) components for removing the gel / transducer array; (iii) components for cleansing the skin after removal of the gel / transducer array; and / or (iv) other components used in the system (i.e., conductive materials, non-conductive materials, soothing gels or creams, bandages, etc.). The properties of these additional components / reagents depend on the specific processing method, and their identification is within the scope of the art for those skilled in the art, so no further explanation is considered necessary. Furthermore, the components / reagents included in the kit may be in separate containers / compartments, or various components / reagents may be combined into one or more containers / compartments depending on their sterility, cross-reactivity, and stability.

[0084] The kit may be placed in any package that enables the components contained therein to function in accordance with this disclosure. In certain non-limiting embodiments, the kit further includes a sealed package in which the components are placed. In certain (but non-limiting) embodiments, the sealed package is substantially impermeable to air and / or substantially impermeable to light.

[0085] Furthermore, the kit may further include a set of written instructions explaining how to use one or more of the kit's components. Kits of this nature can be used in any of the methods described herein or otherwise considered.

[0086] In certain non-limiting embodiments, the shelf life of the kit is at least about 6 months, for example (but not limited to) at least about 9 months, or at least about 12 months.

[0087] Certain non-limiting embodiments of this disclosure are any of the components of AC electric field generating systems disclosed herein or otherwise considered (e.g., U.S. Patents No. 7,016,725, 7,089,054, 7,333,852, 7,565,205, 8,244,345, 8,715,203, 8,764,675, 10,188,851, and 10,441,776, and U.S. Patent Application No. 201 This invention relates to a system comprising one or more transducer arrays and / or one or more hydrogel compositions (such as, but not limited to, those disclosed in Nos. 8 / 0160933, 2019 / 0117956, 2019 / 0307781, and 2019 / 0308016) in combination with at least one of any compositions comprising at least one CAR immune cell disclosed or otherwise considered herein. The system may optionally further comprise one or more of any compositions disclosed or otherwise considered herein. The system may optionally further comprise one or more devices (or one or more components of a device) used in one or more additional therapeutic steps. Examples

[0088] Examples are shown below. However, it should be understood that this disclosure is not limited to the application to the specific experiments, results, and experimental procedures disclosed below herein. Rather, these examples are provided simply as one of various embodiments and are intended to be illustrative rather than exhaustive. Example 1

[0089] Serum from mice treated with TT field therapy and serum from mice treated with pseudoheat (heat sham) were incubated in cytokine array membranes and compared with each other. As shown in Figure 1, these experiments demonstrated that (1) CXCL9 increased after TT field therapy in a mammary cancer mouse model, (2) CXCL9, CCL5, and IL15 increased after TT field therapy in an ovarian cancer mouse model, and (3) CXCL9, CCL5, and IL15 increased after TT field therapy in a small cell lung cancer mouse model, with a partial increase in CXCL10 after TT field therapy. These results suggest that tumors treated with TT field therapy may secrete higher levels of cytokines to attract cytotoxic T cells to the tumor site.

[0090] Next, the tumor is directly analyzed (using tumor lysates, RNA samples, and / or tissue structures) for chemokines found in serum (i.e., CXCL9, CXCL10, CCL5, IL15, etc.) and associated with T cell attraction. It is determined that TT field therapy increases the expression of these chemokines.

[0091] Furthermore, strong adhesion to the tumor is necessary to enable overflow. Increased expression of VCAM-1 and ICAM-1 on endothelial cells binds to VLA-4 and LFA-1 on T cells, respectively, mediating their recruitment to the tumor (Tantalo et al. (2021) J Immunother Cancer, 9:e002555). Therefore, tumor tissue treated in the TT field is stained with antibodies against adhesion molecules (P / E selectin and VCAM1 / ICAM1 (CD31) which suppress T cell proliferation and support T cell adhesion to endothelial cells), and the results are compared with heat-treated control tumor samples. Increased VCAM-1 and ICAM-1 expression is observed in tumor samples treated in the TT field, and staining for CD31 indicates whether there is more VCAM-1 on endothelial cells, showing greater T cell recruitment. Example 2

[0092] This embodiment relates to the use of optimal signaling CAR immune cells and the simultaneous use of TT sites to increase the effectiveness of these cancer therapies and provide a synergistic improvement compared to either treatment alone.

[0093] The subject is treated in a TT field by applying the OPTUNE® device (Novocure Limited, St. Helier, Jersey) to the subject's skin with a pair of arrays positioned to the left and right (LR) and / or front and back (AP) of the tumor. Each subject is then chronically treated in a 150-200 kHz TT field. The device is worn for at least approximately 80% of the time, with brief interruptions between sessions to allow for slight adjustments to the array placement and cell and skin regeneration.

[0094] A composition containing CAR immune cells (e.g., CAR-T cells, but not limited to these) is prepared. In one example, the immune cells are obtained from a subject prior to TT field exposure. Alternatively, the immune cells are obtained from an allogeneic source. In yet another example, pluripotent stem cells are obtained from the autologous or allogeneic source and differentiated into immune cells in vitro. The immune cells are then modified to express CARs. The autologous or allogeneic CAR immune cells are then cultured before transplantation / injection for adoptive cell transfer therapy.

[0095] Two weeks after the start of TT field application, approximately 10 to 10 billion CAR immune cells are transplanted or injected into human subjects approximately 1 to 3 times per week for 6 to 8 weeks.

[0096] Following combination therapy, the effectiveness of the above combination therapy against the tumor(s) will be evaluated.

[0097] Combining the TT field with CAR immunotherapy provides a synergistic antitumor effect that promotes improvement. Non-limiting exemplary embodiments of the concept(s) of the present invention

[0098] Exemplary Embodiment 1. A method for reducing the viability of cancer cells, the method comprising: (1) applying an alternating electric field to the cancer cells for a certain period of time; and (2) administering at least one composition to the cancer cells, wherein the at least one composition comprises at least one chimeric antigen receptor (CAR) immune cell. The method may be in vitro or in vivo.

[0099] Exemplary Embodiment 2. A method for treating cancer in a subject, the method comprising: (1) applying an alternating electric field to a target region of the subject for a certain period of time; and (2) administering to the subject at least one composition comprising at least one CAR immune cell.

[0100] Exemplary Embodiment 3. A method for reducing the volume of a tumor and / or preventing an increase in the volume of a tumor, wherein the tumor is present in the body of a living organism and comprises a plurality of cancer cells, and the method comprises (1) applying an alternating electric field to a target area of ​​the subject for a certain period of time, and (2) administering to the subject at least one composition, wherein the at least one composition comprises at least one CAR immune cell.

[0101] Exemplary Embodiment 4. A method comprising the steps of (1) applying an alternating electric field to a target area of ​​a subject for a certain period of time, and (2) administering to the subject at least one composition, wherein the at least one composition comprises at least one chimeric antigen receptor (CAR) immune cell, the administration of the alternating electric field increases the cytotoxicity of the at least one chimeric antigen receptor (CAR) immune cell against cancer cells in the subject compared to the administration of the at least one chimeric antigen receptor (CAR) immune cell to the subject in the absence of the application of the alternating electric field.

[0102] Exemplary Embodiment 5. The method according to any one of Exemplary Embodiments 1 to 4, wherein the alternating current field is applied at a frequency in the range of about 50 kHz to about 1 MHz; the alternating current field has an electric field intensity of at least about 1 V / cm in at least a portion of the target area of ​​the subject; the alternating current field is induced by an applied voltage of at least 50 V RMS or at least 50 V p2p; and the period during which the alternating current field is applied is at least about 50% of a continuous period of at least about 24 hours.

[0103] Exemplary Embodiment 6. The method according to any one of Exemplary Embodiments 1 to 5, wherein the alternating electric field is applied to the cancer cell / target region for a period of time sufficient to increase the expression of at least one chemoattractant in the cancer cell compared to control cancer cells not exposed to the alternating electric field.

[0104] Exemplary Embodiment 7. The method according to Exemplary Embodiment 6, wherein the chemical attractant is selected from the group consisting of CXCL9, CXCL10, CCL5, and IL-15.

[0105] Exemplary Embodiment 8. The method according to any one of Exemplary Embodiments 1 to 7, wherein the at least one CAR immune cell is selected from the group consisting of CAR-T cells, CAR-NK cells, CAR-macrophages, and combinations thereof.

[0106] Exemplary Embodiment 9. The method according to any one of Exemplary Embodiments 1 to 8, wherein the chimeric antigen receptor comprises an extracellular antigen-binding domain, a hinge or spacer region, a transmembrane portion, and an intracellular domain comprising at least one signaling domain.

[0107] Exemplary Embodiment 10. The method according to any one of Exemplary Embodiments 1 to 9, wherein the chimeric antigen receptor comprises an extracellular antigen-binding domain that recognizes tumor-associated antigens (TAAs).

[0108] Exemplary Embodiment 11. The method according to Exemplary Embodiment 10, wherein the at least one TAA is selected from the group consisting of carcinoembryonic antigen (CEA), epidermal growth factor receptor (EGFR), human epidermal growth factor receptor 2 (HER2), mesoserine (MSLN), prostate stem cell antigen (PSCA), mucin 1 (MUC1), tyrosine kinase-like orphan receptor 1 (ROR1), programmed death ligand 1 (PD-L1), and CD80 / CD86.

[0109] Exemplary Embodiment 12. The method according to any one of the exemplary embodiments 1 to 11, wherein the CAR immune cells express at least one of lymphocyte function-associated antigen-1 (LFA-1) and late-stage antigen 4 (VLA-4).

[0110] Exemplary Embodiment 13. The method according to any one of Exemplary Embodiments 1 to 12, wherein the composition further comprises at least one substance selected from the group consisting of an immune checkpoint inhibitor, a steroid (e.g., dexamethasone), tocilizumab, interleukin-1 (IL-1), IL-1R, IL-2, IL-11, IL-15, IL-17, IL-18, IL-21, docetaxel, and combinations thereof.

[0111] Exemplary Embodiment 14. The method according to any one of Exemplary Embodiments 1 to 13, wherein the composition further comprises at least one substance selected from the group consisting of anti-PD-1 therapeutic agents, anti-PD-L1 therapeutic agents, chemotherapeutic agents, paclitaxel, ifosfamide, etoposide, gemcitabine, lomustine, nab-paclitaxel, temozolomide, carboplatin, TKI inhibitors, mTOR inhibitors, Akt inhibitors, PI3K inhibitors, PARP inhibitors, FGF inhibitors, anti-LAB3 agents, anti-CTLA-4 therapeutic agents, aromatase inhibitors, denosumab, pembrolizumab, lirirumab, monoarizumab, and combinations thereof.

[0112] Exemplary Embodiment 15. The method according to any one of Exemplary Embodiments 1 to 14, further comprising the step of (3) administering at least one additional composition to the cancer cells / subject, wherein the at least one additional composition comprises a substance selected from the group consisting of an immune checkpoint inhibitor, a steroid (e.g., dexamethasone), tocilizumab, interleukin-1 (IL-1), IL-1R, IL-2, IL-11, IL-15, IL-17, IL-18, IL-21, docetaxel, and combinations thereof.

[0113] Exemplary Embodiment 16. The method according to any one of Exemplary Embodiments 1 to 15, further comprising the step of administering at least one additional composition to the cancer cells, wherein the at least one additional composition comprises a substance selected from the group consisting of anti-PD-1 therapeutic agents, anti-PD-L1 therapeutic agents, chemotherapeutic agents, paclitaxel, docetaxel, ifosfamide, etoposide, gemcitabine, lomustine, nab-paclitaxel, temozolomide, carboplatin, TKI inhibitors, mTOR inhibitors, Akt inhibitors, PI3K inhibitors, PARP inhibitors, FGF inhibitors, anti-LAB3 agents, anti-CTLA-4 therapeutic agents, aromatase inhibitors, denosumab, pembrolizumab, lirirumab, monoarizumab, and combinations thereof.

[0114] Exemplary Embodiment 17. The method according to any one of the exemplary embodiments 1 to 16, wherein steps (1) and (2) are performed substantially simultaneously.

[0115] Exemplary Embodiment 18. The method according to any one of claims 1 to 17, wherein steps (1) and (2) are carried out in whole or in part sequentially, and the at least one composition is administered after the application of the alternating electric field has been initiated.

[0116] Exemplary Embodiment 19. The method according to Exemplary Embodiment 18, wherein the at least one composition is administered before the period during which the alternating electric field is applied has elapsed.

[0117] Exemplary Embodiment 20. The method according to Exemplary Embodiment 18, wherein the at least one composition is administered after the period of time has elapsed.

[0118] Exemplary Embodiment 21. The method according to any one of the exemplary embodiments 1 to 20, wherein steps (1) and (2) are repeated one or more times.

[0119] Exemplary Embodiment 22. The method according to any one of the exemplary embodiments 1 to 21, wherein the cancer cells are in the form of at least one solid tumor.

[0120] Exemplary Embodiment 23. The method according to any one of Exemplary Embodiments 1 to 22, wherein the cancer cells are selected from the group consisting of hepatocellular carcinoma cells, glioblastoma cells, pleural mesothelioma cells, differentiated thyroid cancer cells, advanced renal cell carcinoma cells, ovarian cancer cells, pancreatic cancer cells, lung cancer cells, breast cancer cells, and combinations thereof.

[0121] Exemplary Embodiment 24. A kit for reducing the viability of cancer cells, the kit comprising components for preparing a composition containing CAR immune cells by isolating at least one immune cell or its precursor from a subject or an allogeneic donor, modifying the at least one immune cell to express the chimeric antigen receptor, and culturing the at least one chimeric antigen receptor (CAR) immune cell, the kit optionally further comprising an electric field generator configured to apply an alternating electric field to the cancer cells for a certain period of time.

[0122] This invention describes the concept(s) of the present invention in conjunction with the specific experiments, results, and terms set forth below, but it will be apparent to those skilled in the art that many alternatives, modifications, and variations are obvious. Accordingly, this invention is intended to encompass all alternatives, modifications, and variations that fall within the spirit and broad scope of this disclosure.

Claims

1. A composition comprising at least one chimeric antigen receptor (CAR) immune cell for use in a method of treating cancer in a subject, wherein the method is (1) The step of applying an alternating electric field to the target area of ​​the subject for a certain period of time, (2) A composition comprising the step of administering the composition to the subject.

2. A chemoattractant for use in a method of treating cancer in a subject, wherein the method is (1) A step of applying an alternating electric field to a target region of the subject containing cancer cells to cause the chemotactic substance to be expressed by the cancer cells of the subject, (2) A chemoattractant comprising the step of administering at least one composition to the subject, wherein the at least one composition comprises at least one chimeric antigen receptor (CAR) immune cell.

3. A method for treating cancer in a subject, wherein the method is (1) The step of applying an alternating electric field to the target area of ​​the subject for a certain period of time, (2) A method comprising the step of administering at least one composition to the subject, wherein the at least one composition comprises at least one CAR immune cell.

4. The aforementioned AC electric field, The signal is applied at a frequency in the range of approximately 50 kHz to approximately 1 MHz. The alternating electric field has an electric field intensity of at least about 1 V / cm in at least a portion of the target area of ​​the subject. The AC electric field is induced by an applied voltage of at least 50V p2p, and The composition for use according to claim 1, the chemical attractant for use according to claim 2, or the method according to claim 3, wherein the certain period during which the alternating electric field is applied is at least about 50% of a continuous period of at least about 24 hours.

5. The composition for use according to claim 1 or 4, the chemoattractant for use according to claim 2 or 4, or the method according to claim 3 or 4, wherein the aforementioned period is sufficient to increase the expression of at least one chemoattractant in cancer cells in a subject by applying the alternating electric field, compared to control cancer cells in a subject not exposed to the alternating electric field.

6. The composition for use according to any one of claims 1, 4, or 5, the chemoattractant for use according to any one of claims 2, 4, or 5, or the method according to any one of claims 3 to 5, wherein the at least one CAR immune cell is selected from the group consisting of CAR-T cells, CAR-NK cells, CAR-macrophages, and combinations thereof.

7. The chimeric antigen receptor comprises an extracellular antigen-binding domain that recognizes tumor-associated antigens (TAAs), a composition for use according to any one of claims 1 or 4 to 6, a chemoattractant for use according to any one of claims 2 or 4 to 6, or the method according to any one of claims 3 to 6.

8. The composition for use, chemoattractant for use, or method for use according to claim 7, wherein the at least one TAA is selected from the group consisting of carcinoembryonic antigen (CEA), epidermal growth factor receptor (EGFR), human epidermal growth factor receptor 2 (HER2), mesoserine (MSLN), prostate stem cell antigen (PSCA), mucin 1 (MUC1), tyrosine kinase-like orphan receptor 1 (ROR1), programmed death ligand 1 (PD-L1), and CD80 / CD86.

9. Steps (1) and (2) are carried out substantially simultaneously, comprising a composition for use according to claim 1 or any one of claims 4 to 8, a chemical attractant for use according to claim 2 or any one of claims 4 to 8, or the method according to any one of claims 3 to 8.

10. Steps (1) and (2) are carried out in whole or in part sequentially, wherein the at least one composition is administered after the application of the alternating electric field has started, the composition for use according to any one of claims 1 or 4 to 8, the chemical attractant for use according to any one of claims 2 or 4 to 8, or the method according to any one of claims 3 to 8.

11. Steps (1) and (2) are repeated once or more times, comprising the composition for use according to claim 1 or any one of claims 4 to 10, the chemical attractant for use according to claim 2 or any one of claims 4 to 10, or the method according to any one of claims 3 to 10.

12. The cancer is a solid tumor, the composition for use according to any one of claims 1 or 4 to 11, the chemical attractant for use according to any one of claims 2 or 4 to 11, or the method according to any one of claims 3 to 11.

13. The cancer is selected from the group consisting of hepatocellular carcinoma, glioblastoma, pleural mesothelioma, differentiated thyroid cancer, advanced renal cell carcinoma, ovarian cancer, pancreatic cancer, lung cancer, breast cancer, and combinations thereof, and is a composition for use according to any one of claims 1 or 4 to 12, a chemical attractant for use according to any one of claims 2 or 4 to 12, or the method according to any one of claims 3 to 12.

14. The method is further defined as a method for reducing the viability of cancer cells and / or reducing the volume of a tumor, wherein the tumor is present in the body of the subject and comprises a plurality of cancer cells and / or prevents an increase in the volume of the tumor, wherein the tumor is present in the body of the subject and comprises a plurality of cancer cells, a composition for use according to claim 1 or any one of claims 4 to 13, a chemical attractant for use according to claim 2 or any one of claims 4 to 13, or the method according to any one of claims 3 to 13.

15. A kit for reducing the viability of cancer cells, the kit is A composition comprising at least one chimeric antigen receptor (CAR) immune cell, A kit comprising an electric field generator configured to apply an alternating electric field to the aforementioned cancer cells for a certain period of time.

16. An in vitro method for reducing the viability of cancer cells, wherein the method is (1) The step of applying an alternating electric field to the cancer cells for a certain period of time, (2) A method comprising the step of administering at least one composition to cancer cells, wherein the at least one composition comprises at least one chimeric antigen receptor (CAR) immune cell.

17. A method for reducing the viability of cancer cells, wherein the method is (1) The step of applying an alternating electric field to the cancer cells for a certain period of time, (2) A method comprising the step of administering at least one composition to cancer cells, wherein the at least one composition comprises at least one chimeric antigen receptor (CAR) immune cell.

18. The aforementioned AC electric field, The signal is applied at a frequency in the range of approximately 50 kHz to approximately 1 MHz. The alternating electric field has an electric field intensity of at least about 1 V / cm in at least a portion of the cancer cells. The AC electric field is induced by an applied voltage of at least 50V p2p, and The composition for use according to claim 1, the chemical attractant for use according to claim 2, or the method according to claim 3, wherein the certain period during which the alternating electric field is applied is at least about 50% of a continuous period of at least about 24 hours.

19. The aforementioned electric field generating device, The AC electric field is configured to be applied to the cancer cells at a frequency in the range of approximately 50 kHz to approximately 1 MHz. The electric field generator is configured to apply the alternating electric field to the cancer cells at least a portion of the cancer cells with an electric field intensity of at least about 1 V / cm. The electric field generator is configured to apply the alternating current electric field to the cancer cells by applying an applied voltage of at least 50V p2p to one or more electrodes, thereby inducing the alternating current electric field, and The kit according to claim 15, wherein the aforementioned period is at least one of the following: at least about 50% of a continuous period of at least about 24 hours.

20. The kit according to claim 15 or 19, or the method according to any one of claims 16 to 18, wherein the aforementioned period is sufficient to increase the expression of at least one chemoattractant in the cancer cells by applying the alternating electric field, compared to control cancer cells that have not been exposed to the alternating electric field.

21. The kit according to any one of claims 15, 19, or 20, or the method according to any one of claims 16 to 18 or 20, wherein the at least one CAR immune cell is selected from the group consisting of CAR-T cells, CAR-NK cells, CAR-macrophages, and combinations thereof.

22. The kit according to any one of claims 15 or 19-21, or the method according to any one of claims 16-18, 20 or 21, wherein the chimeric antigen receptor comprises an extracellular antigen-binding domain that recognizes tumor-associated antigens (TAAs).

23. The kit or method according to claim 22, wherein the at least one TAA is selected from the group consisting of carcinoembryonic antigen (CEA), epidermal growth factor receptor (EGFR), human epidermal growth factor receptor 2 (HER2), mesoserine (MSLN), prostate stem cell antigen (PSCA), mucin 1 (MUC1), tyrosine kinase-like orphan receptor 1 (ROR1), programmed death ligand 1 (PD-L1), and CD80 / CD86.

24. The method according to any one of claims 16-18 or 20-23, wherein steps (1) and (2) are carried out substantially simultaneously.

25. The method according to any one of claims 16 to 18 or 20 to 23, wherein steps (1) and (2) are carried out in whole or in part sequentially, and the at least one composition is administered after the application of the alternating electric field has started.

26. The method according to any one of claims 16 to 18 or 20 to 25, wherein steps (1) and (2) are repeated once or more times.

27. The cancer cells are selected from the group consisting of hepatocellular carcinoma cells, glioblastoma cells, pleural mesothelioma cells, differentiated thyroid cancer cells, advanced renal cell carcinoma cells, ovarian cancer cells, pancreatic cancer cells, lung cancer cells, breast cancer cells, and combinations thereof, according to the kit according to any one of claims 15 or 19-23, or according to any one of claims 16-18 or 20-26.

28. The kit according to any one of claims 15, 19-23, or 27, or the method according to any one of claims 16-18, or 20-27, wherein the cancer cells are cells of a solid tumor.

29. (1) The step of applying an alternating electric field to the target area of ​​the subject for a certain period of time, (2) The step of administering at least one composition to the subject, wherein the at least one composition comprises at least one chimeric antigen receptor (CAR) immune cell, A method wherein the administration of the alternating electric field increases the cytotoxicity of at least one chimeric antigen receptor (CAR) immune cell against cancer cells in the subject, compared to the administration of at least one chimeric antigen receptor (CAR) immune cell to the subject in the absence of the application of the alternating electric field.

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