Combination therapies with NK cell-based immunotherapies

Abstract

Embodiments of the disclosure include methods and compositions in which NK cells and one or more additional agents are provided to an individual in need thereof. In specific embodiments, the additional agent(s) comprises one or more KRAS inhibitors and / or one or more inhibitors of PARP.

Description

COMBINATION THERAPIES WITH NK CELL-BASED IMMUNOTHERAPIESCROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims priority to U.S. Provisional Patent Application Serial No.63 / 773,779, filed March 18, 2025, which is incorporated by reference herein in its entirety.I. Technical Field

[0002] This disclosure relates at least to the fields of immunology, cell biology, molecular biology, and medicine, including at least cancer medicine.II. Background

[0003] Natural killer (NK) cells are an emerging cellular immunotherapy for patients with malignant hematologic disease as well as solid tumors. The present disclosure satisfies a long-felt need in the art to improve upon immunotherapies including those that utilize NK cells.BRIEF SUMMARY

[0004] Embodiments of the disclosure include methods and compositions for treatment of an individual using adoptive cell therapy. In some embodiments, the individual is in need of a treatment that utilizes adoptive cell therapy. In certain embodiments, the individual is in need of adoptive cell therapy for cancer or an autoimmune disorder. In specific embodiments, the individual is provided a therapeutically effective amount of adoptive cell therapy that includes both non-engineered and / or engineered NK cells and one or more additional agents. In specific embodiments, the one or more additional agents may be capable of being able to directly or indirectly enhance the NK cells to initiate signaling, activation, and killing of target cells. The adoptive cell therapy and the one or more additional agents may or may not have a synergistic therapeutic effect for the individual.

[0005] Embodiments of the disclosure combine NK cells or engineered NK cells (including CAR NK cells, as one example) with one or more agents in order to improve anti-cancer activity of the cells. The one or more agents used in combination with the NK cells or engineered NK cells may themselves have anti-cancer activity.

[0006] Particular embodiments of the disclosure utilize NK cell-based immunotherapies with one or more other agents that may or may not be small molecule inhibitors, proteins, peptides, nucleic acid, carbohydrates, a combination thereof, and so forth. In certain302140458.1 - 1 -embodiments, the disclosure encompasses combinations of NK cell-based immunotherapies with one or more inhibitors of poly ADP -ribose polymerase (PARP) and / or one or more agents that target the Kirsten ras oncogene homolog from the mammalian ras gene family (KRAS) pathways, such as one or more KRAS inhibitors. Such embodiments of combinations therapy may be utilized for treating cancer and / or overcoming therapy resistance mechanisms. It is contemplated that the adoptive cell therapy and the other agent(s) may or may not be administered at the same time and / or may or may not be in the same formulation.

[0007] Aspects of the disclosure encompass integration of targeted molecular therapies and immune cell-based approaches. The targeted molecular therapies may comprise one or more agents that may be inhibitors, such as agents that have the ability to reduce or eliminate any function of a target whose activity is deleterious to an individual.

[0008] In embodiments, the one or more additional agents utilized in combination with NK cell therapy comprise one or more inhibitors of KRAS. In some cases, the NK cells (of any kind, including engineered) and / or the one or more additional agents for the combination therapy may target certain cells, such as cancer cells. The disclosure encompasses NK cells that may have been modified to express one or multiple heterologous proteins that are not naturally expressed in NK cells. In certain embodiments, the heterologous protein(s) are present on the surface of the NK cells.

[0009] In embodiments wherein a recipient of the encompassed combination there is provided, the individual may have cancer. The individual may or may not have a cancer in which the cells comprise a RAS mutation. In some embodiments, the individual has cancer but does not have cancer harboring one or more specific RAS mutations, and in specific embodiments the NK cells and the additional agent(s) exert synergy for the patient. In some embodiments, the individual has cancer and the cancer cells do harbor one or more specific RAS mutations, and in specific embodiments the NK cells and the additional agent(s) exert synergy for the patient.

[0010] In embodiments wherein a recipient of the encompassed combination there is provided, the individual may have cancer. The individual may or may not have a cancer in which the cells comprise one or more gene mutations that affect DNA repair. In some embodiments, the individual has cancer but does not have cancer harboring one or more gene mutations that affect DNA repair, and in specific embodiments the NK cells and the additional agent(s) exert synergy for the patient. In some embodiments, the individual has cancer and the cancer cells do harbor one or more specific mutations gene mutations that affect DNA repair,302140458.1 - 2 -and in specific embodiments the NK cells and the additional agent(s) exert synergy for the patient.

[0011] In particular embodiments, there is a method of treating cancer in an individual, comprising the step of administering to the individual a therapeutically effective amount of any one of the compositions encompassed herein. In particular embodiments, there is a method of treating cancer in an individual, comprising the step of administering to the individual a therapeutically effective amount of any one of the regimens encompassed herein. In some embodiments, the NK cells and the one or more additional agent(s) (inhibitors) may or may not be administered to the individual at the same time. The NK cells and the additional agent(s) (inhibitors) may or may not be administered in the same formulation. In specific embodiments, the NK cells and the one or more additional agent(s) (inhibitors) are administered to the individual at different times. In specific embodiments, the NK cells are autologous or allogeneic with respect to the individual.

[0012] Certain embodiments of the present disclosure are characterized through the following enumerated aspects.

[0013] Aspect 1 is a regimen, comprising: (a) a natural killer (NK) cell-based immunotherapy; and (b) one or more inhibitors of poly ADP-ribose polymerase (PARP) and / or one or more inhibitors of a Kirsten ras oncogene homolog from the mammalian ras gene family (KRAS) pathway.

[0014] Aspect 2 is a regimen of aspect 1, wherein the NK cells of the NK cell-based immunotherapy are wild type.

[0015] Aspect 3 is a regimen of aspect 1 or 2, wherein the NK cells of the NK cell-based immunotherapy are not engineered by the hand of man.

[0016] Aspect 4 is a regimen of aspect 1 or 2, wherein the NK cells of the NK cell-based immunotherapy are engineered by the hand of man.

[0017] Aspect 5 is a regimen of aspect 4, wherein the NK cells are engineered to express one or more of one or more chimeric antigen receptors, one or more T cell receptors or one or more chains thereof, one or more antibodies, one or more fusion proteins, one or more cytokines, one or more chemokine receptors, one or more CD3 chains, or a combination thereof, optionally wherein the NK cells are engineered to express a) a CD3 protein complex comprising part or all of a single chain or any combination of CD3(^, CD3d, CD3e, or CD3g, b) optionally at least one cytokine, c) at least one TCRb and TCRa chain and / or a TCRg and TCRd chain, and d) a polypeptide comprising a CD 16 Fc binding domain..302140458.1 - 3 -

[0018] Aspect 6 is a regimen of aspect 5, wherein the antibody is a bi-specific or multispecific antibody.

[0019] Aspect 7 is a regimen of any one of aspects 1-6, wherein the PARP is PARP1, PARP2, PARP3, PARP4, PARP5a, PARP5b, PARP7, PARP9, PARP11, PARP14, and / or PARP15.

[0020] Aspect 8 is a regimen of any one of aspects 1-7, wherein the PARP inhibitor is selected from the group consisting of olaparib, niraparib, rucaparib, talazoparib, veliparib, pamiparib, saruparib, Fluzoparib, INO-1001, CEP-9722, or a combination thereof.

[0021] Aspect 9 is a regimen of any one of aspects 1-8, wherein the one or more inhibitors of a KRAS pathway is an inhibitor of Kirsten RAS (KRAS), Harvey RAS (HRAS), Neuroblastoma RAS (N-RAS), R-RAS, RAP, RAL, RHEB, RIN, and / or RIT.

[0022] Aspect 10 is a regimen of any one of aspects 1 -9, wherein the one or more inhibitors of a KRAS pathway is an inhibitor of KRAS.

[0023] Aspect 11 is a regimen of any one of aspects 1-10, wherein (a) and (b) are in the same formulation.

[0024] Aspect 12 is a regimen of any one of aspects 1-10, wherein (a) and (b) are not in the same formulation.

[0025] Aspect 13 is a composition, comprising: (a) a natural killer (NK) cell-based immunotherapy; and (b) one or more inhibitors of poly ADP-ribose polymerase (PARP) and / or one or more inhibitors of a KRAS pathway, optionally wherein the one or more inhibitors of poly ADP-ribose polymerase (PARP) and / or one or more inhibitors of a KRAS pathway are in the NK cells.

[0026] Aspect 14 is a composition of aspect 13, wherein the composition is comprised in a pharmaceutically acceptable carrier.

[0027] Aspect 15 is a composition of aspect 13 or 14, wherein the NK cells of the NK cellbased immunotherapy are wild type.

[0028] Aspect 16 is a composition of any one of aspects 13-15, wherein the NK cells of the NK cell-based immunotherapy are not engineered by the hand of man.

[0029] Aspect 17 is a composition of any one of aspects 13-15, wherein the NK cells of the NK cell-based immunotherapy are engineered by the hand of man.

[0030] Aspect 18 is a composition of aspect 17, wherein the NK cells are engineered to express one or more of one or more chimeric antigen receptors, one or more T cell receptors or one or more chains thereof, one or more antibodies, one or more fusion proteins, one or more302140458.1 - 4 -cytokines, one or more chemokine receptors, one or more CD3 chains, or a combination thereof.

[0031] Aspect 19 is a composition of aspect 18, wherein the antibody is a bi-specific or multi-specific antibody.

[0032] Aspect 20 is a composition of any one of aspects 13-19, wherein the PARP is PARP1, PARP2, PARP3, PARP4, PARP5a, PARP5b, PARP7, PARP9, PARP11, PARP14, and / or PARP15.

[0033] Aspect 21 is a composition of any one of aspects 13-20, wherein the PARP inhibitor is selected from the group consisting of olaparib, niraparib, rucaparib, talazoparib, veliparib, pamiparib or a combination thereof.

[0034] Aspect 22 is a composition of any one of aspects 13-32, wherein the one or more inhibitors of a KRAS pathway is an inhibitor of Kirsten RAS (KRAS), Harvey RAS (HRAS), Neuroblastoma RAS (N-RAS), R-RAS, RAP, RAL, RHEB, RIN, and / or RIT.

[0035] Aspect 23 is a composition of any one of aspects 13-22, wherein the one or more inhibitors of a KRAS pathway is an inhibitor of KRAS.

[0036] Aspect 24 is a composition of any one of aspects 13-23, wherein (a) and (b) are in the same formulation.

[0037] Aspect 25 is a composition of any one of aspects 13-24, wherein (a) and (b) are not in the same formulation

[0038] Aspect 26 is a kit, comprising the composition of any one of aspects 13-25.

[0039] Aspect 27 is a method of treating an individual in need thereof, comprising the step of: subjecting the individual to a therapeutically effective amount of the regimen of any one of claims 1-12; or administering to the individual a therapeutically effective amount of the composition of any one of claims 13-25.

[0040] Aspect 28 is a method of aspect 27, wherein the individual has cancer or has an autoimmune disease.

[0041] Aspect 29 is a method of aspect 27 or 28, wherein there is a synergistic effect from (a) natural killer (NK) cell-based immunotherapy; and the (b) one or more inhibitors of poly ADP-ribose polymerase (PARP) and / or one or more inhibitors of a KRAS pathway.

[0042] Aspect 30 is a method of any one of aspects 27-29, wherein the NK cell-based immunotherapy is administered to the individual prior to the one or more inhibitors of PARP and / or the one or more inhibitors of a KRAS pathway.302140458.1 - 5 -

[0043] Aspect 31 is a method of any one of aspects 27-29, wherein the NK cell-based immunotherapy is administered to the individual subsequent to the one or more inhibitors of PARP and / or the one or more inhibitors of a KRAS pathway.

[0044] Aspect 32 is a method of any one of aspects 27-29, wherein the NK cell-based immunotherapy is administered to the individual at the same time as the one or more inhibitors of PARP and / or the one or more inhibitors of a KRAS pathway.

[0045] Aspect 33 is a method of treating an individual in need thereof, comprising: in any order, administering a therapeutically effective amount of a NK cell-based immunotherapy; and one or both of (1) one or more inhibitors of PARP and (2) one or more inhibitors of a KRAS pathway.

[0046] Aspect 34 is a method of aspect 33, wherein the NK cell-based immunotherapy and one or more inhibitors of PARP are administered to the individual.

[0047] Aspect 35 is a method of aspect 33, wherein the NK cell-based immunotherapy and one or more inhibitors of a KRAS pathway are administered to the individual.

[0048] Other objects, features and advantages of the present invention will become apparent from the following detailed description. It should be understood, however, that the detailed description and the specific examples, while indicating specific embodiments of the invention, are given by way of illustration only, since various changes and modifications within the spirit and scope of the invention will become apparent to those skilled in the art from this detailed description.BRIEF DESCRIPTION OF THE DRAWINGS

[0049] The following drawings form part of the present specification and are included to further demonstrate certain aspects of the present subject matter of the disclosure. The subject matter of the disclosure may be better understood by reference to one or more of these drawings in combination with the detailed description of specific embodiments presented herein.

[0050] FIG. 1. Combining NK cells with an example of a KRAS inhibitor ((1S,2S)-N-((63 S,4S,Z)- 11 -ethyl- 12-(2-((S)- 1 -m ethoxy ethyl)-5-(4-methylpiperazin- 1 -yljpyri din-3 -y 1)-10,10-dimethyl-5,7-dioxo-61,62,63,64,65,66-hexahydro-llH-8-oxa-2(4,2)-thiazola-l(5,3)-indola-6(l,3)-pyridazinacycloundecaphane-4-yl)-2-methylcyclopropane-l-carboxamide (also referred to as (RMC-6236)) enhances the antitumor activity of NK cells. FIG. 1 depicts representative images of 3D tumor spheroids treated with NK cells, KRAS inhibitor, or both combinations. The spheroids were cultured for 7 days with NK cells and / or KRAS inhibitor.302140458.1 - 6 -

[0051] FIG. 2. Combining NK cells with an example of a KRAS inhibitor (6-Fluoro-7-(2-fluoro-6-hydroxyphenyl)-(lM)-l-[4-methyl-2-(propan-2-yl)pyridin-3-yl]-4-[(2S)-2- methyl-4-(prop-2-enoyl)piperazin-l-yl]pyrido[2,3-d]pyrimidin-2(lH)-one (also known as Sotorasib) enhances the antitumor activity of NK cells. FIG. 2 depicts representative images of 3D tumor spheroids treated with NK cells, KRAS inhibitor, or both combinations. The spheroids were cultured for 7 days with NK cells and / or KRAS inhibitor.

[0052] FIG. 3. Combining NK cells with Sotorasib or RMC-6236 enhances the antitumor activity of NK cells. FIG. 3 depicts representative images of 3D tumor spheroids treated with NK cells, KRAS inhibitor, or both combinations. The spheroids were cultured for 7 days with NK cells and / or KRAS inhibitor.

[0053] FIG. 4. Combining NK cells with Sotorasib enhances the antitumor activity of NK cells. FIG. 4 depicts representative real-time impedance data from xCELLigence assay of HCT-116 (CRC) cells that were treated with Sotorasib (1 pM) for 24 hours, followed by treatment with NK cells or no treatment. SDS (sodium dodecyl sulfate) was used as a full-lysis positive control.

[0054] FIG. 5. Combining CAR-NK cells with Sotorasib enhances the antitumor activity of NK cells. FIG. 5 depicts representative real-time impedance data from xCELLigence assay of HCT-116 (CRC) cells that were treated with Sotorasib (1 pM) for 24 hours, followed by treatment with CAR-NK cells or no treatment. SDS (sodium dodecyl sulfate) was used as a full-lysis positive control.

[0055] FIG. 6. Combining NK cells with RMC-6236 enhances the antitumor activity of NK cells. FIG. 6 depicts representative real-time impedance data from xCELLigence assay of HCT-116 (CRC) cells that were treated with RMC-6236 (1 pM) for 24 hours, followed by treatment with NK cells or no treatment. SDS (sodium dodecyl sulfate) was used as a full-lysis positive control.

[0056] FIG. 7. Combining CAR-NK cells with RMC-6236 enhances the antitumor activity of NK cells. FIG. 7 depicts representative real-time impedance data from xCELLigence assay of HCT-116 (CRC) cells that were treated with RMC-6236 (1 pM) for 24 hours, followed by treatment with TROP2 / CARNK cells or no treatment. SDS (sodium dodecyl sulfate) was used as a full-lysis positive control.

[0057] FIG. 8. Combining NT or CAR-NK cells with RMC-6236 enhances the antitumor activity of NK cells. FIG. 8 depicts representative real-time impedance data from xCELLigence assay of MIA-Paca-2 (PDAC) cells that were treated with RMC-6236 (1 pM) for 24 hours,302140458.1 - 7 -followed by treatment with TROP2 / CAR NK cells or no treatment. SDS (sodium dodecyl sulfate) was used as a full-lysis positive control.

[0058] FIG. 9. Combining NT or CAR-NK cells with RMC-6236 enhances the antitumor activity of NK cells. FIG. 9 depicts representative real-time impedance data from xCELLigence assay of Hep-G2(HCC) cells that were treated with RMC-6236 (1 pM) for 24 hours, followed by treatment with TROP2 / CAR NK cells or no treatment. SDS (sodium dodecyl sulfate) was used as a full-lysis positive control.

[0059] FIGS. 10A and 10B. Combining CAR-NK cells with an example of a PARPi (Olaparib) enhances the antitumor activity of NK cells. FIG. 10A depicts representative realtime impedance data from xCELLigence assay of SKOV3 (Ovarian Cancer) cells that were treated with Olaparib (lOuM) for 24 hours, followed by treatment with TROP2 / CARNK cells or no treatment. FIG. 10B depicts representative real-time impedance data from xCELLigence assay of PATC148 (PDAC) cells that were treated with Olaparib (lOuM) for 24 hours, followed by treatment with TROP2 / CAR NK cells or no treatment.

[0060] FIG. 11. Combining CAR-NK cells with Olaparib enhances the antitumor activity of NK cells. FIG. 11 depicts representative image data from a spheroid assay of SKOV3 (Ovarian Cancer) cells that were treated with Olaprib (lOOng / ml) for 24 hours, followed by treatment with CAR-NK cells or no treatment.

[0061] FIGS. 12A-12E. RMC-6236 synergizes with CARNK cells to enhance antitumor activity in KRAS-mutant pancreatic cancer models. FIG. 12A depicts an experimental schematic. FIG. 12B depicts real-time cytotoxicity curves demonstrating the effects of RMC-6236 alone, CARNK cells alone, and combination treatment in MIA-Paca-2 (KRASAG12C) model. FIG. 12C depicts real-time cytotoxicity curves demonstrating the effects of RMC-6236 alone, CAR NK cells alone, and combination treatment in PATC148 (KRASAG12D) model. FIG. 12D depicts representative fluorescence images of tumor spheroids (MIA-Paca-2 (KRASAG12C)) at Day 0 and Day 6 under treatment with CAR / IL15 NK cells alone or in combination with RMC-6236. FIG. 12E depicts representative fluorescence images of tumor spheroids at Day 0 and Day 6 under treatment with TROP2 / IL15 NK cells alone or in combination with RMC-6236.

[0062] FIGS. 13A-13D. RMC-6236 enhances tumor susceptibility to CAR NK cells and preserves NK cell functional states. FIG. 13 A depicts representative flow cytometry histograms showing surface expression of stress ligands (MICA / B, ULBP2 / 5 / 6, and B7-H6) on tumor cells following treatment with RMC-6236. RMC-6236-treated tumor cells (ii) and untreated controls (i) are shown. RMC-6236 treatment promotes the expression of stress-induced ligands302140458.1 - 8 -on tumor cells, thereby enhancing their recognition and elimination by NK cells. Numbers indicate mean fluorescence intensity (MFI). FIG. 13B depicts CyTOF -based dimensionality reduction (FlowSOM clustering over UMAP projection) of NK cells co-cultured with tumor cells in the absence (top row) or presence (bottom row) of RMC-6236. Each color represents a distinct metacluster. FIG. 13C depicts quantification of FlowSOM metacluster distribution across experimental conditions. FIG. 13D depicts a heatmap of CyTOF marker expression across NK cell metaclusters. These data suggest that RMC-6236 in combination with NK cells protects NK cells from tumor-induced dysfunction and preserves functional NK cell subsets, including metaclusters 2-10, characterized by high expression of activation and cytotoxic markers such as NKG2D, DNAM-1, and Granzyme B.

[0063] FIGS. 14A-14B. RMC-6236 enhances tumor immunogenicity while reprogramming CAR NK cell transcriptional states. FIG. 14A depicts differential gene expression analysis of PATC148 tumor cells in NK-tumor co-culture following RMC-6236 treatment at 3 and 6 days. Lollipop plots depict A mean variance-stabilized expression (RMC - Control), with lollipop plots on right of value 0 for A mean VST indicating upregulation and lollipop plots on left with negative A mean VST values indicating downregulation. FIG. 14B depicts a heatmap of row-scaled (z-score) transcript expression in CAR NK cells treated with RMC-6236. Columns represent biological replicates, and genes cluster into functional modules. RNA-seq analysis demonstrated that RMC-6236 treatment of tumor cells downregulates inhibitory ligands associated with NK cell suppression, including HLA-ABC and HLA-E, while upregulating stress-induced ligands that promote NK cell activation, such as ULBP2 and MICA / B.

[0064] FIG. 15. KRAS inhibition reprograms ERK-STAT signaling to enhance CAR NK cell antitumor activity. FIG. 15 depicts immunoblot analyses of key signaling nodes in CAR NK cells following KRAS pathway inhibition. Representative Western blots show phosphorylation and total protein levels across indicated experimental conditions. The dashed line separates control and KRAS inhibitor-treated samples. Loading controls are shown at the bottom of each panel. In summary, Western blot analysis of NK cells demonstrated that RMC-6236 suppresses MAPK and mTOR signaling, as evidenced by reduced ERK and S6 phosphorylation, while enhancing STAT signaling pathways, including STAT3 and STAT5. This signaling shift suggests that RMC-6236 reprograms NK cells toward a cytokine-responsive state that may promote sustained NK-cell functionality.

[0065] FIGS. 16A-16B. Combination of CAR NK cells with the KRAS inhibitor RMC-6236 enhances antitumor activity and tumor infiltration in a PATC148 pancreatic ductal302140458.1 - 9 -adenocarcinoma (PDAC) model. FIG. 16A depicts representative longitudinal bioluminescence imaging (BLI) of NSG mice bearing luciferase-expressing PATC148 tumors treated with: vehicle control (PATC148 alone), RMC-6236 alone, TROP2 / IL-15 CAR-NK cells alone, or the combination of TROP2 / IL-15 CAR-NK cells with RMC-6236. Images were acquired at Days -4 pre NK treatment, and Days 6, 14, 20, and 27 post-treatment. Tumor burden is shown as pseudocolor radiance (photons / sec / cm2 / sr *106). FIG. 16B depicts quantification of tumor burden over time measured by total flux (photons / sec). Data are presented as mean ± SEM. These data demonstrate that the combination of RMC-6236 with CAR NK cells significantly enhances antitumor efficacy in vivo in the PATC148 tumor model.

[0066] FIGS. 17A-17D. KRAS inhibition enhances CAR NK cell antitumor activity and intratumoral immune infiltration in vivo. FIG. 17A depicts representative low-magnification H&E and CD45 immunohistochemistry of PATC148 PDAC xenografts treated with vehicle control (Tumor Alone), CAR NK cells (TROP2 / IL15), or the combination with RMC-6236. Dashed outlines demarcate tumor regions (scale bar, 1 mm). FIG. 17B depicts higher-magnification of H&E and CD45 staining (scale bar, 100 pm). FIG. 17C depicts quantification of intratumoral CD45+immune infiltration across treatment groups. Violin plots depict distribution of CD45+cell density per tumor section. FIG. 17D depicts quantification of tumor burden across groups at endpoint. Violin plots show tumor volume distribution. Immunohistochemical analysis demonstrated increased CD45+immune cell infiltration within tumors following combination therapy with RMC-6236 and CAR NK cells. This enhanced immune infiltration correlated with a significantly reduced tumor burden in the PATC148 model, supporting improved antitumor activity of the combination treatment.

[0067] FIGS. 18A-18C. RAS inhibition in combination therapy preserves NK cell function and prevents exhaustion-induced dysfunction in vivo. FIG. 18A depicts longitudinal analysis of NK cell functional capacity following exposure to tumor-bearing environments in vivo. NK cells isolated from mice treated with CAR NK cells alone (blue) or CAR NK cells plus RMC-6236 (red) were assessed for effector function over time. FIG. 18B depicts representative images of tumor spheroid co-culture assays at day 0 and day 6. NK cells were derived from combination-treated mice. FIG. 18C depicts quantification of spheroid area or viability over time following exposure to NK cells derived from treated mice. These findings suggest that RMC-6236 preserves NK-cell function by protecting NK cells from tumor- and TME-induced dysfunction, thereby enhancing immune infiltration and antitumor activity.

[0068] FIGS. 19A-19B. CAR NK cells combined with KRAS inhibition acquire a distinct metabolic and inflammatory transcriptional phenotype in vivo. FIG. 19A depicts gene set302140458.1 - 10 -enrichment analysis (GSEA) of CARNK cells isolated from tumors in mice treated with CAR NK cells alone versus CAR NK cells plus RMC-6236. Normalized enrichment scores (NES) are shown for hallmark pathways, with color indicating -log 10 FDR. FIG. 19B depicts Unsupervised hierarchical clustering heatmap of differentially expressed genes in CAR NK cells isolated from tumors of non-treated versus RMC-6236-treated mice. Columns represent individual biological replicates, and rows represent differentially expressed genes. Pathway enrichment analysis demonstrates that RMC-6236 promotes metabolic fitness and immune activation pathways while suppressing tumor-promoting pathways such as KRAS, TGF-P, and Wnt signaling.DETAILED DESCRIPTION

[0069] PCT Application No. PCT / US2024 / 027601 is incorporated by reference herein in its entirety.

[0070] In keeping with long-standing patent law convention, the words “a” and “an” when used in the present specification in concert with the word comprising, including the claims, denote “one or more.” Some embodiments of the disclosure may consist of or consist essentially of one or more elements, method steps, and / or methods of the disclosure. It is contemplated that any method or composition described herein can be implemented with respect to any other method or composition described herein and that different embodiments may be combined.

[0071] Throughout this specification, unless the context requires otherwise, the words “comprise”, “comprises” and “comprising” will be understood to imply the inclusion of a stated step or element or group of steps or elements but not the exclusion of any other step or element or group of steps or elements. By “consisting of’ is meant including, and limited to, whatever follows the phrase “consisting of.” Thus, the phrase “consisting of’ indicates that the listed elements are required or mandatory, and that no other elements may be present. By “consisting essentially of’ is meant including any elements listed after the phrase, and limited to other elements that do not interfere with or contribute to the activity or action specified in the disclosure for the listed elements. Thus, the phrase “consisting essentially of’ indicates that the listed elements are required or mandatory, but that no other elements are optional and may or may not be present depending upon whether or not they affect the activity or action of the listed elements.302140458.1 - 11 -

[0072] Reference throughout this specification to “one embodiment,” “an embodiment,” “a particular embodiment,” “a related embodiment,” “a certain embodiment,” “an additional embodiment,” or “a further embodiment” or combinations thereof means that a particular feature, structure or characteristic described in connection with the embodiment is included in at least one embodiment of the present invention. Thus, the appearances of the foregoing phrases in various places throughout this specification are not necessarily all referring to the same embodiment. Furthermore, the particular features, structures, or characteristics may be combined in any suitable manner in one or more embodiments.

[0073] As used herein, the terms “or” and “and / or” are utilized to describe multiple components in combination or exclusive of one another. For example, “x, y, and / or z” can refer to “x” alone, “y” alone, “z” alone, “x, y, and z,” “(x and y) or z,” “x or (y and z),” or “x or y or z.” It is specifically contemplated that x, y, or z may be specifically excluded from an embodiment.

[0074] Throughout this application, the term “about” is used according to its plain and ordinary meaning in the area of cell and molecular biology to indicate that a value includes the standard deviation of error for the device or method being employed to determine the value.

[0075] As used herein, the term “CD3 receptor complex” or “CD3 co-receptor complex” refers to the protein complex that in nature acts as a T cell co-receptor and is comprised of CD3 chain, CD3y chain, a CD36 chain, and two CD3s chains (although in alternatives only one CD3e chain is used).

[0076] The term “combination therapy” and similar terms, as used herein, may refer to at least two different entities that are utilized as a therapy for an individual in need of the therapy. The two or more different entities may or may not be administered to an individual as a physical combination or mixture. The two different entities may or may not be administered to the individual at the same time and / or by the same administration route. One of the entities of the combination therapy may be administered more frequently and / or in greater quantity than the other entity. In some embodiments, the two different entities have an additive effect as the therapy for the individual, whereas in some embodiments the two different entities have a synergistic effect as the therapy for the individual.

[0077] The term “engineered” as used herein refers to an entity that is generated by the hand of man, including a cell, nucleic acid, polypeptide, vector, and so forth. In at least some cases, an engineered entity is synthetic and comprises elements that are not naturally present or configured in the manner in which it is utilized in the disclosure. In specific embodiments,302140458.1 - 12 -a vector is engineered through recombinant nucleic acid technologies, and a cell is engineered through transfection or transduction of an engineered vector. Cells may be engineered to express heterologous proteins that are not naturally expressed by the cells, either because the heterologous proteins are recombinant or synthetic or because the cells do not naturally express the proteins.

[0078] The phrases “pharmaceutical or pharmacologically acceptable” refers to molecular entities and compositions that do not produce an adverse, allergic, or other untoward reaction when administered to an animal, such as a human, as appropriate. The preparation of a pharmaceutical composition comprising an antibody or additional active ingredient will be known to those of skill in the art in light of the present disclosure. Moreover, for animal (e.g., human) administration, it will be understood that preparations should meet sterility, pyrogenicity, general safety, and purity standards as required by FDA Office of Biological Standards.

[0079] As used herein, “pharmaceutically acceptable carrier” includes any and all aqueous solvents (e.g., water, alcoholic / aqueous solutions, saline solutions, parenteral vehicles, such as sodium chloride, Ringer's dextrose, efc.), non-aqueous solvents (e.g., propylene glycol, polyethylene glycol, vegetable oil, and injectable organic esters, such as ethyloleate), dispersion media, coatings, surfactants, antioxidants, preservatives (e.g., antibacterial or antifungal agents, anti-oxidants, chelating agents, and inert gases), isotonic agents, absorption delaying agents, salts, drugs, drug stabilizers, gels, binders, excipients, disintegration agents, lubricants, sweetening agents, flavoring agents, dyes, fluid and nutrient replenishers, such like materials and combinations thereof, as would be known to one of ordinary skill in the art. The pH and exact concentration of the various components in a pharmaceutical composition are adjusted according to well-known parameters.

[0080] The term “subject,” as used herein, generally refers to an individual having a that has or is suspected of having cancer. The subject can be any organism or animal subject that is an object of a method or material, including mammals, e.g., humans, laboratory animals (e.g., primates, rats, mice, rabbits), livestock (e.g., cows, sheep, goats, pigs, turkeys, and chickens), household pets (e.g., dogs, cats, and rodents), horses, and transgenic non-human animals. The subject can be a patient, e.g., have or be suspected of having a disease (that may be referred to as a medical condition), such as benign or malignant neoplasias, or cancer. The subject may being undergoing or having undergone treatment. The subject may be asymptomatic. The subject may be healthy individuals but that are desirous of prevention of cancer. The term “individual” may be used interchangeably, in at least some cases. The “subject” or "individual",302140458.1 - 13 -as used herein, may or may not be housed in a medical facility and may be treated as an outpatient of a medical facility. The individual may be receiving one or more medical compositions via the internet. An individual may comprise any age of a human or non-human animal and therefore includes both adult and juveniles ( / .<?., children) and infants and includes in utero individuals. It is not intended that the term connote a need for medical treatment, therefore, an individual may voluntarily or involuntarily be part of experimentation whether clinical or in support of basic science studies.

[0081] As used herein “treatment” or “treating,” includes any beneficial or desirable effect on the symptoms or pathology of a disease or pathological condition, and may include even minimal reductions in one or more measurable markers of the disease or condition being treated, e.g., cancer. Treatment can involve optionally either the reduction or amelioration of one or more symptoms of the disease or condition, or the delaying of the progression of the disease or condition. “Treatment” does not necessarily indicate complete eradication or cure of the disease or condition, or associated symptoms thereof. Treating may mean alleviation of at least one symptom of the disease or condition.I. General Embodiments of the Disclosure

[0082] The present disclosure specifically relates to NK cells, whether or not they have been engineered in any manner by the hand of man, that are used with another one or more therapies (not necessarily at the same time or in the same formulation) as a therapy for an individual in need of such a therapy. The NK cells may or may not have been modified to express one or more heterologous proteins. The NK cells may or may not comprise one or more vectors that express one or more gene products, whether or not the cells also naturally express the one or more gene products. In some embodiments, wild-type (including that have not been engineered) or engineered NK cells are given as a therapy in conjunction with one or more other agents, including one or more inhibitors of any kind. The inhibitor(s) may target a particular protein, group of similar proteins, class of proteins, proteins with the same or similar function, and so forth.

[0083] In some embodiments, the subject matter of the disclosure relates to novel therapeutic strategies combining natural killer (NK) cells (whether engineered or not) with one or more KRAS inhibitors and / or one or more PARP inhibitors, and in specific embodiment the one or more KRAS inhibitors and / or one or more PARP inhibitors enhance the antitumor activity of NK cells against cancer. The combination treatment demonstrates a synergistic302140458.1 - 14 -effect, in some cases, such as significantly reducing tumor viability and / or size compared to treatment with either the NK cells or one or more KRAS inhibitors and / or one or more PARP inhibitors alone.

[0084] In certain embodiments, the combination therapy targets cancer cells of any kind, including tumor spheroids and leverages the effects of any inhibitors encompassed herein on cancer cells, for example to improve their susceptibility to NK cell-mediated cytotoxicity. As shown elsewhere herein, studies conducted on PANCI pancreatic cancer cells and three-dimensional (3D) tumor spheroids showed that pretreatment with KRAS inhibitors enhanced NK cell anti-cancer activity, as evidenced by a marked reduction in spheroid viability and size over seven days of treatment (as one example). Additionally, the combination of KRAS inhibitors with either non-transduced (NT) NK cells or CAR NK cells demonstrated enhanced efficacy in various cancer models, including MIA-Paca-2 (PDAC) and Hep-G2 (HCC). Furthermore, the synergistic potential of other agents, such as the PARP inhibitor Olaparib, showed enhanced CAR NK cell activity against ovarian cancer as an example of cancer. Embodiments of combining NK cell-based immunotherapies with other agent(s) (including at least small molecule inhibitors), including those that target KRAS pathways and / or that are PARP inhibitors, are an embodiment treatment of cancers, including hematological and solid tumors, and overcoming resistance mechanisms. This disclosure offers a promising therapeutic solution to address the unmet need in treating cancers, facilitating outcomes through the integration of targeted molecular therapies and immune cell-based approaches.

[0085] The disclosure concerns compositions that at least include engineered and / or nonengineered NK cells and one or both of (a) one or more KRAS inhibitors and (b) one or more PARP inhibitors. In some cases, the NK cells and any of the inhibitor(s) are in the same formulation, although in alternative embodiments the NK cells and any of the inhibitor(s) are utilized as physically separate compositions.

[0086] In particular embodiments, the NK cells and the one or more KRAS inhibitors exert a synergistic therapeutic effect for a recipient individual. In particular embodiments, the NK cells and the one or more PARP inhibitors exert a synergistic therapeutic effect for a recipient individual. In particular embodiments, the NK cells, the one or more KRAS inhibitors, and the one or more KRAS inhibitors exert a synergistic therapeutic effect for a recipient individual.302140458.1 - 15 -II. Inhibitors of a KRAS Pathway

[0087] In embodiments of the disclosure one or more inhibitors of a KRAS pathway are utilized. In some embodiments, one or more KRAS inhibitors may form a covalent bond with KRAS. In some cases, the inhibitor irreversibly engages the KRAS protein through covalent bond formation. In certain embodiments, one or more KRAS inhibitors are panKRAS inhibitors, able to bind multiple members of the RAS family, such as inhibitors of two or more of Harvey RAS (HRAS), Kirsten RAS (KRAS), Neuroblastoma RAS (N-RAS), R-RAS, RAP, RAL, RHEB, RIN, and / or RIT.

[0088] In some aspects, one or more KRAS inhibitors bind a particular mutation in the KRAS protein, such as the G12 or G13 amino acid site. In some aspects, one or more KRAS inhibitors bind a particular mutation in the KRAS protein, such as a single amino acid change, a deletion, an inversion, and so forth. In some aspects, one or more KRAS inhibitors bind a particular mutation in the KRAS protein, such as the G12C genetic mutation (which occurs in several types of cancer, including at least lung, colorectal, and pancreatic cancer) that is a single point mutation that occurs when glycine is replaced by cysteine at codon 12. In some embodiments, the inhibitor may inhibit G12D, G12S and / or G12R, merely as examples. Some KRAS inhibitors may or may not target only a mutant KRAS protein and not a wild-type KRAS protein. In some embodiments, the KRAS inhibitor targets any mutation at G12, including at least G12A, G12R, G12C, G12V, G12D, or G12S. In certain embodiments, the inhibitor targets any G13 mutation, including at least G13D. The inhibitor may target a KRAS mutation that is not at G12 and / or G13. The inhibitor may not target a mutation at G12 and / or G13.

[0089] In some embodiments, an agent utilized as part of a combination therapy is a selective inhibitor that targets one or more KRAS mutations while sparing RAS signaling in normal cells. In some embodiments, an inhibitor is a pan-KRAS inhibitor that may also be a pan-KRAS selective inhibitor. In certain embodiments, the inhibitor inhibits wild-type and mutant KRAS isoforms, but not NRAS and HRAS.

[0090] The inhibitor may be a mutation-selective KRAS inhibitor that inhibits wild-type KRAS, NRAS and HRAS. The inhibitor may be a pan-KRAS inhibitor that does not inhibit wild-type NRAS and HRAS. The inhibitor may be a pan-KRAS inhibitor that targets all KRAS, NRAS and HRAS mutations.

[0091] The inhibitor may inhibit one or more isoforms of a member of the RAS family, including at least KRAS having two splice variants, KRAS4A and KRAS4B. In specific embodiments, the inhibitor of a KRAS pathway comprises Otorasib, adagrasib (also referred302140458.1 - 16 -to as MRTX-849), MRTX1133, JNJ-74699157 (also known as ARS-3248), LY3499446, LY3537982, JDQ-443, GDC-6036, TG6330, D-1553, BPI-421286, GH35, BEBT-607, JAB-21000, IBI351, a selective KRASG12D inhibitor, (e.g., MRTX1133), one or more of the following from Zhang, et al., Nat. Chem. Biol. 18, 1177-1183 (2022): G12Si-l (3-(7-(8-chloronaphthalen-l-yl)-2-(((S)-l-methylpyrrolidin-2-yl)methoxy)-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidin-4-yl)-8-oxa-3-azabicyclo[4.2.0]octan-7-one);i3t25i-3 1-((lR,5S)-3-(7-(8-chloronaphthalen-l-yl)-8-fluoro-2-((tetrahydro-lH-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octan-8-yl)-2,2-dihydroxybutane- 1,3 -di one (Zhang, et al., J. Am. Chem. Soc. 144, 15916-15921 (2022)), (S)- 2-(4-(7-(8-chloronaphthalen-l-yl)-8-fluoro-2-((tetrahydro-lH-pyrrolizin-7a(5H)-yl)methoxy)pyrido[4,3-d]pyrimidin-4-yl)-l-(2,2-dihydroxy-3-oxobutanoyl)piperazin-2-yl)acetonitrile (Zhang, et al., J. Am. Chem. Soc. 144, 15916-15921 (2022)), or a combination thereof.III. Inhibitors of PARP

[0092] In embodiments of the disclosure, one or more PARP inhibitors (that also may be referred to as poly (ADP-ribose) polymerase inhibitors) are utilized for therapy. The PARP protein helps repair damaged DNA in cells, and PARP inhibitors trap the PARP protein at DNA damage sites resulting in an accumulation of DNA damage and ultimately leading to death of the cell. PARP inhibitors for treating cancer cells keep the cancer cells from repairing damaged DNA, leading to cancer cell death. In specific embodiments, PARP inhibitors are a targeted therapy. In some embodiments, the PARP inhibitor(s) are utilized against cancers with DNA repair defects, such as those caused by BRCA gene mutations. The inhibitor may inhibit one or more of the following: PARP1, PARP2, PARP3, PARP4, PARP5a, PARP5b, PARP7, PARP9, PARP11, PARP 14, and / or PARP15.302140458.1 - 17 -

[0093] A PARP inhibitor for use in embodiments of the present disclosure may be selected from the group consisting of olaparib, niraparib, rucaparib, talazoparib, veliparib, pamiparib, saruparib, Fluzoparib, INO-1001, CEP-9722, and a combination thereof.IV. NK Cells

[0094] The NK cells that are utilized may be obtained from any suitable source, including fresh or frozen. In certain embodiments, NK cells are not NK cells obtained from iPSC differentiation. In certain embodiments, NK cells are derived from human peripheral blood mononuclear cells (PBMC), unstimulated leukapheresis products (PBSC), NK cell lines (e.g., NK-92), human embryonic stem cells (hESCs), induced pluripotent stem cells (iPSCs), bone marrow, or umbilical cord blood by methods well known in the art. Specifically, the NK cells may be isolated from cord blood (CB), peripheral blood (PB), bone marrow, stem cells, NK cell lines, or a mixture thereof. In particular embodiments, the NK cells are isolated from pooled CB. The CB may be pooled from 2, 3, 4, 5, 6, 7, 8, 9, 10, or more units. The NK cells may be autologous or allogeneic with respect to a recipient individual. The isolated NK cells may or may not be haplotype matched for the subject to be administered the cell therapy. NK cells can be detected by specific surface markers, such as CD 16 and CD56 in humans, for example. In some cases, the source of the NK cells is cord blood and the NK cells may be in the cord blood in a heterogeneous mixture of cells and may be depleted of certain cells expressing CD3. In other methods, umbilical CB is used to derive NK cells by the isolation of CD34+ cells.

[0095] In certain embodiments, the NK cells may be pre-activated, such as with one or more inflammatory cytokines, and they may be expanded or not be expanded. In some cases, the NK cells are pre-activated either prior to engineering of any kind or following engineering of any kind. In specific embodiments, pre-activation of the NK cells may comprise culturing the isolated NK cells in the presence of one or more cytokines. The NK cells may be stimulated with IL-2, or other cytokines that bind the common gamma-chain (e.g., IL-7, IL-12, IL-15, IL-18, IL-21, and others). In particular embodiments, the pre-activation cytokines may be selected from the group consisting of IL-12, IL-15, IL-18, and a combination thereof. One or more additional cytokines may be used for the pre-activation step. The pre-activation may be for a short period of time such as 5-72 hours, such as 10-50 hours, particularly 10-20 hours, such as 12, 13, 14, 15, 16, 17, 18, 19, or 20 hours, specifically about 16 hours. The pre-activation culture may comprise IL-12 at a concentration of 0.1-150 ng / mL, such as 0.5-50 ng / mL,302140458.1 - 18 -particularly 1-20 ng / mL, such as 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 ng / mL, specifically about lOng / mL. The pre-activation culture may comprise IL-18 and / or IL-15 at a concentration of 10-100 ng / mL, such as 40-60 ng / mL, particular 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, or 55 ng / mL, specifically about 50 ng / mL.

[0096] In some cases, the NK cells are expanded either prior to engineering of any kind or following engineering of any kind. Pre-activated NK cells may be expanded in the presence of artificial antigen presenting cells (aAPCs) and / or feeders / fragments or NK activating beads. The pre-activated NK cells may be washed prior to expansion, such as 2, 3, 4, or 5 times, specifically 3 times. The aAPCs may be engineered to express CD137 ligand and / or a membrane-bound cytokine. The membrane-bound cytokine may be membrane-bound IL-21 (mIL-21) or membrane-bound IL-15 (mIL-15). In particular embodiments, the aAPCs are engineered to express CD137 ligand and mIL-21. The aAPCs may be derived from cancer cells, such as leukemia cells. The aAPCs may not express endogenous HLA class I, II, or CD Id molecules. They may express ICAM-1 (CD54) and LFA-3 (CD58). In particular, the aAPCs may be K562 cells, such as K562 cells engineered to express CD137 ligand and mIL-21. The aAPCs may be irradiated. In some embodiments, fragments of APC can be used to expand the NK cells. The engineering may be by any method known in the art, such as retroviral transduction. Retroviral transduction may be at least, at most, or about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 days following NK co-culturing with an antigen presenting cell. In some embodiments, retroviral transduction comprises co-transduction of more than one construct. In some embodiments, retroviral transduction occurs after or at about 5 days of co-culturing with an antigen presenting cell. In some embodiments, co-culturing with an antigen presenting cell continues following transduction of an NK cell. The expansion may be for about 2-30 days, such as 3-20 days, particularly 12-16 days, such as 12, 13, 14, 15, 16, 17, 18, or 19 days, specifically about 14 days. The pre-activated NK cells and aAPCs may be present at a ratio of about 3: 1-1:3, such as 2:1, 1:1, 1:2, specifically about 1:2. The expansion culture may further comprise cytokines to promote expansion, such as IL-2. The IL-2 may be present at a concentration of about 10-500 U / mL, such as 100-300 U / mL, particularly about 200 U / mL. The IL-2 may be replenished in the expansion culture, such as every 2-3 days. The aAPCs may be added to the culture at least a second time, such as at about 7 days of expansion.

[0097] In particular embodiments, the NK cells are transfected or transduced with one or more membrane bound cytokines, including IL-21, IL-12, IL-18, IL-23, IL-7, and / or IL-15, either secreted by NK cells or tethered to the NK cell membrane. In such cases, the membrane bound cytokine may be tethered to the NK cell membrane with a particular transmembrane302140458.1 - 19 -domain, such as the transmembrane domain of CD8, CD28, CD27, B7H3, IgGl, IgG4, CD4, DAP 10, DAP 12, for example.

[0098] Following preparation, the engineered NK cells may be immediately infused into an recipient individual, or the NK cells may be stored, such as by cryopreservation. In some cases, when the NK cells are going to be subject to cry opreservation, the NK cells are deactivated pre-cryopreservation using a deactivating agent (e.g., a kinase inhibitor, e.g., Dasatinib, nilotinib, rapamycin, etc.). In certain aspects, the cells may be propagated for days, weeks, or months ex vivo as a bulk population within about 1, 2, 3, 4, or 5 days.

[0099] The NK cells may be employed as therapy without engineering, including without modification(s) by the hand of man, although in some cases the non-engineered NK cells are expanded first to provide sufficient amounts of cells.

[0100] In embodiments wherein engineered NK cells are utilized as therapy, the engineering may be of any kind. They may be transduced or transfected with a heterologous protein, a synthetic protein, an antibody, or a combination thereof. NK cells may be engineered to express one or more of the following: one or more chimeric antigen receptors, one or more T cell receptors or one or more chains thereof, one or more antibodies, one or more fusion proteins, one or more cytokines, one or more chemokine receptors, one or more CD3 chains, or a combination thereof.

[0101] In particular embodiments, the NK cells are loaded with antibodies prior to use. The NK cells may be loaded in any specific manner, including in culture or immediately before infusion, for example, to produce a complex of NK cells with the antibodies. The conditions are suitable enough to allow for an effective amount of antibody to bind to the surface of the NK cells. In the case of use of monospecific antibodies, the Fc region of the monospecific antibody binds the NK cell while the antigen binding domain of the monospecific antibody is free to bind its target antigen. In certain embodiments of cases of use of multispecific antibodies, one or more antigen binding domains of the antibody can bind the surface of the NK cells, such as through an antigen on the surface of the NK cells, (for example but not limited to, CD3, NKp30, NKp44, NKp46, CD16, CD32, CD64, KIRs, and the like), and the other antigen binding domain is free to bind its target antigen. In certain embodiments of cases of use of multispecific antibodies, one or more antigen binding domains of the antibody can bind one or more target antigens.

[0102] The culture conditions by which the NK cells become loaded may or may not be of a particular type having one or more specific parameters. In particular embodiments, the loading of the NK cells occurs in culture at a specific temperature, such as 37 °C, although in302140458.1 - 20 -alternative embodiments the temperature is 36 °C or 38 °C, or lower or higher. The duration of the loading step may be for any suitable amount of time, such as in a range of one minute to 24 hours or longer. For example, the range may be in the range of 1 min to 24 hrs, 1 min to 18 hrs, 1 min to 12 hours, 1 min to 6 hrs, 1 min to 1 hr, 30 min to 24 hrs, 30 min to 18 hrs, 30 min to 12 hrs, 30 min to 6 hrs, 30 min to 1 hr, 1-24 hrs, 1-18 hrs, 1-12 hrs, 1-6 hrs, 6-24 hrs, 6-18 hrs, 6-12 hrs, 12-24 hrs, 12-18 hrs, or 18-24 hrs. In some embodiments, the duration of the loading step may be greater than or equal to approximately 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, or 48 hours, or any range derivable therein. In specific embodiments, the cell culture media is basal media or complex media. In some cases, the culture comprises one or more reagents that were utilized during pre-activation and / or expansion steps, while in other cases the culture does not. In specific embodiments, the culture comprises one or more cytokines, including one or more of IL-12, IL-15, IL-2, and IL-18, for example. In some embodiments, the culture comprises APCs of any kind.

[0103] In certain embodiments, antibodies of compositions described herein are subjected in an effective amount to an effective amount of NK cells of the disclosure, thereby producing a complex that is “chimeric antigen receptor-like.” In particular, an antigen binding domain of the antibody binds to the NK cells, such as through the antigen that is a cell surface protein. A plurality of antibodies may be subjected to a plurality of NK cells such that there are multiple complexes of cell / antibody. The antibodies may be of any type, including monospecific, bi specific, or multi specific, and in specific cases the antibody engages both the NK cell and a target antigen through an antigen binding domain of the antibody (such as with engagers in the art that are fusion proteins consisting of two single-chain variable fragments (scFvs) of different antibodies). In cases wherein the antibody is monospecific, an antigen binding domain of the antibody binds a target antigen, such as a cancer antigen, and another part of the antibody binds the NK cells, such as an Fc region of the antibody. In cases wherein the antibody is multi specific, one or more antigen binding domains of the antibody binds the NK cell (such as through an NK cell surface antigen, either naturally occurring, or transgenic, e.g., CD3) and one or more antigen binding domains of the antibody binds one or more target antigens. The multispecific antibody may be bispecific, trispecific, or tetraspecific, for example. In cases wherein the antibody is trispecific or tetraspecific, the additional antigen binding domains may bind other cells, such as stem cells.

[0104] In particular embodiments, the antibodies may bind any NK cell surface antigen (that may or may not be receptors) on NK cells, such as CD 16 (including CD 16a or CD 16b), CD32, CD56, CD64, a c-type lectin such as NKG2D, NKG2C, a costimulatory molecule such302140458.1 - 21 -as CS1, DNAM, 2B4, CD2, an NCR, NKp30, NKp44, NKp46, or KIR, and redirect the NK cells to a target, thus increasing the response and specificity against different tumors. In certain embodiments, the antibodies may bind to a transgenic NK cell surface antigen, such as CD3.

[0105] In some embodiments, the antibodies may bind any suitable antigen (e.g., antigens described herein, such as those that are described as targets of CARs and / or TCRs). In particular embodiments, an antibody targets CD 19. In particular embodiments, an antibody targets CD20. In particular embodiments, an antibody targets CD123. In particular embodiments, an antibody targets EGFR. In particular embodiments, an antibody targets EGFR2. In particular embodiments, an antibody targets C-met (tyrosine-protein kinase MET, aka mesenchymal epithelial transition factor). In particular embodiments, an antibody targets EGFR and C-met. In particular embodiments, an antibody targets TROP-2 (tumor-associated calcium signal transducer 2).

[0106] In certain embodiments, generation of loaded NK cells may be by any suitable means, such that the conditions are sufficient for the appropriate region of the antibody to bind the appropriate surface region of the NK cell. Any particular medium may be utilized, in certain instances. In specific cases, Plasma-Lyte A and / or human serum albumin are utilized, wherein in other cases they are not. Once the complexes are formed in culture, they may or may not be washed prior to administration to the subject, such as through infusion. In alternative embodiments, the NK cells and the antibodies are administered separately, and the complexes form in vivo.V. NK Cell Expansion

[0107] In particular embodiments, NK cells are expanded to increase their quantity prior to administration to an individual in need thereof. The expanded cells may or may not be derived from pre-activated NK cells such that a pre-activation step may occur before an expansion step. The NK cell expansion step may be of any suitable such that the NK cell population is expanded, but in specific cases the expansion step utilizes particular one or more reagents, such as in culture, to enhance their expansion. In certain cases the NK cells may not be expanded. IL-2 or IL- 15 or IL- 18 or any combination of the cytokines may be added to the expansion culture before or during expansion. The NK cells can be expanded ex vivo in flasks or in one of several different bioreactor configurations with continuous perfusion of media / additives, in specific embodiments.302140458.1 - 22 -

[0108] In specific cases, the NK cells (whether pre-activated or not) may be washed (e.g., with PBS or Plasma Lyte or human serum albumin or culture media or combinations thereof) prior to and / or after expansion, such as 1, 2, 3, 4, or 5 times. In some embodiments, cells are washed specifically 3 times. In particular embodiments, the NK cells are expanded in the presence of artificial antigen presenting cells (aAPCs). In particular embodiments, the NK cells are expanded in the presence of fragments of aAPCs. The aAPCs may be engineered to express CD137 ligand and / or a membrane-bound cytokine. The membrane-bound cytokine may be membrane-bound IL-21 (mIL-21) or membrane-bound IL- 15 (mIL-15). In particular embodiments, the aAPCs are engineered to express CD137 ligand and mIL-21. The aAPCs may be derived from cancer cells, such as leukemia cells. The aAPCs may not express endogenous HLA class I, II, or CD Id molecules. They may express ICAM-1 (CD54) and LFA-3 (CD58) or CD48. In particular, the aAPCs may be K562 cells, such as K562 cells engineered to express CD137 ligand and mIL-21. The engineering may be by any method known in the art, such as retroviral transduction, although any viral or non-viral vector may be utilized. The aAPCs may or may not be irradiated. The expansion may be for a particular duration in time, such as for about 2-30 days, such as 3-20 days, particularly 12-16 days, such as 12, 13, 14, 15, 16, 17, 18, or 19 days, specifically about 14 days. The pre-activated NK cells and aAPCs may be present at a ratio ofabout 3: 1-1:3, such as 2:1, 1:1, 1:2, specifically about 1:2. The expansion culture may further comprise one or more cytokines to promote expansion, such as IL-2. The IL-2 may be present at a concentration of about 10-500 U / mL, such as 100-300 U / mL, particularly about 200 U / mL. The IL-2 may be replenished in the expansion culture, including at a certain frequency, such as every 2-3 days. The aAPCs may be added to the culture at least a second time, such as at about 7 days of expansion. Any cytokine(s) used in the pre-activation and / or expansion steps may be recombinant human cytokines.

[0109] In some embodiments, following expansion, the NK cells may be immediately utilized in any manner, such as complexed with one or more antibodies, or they may be stored, such as by cryopreservation. In certain aspects, the cells may be propagated for days, weeks, or months ex vivo as a bulk population within about 1, 2, 3, 4, or 5 days.

[0110] Activated and / or expanded NK cells can secrete type I cytokines, such as interferon-y, tumor necrosis factor-a and granulocyte-macrophage colony-stimulating factor (GM-CSF), which activate both innate and adaptive immune cells as well as other cytokines and chemokines. The measurement of these cytokines can be used to determine the activation status of NK cells. In addition, other methods known in the art for determination of NK cell activation may be used for characterization of the NK cells of the present disclosure.302140458.1 - 23 -[OHl] Thus, with respect to particular pre-activation and expansion aspects of the disclosure, in specific embodiments the NK cells pre-activated with any combination of IL-12, IL15, and / or IL-18 followed by expansion with aAPCs, such as K562 cells expressing mIL-21 and CD 137 ligand, provide a highly potent cellular product. Thus, methods are provided using the present NK cells for the treatment of various diseases, such as immunotherapy of patients with cancer. In an exemplary method, the isolated NK cells may be subjected to a brief period, such as about 16 hours, of pre-activation with a combination of cytokines, such as interleukin-12 (IL-12), IL-15, and / or IL-18, followed by expansion using artificial antigen presenting cells (aAPCs), such as K562 feeder cells expressing membrane-bound IL-21 and CD137 ligand, and / or exogenous IL-2. IL-2 or IL- 15 or IL- 18 or any combination of the cytokines may be added to the expansion culture at least a second time.VI. Heterologous Proteins and Mutations

[0112] In specific embodiments, the NK cells are engineered, such as to express one or more heterologous proteins. A heterologous protein may or may not be a synthetic protein, such as one not normally found in nature. The heterologous proteins may facilitate activity of the NK cells in any manner, including at least their activation, persistence, expansion, homing, and / or cytotoxicity.A. Monospecific, Bispecific or Multi-specific Antibodies

[0113] In some embodiments, the NK cells are modified to express one or more monospecific, bispecific or multi-specific antibodies, although in other cases the NK cells do not express the antibodies but the antibodies are utilized in conjunction with the NK cells.

[0114] In cases wherein the NK cells are engineered to express the antibodies, the antibodies may be engagers that bridge a particular immune effector cell with a particular target cell for destruction of the target cell. In specific embodiments, the engineered NK cells to be used with standard T-cell engagers (BiTEs) because they have been modified to express CD3 that in many cases is the T cell antigen to which the BiTE engager binds. In such cases, the BiTE used may also target a cancer or viral antigen that may be tailored to the medical condition of an intended recipient individual. For example, the BiTE may be tailored to bind a cancer antigen that is characteristic of the cancer cells of a cancer of the individual. The anti-CD3 antibody of the BiTE may target the CD3y chain, CD3b chain, CD3e chain, or CD3 chain.302140458.1 - 24 -

[0115] In some cases, the NK cells may express one or more of the CD3 chains. The NK cells may express the CD3 complex (with or without TCR) that allows the NK cells to be utilized as a therapy with BiTEs, the NK cells may be modified to express (or not to express but instead used in conjunction with) one or more bispecific NK engagers (BiKEs). The BiKE comprises an antibody that binds a surface protein on the NK cell, including a naturally expressed surface protein on NK cells (for example but not limited to, NKp30, NKp44, NKp46, CD 16, CD32, CD64, KIRs, and the like), and also comprises an antibody that binds a desired target antigen. The BiKE may target the NK cells through an antibody an NK surface protein such as CD16, CS1, CD32, CD64, CD56, NKG2D, NKG2C, DNAM, 2B4, CD2, an NCR, NKp30, NKp44, NKp46, or KIR, for example. In such cases, the BiKE used in the invention may also target a cancer or viral antigen that may be tailored to the medical condition of an intended recipient individual. For example, the BiKE may be tailored to bind a cancer antigen that is characteristic of the cancer cells of a cancer of the individual.

[0116] In certain embodiments, an antibody is Blinatumomab. In certain embodiments, an antibody is Tebentafusp. In certain embodiments, an antibody is Mosunetuzumab. In certain embodiments, an antibody is Teclistamab. In certain embodiments, an antibody is Glofitamab. In certain embodiments, an antibody is Epcoritamab. In some embodiments, an antibody is Flotetuzumab. In some embodiments, an antibody is APV0436. In some embodiments, an antibody is TNB383B. In certain embodiments of cases of use of multispecific antibodies, one or more antigen binding domains of the antibody can bind one or more target antigens. In certain embodiments, an antibody is Amivantamab. In certain embodiments, an antibody is Cetuximab. In certain embodiments, an antibody is Imgatuzumab.

[0117] In embodiments wherein an NK cell expresses the CD3 complex (with or without TCR) and one or more BiKEs, one or more vectors may be utilized to transfect or transduce the cells with the CD3 complex components (with or without TCR) and one or more BiKEs. In some cases, one or more of the CD3 complex components (with or without TCR) and the BiKE may or may not be on the same multi ci str onic vector.B. Engineered Receptors

[0118] In specific embodiments, the NK cells are engineered to express one or more engineered receptors. In some cases, the engineered receptors are engineered antigen receptors that target a cancer or viral antigen of any kind. The receptor may be tailored to target a desired antigen based on a medical condition of an intended recipient individual.302140458.1 - 25 -

[0119] In some embodiments, the engineered antigen receptor is a chimeric antigen receptor (CAR). The NK cells may be modified to encode at least one CAR, and the CAR may be first generation, second generation, or third or a subsequent generation, for example. The CAR may or may not be bispecific for two or more different antigens. The CAR may comprise one or more costimulatory domains. Each costimulatory domain may comprise the costimulatory domain of any one or more of, for example, members of the TNFR superfamily, CD28, CD137 (4-1BB), CD134 (0X40), DAP10, DAP12, CD27, CD2, CD5, ICAM-1, LFA-1 (CDlla / CD18), Lek, TNFR-I, TNFR-II, Fas, CD30, CD27, NKG2D, 2B4M, CD40 or combinations thereof, for example. In specific embodiments, the CAR comprises CD3zeta. In certain embodiments, the CAR lacks one or more specific costimulatory domains; for example, the CAR may lack 4-1BB and / or lack CD28.

[0120] In particular embodiments, the CAR polypeptide in the cells comprises an extracellular spacer domain that links the antigen binding domain and the transmembrane domain, and this may be referred to as a hinge. Extracellular spacer domains may include, but are not limited to, Fc fragments of antibodies or fragments or derivatives thereof, hinge regions of antibodies or fragments or derivatives thereof, CH2 regions of antibodies, CH3 regions antibodies, artificial spacer sequences or combinations thereof. Examples of extracellular spacer domains include but are not limited to CD8-alpha hinge, CD28, artificial spacers made of polypeptides such as Gly3, or CHI, CH3 domains of IgGs (such as human IgGl or IgG4). In specific cases, the extracellular spacer domain may comprise (i) a hinge, CH2 and CH3 regions of IgG4, (ii) a hinge region of IgG4, (iii) a hinge and CH2 of IgG4, (iv) a hinge region of CD8-alpha or CD4, (v) a hinge, CH2 and CH3 regions of IgGl, (vi) a hinge region of IgGl or (vii) a hinge and CH2 of IgGl, (viii) a hinge region of CD28, or a combination thereof. In specific embodiments, the hinge is from IgGl and in certain aspects the CAR polypeptide comprises a particular IgGl hinge amino acid sequence or is encoded by a particular IgGl hinge nucleic acid sequence.

[0121] The transmembrane domain in the CAR may be derived either from a natural or from a synthetic source. Where the source is natural, the domain in some aspects is derived from any membrane-bound or transmembrane protein. Transmembrane regions include those derived from ( / .<?., comprise at least the transmembrane region(s) of) the alpha, beta or zeta chain of the T- cell receptor, CD28, CD3 zeta, CD3 epsilon, CD3 gamma, CD3 delta, CD45, CD4, CD5, CD8, CD9, CD 16, CD22, CD33, CD37, CD64, CD80, CD86, CD 134, CD137, CD 154, ICOS / CD278, GITR / CD357, NKG2D, and DAP molecules, such as DAP 10 or DAP12. Alternatively the transmembrane domain in some embodiments is synthetic. In some302140458.1 - 26 -aspects, the synthetic transmembrane domain comprises predominantly hydrophobic residues such as leucine and valine. In some aspects, a triplet of phenylalanine, tryptophan and valine may be found at each end of a synthetic transmembrane domain.

[0122] In some embodiments, the engineered receptors utilize one or more homing receptors (that can home to a target not necessarily because of a signal release, such as in the event that they utilize adhesion molecules) and / or one or more chemokine receptors. Examples of chemokine receptors include CXC chemokine receptors, CC chemokine receptors, CX3C chemokine receptors and XC chemokine receptors. In specific cases, the chemokine receptor is a receptor for CCR2, CCR3, CCR5, CCR8, CCR7, CXCR3, L-selectin (CD62L) CXCR1, CXCR2, or CX3CR1.C. Cytokines

[0123] In some embodiments, the cells expressing the NK cells are engineered to express one or more heterologous cytokines and / or are engineered to upregulate normal expression of one or more heterologous cytokines. The cells may or may not be transduced or transfected for one or more cytokines on the same vector as other genes. In certain embodiments, NK cells may be modified to express one or more cytokines, cytokine receptors, chemokines, chemokine receptors, and / or suicide genes.

[0124] One or more cytokines may be co-expressed from a vector, including as a separate polypeptide from any component of the TCR / CD3 complex. Interleukin- 15 (IL- 15), for example, is tissue restricted and only under pathologic conditions is it observed at any level in the serum, or systemically. IL- 15 possesses several attributes that are desirable for adoptive therapy. IL-15 is a homeostatic cytokine that induces development and cell proliferation of natural killer cells, promotes the eradication of established tumors via alleviating functional suppression of tumor-resident cells, and inhibits activation-induced cell death (AICD). In addition to IL- 15, other cytokines are envisioned. These include, but are not limited to, cytokines, chemokines, and other molecules that contribute to the activation and proliferation of cells used for human application. NK cells expressing IL- 15 are capable of continued supportive cytokine signaling, which is useful for their survival post-infusion.

[0125] In some embodiments, the cells express one or more exogenously provided engineered receptors, wherein the engineered receptor comprises a chemokine receptor and / or a cytokine receptor. In some embodiments, a cytokine receptor is an IL- 15 receptor. In some embodiments, a cytokine receptor is a non-naturally occurring variant of a cytokine receptor.302140458.1 - 27 -In some embodiments, a cytokine receptor is an IL-15, IL-9, IL-7, IL-10, IL-27, IL-12, IL-2, IL-18, IL-21, IL-23, or GMCSF receptor, or a combination thereof.

[0126] In specific embodiments, the cells express one or more exogenously provided cytokines. As one example, the cytokine is IL-15, IL-10, IL-7, IL-9, IL-27, IL-12, IL-2, IL-18, IL-21, IL-23, GMCSF, or a combination thereof. The cytokine may be exogenously provided to the NK cells because it is expressed from an expression vector within the cell. In an alternative case, an endogenous cytokine in the cell is upregulated upon manipulation of regulation of expression of the endogenous cytokine, such as genetic recombination at the promoter site(s) of the cytokine. In cases wherein the cytokine is provided on an expression construct to the cell, the cytokine may be encoded from the same vector as one or more components of the CD3 complex with or without the TCR complex.D. Antigens and Antibodies

[0127] The NK cells may be utilized with monospecific, bispecific or multi-specific antibodies that target one or more particular antigens. In addition, the NK cells may be modified with engineered antigen receptors that target one or more particular antigens. In cases wherein the NK cells are modified with one or more engineered antigen receptors, the antigen targeted by the monospecific, bispecific or multi-specific antibody, and the antigen targeted by the one or more engineered antigen receptors may or may not be the same antigen. In some cases, the antigen targeted by the monospecific, bispecific or multi-specific antibody, and the antigen targeted by the one or more engineered antigen receptors are different antigens but are associated with the same type of cancer. In some embodiments, the antigen targeted by the monospecific, bispecific or multispecific antibody, and the antigen targeted by the one or more engineered antigen receptors are different antigens, but are each associated with solid tumors.

[0128] Among the antigens targeted by the antibodies and / or engineered antigen receptors are those expressed in the context of a disease, condition, or cell type to be targeted via the adoptive cell therapy. Among the diseases and conditions are proliferative, neoplastic, and malignant diseases and disorders, including cancers and tumors, including hematologic cancers, cancers of the immune system, such as lymphomas, leukemias, and / or myelomas, such as B, T, and myeloid leukemias, lymphomas, and multiple myelomas. In some embodiments, the antigen is selectively expressed or overexpressed on cells of the disease or condition, e.g., the tumor or pathogenic cells, as compared to normal or non-targeted cells or tissues. In other302140458.1 - 28 -embodiments, the antigen is expressed on normal cells and / or is expressed on the engineered cells.

[0129] Any suitable antigen may be targeted in the present method. The antigen may be associated with certain cancer cells but not associated with non-cancerous cells, in some cases. Exemplary antigens include, but are not limited to, antigenic molecules from infectious agents, auto- / self-antigens, tumor- / cancer-associated antigens, and tumor neoantigens (Linnemann et al., 2015). In particular aspects, the antigen includes KRAS, TROP-2, NY-ESO, CD19, EBNA, CD123, HER2, CA-125, TRAIL / DR4, CD20, CD22, CD70, CD38, CD123, CLL1, carcinoembryonic antigen, alphafetoprotein, CD56, AKT, Her3, epithelial tumor antigen, CD319 (CS1), ROR1, folate binding protein, HIV-1 envelope glycoprotein gpl20, HIV-1 envelope glycoprotein gp41, CD5, CD23, CD30, HERV-K, IL-1 IRalpha, kappa chain, lambda chain, CSPG4, CD33, CD47, CLL-1, U5snRNP200, CD200, BAFF-R, BCMA, CD99, p53, mutated p53, Ras, mutated ras, c-Myc, cytoplasmic serine / threonine kinases (e.g., A-Raf, B-Raf, and C-Raf, cyclin-dependent kinases), MAGE-A1, MAGE-A2, MAGE-A3, MAGE-A4, MAGE-A6, MAGE-A10, MAGE-A12, MART-1, melanoma-associated antigen, BAGE, DAM-6, -10, GAGE-1, -2, -8, GAGE-3, -4, -5, -6, -7B, NA88-A, MC1R, mda-7, gp75, GplOO, PSA, PSM, Tyrosinase, tyrosinase-related protein, TRP-1, TRP-2, ART -4, CAMEL, CEA, Cyp-B, hTERT, hTRT, iCE, MUC1, MUC2, Phosphoinositide 3 -kinases (PI3Ks), TRK receptors, PRAME, P15, RU1, RU2, SART-1, SART-3, Wilms' tumor antigen (WT1), AFP, -catenin / m, Caspase-8 / m, CDK-4 / m, ELF2M, GnT-V, G250, HAGE, HSP70-2M, HST-2, KIAA0205, MUM-1, MUM-2, MUM-3, Myosin / m, RAGE, SART-2, TRP-2 / INT2, 707-AP, Annexin II, CDC27 / m, TPI / mbcr-abl, BCR-ABL, interferon regulatory factor 4 (IRF4), ETV6 / AML, LDLR / FUT, Pml / RAR, Tumor-associated calcium signal transducer 1 (TACSTD1) TACSTD2, receptor tyrosine kinases (e.g., Epidermal Growth Factor receptor (EGFR) (in particular embodiments, EGFRvIII), platelet derived growth factor receptor (PDGFR), vascular endothelial growth factor receptor (VEGFR)), VEGFR2, cytoplasmic tyrosine kinases (e.g., src-family, syk-ZAP70 family), integrin-linked kinase (ILK), signal transducers and activators of transcription STAT3, STATS, and STATE, hypoxia inducible factors (e.g., HIF-1 and HIF-2), Nuclear Factor-Kappa B (NF-B), Notch receptors (e.g., Notchl-4), NY ESO 1, c-Met, mammalian targets of rapamycin (mTOR), WNT, extracellular signal-regulated kinases (ERKs), and their regulatory subunits, PMSA, PR-3, MDM2, Mesothelin, renal cell carcinoma-5T4, SM22-alpha, carbonic anhydrases I (CAI) and IX (CAIX) (also known as G250), STEAD, TEL / AML1, GD2, proteinase3, hTERT, sarcoma translocation breakpoints, EphA2, ML-IAP, EpCAM, ERG (TMPRSS2 ETS fusion gene),302140458.1 - 29 -NA17, PAX3, ALK, androgen receptor, cyclin Bl, polysialic acid, MYCN, RhoC, GD3, fucosyl GM1, mesothelian, PSCA, sLe, PLAC1, GM3, BORIS, Tn, GLoboH, NY-BR-1, RGsS, SAGE, SART3, STn, PAX5, OY-TES1, sperm protein 17, LCK, HMWMAA, AKAP-4, SSX2, XAGE 1, B7H3, legumain, TIE2, Page , MAD-CT-1, FAP, MAD-CT-2, fos related antigen 1, CBX2, CLDN6, SPANX, TPTE, ACTL8, ANKRD30A, CDKN2A, MAD2L1, CTAG1B, SUNCI, and LRRN1. Examples of sequences for antigens are known in the art, for example, in the GENBANK® database: KRAS (Accession No. NC_000012.12), CD19 (Accession No. NG_007275.1), EBNA (Accession No. NG_002392.2), WT1 (Accession No. NG_009272.1), CD123 (Accession No. NC_000023.11), NY-ESO (Accession No. NC_000023.11), EGFRvIII (Accession No. NG_007726.3), MUC1 (Accession No. NG_029383.1), HER2 (Accession No. NG_007503.1), CA-125 (Accession No. NG_055257.1), WT1 (Accession No. NG_009272.1), Mage-A3 (Accession No. NG_013244.1), Mage-A4 (Accession No. NG_013245.1), Mage-AlO (Accession No. NC_000023.11), TRAIL / DR4 (Accession No. NC_000003.12), and / or CEA (Accession No. NC_000019.10).

[0130] Tumor-associated antigens may be derived from prostate, breast, colorectal, lung, pancreatic, renal, mesothelioma, ovarian, liver, brain, bone, stomach, spleen, testicular, cervical, anal, gall bladder, thyroid, or melanoma cancers, as examples. Exemplary tumor-associated antigens or tumor cell-derived antigens include MAGE 1, 3, and MAGE 4 (or other MAGE antigens such as those disclosed in International Patent Publication No. WO 99 / 40188); PRAME; BAGE; RAGE, Lage (also known as NY ESO 1); KRAS (such as those disclosed in US patent 10,611,816); SAGE; and HAGE or GAGE. These non-limiting examples of tumor antigens are expressed in a wide range of tumor types such as melanoma, lung carcinoma, sarcoma, and bladder carcinoma. See, e.g., U.S. Patent No. 6,544,518. Prostate cancer tumor-associated antigens include, for example, prostate specific membrane antigen (PSMA), prostate-specific antigen (PSA), prostatic acid phosphates, NKX3.1, and six-transmembrane epithelial antigen of the prostate (STEAP).

[0131] Other tumor associated antigens include Plu-1, HASH-1, HasH-2, Cripto and Criptin. Additionally, a tumor antigen may be a self-peptide hormone, such as whole length gonadotrophin hormone releasing hormone (GnRH), a short 10 amino acid long peptide, useful in the treatment of many cancers.

[0132] Antigens may include epitopic regions or epitopic peptides derived from genes mutated in tumor cells or from genes transcribed at different levels in tumor cells compared to normal cells, such as telomerase enzyme, survivin, mesothelin, mutated ras, mutated KRAS,302140458.1 - 30 -bcr / abl rearrangement, Her2 / neu, mutated or wild-type p53, cytochrome P450 1B1, and abnormally expressed intron sequences such as N-acetylglucosaminyltransf erase- V; clonal rearrangements of immunoglobulin genes generating unique idiotypes in myeloma and B-cell lymphomas; tumor antigens that include epitopic regions or epitopic peptides derived from oncoviral processes, such as human papilloma virus proteins E6 and E7; Epstein bar virus protein LMP2; nonmutated oncofetal proteins with a tumor- selective expression, such as carcinoembryonic antigen and alpha-fetoprotein.E. Suicide Gene

[0133] In particular embodiments, a suicide gene is utilized in conjunction with the NK cell therapy to control its use and allow for termination of the cell therapy at a desired event and / or time. The suicide gene is employed in transduced cells for the purpose of eliciting death for the transduced cells when needed. The cells of the present disclosure that have been modified to harbor one or more vectors encompassed by the disclosure that may comprise one or more suicide genes. In some embodiments, the term “suicide gene” as used herein is defined as a gene which, upon administration of a prodrug or other agent, effects transition of a gene product to a compound which kills its host cell. In other embodiments, a suicide gene encodes a gene product that is, when desired, targeted by an agent (such as an antibody) that targets the suicide gene product.

[0134] In some cases, the cell therapy may be subject to utilization of one or more suicide genes of any kind when an individual receiving the cell therapy and / or having received the cell therapy shows one or more symptoms of one or more adverse events, such as cytokine release syndrome, neurotoxicity, anaphylaxis / allergy, and / or on-target / off tumor toxicities (as examples) or is considered at risk for having the one or more symptoms, including imminently. The use of the suicide gene may be part of a planned protocol for a therapy or may be used only upon a recognized need for its use. In some cases the cell therapy is terminated by use of agent(s) that targets the suicide gene or a gene product therefrom because the therapy is no longer required.

[0135] Utilization of the suicide gene may be instigated upon onset of at least one adverse event for the individual, and that adverse event may be recognized by any means, including upon routine monitoring that may or may not be continuous from the beginning of the cell therapy. The adverse event(s) may be detected upon examination and / or testing. In cases wherein the individual has cytokine release syndrome (which may also be referred to as302140458.1 - 31 -cytokine storm), the individual may have elevated inflammatory cytokine(s) (merely as examples: interferon-gamma, granulocyte macrophage colony-stimulating factor, IL- 10, IL-6 and TNF-alpha); fever; fatigue; hypotension; hypoxia, tachycardia; nausea; capillary leak; cardiac / renal / hepatic dysfunction; or a combination thereof, for example. In cases wherein the individual has neurotoxicity, the individual may have confusion, delirium, aplasia, and / or seizures. In some cases, the individual is tested for a marker associated with onset and / or severity of cytokine release syndrome, such as C-reactive protein, IL-6, TNF-alpha, and / or ferritin.

[0136] Examples of suicide genes include engineered nonsecretable (including membrane bound) tumor necrosis factor (TNF)-alpha mutant polypeptides (see PCT / US19 / 62009, which is incorporated by reference herein in its entirety), and they may be affected by delivery of an antibody that binds the TNF-alpha mutant. Examples of suicide gene / prodrug combinations that may be used are Herpes Simplex Virus-thymidine kinase (HSV-tk) and ganciclovir, acyclovir, or FIAU; oxidoreductase and cycloheximide; cytosine deaminase and 5-fluorocytosine; thymidine kinase thymidylate kinase (Tdk::Tmk) and AZT; and deoxycytidine kinase and cytosine arabinoside. The E.coli purine nucleoside phosphorylase, a so-called suicide gene that converts the prodrug 6-methylpurine deoxyriboside to toxic purine 6-methylpurine, may be utilized. Other suicide genes include CD20, CD52, inducible caspase 9, purine nucleoside phosphorylase (PNP), Cytochrome p450 enzymes (CYP), Carboxypeptidases (CP), Carboxylesterase (CE), Nitroreductase (NTR), Guanine Ribosyltransferase (XGRTP), Glycosidase enzymes, Methionine-a,y-lyase (MET), EGFRv3, and Thymidine phosphorylase (TP), as examples.F. Knockout or Knockdown of Endogenous Genes

[0137] In certain embodiments, NK cells of the disclosure may include gene editing of the NK cells to remove 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more endogenous genes in the NK cells. In some cases the gene editing occurs in NK cells expressing one or more heterologous transgenes, whereas in other cases the gene editing occurs in NK cells that do not express a heterologous transgene but that ultimately will express one or more heterologous transgenes, in at least some cases. In particular embodiments, the NK cells that are gene edited are expanded NK cells.

[0138] In particular cases, one or more endogenous genes of the NK cells are modified, such as disrupted in expression where the expression is reduced in part or in full. In specific302140458.1 - 32 -cases, one or more genes are knocked down or knocked out using processes of the disclosure. In specific cases, multiple genes are knocked down or knocked out in the same step as processes of the disclosure. The genes that are edited in the NK cells may be of any kind, but in specific embodiments the genes are genes whose gene products inhibit activity and / or proliferation of NK cells. In specific cases the genes that are edited in the NK cells allow the NK cells to work more effectively in a tumor microenvironment. In specific cases, the genes are one or more of NKG2A, SIGLEC-7, LAG3, TIM3, CISH, FOXO1, TGFBR2, TIGIT, CD96, ADORA2, NR3C1, PD1, PDL-1, PDL-2, CD47, SIRPA, SHIP1, ADAM17, RPS6, 4EBP1, CD25, CD40, IL21R, ICAM1, CD95, CD80, CD86, IL10R, TDAG8, CD5, CD7, SLAMF7, CD38, LAG3, TCR, beta2-microglobulin, HLA, CD73, CREB, CREM, ICER, and CD39. In specific embodiments, the TGFBR2 gene is knocked out or knocked down in the NK cells. In specific embodiments, the CISH gene is knocked out or knocked down in the NK cells. In specific embodiments, the CD38 gene is knocked out or knocked down in the NK cells.

[0139] In some embodiments, the gene editing is carried out using one or more DNA-binding nucleic acids, such as alteration via an RNA-guided endonuclease (RGEN). For example, the alteration can be carried out using clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated (Cas) proteins. In general, "CRISPR system" refers collectively to transcripts and other elements involved in the expression of or directing the activity of CRISPR-associated ("Cas") genes, including sequences encoding a Cas gene, a tracr (trans-activating CRISPR) sequence (e.g., tracrRNA or an active partial tracrRNA), a tracr-mate sequence (encompassing a "direct repeat" and a tracrRNA-processed partial direct repeat in the context of an endogenous CRISPR system), a guide sequence (also referred to as a "spacer" in the context of an endogenous CRISPR system), and / or other sequences and transcripts from a CRISPR locus. Methods of utilizing a CRISPR system are well known in the artVII. Administration of Therapeutic Compositions

[0140] The NK cells and the one or more inhibitors of the KRAS pathway and / or the one or more PARP inhibitors are administered to an individual in need thereof. In some cases, the two or three components are administered separately to an individual, whereas in other cases the two or three components are combined together prior to administration, such as in an ex vivo manner. In some cases, the two components are not combined prior to administration, but302140458.1 - 33 -are co-administered by any suitable route of administration, such as by co-infusion to the patient.

[0141] Embodiments of the present disclosure concern methods for the use of the compositions comprising NK cells and the one or more inhibitors of the KRAS pathway and / or the one or more PARP inhibitors are provided herein for treating or preventing a medical disease or disorder. The method includes administering to the subject a therapeutically effective amount of the NK cells with the the one or more inhibitors of the KRAS pathway and / or the one or more PARP inhibitors, thereby treating or preventing the disease in the subject, including reducing the risk of, reducing the severity of (including the severity of one or more symptoms), and / or delaying the onset of the disease. In certain embodiments of the present disclosure, cancer or autoimmune disease is treated by transfer of a composition comprising the NK cell population and corresponding one or more inhibitors of the KRAS pathway and / or the one or more PARP inhibitors. In at least some cases, because of their release of pro-inflammatory cytokines, NK cells may reverse the anti-inflammatory tumor microenvironment and increase adaptive immune responses by promoting differentiation, activation, and / or recruitment of accessory immune cell to sites of malignancy. In certain embodiments, a providing step may comprise culturing the NK cells with the one or more inhibitors of the KRAS pathway and / or the one or more PARP inhibitors for a specific duration of time (e.g., about 5 minutes to about 24 hours or more) and storing the NK cells and the one or more inhibitors of the KRAS pathway and / or the one or more PARP inhibitors for a period of time (e.g., about 1, 2, 3, 4, 5 days, or greater than 5 days) prior to infusion / administration.

[0142] Cancers for which the present treatment methods are useful include any malignant cell type, such as those found in a solid tumor or a hematological tumor. Exemplary solid tumors can include, but are not limited to, a tumor of an organ selected from the group consisting of pancreas, colon, cecum, stomach, brain, head, neck, ovary, kidney, larynx, sarcoma, lung, bladder, melanoma, prostate, and breast. Exemplary hematological tumors include tumors of the bone marrow, T or B cell malignancies, leukemias, lymphomas, blastomas, myelomas, and the like. Further examples of cancers that may be treated using the methods provided herein include, but are not limited to, lung cancer (including small-cell lung cancer, non-small cell lung cancer, adenocarcinoma of the lung, and squamous carcinoma of the lung), cancer of the peritoneum, gastric or stomach cancer (including gastrointestinal cancer and gastrointestinal stromal cancer), pancreatic cancer, cervical cancer, ovarian cancer, liver cancer, bladder cancer, breast cancer, colon cancer, colorectal cancer, endometrial or uterine carcinoma, salivary gland carcinoma, kidney or renal cancer, prostate cancer, vulval cancer,302140458.1 - 34 -thyroid cancer, various types of head and neck cancer, and melanoma. In certain embodiments, the cancer is a solid tumor cancer. In certain embodiments, the cancer is pancreatic cancer. In certain embodiments, the cancer is ovarian cancer. In certain embodiments, the cancer is lung cancer. In certain embodiments, the cancer is colorectal cancer. In certain embodiments, the cancer is a cancer associated with KRAS positive tumors.

[0143] The cancer may specifically be of the following histological type, though it is not limited to these: neoplasm, malignant; carcinoma; carcinoma, undifferentiated; giant and spindle cell carcinoma; small cell carcinoma; papillary carcinoma; squamous cell carcinoma; lymphoepithelial carcinoma; basal cell carcinoma; pilomatrix carcinoma; transitional cell carcinoma; papillary transitional cell carcinoma; adenocarcinoma; gastrinoma, malignant; cholangiocarcinoma; hepatocellular carcinoma; combined hepatocellular carcinoma and cholangiocarcinoma; trabecular adenocarcinoma; adenoid cystic carcinoma; adenocarcinoma in adenomatous polyp; adenocarcinoma, familial polyposis coli; solid carcinoma; carcinoid tumor, malignant; branchiolo-alveolar adenocarcinoma; papillary adenocarcinoma; chromophobe carcinoma; acidophil carcinoma; oxyphilic adenocarcinoma; basophil carcinoma; clear cell adenocarcinoma; granular cell carcinoma; follicular adenocarcinoma; papillary and follicular adenocarcinoma; nonencapsulating sclerosing carcinoma; adrenal cortical carcinoma; endometroid carcinoma; skin appendage carcinoma; apocrine adenocarcinoma; sebaceous adenocarcinoma; ceruminous adenocarcinoma; mucoepidermoid carcinoma; cystadenocarcinoma; papillary cystadenocarcinoma; papillary serous cystadenocarcinoma; mucinous cystadenocarcinoma; mucinous adenocarcinoma; signet ring cell carcinoma; infiltrating duct carcinoma; medullary carcinoma; lobular carcinoma; inflammatory carcinoma; paget's disease, mammary; acinar cell carcinoma; adenosquamous carcinoma; adenocarcinoma w / squamous metaplasia; thymoma, malignant; ovarian stromal tumor, malignant; thecoma, malignant; granulosa cell tumor, malignant; androblastoma, malignant; sertoli cell carcinoma; leydig cell tumor, malignant; lipid cell tumor, malignant; paraganglioma, malignant; extra-mammary paraganglioma, malignant; pheochromocytoma; glomangiosarcoma; malignant melanoma; amelanotic melanoma; superficial spreading melanoma; lentigo malignant melanoma; acral lentiginous melanomas; nodular melanomas; malignant melanoma in giant pigmented nevus; epithelioid cell melanoma; blue nevus, malignant; sarcoma; fibrosarcoma; fibrous histiocytoma, malignant; myxosarcoma; liposarcoma; leiomyosarcoma; rhabdomyosarcoma; embryonal rhabdomyosarcoma; alveolar rhabdomyosarcoma; stromal sarcoma; mixed tumor, malignant; mullerian mixed tumor; nephroblastoma; hepatoblastoma; carcinosarcoma; mesenchymoma, malignant; brenner tumor,302140458.1 - 35 -malignant; phyllodes tumor, malignant; synovial sarcoma; mesothelioma, malignant; dysgerminoma; embryonal carcinoma; teratoma, malignant; struma ovarii, malignant; choriocarcinoma; mesonephroma, malignant; hemangiosarcoma; hemangioendothelioma, malignant; kaposi's sarcoma; hemangiopericytoma, malignant; lymphangiosarcoma; osteosarcoma; juxtacortical osteosarcoma; chondrosarcoma; chondroblastoma, malignant; mesenchymal chondrosarcoma; giant cell tumor of bone; ewing's sarcoma; odontogenic tumor, malignant; ameloblastic odontosarcoma; ameloblastoma, malignant; ameloblastic fibrosarcoma; pinealoma, malignant; chordoma; glioma, malignant; ependymoma; astrocytoma; protoplasmic astrocytoma; fibrillary astrocytoma; astroblastoma; glioblastoma; oligodendroglioma; oligodendroblastoma; primitive neuroectodermal; cerebellar sarcoma; ganglioneuroblastoma; neuroblastoma; retinoblastoma; olfactory neurogenic tumor; meningioma, malignant; neurofibrosarcoma; neurilemmoma, malignant; granular cell tumor, malignant; malignant lymphoma; hodgkin's disease; hodgkin's; paragranuloma; malignant lymphoma, small lymphocytic; malignant lymphoma, large cell, diffuse; malignant lymphoma, follicular; mycosis fungoides; other specified non-hodgkin's lymphomas; B-cell lymphoma; low grade / follicular non-Hodgkin's lymphoma (NHL); small lymphocytic (SL) NHL; intermediate grade / follicular NHL; intermediate grade diffuse NHL; high grade immunoblastic NHL; high grade lymphoblastic NHL; high grade small non-cleaved cell NHL; bulky disease NHL; mantle cell lymphoma; AIDS-related lymphoma; Waldenstrom's macroglobulinemia; malignant histiocytosis; multiple myeloma; mast cell sarcoma; immunoproliferative small intestinal disease; leukemia; lymphoid leukemia; plasma cell leukemia; erythroleukemia; lymphosarcoma cell leukemia; myeloid leukemia; basophilic leukemia; eosinophilic leukemia; monocytic leukemia; mast cell leukemia; megakaryoblastic leukemia; myeloid sarcoma; hairy cell leukemia; chronic lymphocytic leukemia (CLL); acute lymphoblastic leukemia (ALL); acute myeloid leukemia (AML); and chronic myeloblastic leukemia.

[0144] In some embodiments, the medical condition treated or prevented (or delayed in onset and / or reduced in severity of one or more symptoms) with methods and compositions encompassed herein is an autoimmune disorder, such as at least Systemic Lupus Erythematosus, Rheumatoid Arthritis, Systemic Sclerosis, Antiphospholipid Syndrome, Type 1 Diabetes Mellitus, Multiple Sclerosis, Myasthenia Gravis, Autoimmune Hemolytic Anemia, Immune Thrombocytopenia, Primary Biliary Cholangitis, Autoimmune Hepatitis, Crohn’s Disease, Ulcerative Colitis, Sjogren’s Syndrome, Hashimoto’s Thyroiditis, Graves’ Disease, Pemphigus Vulgaris, Bullous Pemphigoid, Dermatomyositis, Polymyositis, Ankylosing Spondylitis, Psoriatic Arthritis, Autoimmune Encephalitis, Guillain-Barre Syndrome, Chronic302140458.1 - 36 -Inflammatory Demyelinating Polyneuropathy, Vasculitis (e.g., ANCA-associated vasculitis), Autoimmune Uveitis, Addison’s Disease, Celiac Disease, and / or Behget’s Disease.

[0145] The therapy provided herein may comprise administration of a combination of therapeutic agents, such as a first cancer therapy and a second cancer therapy. The first cancer therapy may be the NK cells and the one or more inhibitors of the KRAS pathway and / or the one or more PARP inhibitors. The therapies may be administered in any suitable manner known in the art. For example, the first and second cancer treatment may be administered sequentially (at different times) or concurrently (at the same time). In some embodiments, the first and second cancer treatments are administered in a separate composition. In some embodiments, the first and second cancer treatments are in the same composition. Embodiments of the disclosure relate to compositions and methods comprising therapeutic compositions. The different therapies may be administered in one composition or in more than one composition, such as 2 compositions, 3 compositions, or 4 compositions. Various combinations of the agents may be employed. Examples of therapies other than those of the present disclosure include surgery, chemotherapy, drug therapy, radiation, hormone therapy, immunotherapy (other than that of the present disclosure), or a combination thereof.

[0146] The therapeutic agents of the disclosure may be administered by the same route of administration or by different routes of administration. In some embodiments, the cancer therapy is administered intravenously, intramuscularly, subcutaneously, topically, orally, transdermally, intraperitoneally, intraorbitally, by implantation, by inhalation, intrathecally, intraventricularly, or intranasally. In some embodiments, the antibiotic is administered intravenously, intramuscularly, subcutaneously, topically, orally, transdermally, intraperitoneally, intraorbitally, by implantation, by inhalation, intrathecally, intraventricularly, or intranasally. The appropriate dosage may be determined based on the type of disease to be treated, severity and course of the disease, the clinical condition of the individual, the individual's clinical history and response to the treatment, and the discretion of the attending physician.

[0147] The treatments may include various “unit doses.” Unit dose is defined as containing a predetermined-quantity of the therapeutic composition. The quantity to be administered, and the particular route and formulation, is within the skill of determination of those in the clinical arts. A unit dose need not be administered as a single injection but may comprise continuous infusion over a set period of time. In some embodiments, a unit dose comprises a single administrable dose.302140458.1 - 37 -

[0148] The quantity to be administered, both according to number of treatments and unit dose, depends on the treatment effect desired. An effective dose is understood to refer to an amount necessary to achieve a particular effect. In the practice in certain embodiments, it is contemplated that doses in the range from 10 mg / kg to 200 mg / kg can affect the protective capability of these agents. Thus, it is contemplated that doses include doses of about 0.1, 0.5, 1, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 100, 105, 110, 115, 120, 125, 130, 135, 140, 145, 150, 155, 160, 165, 170, 175, 180, 185, 190, 195, and 200, 300, 400, 500, 1000 pg / kg, mg / kg, pg / day, or mg / day or any range derivable therein. Furthermore, such doses can be administered at multiple times during a day, and / or on multiple days, weeks, or months.

[0149] In certain embodiments, the effective dose of the pharmaceutical composition is one which can provide a blood level of about 1 pM to 150 pM. In another embodiment, the effective dose provides a blood level of about 4 pM to 100 pM.; or about 1 pM to 100 pM; or about 1 pM to 50 pM; or about 1 pM to 40 pM; or about 1 pM to 30 pM; or about 1 pM to 20 pM; or about 1 pM to 10 pM; or about 10 pM to 150 pM; or about 10 pM to 100 pM; or about 10 pM to 50 pM; or about 25 pM to 150 pM; or about 25 pM to 100 pM; or about 25 pM to 50 pM; or about 50 pM to 150 pM; or about 50 pM to 100 pM (or any range derivable therein). In other embodiments, the dose can provide the following blood level of the agent that results from a therapeutic agent being administered to a subject: about, at least about, or at most about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100 pM or any range derivable therein. In certain embodiments, the therapeutic agent that is administered to a subject is metabolized in the body to a metabolized therapeutic agent, in which case the blood levels may refer to the amount of that agent. Alternatively, to the extent the therapeutic agent is not metabolized by a subject, the blood levels discussed herein may refer to the unmetabolized therapeutic agent.

[0150] Precise amounts of the therapeutic composition also depend on the judgment of the practitioner and are peculiar to each individual. Factors affecting dose include physical and clinical state of the patient, the route of administration, the intended goal of treatment (alleviation of symptoms versus cure) and the potency, stability and toxicity of the particular therapeutic substance or other therapies a subject may be undergoing.302140458.1 - 38 -

[0151] It will be understood by those skilled in the art and made aware that dosage units of pg / kg or mg / kg of body weight can be converted and expressed in comparable concentration units of pg / ml or mM (blood levels), such as 4 pM to 100 pM. It is also understood that uptake is species and organ / tissue dependent. The applicable conversion factors and physiological assumptions to be made concerning uptake and concentration measurement are well-known and would permit those of skill in the art to convert one concentration measurement to another and make reasonable comparisons and conclusions regarding the doses, efficacies and results described herein.VIII. Kits

[0152] Certain aspects of the present disclosure also concern kits comprising compositions of the disclosure or compositions to implement methods of the disclosure. In particular embodiments, the kit comprises NK cells, fresh or frozen, and that may or may not have been pre-activated or expanded. The NK cells may or may not already express one or more heterologous proteins, may or may not have the expression of one or more endogenous genes disrupted, or both. In cases wherein the NK cells do not already express one or more heterologous components, the kit may comprise reagents for corresponding transfection or transduction of the NK cells, including reagents such as vectors that express the component(s), primers for amplification of the component(s), and so forth. In some cases, the NK cells may or may not also express one or more heterologous proteins as defined herein, and when they do not, the kit may comprise vectors that express the heterologous protein(s), primers for amplification of the heterologous protein(s), and so forth.

[0153] Kits may comprise components which may be individually packaged or placed in a container, such as a tube, bottle, vial, syringe, or other suitable container means. Individual components may also be provided in a kit in concentrated amounts; in some embodiments, a component is provided individually in the same concentration as it would be in a solution with other components. Concentrations of components may be provided as lx, 2x, 5x, lOx, or 20x or more.IX. Examples

[0154] The following examples are included to demonstrate preferred embodiments of the invention. It should be appreciated by those of skill in the art that the techniques disclosed in the examples which follow represent techniques discovered by the inventor to function well in302140458.1 - 39 -the practice of the invention, and thus can be considered to constitute preferred modes for its practice. However, those of skill in the art should, in light of the present disclosure, appreciate that many changes can be made in the specific embodiments which are disclosed and still obtain a like or similar result without departing from the spirit and scope of the invention.EXAMPLE 1COMBINING NK CELLS OR CAR-NK CELLS WITH AGENT(S) TO IMPROVE ANTITUMOR ACTIVITY

[0155] The outcome of combining NK cells or CAR-NK cells with one or more other agents was characterized by measuring for improvement in antitumor activity.

[0156] FIG. 1 shows that combining NK cells with a KRAS inhibitor (RMC-6236) enhanced the antitumor activity of NK cells. PANCI cells were treated with RMC-6236 (1 pM) for 24 hours, followed by treatment of the spheroids with NK cells or leaving them untreated. In FIG. 1, representative images of 3D tumor spheroids treated with NK cells, KRAS inhibitor, or both combinations are shown. The spheroids were cultured for 7 days with NK cells and / or the KRAS inhibitor. Both spheroid viability and size significantly decreased with the combination treatment compared to treatment with either agent alone.

[0157] Combining NK cells with the KRAS inhibitor Sotorasib enhanced the antitumor activity of NK cells (FIG. 2). PANCI cells were treated with Sotorasib (1 pM) for 24 hours, followed by treatment of the spheroids with NK cells or leaving them untreated. Representative images are shown in FIG. 2 of 3D tumor spheroids treated with NK cells, KRAS inhibitor, or both combinations. The spheroids were cultured for 7 days with NK cells and / or KRAS inhibitor. Spheroid viability and size significantly decreased with the combination treatment compared to treatment with either agent alone.

[0158] In FIG. 3, it is shown that combining NK cells with Sotorasib or RMC-6236 enhanced the antitumor activity of NK cells. HCT-116 cells were treated with Sotorasib or RMC-6236 (1 pM) for 24 hours, followed by treatment of the spheroids with NK cells or leaving them untreated. Representative images of 3D tumor spheroids treated with NK cells, KRAS inhibitor, or both combinations are shown. The spheroids were cultured for 7 days with NK cells and / or KRAS inhibitor. Spheroid viability and size significantly decreased with the combination treatment compared to treatment with either agent alone.302140458.1 - 40 -

[0159] Combining NK cells with a KRAS (Sotorasib) inhibitor enhanced the antitumor activity of NK cells (FIG. 4). An xCELLigence assay demonstrated the enhanced antitumor activity of NK cells combined with Sotorasib in HCT-116 (colorectal cancer) cells. HCT-116 (CRC) cells were treated with Sotorasib (1 pM) for 24 hours, followed by treatment with NK cells or leaving the cells untreated. SDS (sodium dodecyl sulfate) was used as a full-lysis positive control. Real-time impedance data from the xCELLigence system illustrated the effects of NK cells, KRAS inhibitor, or their combination on cell growth and viability. Images showed a significant reduction in cell viability with the combination treatment compared to single-agent treatments.

[0160] In FIG. 5, combining CAR-NK cells with Sotorasib enhanced the antitumor activity of NK cells. The xCELLigence assay demonstrated the enhanced antitumor activity of NK cells combined with Sotorasib in HCT-116 (CRC) cells. HCT-116 (CRC) cells were treated with Sotorasib (1 pM) for 24 hours, followed by treatment with CAR-NK cells or leaving the cells untreated. SDS was used as a full-lysis positive control. Real-time impedance data from the xCELLigence system illustrated the effects of TROP2 / CARNK cells, KRAS inhibitor, or their combination on cell growth and viability. The images showed a significant reduction in cell viability with the combination treatment compared to single-agent treatments.

[0161] Combining NK cells with a KRAS(RMC-6236) inhibitor enhanced the antitumor activity of NK cells (FIG. 6). An xCELLigence assay demonstrated the enhanced antitumor activity of NK cells combined with RMC-6236 in HCT-116 (CRC) cells. HCT-116 (CRC) cells were treated with RMC-6236 (1 pM) for 24 hours, followed by treatment with NK cells or leaving the cells untreated. SDS was used as a full-lysis positive control. Representative realtime impedance data from the xCELLigence system illustrated the effects of NK cells, KRAS inhibitor, or their combination on cell growth and viability. The images showed a significant reduction in cell viability with the combination treatment compared to single-agent treatments.

[0162] In FIG. 7, it is shown that combining CAR-NK cells with a KRAS (RMC-6236) inhibitor enhanced the antitumor activity of NK cells. xCELLigence assay demonstrated the enhanced antitumor activity of NK cells combined with RMC-6236 in HCT-116 (CRC) cells. HCT-116 (CRC) cells were treated with RMC6236 (1 pM) for 24 hours, followed by treatment with TROP2 / CARNK cells or leaving the cells untreated. SDS was used as a full-lysis positive control. Representative real-time impedance data from the xCELLigence system illustrated the effects of TROP2 / CAR NK cells, KRAS inhibitor, or their combination on cell growth and302140458.1 - 41 -viability. Images showed a significant reduction in cell viability with the combination treatment compared to single-agent treatments.

[0163] Shown in FIG. 8, combining NT or CAR-NK cells with RMC-6236 enhanced the antitumor activity of NK cells. An xCELLigence assay demonstrated the enhanced antitumor activity of NK cells treated with RMC-6236 in MIA-Paca-2 (PDAC) cells. MIA-Paca-2 (pancreatic ductal adenocarcinoma, PDAC) cells were treated with RMC6236 (1 pM) for 24 hours, followed by treatment with TROP2 / CAR NK cells or leaving the cells untreated. SDS was used as a full-lysis positive control. Representative real-time impedance data from the xCELLigence system illustrated the effects of both NT NK cells and TROP2 / CAR NK cells, KRAS inhibitor, or their combination on cell growth and viability. Images showed a significant reduction in cell viability with the combination treatment compared to single-agent treatments.

[0164] Combining NT or CAR-NK cells with RMC-6236 enhanced the antitumor activity of NK cells (FIG. 9). xCELLigence assay demonstrated the enhanced antitumor activity of NK cells combined with RMC-6236 in Hep-G2(HCC) cells. Hep-G2(HCC) cells were treated with RMC6236 (1 pM) for 24 hours, followed by treatment with TROP2 / CAR NK cells or leaving the cells untreated. SDS was used as a full-lysis positive control. Representative real-time impedance data from the xCELLigence system illustrated the effects of both NT NK cells and TROP2 / CAR NK cells, KRAS inhibitor, or their combination on cell growth and viability. Images showed a significant reduction in cell viability with the combination treatment compared to single-agent treatments. (FIG. 9).

[0165] Shown in FIGS. 10A and 10B, combining CAR-NK cells with a poly ADP-ribose polymerase (PARP) inhibitor (PARPi) (Olaparib) enhanced the antitumor activity of NK cells. An xCELLigence assay demonstrated the enhanced antitumor activity of NK cells combined with Olaparib in PATC148 (PDAC) and SKOV3 (ovarian cancer) cells. PATC148 (PDAC) and SKOV3 (ovarian cancer) cells were treated with Olaparib (lOuM) for 24 hours, followed by treatment with TROP2 / CAR NK cells or leaving the cells untreated. Representative realtime impedance data from the xCELLigence assay illustrated the effects of both NT NK cells and TROP2 / CARNK cells, or their combination on cell growth and viability. Images showed a significant reduction in cell viability with the combination treatment compared to singleagent treatments (FIGS. 10A and 10B).302140458.1 - 42 -

[0166] Shown in FIG. 11, combining CAR-NK cells with the PARPi Olaparib enhanced the antitumor activity of NK cells. A spheroid assay demonstrated the enhanced antitumor activity of TROP2 / CARNK cells combined with Olaparib in SKOV3 cells. SKOV3 cells were treated with Olaparib (lOOmM) for 24 hours, followed by treatment with CAR-NK cells or leaving the cells untreated. Representative image data from the spheroid assay illustrated the effects of TROP2 / CAR-NK cells, Olaparib, or their combination on cell growth and viability. Images showed a significant reduction in cell viability with the combination treatment compared to single-agent treatments.EXAMPLE 2SYNERGISTIC COMBINATION OF KRAS INHIBITOR WITH CAR-NK CELLS FOR ANTITUMOR RESPONSE

[0167] The outcome of combining CAR NK cells with KRAS inhibition was evaluated for antitumor activity.

[0168] Shown in FIGS. 12A-12E, RMC-6236 can synergize with CARNK cells to enhance antitumor activity in KRAS-mutant pancreatic cancer models. First, tumor cells were pretreated with the pan-RAS inhibitor RMC-6236 for 24 hours prior to co-culture with CAR NK cells, then CAR NK cells were subsequently added, and tumor killing was assessed longitudinally (FIG. 12A). Real-time cytotoxicity curves illustrated the effects of RMC-6236 alone, CARNK cells alone, and combination treatment in MIA-Paca-2 (KRASAG12C) and PATC148 (KRASAG12D) models (FIGS. 12B-12C). It was found that combination treatment resulted in enhanced tumor control compared to monotherapy conditions. As shown in FIGS 12D-12E, fluorescence images of tumor spheroids (MIA-Paca-2 (KRASAG12C) and PATC148 (KRASAG12D), respectively) at Day 0 and Day 6 under indicated treatment conditions were also acquired. It was found that tumor-alone controls demonstrated progressive growth, CAR NK cells reduced tumor burden, and the combination of CAR NK cells with RMC-6236 led to the most pronounced tumor regression (FIGS. 12D-12E). Together, these data suggested that pharmacologic RAS inhibition may sensitize KRAS-mutant tumor cells to CAR NK-mediated cytotoxicity and result in improved antitumor efficacy.

[0169] FIGS. 13A-13D show that RMC-6236 can enhance tumor susceptibility to CAR NK cells and preserve NK cell functional states. When evaluated for surface expression of stress ligands (MICA / B, ULB P2 / 5 / 6, and B7-H6) on tumor cells following treatment with RMC-6236, it was found that RMC-6236 treatment increased expression of multiple NK-activating ligands, which was consistent with enhanced tumor vulnerability to NK cell-302140458.1 - 43 -mediated cytotoxicity (FIG. 13A). FIG. 13B shows CyTOF-based dimensionality reduction (FlowSOM clustering over UMAP projection) of NK cells co-cultured with tumor cells in the absence (top row) or presence (bottom row) of RMC-6236. It was found that RMC-6236 exposure altered NK cell phenotypic distribution, with enrichment of clusters associated with activation and cytotoxic function (FIG. 13B), and quantification of FlowSOM metacluster distribution across experimental conditions indicated a shift toward cytotoxic NK cell subsets in the presence of RMC-6236 (FIG. 13C). In FIG. 13D, a heatmap of CyTOF marker expression across NK cell metaclusters is shown. RMC-6236 treatment was found to be associated with increased expression of cytotoxic and activation markers (e.g., Granzyme B, Perforin, CD69, NKG2D) and reduced expression of exhaustion-associated markers, which indicated preservation of NK cell functional fitness. Together, these data indicated that RMC-6236 may enhance tumor cell susceptibility by upregulating stress ligands while concurrently protecting NK cells from tumor-induced functional exhaustion.

[0170] In FIGS. 14A-14B, it is shown that RMC-6236 can enhance tumor immunogenicity while reprogramming CARNK cell transcriptional states. Differential gene expression analysis was conducted on PATC148 tumor cells in NK-tumor co-culture following RMC-6236 treatment at 3 and 6 days. It was found that RMC treatment induced multiple NK-activating stress ligands, including ULBP2 / 3, MICA / B, and NCR3LG1, which was consistent with enhanced tumor susceptibility to NK-mediated cytotoxicity. Notably, RMC also reduced expression of classical HLA class I molecules (HLA-A, HLA-B, HLA-C) as well as the non-classical inhibitory ligand HLA-E, particularly at later timepoints. This coordinated upregulation of activating ligands together with attenuation of inhibitory HLA signals may shift the tumor cell surface balance toward enhanced NK-cell activation. FIG. 14B illustrates a heatmap of row-scaled (z-score) transcript expression in CAR NK cells treated with RMC-6236. It was found that RMC induced transcriptional programs associated with inflammatory and cytokine signaling (IL6R, IKBIP, CX3CR1, GADD45A), cellular stress adaptation (SOD2-OT1, PDCD6-AHRR), cytoskeletal organization and motility (FLNB, FRMD6, LSAMP), metabolic and transporter pathways (SLC2A2, SLC5A1-AS1, CYP26B1), and epithelial / differentiation-associated markers (KRT5, KRTAP12-3). These changes indicated that RMC may remodel NK cell functional states, promoting adaptive stress responses and maintaining effector-associated transcriptional programs. Together, these data indicated that RMC-6236 may enhance tumor vulnerability through stress ligand upregulation and reduction of inhibitory HLA signals, while simultaneously reshaping CARNK cell transcriptional states in a manner compatible with sustained antitumor function.302140458.1 - 44 -

[0171] KRAS inhibition can reprogram ERK-STAT signaling to enhance CAR NK cell antitumor activity (FIG. 15). Immunoblot analysis was conducted for key signaling nodes in CAR NK cells following KRAS pathway inhibition. Western blots were performed for phosphorylation and total protein levels across indicated experimental conditions (FIG. 15). KRAS inhibition suppressed ERK pathway activation while modulating downstream STAT signaling, which was consistent with signaling rewiring that favors NK effector function. Furthermore, reduced phospho-ERK was accompanied by rebalancing of STAT pathway activity and preservation of effector-associated signaling programs. Collectively, these data indicated that KRAS inhibition may reshape intracellular signaling in CAR NK cells and attenuate ERK-driven stress signaling while promoting STAT-dependent pathways associated with enhanced cytotoxic fitness.

[0172] In FIGS. 16A-16B, it is shown that combination of CAR NK cells with the KRAS inhibitor RMC-6236 can enhance antitumor activity and tumor infiltration in a PATC148 pancreatic ductal adenocarcinoma (PDAC) model. Longitudinal bioluminescence imaging (BLI) was conducted on NSG mice bearing luciferase-expressing PATC148 tumors treated with: vehicle control (PATC148 alone), RMC-6236 alone, TROP2 / IL-15 CAR-NK cells alone, or the combination of TROP2 / IL-15 CAR-NK cells with RMC-6236. Images were acquired on days -4 pre- NK treatment, and days, 6, 14, 20, and 27 post-treatment. It was found that combination therapy resulted in marked tumor regression compared with monotherapy groups (FIG. 16A). Tumor burden was also quantified over time by measuring total flux (photons / sec). While RMC-6236 or CAR-NK cells alone moderately delayed tumor progression, the combination therapy significantly suppressed tumor growth and sustained tumor control through the study period (FIG. 16B).

[0173] KRAS inhibition can enhance CAR NK cell antitumor activity and intratumoral immune infiltration in vivo (FIGS. 17A-17D). H&E staining and CD45 immunohistochemistry were performed on PATC148 PDAC xenografts that were treated with vehicle control (Tumor Alone), CAR NK cells (TROP2 / IL15), or the combination with RMC-6236. It was found that combination treatment reduced viable tumor area and was associated with increased CD45+immune infiltration compared with monotherapy or control groups (FIG. 17A). FIG. 17B shows higher-magnification of the H&E and CD45 staining (scale bar, 100 pm). It was found that tumors that received CARNK cells plus RMC-6236 showed marked CD45+immune cell accumulation and architectural disruption consistent with enhanced cytotoxic activity (FIG.17B).302140458.1 - 45 -

[0174] Intratumoral CD45+immune infiltration was quantified and evaluated across treatment groups. FIG. 17C shows violin plots that depict distribution of CD45+cell density per tumor section. It was found that combination therapy significantly increased immune infiltration compared with CAR NK cells alone (FIG. 17C). Tumor burden was also quantified across groups at endpoint and shown as violin plots of tumor volume distribution in FIG. 17D. The results demonstrated superior tumor control in the combination group relative to monotherapy arms. Together, these data indicated that KRAS inhibition can synergize with CAR NK cell therapy in vivo, and promote enhanced immune infiltration and improved tumor control.

[0175] FIGS. 18A-18C show that RAS inhibition in combination therapy can preserve NK cell function and prevent exhaustion-induced dysfunction in vivo. Longitudinal analysis was conducted for NK cell functional capacity following exposure to tumor-bearing environments in vivo. NK cells isolated from mice treated with CAR NK cells alone or CAR NK cells plus RMC-6236 were assessed for effector function over time. While NK cells derived from the CAR NK-alone arm exhibited progressive functional decline consistent with exhaustion, NK cells from the combination arm demonstrated sustained functional activity, which indicated protection from tumor-induced dysfunction (FIG. 18 A). Images of tumor spheroid co-culture assays were taken on day 0 and day 6. It was found that NK cells derived from combination-treated mice showed enhanced tumor cell clearance compared with NK cells derived from the CAR NK-alone arm, while tumor-alone controls demonstrated continued spheroid growth (FIG. 18B). Spheroid area or viability over time was quantified following exposure to NK cells derived from treated mice. NK cells from the combination arm maintained cytotoxic activity, which resulted in sustained tumor reduction, whereas NK cells from the monotherapy arm exhibited diminished antitumor efficacy (FIG. 18C). Together, these findings indicated that RAS pathway inhibition can protect CAR NK cells from exhaustion-induced functional impairment in vivo, thereby enhance durable antitumor activity.

[0176] In FIGS. 19A-19B, it is shown that CARNK cells combined with KRAS inhibition can acquire a distinct metabolic and inflammatory transcriptional phenotype in vivo. Gene set enrichment analysis (GSEA) was performed on CAR NK cells isolated from tumors in mice treated with CAR NK cells alone versus CAR NK cells plus RMC-6236 (FIG. 19A). It was found that combination therapy was associated with enrichment of oxidative phosphorylation, MYC targets, DNA repair, fatty acid metabolism, interferon responses, glycolysis, and reactive oxygen species pathways. In contrast, pathways linked to KRAS signaling, TNF-NF-KB signaling, TGF-P signaling, Wnt / p-catenin signaling, and mitotic progression were relatively302140458.1 - 46 -suppressed. These findings indicated metabolic and inflammatory rewiring of CAR NK cells in the combination setting. FIG. 19B shows an unsupervised hierarchical clustering heatmap of differentially expressed genes in CAR NK cells isolated from tumors of non-treated versus RMC-6236-treated mice. It was found that RMC treatment induced a distinct transcriptional cluster characterized by upregulation of metabolic, mitochondrial, and interferon-associated genes and coordinated downregulation of KRAS- and stress-associated programs. Together, these data indicated that KRAS inhibition may reprogram CAR NK cells in vivo toward a metabolically optimized and less exhaustion-prone molecular state, consistent with enhanced functional persistence and antitumor activity.* * *

[0177] All of the methods disclosed and claimed herein can be made and executed without undue experimentation in light of the present disclosure. While the compositions and methods of this invention have been described in terms of preferred embodiments, it will be apparent to those of skill in the art that variations may be applied to the methods and in the steps or in the sequence of steps of the method described herein without departing from the concept, spirit and scope of the invention. More specifically, it will be apparent that certain agents which are both chemically and physiologically related may be substituted for the agents described herein while the same or similar results would be achieved. All such similar substitutes and modifications apparent to those skilled in the art are deemed to be within the spirit, scope and concept of the invention as defined by the appended claims.302140458.1 - 47 -

Claims

CLAIMSWhat is claimed is:

1. A regimen, comprising:(a) a natural killer (NK) cell-based immunotherapy; and(b) one or more inhibitors of poly ADP-ribose polymerase (PARP) and / or one or more inhibitors of a Kirsten ras oncogene homolog from the mammalian ras gene family (KRAS) pathway.

2. The regimen of claim 1, wherein the NK cells of the NK cell-based immunotherapy are wild type.

3. The regimen of claim 1, wherein the NK cells of the NK cell-based immunotherapy are not engineered by the hand of man.

4. The regimen of claim 1, wherein the NK cells of the NK cell-based immunotherapy are engineered by the hand of man.

5. The regimen of claim 4, wherein the NK cells are engineered to express one or more of one or more chimeric antigen receptors, one or more T cell receptors or one or more chains thereof, one or more antibodies, one or more fusion proteins, one or more cytokines, one or more chemokine receptors, one or more CD3 chains, or a combination thereof, optionally wherein the NK cells are engineered to express a) a CD3 protein complex comprising part or all of a single chain or any combination of CD3(^, CD3b, CD3e, or CD3y, b) optionally at least one cytokine, c) at least one TCR[3 and TCRoc chain and / or a TCRy and TCRb chain, and d) a polypeptide comprising a CD 16 Fc binding domain..

6. The regimen of claim 5, wherein the antibody is a bi-specific or multi-specific antibody.

7. The regimen of claim 1, wherein the PARP is PARP1, PARP2, PARP3, PARP4, PARP5a, PARP5b, PARP7, PARP9, PARP11, PARP14, and / or PARP15.

8. The regimen of claim 1, wherein the PARP inhibitor is selected from the group consisting of olaparib, niraparib, rucaparib, talazoparib, veliparib, pamiparib, saruparib, Fluzoparib, INO-lOOl, CEP-9722, or a combination thereof.302140458.1 - 48 -9. The regimen of claim 1, wherein the one or more inhibitors of a KRAS pathway is an inhibitor of Kirsten RAS (KRAS), Harvey RAS (HRAS), Neuroblastoma RAS (N- RAS), R-RAS, RAP, RAL, RHEB, RIN, and / or RIT.

10. The regimen of claim 1, wherein the one or more inhibitors of a KRAS pathway is an inhibitor of KRAS.

11. The regimen of claim 1, wherein (a) and (b) are in the same formulation.

12. The regimen of claim 1, wherein (a) and (b) are not in the same formulation.

13. A composition, comprising:(a) a natural killer (NK) cell-based immunotherapy; and(b) one or more inhibitors of poly ADP-ribose polymerase (PARP) and / or one or more inhibitors of a KRAS pathway, optionally wherein the one or more inhibitors of poly ADP-ribose polymerase (PARP) and / or one or more inhibitors of a KRAS pathway are in the NK cells.

14. The composition of claim 13, wherein the composition is comprised in a pharmaceutically acceptable carrier.

15. The composition of claim 13, wherein the NK cells of the NK cell-based immunotherapy are wild type.

16. The composition of claim 13, wherein the NK cells of the NK cell-based immunotherapy are not engineered by the hand of man.

17. The composition of claim 13, wherein the NK cells of the NK cell-based immunotherapy are engineered by the hand of man.

18. The composition of claim 17, wherein the NK cells are engineered to express one or more of one or more chimeric antigen receptors, one or more T cell receptors or one or more chains thereof, one or more antibodies, one or more fusion proteins, one or more cytokines, one or more chemokine receptors, one or more CD3 chains, or a combination thereof.302140458.1 - 49 -19. The composition of claim 18, wherein the antibody is a bi-specific or multi-specific antibody.

20. The composition of claim 13, wherein the PARP is PARP1, PARP2, PARP3, PARP4, PARP5a, PARP5b, PARP7, PARP9, PARP11, PARP14, and / or PARP15.

21. The composition of claim 13, wherein the PARP inhibitor is selected from the group consisting of olaparib, niraparib, rucaparib, talazoparib, veliparib, pamiparib or a combination thereof.

22. The composition of claim 13, wherein the one or more inhibitors of a KRAS pathway is an inhibitor of Kirsten RAS (KRAS), Harvey RAS (HRAS), Neuroblastoma RAS (N-RAS), R-RAS, RAP, RAL, RHEB, RIN, and / or RIT.

23. The composition of claim 13, wherein the one or more inhibitors of a KRAS pathway is an inhibitor of KRAS.

24. The composition of claim 13, wherein (a) and (b) are in the same formulation.

25. The composition of claim 13, wherein (a) and (b) are not in the same formulation26. A kit, comprising the composition of claim 13.

27. A method of treating an individual in need thereof, comprising the step of:subjecting the individual to a therapeutically effective amount of the regimen of claim 1; oradministering to the individual a therapeutically effective amount of the composition of claim 13.

28. The method of claim 27, wherein the individual has cancer or has an autoimmune disease.

29. The method of claim 27, wherein there is a synergistic effect from (a) natural killer (NK) cell-based immunotherapy; and the (b) one or more inhibitors of poly ADP-ribose polymerase (PARP) and / or one or more inhibitors of a KRAS pathway.302140458.1 - 50 -30. The method of claim 27, wherein the NK cell-based immunotherapy is administered to the individual prior to the one or more inhibitors of PARP and / or the one or more inhibitors of a KRAS pathway.

31. The method of claim 27, wherein the NK cell-based immunotherapy is administered to the individual subsequent to the one or more inhibitors of PARP and / or the one or more inhibitors of a KRAS pathway.

32. The method of claim 27, wherein the NK cell-based immunotherapy is administered to the individual at the same time as the one or more inhibitors of PARP and / or the one or more inhibitors of a KRAS pathway.

33. A method of treating an individual in need thereof, comprising:in any order, administering a therapeutically effective amount of a NK cell-based immunotherapy; and one or both of (1) one or more inhibitors of PARP and (2) one or more inhibitors of a KRAS pathway.

34. The method of claim 33, wherein the NK cell-based immunotherapy and one or more inhibitors of PARP are administered to the individual.

35. The method of claim 33, wherein the NK cell-based immunotherapy and one or more inhibitors of a KRAS pathway are administered to the individual.302140458.1 - 51 -

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