METHOD FOR TREATING A TUMOR AND PHARMACEUTICAL COMPOSITION

Description

1 / 39 METHOD FOR TREATING A TUMOR AND PHARMACEUTICAL COMPOSITION CROSS-REFERENCES TO RELATED REQUESTS

[0001] This application claims the benefit of U.S. Provisional Patent Application No. 63 / 458,843, filed April 12, 2023, which is incorporated herein by reference in its entirety. 1. FIELD OF DISSEMINATION

[0002] The present disclosure is in the field of compositions comprising xenogeneic antigen-presenting cells (e.g., from one or more pigs), methods of producing such compositions and uses of such compositions for treating tumors. 2. FUNDAMENTALS OF DISCLOSURE

[0003] Traditional cancer therapies, such as chemotherapy, surgery, radiation, etc., are often inadequate for treating cancer patients and are generally associated with serious side effects. In this sense, cancer immunotherapy offers several advantages over traditional cancer therapies. Several different immunotherapy approaches have been investigated. One such immunotherapy strategy involved the intratumoral administration of allogeneic inflammatory dendritic cells, usually in combination with another anticancer therapy, for the treatment of certain types of cancer (see, for example, Jin et al. (2022), Oncoimmunology, 11(1):e2099642; Frobom et al. (2020), Cancer Immunol. Imunoter., 69(11):23932401; Karlsson-Parra et al. (2018), Pharm. Res., 35:156).For example, ilixadencel, an allogeneic dendritic cell product derived from human monocytes stimulated with potent activators, has been investigated in human clinical trials (see, for example, Frobom et al. (2020), Cancer Immunol. Imunooutros., 69(11):2393-2401). There is still a need for cancer immunotherapies that demonstrate a robust immune response.

[0004] Citation of a reference in this document cannot be interpreted as an admission that it is prior art for the Petition 870250092182, dated 08 / 10 / 2025, page 12 / 72 2 / 39 present disclosure. 3. SUMMARY OF THE DISCLOSURE

[0005] In one aspect, methods are provided here for treating a tumor in a subject in need thereof, comprising administering a composition comprising antigen-presenting cells to the subject, wherein the antigen-presenting cells are obtained from a species different from the subject.

[0006] In one embodiment, the tumor is a solid tumor, and the administration is made into the subject's tumor by means of an intratumoral injection.

[0007] In one embodiment, the antigen-presenting cells are obtained from a swine species. In one embodiment, the antigen-presenting cells are obtained from a species that is a miniature swine. In one embodiment, the subject is a human. In one embodiment, the swine is an alpha-1,3 galactosyltransferase-deficient swine. In one embodiment, the alpha-1,3 galactosyltransferase-deficient swine is an inbred swine with swine leukocyte antigen (SLA). In one embodiment, the miniature swine is an alpha-1,3 galactosyltransferase-deficient miniature swine. In one embodiment, the swine is an alpha-1,3 galactosyltransferase-deficient miniature swine and is an inbred swine with swine leukocyte antigen (SLA).

[0008] In one embodiment, the method triggers a tumor-specific immune response. In another embodiment, the method produces an abscopal effect.

[0009] In one embodiment, the tumor is a solid cancerous tumor. In one embodiment, the tumor is selected from the group consisting of sarcomas, carcinomas, lymphomas, breast tumors, prostate tumors, head and neck tumors, glioblastomas, bladder tumors, pancreatic tumors, liver tumors, ovarian tumors, colorectal tumors, lung tumors, skin tumors, lymphoid tumors, gastrointestinal tumors, gastrointestinal stromal tumors, cervical tumors, hepatocellular carcinomas, renal cell carcinomas, melanomas, colorectal carcinomas, carcinomas Petition 870250092182, dated 08 / 10 / 2025, page 13 / 72 3 / 39 esophageal, brain tumors, kidney tumors, lung tumors (including non-small cell lung cancer), gastric tumors, bile duct tumors, uterine tumors, and childhood (pediatric) tumors. In one modality, the tumor is resistant to treatment with chemotherapy and / or immunotherapy.

[0010] In one embodiment, antigen-presenting cells are derived from one or more pigs or miniature pigs using a leukapheresis procedure, wherein the leukapheresis procedure generates a leukopace containing peripheral blood mononuclear cells, and the leukopace is further fractionated by counterflow elutriation.

[0011] In one embodiment, the composition comprising antigen-presenting cells is substantially free of pathogens.

[0012] In one embodiment, antigen-presenting cells are obtained from pigs or miniature pigs of different genotypes.

[0013] In one embodiment, the composition comprising antigen-presenting cells is administered in single or multiple doses. In another embodiment, the composition comprising antigen-presenting cells is administered by means of an intratumoral injection of at least about 1 x 10⁶ antigen-presenting cells per dose.In one embodiment, the composition comprising antigen-presenting cells is administered via intratumoral injection of approximately 1 x 10⁶, approximately 5 x 10⁶, approximately 10 x 10⁶, approximately 15 x 10⁶, approximately 20 x 10⁶, approximately 25 x 10⁶, approximately 30 x 10⁶, approximately 35 x 10⁶, approximately 40 x 10⁶, approximately 45 x 10⁶, approximately 50 x 10⁶, approximately 55 x 10⁶, approximately 60 x 10⁶, approximately 65 x 10⁶, approximately 70 x 10⁶, approximately 75 x 10⁶, approximately 80 x 10⁶, approximately 85 x 10⁶, approximately 90 x 10⁶, approximately 95 x 10⁶, approximately 10 x 107, approximately 15 x 107, approximately 20 x 107, approximately 25 x 107, approximately 30 x 107, approximately 35 x 107, approximately 40 x 107, approximately 45 x 107, or approximately 50 x 107 antigen-presenting cells per dose.

[0014] In one embodiment, the antigen-presenting cells are substantially mature antigen-presenting cells. Petition 870250092182, dated 08 / 10 / 2025, p. 14 / 72 4 / 39

[0015] In one embodiment, antigen-presenting cells are not activated or stimulated.

[0016] In one embodiment, the composition comprises peripheral blood mononuclear cells (PBMCs).

[0017] In one embodiment, the composition comprises monocytes.

[0018] In one embodiment, the composition comprises dendritic cells, macrophages, granulocytes, T cells, B cells and / or NK cells.

[0019] In one embodiment, the composition comprises at least about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95% or more of about 95% of PBMCs, wherein the PBMCs are mature, immature or a combination of mature and immature PBMCs.

[0020] In one embodiment, the composition comprises at least about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95% or more than about 95% of monocytes, wherein the monocytes are mature, immature or a combination of mature and immature monocytes.

[0021] In one embodiment, the composition comprises a mixture of at least about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95% or more of about 95% of monocytes and dendritic cells, wherein the monocytes and / or dendritic cells in said mixture may be mature, immature or a combination of mature and immature monocytes and / or dendritic cells.

[0022] In one embodiment, the mixture comprising monocytes and dendritic cells comprises 10% to 95%, 10% to 90%, 10% to 80%, 10% to 70%, 10% to 60%, 10% to 50%, 10% to 40%, 10% to 30%, 10% to 20%, 20% to 95%, 20% to 90%, 20% to 80%, 20% to 70%, 20% to 60%, 20% to 50%, 20% to 40%, 20% to 30%, 30% to 95%, 30% to 90%, 30% to 80%, 30% to 70%, 30% to 60%, 30% to 50%, 30% to 40%, 40% to 95%, 40% to 90%, 40% to 80%, 40% to 70%, 40% to 60%, Petition 870250092182, dated 08 / 10 / 2025, page 15 / 72 5 / 39 40% to 50%, 50% to 95%, 50% to 90%, 50% to 80%, 50% to 70%, 50% to 60%, 60% to 95%, 60% to 90%, 60% to 80%, 60% to 70%, 70% to 95%, 70% to 90%, 70% to 80%, 80% to 95%, 80% to 90%, 90% to 95% or more than 95% of monocytes. In one embodiment, the mixture comprising monocytes and dendritic cells comprises 10% to 30%, 35% to 55%, 60% to 80% or 85% to 95% of monocytes. In one embodiment, the mixture comprises at least about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or more than about 95% of monocytes.

[0023] In one embodiment, the mixture comprising monocytes and dendritic cells comprises 10% to 95%, 10% to 90%, 10% to 80%, 10% to 70%, 10% to 60%, 10% to 50%, 10% to 40%, 10% to 30%, 10% to 20%, 20% to 95%, 20% to 90%, 20% to 80%, 20% to 70%, 20% to 60%, 20% to 50%, 20% to 40%, 20% to 30%, 30% to 95%, 30% to 90%, 30% to 80%, 30% to 70%, 30% to 60%, 30% to 50%, 30% to 40%, 40% to 95%, 40% to 90%, 40% to 80%, 40% to 70%, 40% to 60%, 40% to 50%, 50% to 95%, 50% to 90%, 50% to 80%, 50% to 70%, 50% to 60%, 60% to 95%, 60% to 90%, 60% to 80%, 60% to 70%, 70% to 95%, 70% to 90%, 70% to 80%, 80% to 95%, 80% to 90%, 90% to 95% or more than 95% of dendritic cells. In one embodiment, the mixture comprising monocytes and dendritic cells consists of 10% to 30%, 35% to 55%, 60% to 80%, or 85% to 95% dendritic cells.In one embodiment, the mixture comprising monocytes and dendritic cells comprises at least about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or more of about 95% dendritic cells.

[0024] In one modality, the subject is receiving another anticancer therapy. In one modality, the other anticancer therapy comprises treatment with one or more immune checkpoint inhibitors. In one Petition 870250092182, dated 08 / 10 / 2025, page 16 / 72 In modality 6 / 39, the other anticancer therapy is an anti-CTLA4 therapy, anti-PD1 therapy, anti-PDL1 therapy, anti-LAG-3 therapy, tumor treatment fields (TTFs), cell-based therapy, a tyrosine kinase inhibitor, a VEGF inhibitor, or any combination thereof. In one modality, the other anticancer therapy comprises treatment with imatinib, sunitinib, regorafenib, pazopanib, nilotinib, avapritinib, ripretinib, sorafenib, pimitespib, ipilimumab, tremelimumab, nivolumab, pembrolizumab, cemiplimab, atelizumab, avelumab, durvalumab, relatlimab, or any combination thereof.

[0025] In one embodiment, the subject does not respond to other anticancer therapy in the absence of administration of the composition comprising antigen-presenting cells.

[0026] In another aspect, a pharmaceutical composition suitable for intratumoral injection is provided herein, wherein the pharmaceutical composition comprises antigen-presenting cells obtained from one or more pigs. 4. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] FIG. 1: FIG. 1 shows a general leukapheresis procedure to generate a leukopak containing antigen-presenting cells from pig blood.

[0028] FIG. 2 shows a comparison of tumor size between the treatment group (administration of a composition comprising human PBMCs) and the control group in a murine melanoma model on day 12.

[0029] FIG. 3 shows a comparison of tumor size between the treatment group (administration of a composition comprising human PBMCs) and the control group in a murine bladder cancer model on day 7.

[0030] FIGS. 4A-4C: FIG. 4A shows the human CD4 T cell count on day 7 in an in vitro study evaluating (a) porcine PBMCs plated with human PBMCs in varying ratios of human to porcine cells (h:p), (b) human PBMCs alone, and (c) allogeneic human PBMCs. FIG. 4B shows the human CD8 T cell count on Day 7 in an in vitro study. Petition 870250092182, dated 08 / 10 / 2025, page 17 / 72 Figure 4C shows the human NK cell count on day 7 in an in vitro study evaluating (a) porcine PBMCs plated with human PBMCs in varying ratios of human to porcine cells (h:p), (b) human PBMCs alone, and (c) allogeneic human PBMCs.

[0031] FIGS. 5A-5G: FIG. 5A shows the human CD4 T cell count on Day 7 in an in vitro study evaluating (a) porcine monocytes plated with human PBMCs in varying ratios of human to porcine cells (h:p), (b) human PBMCs alone, and (c) allogeneic human PBMCs. FIG. 5B shows the human CD8 T cell count on Day 7 in an in vitro study evaluating (a) porcine monocytes plated with human PBMCs in varying ratios of human to porcine cells (h:p), (b) human PBMCs alone, and (c) allogeneic human PBMCs. Figure 5C shows the human CD4 T cell count on Day 7 in an in vitro study evaluating (a) porcine PBMCs plated with human PBMCs in varying ratios of human to porcine cells (h:p), (b) human PBMCs alone, and (c) allogeneic human PBMCs.Figure 5D shows the count of proliferating human CD8 T cells on Day 7 in an in vitro study evaluating (a) porcine monocytes plated with human PBMCs in varying ratios of human to porcine cells (h:p), (b) human PBMCs alone, and (c) allogeneic human PBMCs. Figure 5E shows HLA-DR in human CD4 T cells, in terms of mean fluorescence intensity (MFI), evaluated on Day 7 in an in vitro study evaluating (a) porcine monocytes plated with human PBMCs in varying ratios of human to porcine cells (h:p), (b) human PBMCs alone, and (c) allogeneic human PBMCs. Figure 5F shows HLA-DR in human CD8 T cells, in terms of mean fluorescence intensity (MFI), assessed on Day 7 in an in vitro study evaluating (a) porcine monocytes bathed with human PBMCs in varying ratios of human to porcine cells (h:p), (b) human PBMCs alone, and (c) human PBMCs. Petition 870250092182, dated 08 / 10 / 2025, page 18 / 72 8 / 39 allogeneic. FIG. 5G shows the human NK cell count on Day 7 in an in vitro study evaluating (a) porcine monocytes plated with human PBMCs in varying ratios of human to porcine cells (h:p), (b) human PBMCs alone, and (c) allogeneic human PBMCs. 5. DETAILED DESCRIPTION OF THE DISCLOSURE

[0032] Methods are provided here for treating a tumor in a subject by administering a composition comprising antigen-presenting cells obtained from a species different from the subject, i.e., xenogeneic antigen-presenting cells. The source of these xenogeneic antigen-presenting cells, which may be one or more pigs, is described in Section 5.1. Also described in Section 5.1 are the characteristics of the pig and compositions comprising antigen-presenting cells, including antigen-presenting cells derived from pigs. Methods for isolating antigen-presenting cells (e.g., from one or more pigs) are described in Section 5.2. Methods for treating tumors (e.g., a cancerous solid tumor) comprising administering antigen-presenting cells described herein (e.g., by intratumoral injection), optionally in combination with other anticancer therapies, are described in Section 5.3.Furthermore, pharmaceutical compositions comprising the antigen-presenting cells disclosed herein are described in Section 5.4. 5.1 Xenogeneic Compositions

[0033] Compositions for treating a tumor in a human subject are provided herein, wherein the compositions comprise antigen-presenting cells derived from a species different from the subject. In certain embodiments, compositions comprising antigen-presenting cells xenogeneic to the human subject to be treated are provided herein. These compositions are referred to in this document as xenogeneic compositions. The xenogeneic compositions of the present invention may be administered (e.g., by intratumoral injection) alone or by means of Petition 870250092182, dated 08 / 10 / 2025, page 19 / 72 9 / 39 of the formulation in a pharmaceutical composition as described in Section 5.4

[0034] Antigen-presenting cells in the xenogeneic composition can be any cell that displays antigen bound by major histocompatibility complex (MHC) proteins on its surface. In one embodiment, the MHC proteins are MHC Class II molecules. Antigen-presenting cells in the xenogeneic composition may include, but are not limited to, dendritic cells, macrophages, B cells, monocytes, mononuclear phagocytes, endothelial cells, thymic epithelial cells, granulocytes, or any combination thereof.

[0035] The xenogeneic compositions of the present invention may comprise antigen-presenting cells of any non-human species, including, but not limited to, non-human primates, domestic animals, and rodents. The non-human species may be genetically modified. Domestic animals include pigs, rabbits, dogs, cats, horses, sheep, goats, and cows. In one embodiment, the non-human species is a pig or swine. As set forth herein, the terms pig, porcine, or swine may be used interchangeably.

[0036] The antigen-presenting cells in the xenogeneic compositions of the present invention can be obtained by the methods described in Section 5.2. In certain embodiments, the xenogeneic composition comprises antigen-presenting cells derived from one or more pigs. The one or more pigs used to generate the antigen-presenting cells may be genetically modified pigs, for example, to increase the safety and / or efficacy of the antigen-presenting cells. In some embodiments, the pig is an alpha-1,3 galactosyltransferase-deficient pig. In certain embodiments, the antigen-presenting cells are derived from one or more recombinant pigs. In certain embodiments, the antigen-presenting cells are derived from recombinant pigs of different genotypes. In certain embodiments, the antigen-presenting cells are derived from a Petition 870250092182, dated 08 / 10 / 2025, page 20 / 72 10 / 39 or more miniature pigs. The one or more miniature pigs used to generate the antigen-presenting cells may be genetically modified miniature pigs, for example, to increase the safety and / or efficacy of the antigen-presenting cells. In some embodiments, the miniature pig is an alpha-1,3 galactosyltransferase-deficient miniature pig (see, for example, US Patents Nos. 6,153,428, 6,413,769 and 7,547,816). In certain embodiments, the antigen-presenting cells are derived from one or more recombinant miniature pigs. In certain embodiments, the antigen-presenting cells are derived from recombinant miniature pigs of different genotypes. In some embodiments, the alpha-1,3 galactosyltransferase-deficient pig or miniature pig is an inbred pig with porcine leukocyte antigen (SLA).In certain embodiments, the one or more pigs or miniature pigs described herein may contain knockout genes or transgenes in porcine antigen-presenting cell genes.

[0037] The donors for the antigen-presenting cells used in the xenogeneic compositions described herein can be tested for infectious diseases using methods generally known in the art. In certain embodiments, the donors for the antigen-presenting cells used in the xenogeneic compositions described herein have yielded negative results for syphilis, HIV, hepatitis A, hepatitis B, and / or hepatitis C. In certain embodiments, the xenogeneic compositions comprising antigen-presenting cells are substantially free of pathogens.

[0038] In certain embodiments, the xenogeneic composition comprises antigen-presenting cells derived from peripheral blood. In certain embodiments, the xenogeneic composition comprises antigen-presenting cells derived from porcine peripheral blood. In certain embodiments, the xenogeneic composition comprises antigen-presenting cells derived from bone marrow. In certain embodiments, the xenogeneic composition comprises antigen-presenting cells derived from porcine bone marrow. Petition 870250092182, dated 08 / 10 / 2025, page 21 / 72 11 / 39

[0039] In certain embodiments, the xenogeneic composition comprises peripheral blood mononuclear cells (PBMCs). In certain embodiments, the xenogeneic composition comprises monocytes. In certain embodiments, the xenogeneic composition comprises porcine PBMCs. In certain embodiments, the xenogeneic composition comprises porcine monocytes. In certain embodiments, the xenogeneic composition comprises monocytes derived from porcine peripheral blood. In certain embodiments, the xenogeneic composition comprises monocytes derived from porcine bone marrow.

[0040] In certain embodiments, the xenogeneic composition comprises at least about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or more than about 95% of PBMCs. The PBMCs may be mature, immature, or a combination of mature and immature PBMCs.

[0041] In certain embodiments, the xenogeneic composition comprises 50% to 95%, 50% to 90%, 50% to 80%, 50% to 70%, 50% to 60%, 60% to 95%, 60% to 90%, 60% to 80%, 60% to 70%, 70% to 95%, 70% to 90%, 70% to 80%, 80% to 95%, 80% to 90%, 90% to 95% or more than 95% of PBMCs.

[0042] In certain embodiments, the PBMCs present in the xenogeneic composition are at least 10% to 95%, 10% to 90%, 10% to 80%, 10% to 70%, 10% to 60%, 10% to 50%, 10% to 40%, 10% to 30%, 10% to 20%, 20% to 95%, 20% to 90%, 20% to 80%, 20% to 70%, 20% to 60%, 20% to 50%, 20% to 40%, 20% to 30%, 30% to 95%, 30% to 90%, 30% to 80%, 30% to 70%, 30% to 60%, 30% to 50%, 30% to 40%, 40% to 95%, 40% to 90%, 40% to 80%, 40% to 70%, 40% to 60%, 40% to 50%, 50% to 95%, 50% to 90%, 50% to 80%, 50% to 70%, 50% to 60%, 60% to 95%, 60% to 90%, 60% to 80%, 60% to 70%, 70% to 95%, 70% to 90%, 70% to 80%, 80% to 95%, 80% to 90%, 90% to 95% or more than 95% of mature PBMCs.

[0043] In certain embodiments, the PBMCs present in the xenogeneic composition are at least 10% to 30%, 35% to 55%, 60% to 80% or 85% to 95% mature PBMCs.

[0044] In certain modalities, the PBMCs present in the composition Petition 870250092182, dated 08 / 10 / 2025, p. 22 / 72 12 / 39 xenogeneic components are at least about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, or about 95% mature PBMCs. In certain embodiments, the PBMCs present in the xenogeneic composition are greater than about 95% mature PBMCs. In certain embodiments, the PBMCs present in the xenogeneic composition are about 100% mature PBMCs.

[0045] In certain embodiments, the xenogeneic composition comprises at least about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95% or more than about 95% monocytes. The monocytes may be mature, immature, or a combination of mature and immature monocytes.

[0046] In certain embodiments, the xenogeneic composition comprises 50% to 95%, 50% to 90%, 50% to 80%, 50% to 70%, 50% to 60%, 60% to 95%, 60% to 90%, 60% to 80%, 60% to 70%, 70% to 95%, 70% to 90%, 70% to 80%, 80% to 95%, 80% to 90%, 90% to 95% or more than 95% of monocytes.

[0047] In certain embodiments, the monocytes present in the xenogeneic composition are at least 10% to 95%, 10% to 90%, 10% to 80%, 10% to 70%, 10% to 60%, 10% to 50%, 10% to 40%, 10% to 30%, 10% to 20%, 20% to 95%, 20% to 90%, 20% to 80%, 20% to 70%, 20% to 60%, 20% to 50%, 20% to 40%, 20% to 30%, 30% to 95%, 30% to 90%, 30% to 80%, 30% to 70%, 30% to 60%, 30% to 50%, 30% to 40%, 40% to 95%, 40% to 90%, 40% to 80%, 40% to 70%, 40% to 60%, 40% to 50%, 50% to 95%, 50% to 90%, 50% to 80%, 50% to 70%, 50% to 60%, 60% to 95%, 60% to 90%, 60% to 80%, 60% to 70%, 70% to 95%, 70% to 90%, 70% to 80%, 80% to 90%, 80% to 90%, 90% to 95% or more than 95% of mature monocytes.

[0048] In certain embodiments, the monocytes present in the xenogeneic composition are at least 10% to 30%, 35% to 55%, 60% to 80% or 85% to 95% mature monocytes. Petition 870250092182, dated 08 / 10 / 2025, p. 23 / 72 13 / 39

[0049] In certain embodiments, the monocytes present in the xenogeneic composition are at least about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, or about 95% mature monocytes. In certain embodiments, the monocytes present in the xenogeneic composition are more than about 95% mature monocytes. In certain embodiments, the monocytes present in the xenogeneic composition are about 100% mature monocytes.

[0050] In certain embodiments, the xenogeneic composition comprises a mixture of monocytes and dendritic cells. In certain embodiments, the xenogeneic composition comprises a mixture of monocytes and dendritic cells derived from one or more pigs. The composition may further comprise macrophages, granulocytes, B cells, T cells, NK cells, or any combination thereof. The macrophages, granulocytes, B cells, T cells, NK cells, or any combination thereof may be derived from one or more pigs or other non-human species.

[0051] In certain embodiments, the xenogeneic composition comprises a mixture of at least about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95% or more of about 95% of monocytes and dendritic cells. The monocytes and / or dendritic cells in such mixtures may be mature, immature, or a combination of mature and immature monocytes and / or dendritic cells.

[0052] In certain embodiments, the xenogeneic composition comprises a mixture of 50% to 95%, 50% to 90%, 50% to 80%, 50% to 70%, 50% to 60%, 60% to 95%, 60% to 90%, 60% to 80%, 60% to 70%, 70% to 95%, 70% to 90%, 70% to 80%, 80% to 95%, 80% to 90%, 90% to 95% or more than 95% of monocytes and dendritic cells.

[0053] The mixture of monocytes and dendritic cells may comprise Petition 870250092182, dated 08 / 10 / 2025, page 24 / 72 14 / 39 10% to 95%, 10% to 90%, 10% to 80%, 10% to 70%, 10% to 60%, 10% to 50%, 10% to 40%, 10% to 30%, 10% to 20%, 20% to 95%, 20% to 90%, 20% to 80%, 20% to 70%, 20% to 60%, 20% to 50%, 20% to 40%, 20% to 30%, 30% to 95%, 30% to 90%, 30% to 80%, 30% to 70%, 30% to 60%, 30% to 50%, 30% to 40%, 40% to 95%, 40% to 90%, 40% to 80%, 40% to 70%, 40% to 60%, 40% to 50%, 50% to 95%, 50% to 90%, 50% to 80%, 50% to 70%, 50% to 60%, 60% to 95%, 60% to 90%, 60% to 80%, 60% to 70%, 70% to 95%, 70% to 90%, 70% to 80%, 80% to 95%, 80% to 90%, 90% to 95% or more than 95% monocytes.

[0054] In certain embodiments, the mixture of monocytes and dendritic cells comprises 10% to 30%, 35% to 55%, 60% to 80% or 85% to 95% monocytes.

[0055] In certain embodiments, the mixture of monocytes and dendritic cells comprises at least about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95% or more than about 95% of monocytes.

[0056] In certain embodiments, the monocytes present in the mixture of monocytes and dendritic cells are at least 10% to 95%, 10% to 90%, 10% to 80%, 10% to 70%, 10% to 60%, 10% to 50%, 10% to 40%, 10% to 30%, 10% to 20%, 20% to 95%, 20% to 90%, 20% to 80%, 20% to 70%, 20% to 60%, 20% to 50%, 20% to 40%, 20% to 30%, 30% to 95%, 30% to 90%, 30% to 80%, 30% to 70%, 30% to 60%, 30% to 50%, 30% to 40%, 40% to 95%, 40% to 90%, 40% to 80%, 40% to 70%, 40% to 60%, 40% to 50%, 50% to 95%, 50% to 90%, 50% to 80%, 50% to 70%, 50% to 60%, 60% to 95%, 60% to 90%, 60% to 80%, 60% to 70%, 70% to 95%, 70% to 90%, 70% to 80%, 80% to 90%, 80% to 90%, 90% to 95% or more than 95% mature monocytes.

[0057] In certain embodiments, the monocytes present in the mixture of monocytes and dendritic cells are at least 10% to 30%, 35% to 55%, 60% to 80% or 85% to 95% mature monocytes. Petition 870250092182, dated 08 / 10 / 2025, p. 25 / 72 15 / 39

[0058] In certain embodiments, the monocytes present in the monocyte-dendritic cell mixture are at least about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, or about 95% mature monocytes. In certain embodiments, the monocytes present in the monocyte-dendritic cell mixture are more than about 95% mature monocytes. In certain embodiments, the monocytes present in the monocyte-dendritic cell mixture are about 100% mature monocytes.

[0059] The mixture of monocytes and dendritic cells may comprise 10% to 95%, 10% to 90%, 10% to 80%, 10% to 70%, 10% to 60%, 10% to 50%, 10% to 40%, 10% to 30%, 10% to 20%, 20% to 95%, 20% to 90%, 20% to 80%, 20% to 70%, 20% to 60%, 20% to 50%, 20% to 40%, 20% to 30%, 30% to 95%, 30% to 90%, 30% to 80%, 30% to 70%, 30% to 60%, 30% to 50%, 30% to 40%, 40% to 95%, 40% to 90%, 40% to 80%, 40% to 70%, 40% to 60%, 40% to 50%, 50% to 95%, 50% to 90%, 50% to 80%, 50% to 70%, 50% to 60%, 60% to 95%, 60% to 90%, 60% to 80%, 60% to 70%, 70% to 95%, 70% to 90%, 70% to 80%, 80% to 95%, 80% to 90%, 90% to 95% or more than 95% dendritic cells.

[0060] In certain embodiments, the mixture of monocytes and dendritic cells comprises 10% to 30%, 35% to 55%, 60% to 80%, or 85% to 95% dendritic cells.

[0061] In certain embodiments, the mixture of monocytes and dendritic cells comprises at least about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, or about 95% dendritic cells. In certain embodiments, the mixture of monocytes and dendritic cells comprises more than about 95% dendritic cells.

[0062] In certain embodiments, the dendritic cells present in the mixture of monocytes and dendritic cells are at least 10% to 95%, 10% Petition 870250092182, dated 08 / 10 / 2025, page 26 / 72 16 / 39 to 90%, 10% to 80%, 10% to 70%, 10% to 60%, 10% to 50%, 10% to 40%, 10% to 30%, 10% to 20%, 20% to 95%, 20% to 90%, 20% to 80%, 20% to 70%, 20% to 60%, 20% to 50%, 20% to 40%, 20% to 30%, 30% to 95%, 30% to 90%, 30% to 80%, 30% to 70%, 30% to 60%, 30% to 50%, 30% to 40%, 40% to 95%, 40% 90%, 40% to 80%, 40% to 70%, 40% to 60%, 40% to 50%, 50% to 95%, 50% to 90%, 50% to 80%, 50% to 70%, 50% to 60%, 60% to 95%, 60% to 90%, 60% to 80%, 60% to 70%, 70% to 95%, 70% to 90%, 70% to 80%, 80% to 95%, 80% to 90%, 90% to 95% or more than 95% mature dendritic cells.

[0063] In certain embodiments, the dendritic cells present in the mixture of monocytes and dendritic cells are at least 10% to 30%, 35% to 55%, 60% to 80% or 85% to 95% mature dendritic cells.

[0064] In certain embodiments, the dendritic cells present in the mixture of monocytes and dendritic cells are at least about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, or about 95% mature dendritic cells. In certain embodiments, the dendritic cells present in the mixture of monocytes and dendritic cells are more than about 95% mature dendritic cells. In certain modalities, the dendritic cells present in the mixture of monocytes and dendritic cells are approximately 100% mature dendritic cells.

[0065] In certain modalities, antigen-presenting cells in the xenogeneic composition are not activated or stimulated. In certain modalities, antigen-presenting cells in the xenogeneic composition are activated or stimulated. In certain modalities, antigen-presenting cells are stimulated with one or more pro-inflammatory factors (e.g., granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin 4 (IL-4), etc.). For example, porcine monocytes can be cultured in dendritic cells using GM-CSF and IL-4 for 7 days.

[0066] In certain forms, PBMCs in the xenogeneic composition Petition 870250092182, dated 08 / 10 / 2025, page 27 / 72 17 / 39 are not activated or stimulated. In certain modalities, PBMCs in the xenogeneic composition are activated or stimulated. In certain modalities, PBMCs are stimulated with one or more pro-inflammatory factors.

[0067] In certain modalities, monocytes and / or dendritic cells in the xenogeneic composition are not activated or stimulated. In certain modalities, monocytes and / or dendritic cells in the xenogeneic composition are activated or stimulated. In certain modalities, monocytes and / or dendritic cells are stimulated with one or more pro-inflammatory factors.

[0068] In certain embodiments, antigen-presenting cells in the xenogeneic composition are activated or stimulated using porcine B cells as stimulators. In certain embodiments, antigen-presenting cells in the xenogeneic composition are activated or stimulated using porcine T cells as stimulators. In certain embodiments, antigen-presenting cells in the xenogeneic composition are activated or stimulated using porcine monocyte-derived macrophages as stimulators. 5.2 Methods for obtaining antigen-presenting cells

[0069] Methods for obtaining antigen-presenting cells in the xenogeneic compositions described in Section 5.1 are provided here.

[0070] Antigen-presenting cells can be derived from tissue or blood of any non-human species (e.g., one or more pigs). These antigen-presenting cells can be obtained by methods generally known in the art. Antigen-presenting cells can be obtained using a leukapheresis procedure. This leukapheresis procedure can be used to generate a leukopak containing peripheral blood mononuclear cells. The leukopak can be further fractionated by counterflow elutriation to obtain a composition comprising the desired antigen-presenting cells. In some embodiments, antigen-presenting cells are obtained using Petition 870250092182, dated 08 / 10 / 2025, page 28 / 72 18 / 39 Fluorescence-activated cell sorting (FACS) or magnetically activated cell sorting (MACS) techniques. 5.3 Treatment methods

[0071] Methods are provided here for treating a tumor in a subject in need thereof, comprising administering to the subject (for example, by intratumoral injection) a xenogeneic composition described in Section 5.1. In certain embodiments, administration is effected by means of one or more intratumoral injections, as described in Section 5.3.1.

[0072] In certain embodiments, methods are provided here for treating a tumor in a subject in need thereof, comprising administering (e.g., by intratumoral injection) a xenogeneic composition in combination with a composition comprising human antigen-presenting cells. The antigen-presenting cells in the xenogeneic composition and the composition comprising human antigen-presenting cells may or may not be activated or stimulated.

[0073] The treatment methods provided here can be used to treat a wide variety of tumors. In one modality, the tumor is a solid tumor. In one modality, the tumor is a cancerous tumor. In one specific modality, the tumor is a solid cancerous tumor. In certain modalities, the tumor is resistant to treatment with chemotherapy and / or treatment with other immunotherapy. In certain modalities, the xenogeneic composition is administered (e.g., by intratumoral injection) as a first-line therapy to treat one or more tumors. In certain other modalities, the xenogeneic composition is administered to individuals (e.g., by intratumoral injection) who are receiving or may have previously received other anticancer therapy. In one modality, the antigen-presenting cells in the xenogeneic composition are as different as possible from the cells in the tumor tissue.Thus, in one embodiment, the antigen-presenting cells in the xenogeneic composition are distinctly different from the cells in the tumor tissue. For example, in one... Petition 870250092182, dated 08 / 10 / 2025, page 29 / 72 19 / 39 modality, a method of treating human liver cancer comprises administering a porcine xenogeneic composition that does not comprise porcine liver tissue.

[0074] Examples of tumors that can be treated with the methods of the present invention include, but are not limited to, sarcomas, carcinomas, lymphomas, breast tumors, prostate tumors, head and neck tumors, glioblastomas, bladder tumors, pancreatic tumors, liver tumors, ovarian tumors, colorectal tumors, lung tumors, skin tumors, lymphoid tumors, gastrointestinal tumors, gastrointestinal stromal tumors, cervical tumors, hepatocellular carcinomas, renal cell carcinomas, melanomas, colorectal carcinomas, esophageal carcinomas, brain tumors, renal tumors, lung tumors (including non-small cell lung cancer), gastric tumors, bile duct tumors, uterine tumors, and infantile (pediatric) tumors.

[0075] In certain embodiments, methods are provided here for treating a neoplastic disease in a subject in need thereof, the method comprising administering (for example, by intratumoral injection) a xenogeneic composition described in Section 5.1.

[0076] The xenogeneic compositions, as described herein, may be administered to a subject by any route of administration, including, but not limited to, intratumoral, intravenous, intradermal, subcutaneous, intramuscular, or intranodal administration. In one embodiment, administration is via parenteral administration. In another embodiment, administration is via intratumoral administration. In this intratumoral administration, the xenogeneic composition is injected directly into the tumor. In yet another embodiment, administration is via intracerebral administration. 5.3.1 Intratumoral Administration

[0077] In certain embodiments, the xenogeneic composition as described herein is administered to a subject by means of one or more intratumoral injections directly into a tumor. This direct administration of Petition 870250092182, dated 08 / 10 / 2025, page 30 / 72 The 20 / 39 therapeutic composition in targeted tumor lesions offers several advantages over systemic administration, including higher local concentrations and potentially reduced systemic toxicities.

[0078] In one modality, the injection is into a single tumor. In another modality, the injection is made into different sections of the same tumor. In yet another modality, the injection is into multiple tumors. The ability to inject into multiple sites may lead to a more robust adaptive immune response (e.g., in patients with polyclonal metastases). In some modalities, the injection is into a primary tumor. In other modalities, the injection is into one or more metastatic tumors.

[0079] In one modality, intratumoral injection is an image-guided intratumoral injection. Intratumoral injection can be performed using imaging techniques, including, but not limited to, ultrasound, fluoroscopy, and CT scanning.

[0080] The injection technique, including the design and placement of the injection needle used and the rate of administration of the therapeutic composition, used to perform intratumoral injection may influence the therapeutic effect (e.g., immune response). In certain modalities, intratumoral injection is performed by placing the needle (e.g., using image guidance) into the target tumor, ensuring that the therapeutic composition is distributed throughout the target tumor without leakage into the surrounding tissue (i.e., without leakage outside the target). In one modality, intratumoral injection is performed by placing the needle tip (e.g., using image guidance) within the center of the target tumor.

[0081] Intratumoral injection can be performed using a needle of appropriate gauge. The appropriate needle gauge depends on a variety of factors, including, but not limited to, the type of target tumor and its location and size. In certain modalities, intratumoral injection is performed using an 18-gauge, 19-gauge, 20-gauge, 21-gauge, 22-gauge, 23-gauge, 24-gauge, 25-gauge, 26-gauge, 27-gauge, 28-gauge, 29-gauge, or 30-gauge needle. Petition 870250092182, dated 08 / 10 / 2025, page 31 / 72 21 / 39 In one modality, the intratumoral injection is performed using a 20-gauge needle. In one modality, the intratumoral injection is performed using a 21-gauge needle. In one modality, the intratumoral injection is performed using a 22-gauge needle.

[0082] In certain embodiments, intratumoral injection is performed using a hole-in-the-hole needle (EHN) having an appropriate needle gauge. In one embodiment, intratumoral injection is performed using a 20-gauge, 21-gauge, or 22-gauge hole-in-the-hole needle (EHN) (e.g., marketed by Becton Dickinson).

[0083] In certain embodiments, intratumoral injection is performed using a multi-hole needle (MSHN) having an appropriate needle gauge. Placement may be performed under image guidance, ensuring that all lateral holes are positioned within the tumor. In one embodiment, intratumoral injection is performed in such a way that there is no leakage outside the target of the therapeutic composition. In one embodiment, intratumoral injection is performed using a 20-gauge, 21-gauge, or 22-gauge multi-hole needle (MSHN). In one embodiment, intratumoral injection is performed using a 21-gauge multi-hole needle (MSHN) without a hole at the tip (e.g., ProFusion™ therapeutic infusion needle marketed by Cook Regentec). 5.3.2 Dosages and Dosing Frequency

[0084] The xenogeneic compositions of the present invention can be administered (e.g., by intratumoral injection) in a manner appropriate to the disease being treated. Dosage amounts and frequency of administration will be determined by factors such as the route of administration, the patient's condition, and the type and severity of the patient's disease, although appropriate dosages may be determined based on clinical trials.

[0085] In certain embodiments, xenogeneic compositions comprising antigen-presenting cells, as described herein, may be administered (e.g., by intratumoral injection) in doses Petition 870250092182, dated 08 / 10 / 2025, page 32 / 72 22 / 39 single or multiple doses. In one embodiment, the composition is administered as an initial dose. In another embodiment, the composition is administered under a booster regimen (i.e., repeated treatment). In yet another embodiment, the composition is administered under a booster regimen (i.e., repeated treatment), but at a different tumor site.

[0086] In certain modalities, multiple doses may be administered on the same day, essentially at the same time, immediately after the previous dose (i.e., consecutive administration). Alternatively, multiple doses may be administered on the same day, but not at the same time (i.e., non-consecutive administrations). In these cases, multiple doses may be 2 doses, 3 doses, 4 doses, 5 doses, 6 doses, 7 doses, 8 doses, 9 doses, or 10 doses.

[0087] In certain embodiments, administration of the xenogeneic composition as described herein may be repeated and spaced out by at least 1 day, 2 days, 3 days, 4 days, 5 days, 7 days, 10 days, 15 days, 30 days, 45 days, 2 months, 75 days, 3 months, or at least 6 months. In some embodiments, dosing may be discontinued for a predetermined number of days.

[0088] In certain embodiments, the composition as described herein is administered once weekly for a predetermined number of weeks. In one embodiment, the scheduled number of weeks is 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks, 9 weeks, or 10 weeks. In some embodiments, dosing may be discontinued for at least one week. In certain embodiments, the composition is administered as a single dose followed by a second dose one to six weeks later. In certain embodiments, the composition may be administered at intervals of six to twelve months.

[0089] When the composition described herein is administered multiple times, the individual doses may be essentially the same dosage amount or different dosage amounts. Furthermore, the composition may be administered (e.g., by intratumoral injection) in the same Petition 870250092182, dated 08 / 10 / 2025, page 33 / 72 23 / 39 tumor site or in sites other than the tumor.

[0090] The xenogeneic compositions of the present invention can be administered in varying dosage amounts, depending, inter alia, on the route of administration, the patient's condition, and the type and severity of the patient's disease, although appropriate dosages can be determined based on clinical trials.

[0091] In certain embodiments, the xenogeneic composition is administered by means of one or more injections (e.g., intratumoral) of 1 x 10⁶ to 10 x 10⁶, 10 x 10⁶ to 20 x 10⁶, 20 x 10⁶ to 30 x 10⁶, 30 x 10⁶ to 40 x 10⁶, 40 x 10⁶ to 50 x 10⁶, 50 x 10⁶ to 60 x 10⁶, 60 x 10⁶ to 70 x 10⁶, 70 x 10⁶ to 80 x 10⁶, 80 x 10⁶ to 90 x 10⁶, 90 x 10⁶ to 10 x 10⁷, 10 x 10⁷ to 20 x 10⁷, 20 x 10⁷ to 30 x 107, 30 x 107a40 x 107, 40 x 107a 50 x 107, 50 x 107a 60 x 107, 60 x 107a 70 x 107, 70 x 107a80 x 107, 80 x 107a 90 x 107, 90 x 107a 10 x 108, 10 x 108a 20 x 108, 20 x 108a30 x 108, 30 x 108a 40 x 108, 40 x 108a 50 x 108 antigen-presenting cells per dose.

[0092] In certain embodiments, the xenogeneic composition is administered via one or more injections (e.g., intratumoral) of approximately 1 x 10⁶, approximately 5 x 10⁶, approximately 10 x 10⁶, approximately 15 x 10⁶, approximately 20 x 10⁶, approximately 25 x 10⁶, approximately 30 x 10⁶, approximately 35 x 10⁶, approximately 40 x 10⁶, approximately 45 x 10⁶, approximately 50 x 10⁶, approximately 55 x 10⁶, approximately 60 x 10⁶, approximately 65 x 10⁶, approximately 70 x 10⁶, approximately 75 x 10⁶, approximately 80 x 10⁶, approximately 85 x 10⁶, approximately 90 x 10⁶, approximately 95 x 106, about 10 x 107, about 15 x 107, about 20 x 107, about 25 x 107, about 30 x 107, about 35 x 107, about 40 x 107, about 45 x 107, about 50 x 107, about 55 x 107, about 60x107, about 65x107, about 70x107, about 75x107, about 80x107, about 85x107, about 90x107, about 95x107, about 10x108, about 15x108, about 20x 108, about 25 x 108, about 30 x 108, about 35 x 108, about 40 x 108, about 45 x 108,or approximately 50 x 10⁸ antigen-presenting cells per dose. In certain modalities, the xenogeneic composition is administered via one or more injections (e.g., intratumoral) of, Petition 870250092182, dated 08 / 10 / 2025, page 34 / 72 24 / 39 approximately 100,000 and / or approximately 500,000 antigen-presenting cells per dose. In some embodiments, the antigen-presenting cells in such compositions are mature antigen-presenting cells. In some embodiments, the antigen-presenting cells in such compositions are mature antigen-presenting cells in combination with immature antigen-presenting cells.

[0093] A single dose of antigen-presenting cells can be administered via one or more injections (e.g., intratumoral). In certain modalities, the xenogeneic composition is administered via one or more injections (e.g., intratumoral) of 1 x 10⁶ to 10 x 10⁶, 10 x 10⁶ to 20 x 10⁶, 20 x 10⁶ to 30 x 10⁶, 30 x 10⁶ to 40 x 10⁶, 40 x 10⁶ to 50 x 10⁶, 50 x 10⁶ to 60 x 10⁶, 60 x 10⁶ to 70 x 10⁶, 70 x 10⁶ to 80 x 10⁶, 80 x 10⁶ to 90 x 10⁶, 90 x 10⁶ to 10 x 10⁷, 10 x 10⁷ to 20 x 10⁷, 20 x 10⁷ to 30 x 10⁷. 30 x 107a 40 x 107, 40 x 10 to 50 x 107, 50 x 107a 60 x 107, 60 x 107a 70 x 107, 70 x 107a 80 x 107, 80 x 10 to 90 x 107, 90 x 107a 10 x 108, 10 x 108a 20 x 108, 20 x 108a 30 x 108, 30 x 10 to 40 x 108, 40 x 108a 50 x 108 antigen-presenting cells per injection. In certain modalities, the xenogeneic composition is administered via one or more injections (e.g., intratumoral) of approximately 1 x 10⁶, approximately 5 x 10⁶, approximately 10 x 10⁶, approximately 15 x 10⁶, approximately 20 x 10⁶,about 25x106, about 30x106, about 35x106, about 40x106, about 45x106, about 50x106, about 55x106, about 60x106, about 65x106, about 70x106, about 75x 106, about 80 x 106, about 85 x 106, about 90 x 106, about 95 x 106, about 10 x 107, about 15 x 107, about 20 x 107, about 25 x 107, about 30 x 107, about 35 x 107, about 40 x 107, about 45 x 107, approximately 50 x 107, approximately 55 x 107, approximately 60 x 107, approximately 65 x 107, approximately 70 x 107, approximately 75 x 107, approximately 80 x 107, approximately 85 x 107, approximately 90 x 107, approximately 95 x 107, approximately 10 x 108, approximately 15 x 108, approximately 20 x 108, approximately 25 x 108, approximately 30 x 108, approximately 35 x 108, approximately 40 x 108, approximately 45 x 108, or approximately 50 x 108 antigen-presenting cells per injection. In certain embodiments, the xenogeneic composition is administered by, Petition 870250092182, dated 08 / 10 / 2025, page 35 / 72 25 / 39 by means of one or more injections (e.g., intratumoral) of approximately 100,000 and / or approximately 500,000 antigen-presenting cells per injection. 5.3.3 Clinical Results

[0094] The therapeutic effect observed after administration of the xenogeneic composition of the present invention, individually or in combination with other anticancer therapy, can be evaluated by any of the methods commonly used in anticancer therapies. In certain modalities, administration of the xenogeneic composition triggers a specific immune response to the target tumor.

[0095] Furthermore, treatment with the xenogeneic composition described herein, individually or in combination with other anticancer therapy, may reduce not only the target tumor but also lead to the reduction of untreated tumors in other parts of the body. This result is known as the abscopal effect. Thus, treatment with the xenogeneic composition described herein, individually or in combination with other anticancer therapy, may induce a systemic antitumor immune response. In certain modalities, administration of the xenogeneic composition described herein via targeted intratumoral injection leads to an abscopal effect. In certain modalities, administration of the xenogeneic composition described herein via targeted intratumoral injection in combination with other anticancer therapy (e.g., tumor treatment fields (TTFs) or radiation) leads to an abscopal effect.

[0096] In certain embodiments, the therapeutic effect observed after administration of the xenogeneic composition of the present invention, individually or in combination with other anticancer therapy, is evaluated by measuring the immune response. In certain embodiments, the immune response is evaluated by measuring immunological markers in the blood. In certain embodiments, the tumor-specific immune response is evaluated. In certain embodiments, the systemic immune response is evaluated. The immune response can be evaluated through Petition 870250092182, dated 08 / 10 / 2025, page 36 / 72 26 / 39 • screening cells (which may involve staining PBMCs with specific antibodies, for example, for an HLA class I antigen or an HLA class II antigen that is selectively expressed in donor vaccine cells); • to assess dendritic cell-induced alloimmunization (which may involve screening for alloantibodies against, for example, HLA-A, B, C (MHC-class I) and HLA-DR, DQ, DP (MHC-class II) antigens); • monitor autoimmune events (which may involve screening for autoantibodies against autoantigens, including nuclear antigens (e.g., ANA, SSA, SSB, Sm, RNP, Scl-70, centromeres, and Jo-1) and autoantigens associated with liver parenchyma (e.g., liver-kidney microsomal antigens and mitochondrial antigens) • assess complement activation and / or classical / alternative complement function; and / or • assess the occurrence and activation status of immune cells (e.g., CD3+, CD3+4+, and CD3+8+ T cells, CD19+ B cells, CD3-16+56+ NK cells, CD3-16+56+69+ NK cells, CD3+16+56+ NKT cells, CD3+16+56+69+ NKT cells, and CD3+HLA-DR+ T cells).

[0097] In certain embodiments, the therapeutic effect observed after administration of the xenogeneic composition of the present invention, individually or in combination with other anticancer therapy, is evaluated by assessing the size of the tumor(s). This can be done after 3 and / or 6 months of treatment administration, or as deemed necessary based on tumor progression.

[0098] In certain embodiments, the therapeutic effect observed after administration of the xenogeneic composition of the present invention, individually or in combination with other anticancer therapy, is evaluated by assessing tumor control. In some embodiments, tumor control is evaluated by CT and / or MRI scans, measuring the number of tumor-specific T cells, measuring AFP (alpha-fetoprotein) levels in the blood, Petition 870250092182, dated 08 / 10 / 2025, page 37 / 72 27 / 39 measuring the level of circulating tumor cells or any combination thereof.

[0099] In certain embodiments, the therapeutic effect observed after administration of the xenogeneic composition of the present invention, individually or in combination with other anticancer therapy, is evaluated by assessing the systemic inflammatory response. This can be done by evaluating, for example, the systemic release of relevant cytokines, chemokines and other inflammatory parameters in the blood (e.g., IL-1R, IL-2, IL-4, IL-5, IL-6, IL-7, IL-8, IL-10, IL-12p70, IL-13, IL-17A, G-CSF, GM-CSF, IFN-gamma, MCP-1, MIP1 beta and TNF-alpha).

[0100] In certain modalities, the therapeutic effect is assessed by evaluating long-term changes in Eastern Cooperative Oncology Group (ECOG) scores and / or Karnofsky Performance Status (KPS) scores. In certain modalities, the therapeutic effect is assessed by evaluating long-term changes in Quality of Life scores.

[0101] In certain modalities, the therapeutic effect is evaluated in terms of partial response and / or complete response. In certain modalities, the therapeutic effect is evaluated in terms of progression-free survival and / or overall survival. In certain modalities, progression-free survival and / or overall survival is evaluated by measuring blood parameters, for example, by measuring lactate dehydrogenase (LDH) levels and the neutrophil-to-lymphocyte-derived ratio (dNLR). Elevated LDH and dNLR levels may be associated with worse survival outcomes in patients treated with immunotherapy. In certain modalities, administration of the xenogeneic composition of the present invention, individually or in combination with other anticancer therapy, results in reduced LDH and dNLR levels relative to baseline.

[0102] In certain modalities, the therapeutic effect is evaluated in terms of adverse events, recorded as a measure of safety and tolerability, which may include changes in baseline vital signs (e.g., heart rate, blood pressure, body temperature), changes in baseline laboratory parameters, etc. In a Petition 870250092182, dated 08 / 10 / 2025, page 38 / 72 28 / 39 In the 28 / 39 modality, the administration of the xenogeneic composition of the present invention, individually or in combination with other anticancer therapy, is not associated with serious adverse events. 5.3.4 Combination Treatment

[0103] When used in cancer treatment, the xenogeneic compositions described herein may be combined with other therapies. In some modalities, the xenogeneic composition is administered in combination with therapies that decrease the immunosuppressive tumor environment or activate the immune system.

[0104] In certain embodiments, the xenogeneic composition is administered (e.g., by intratumoral injection) in combination with one or more other anticancer therapies. Examples of other anticancer therapies include, but are not limited to, anti-CTLA4 therapy, anti-PD1 therapy, anti-PDL1 therapy, anti-LAG-3 therapy, tumor treatment fields (TTFs), cell-based therapy, a tyrosine kinase inhibitor, a VEGF inhibitor, or any combination thereof. In one embodiment, the other anticancer therapy comprises treatment with one or more biological products. In one embodiment, the other anticancer therapy comprises treatment with cell therapy products or gene therapy products. In one embodiment, the other anticancer therapy is CAR T-cell-based therapy. In one embodiment, the other anticancer therapy comprises treatment with T-cell couplers.In one embodiment, the other anticancer therapy comprises treatment with one or more immune checkpoint inhibitors. In one embodiment, the other anticancer therapy comprises treatment with one or more small molecule drugs. In one embodiment, the other anticancer therapy comprises treatment with one or more protein kinase inhibitors. In one embodiment, the other anticancer therapy comprises treatment with one or more tyrosine kinase inhibitors. In one embodiment, the other anticancer therapy comprises treatment with one or more alkylating agents, antimetabolites, natural products, or hormones. In one embodiment, the other. Petition 870250092182, dated 08 / 10 / 2025, page 39 / 72 29 / 39 anticancer therapy comprises treatment with gemcitabine or 5-fluorouracil. In one modality, the other anticancer therapy comprises surgery or radiotherapy. In one modality, the other anticancer therapy comprises field tumor treatment.

[0105] In certain modalities, the other anticancer therapy comprises treatment with imatinib, sunitinib, regorafenib, pazopanib, nilotinib, avapritinib, ripretinib, sorafenib, pimitespib, ipilimumab, tremelimumab, nivolumab, pembrolizumab, cemiplimab, atelizumab, avelumab, durvalumab, relatlimab, or any combination thereof.

[0106] The combined treatments described herein may provide “synergy” and prove to be “synergistic,” meaning that the therapeutic effect achieved after administering two or more individual therapies used together is greater than the sum of the therapeutic effects achieved after administering the individual therapies separately. Such synergy can be determined by methods commonly known in the art. In certain modalities, the combined treatment (i.e., administration of the xenogeneic composition of the present invention in combination with another anticancer therapy) provides a synergistic antitumor effect.

[0107] Appropriate dosages of the therapies and / or individual compositions may be reduced due to combined action (synergy), in order to increase the therapeutic index or attenuate toxicity or other side effects or consequences. 5.4 Pharmaceutical Compositions

[0108] Pharmaceutical compositions comprising the xenogeneic compositions described in Section 5.1 are provided here.

[0109] In certain embodiments, pharmaceutical compositions comprising xenogeneic compositions in combination with one or more pharmaceutically or physiologically acceptable vehicles, diluents and / or excipients are provided herein. Such pharmaceutical compositions may be formulated in accordance with standard procedures in the art. In a Petition 870250092182, dated 08 / 10 / 2025, page 40 / 72 30 / 39 modality, the pharmaceutical composition is in the form of an aqueous solution.

[0110] In certain embodiments, the pharmaceutical composition comprises one or more pharmaceutically acceptable carriers. In some embodiments, the pharmaceutically acceptable carrier is a phosphate-buffered saline solution, water, or an emulsion, such as an oil-in-water or water-in-oil emulsion. In certain embodiments, the pharmaceutical composition is formulated as an aqueous solution, for example, in physiologically compatible buffers such as Hanks' solution, Ringer's solution, or physiological saline buffer.

[0111] In certain embodiments, the pharmaceutical composition comprises adjuvants, excipients, stabilizers, preservatives and / or other pharmaceutically acceptable components known in the art. In one embodiment, the pharmaceutical composition comprises a humectant. In some embodiments, the pharmaceutical composition comprises one or more suspending, stabilizing and / or dispersing agents. In some embodiments, the pharmaceutical composition comprises one or more emulsifiers. In certain embodiments, the pharmaceutical composition is sterilized. In one embodiment, the pharmaceutical composition comprises one or more preservatives. In another embodiment, the pharmaceutical composition does not contain a preservative.

[0112] In certain embodiments, the pharmaceutical compositions provided herein may comprise buffers, such as neutral buffered saline solution, phosphate-buffered saline solution and the like; carbohydrates, such as glucose, mannose, sucrose or dextrans, mannitol; proteins; polypeptides or amino acids, such as glycine; antioxidants; chelating agents, such as EDTA or glutathione; adjuvants (e.g., aluminum hydroxide); and / or preservatives.

[0113] In certain embodiments, the pharmaceutical composition comprising the antigen-presenting cells, as described herein, is in the form of a suspension or dispersion. Such a suspension or dispersion may be prepared by conventional dispersion and suspension processes and may contain one or more pharmaceutically acceptable excipients. In one embodiment, the pharmaceutical composition is a suspension or dispersion. Petition 870250092182, dated 08 / 10 / 2025, page 41 / 72 31 / 39 aqueous isotonic. In one embodiment, the suspension or dispersion comprises one or more viscosity regulating agents.

[0114] In certain embodiments, the pharmaceutical composition comprising the xenogeneic composition as described herein is formulated in the form of a hydrogel. In some embodiments, the hydrogel is a multidomain peptide-based hydrogel. These multidomain peptide-based hydrogels can be prepared by methods known in the art. In certain embodiments, antigen-presenting cells incorporated into a multidomain peptide-based hydrogel exhibit significantly improved delivery and retention within tumors. In certain embodiments, the pharmaceutical composition is a sustained-release hydrogel-based composition.

[0115] In certain embodiments, the xenogeneic compositions described herein are formulated in a pharmaceutical composition with diluents and / or with other components, such as IL-2 or other cytokines or cell populations.

[0116] The pharmaceutical compositions described herein can be maintained at temperatures around 2-8 °C, or can be frozen and thawed shortly before use. In certain embodiments, the pharmaceutically acceptable vehicle may be a frozen cell preservation medium. For example, the xenogeneic compositions comprising antigen-presenting cells described herein may be frozen in heat-inactivated universal donor plasma comprising dimethyl sulfoxide (DMSO) to enable their storage. Such a medium comprising the antigen-presenting cells may be used directly, for example, injected intratumorally, once thawed. Alternatively, cells frozen in such a medium may be thawed, washed, and resuspended in an appropriately buffered saline solution or in a saline solution containing human serum albumin before being administered, for example, injected intratumorally.In one embodiment, the vehicle is pharmaceutically acceptable. Petition 870250092182, dated 08 / 10 / 2025, page 42 / 72 32 / 39 is a saline solution comprising human serum albumin. In another embodiment, the pharmaceutically acceptable vehicle is a physiological sodium chloride solution comprising 2% human serum albumin.

[0117] The pharmaceutical compositions provided herein may be formulated for any route of administration, including, but not limited to, intratumoral, intravenous, intradermal, subcutaneous, intramuscular, or intranodal administration. In one embodiment, the pharmaceutical composition is formulated for parenteral administration. In another embodiment, the pharmaceutical composition is formulated for intratumoral administration. In intratumoral administration, the pharmaceutical composition is injected directly into the tumor. In a specific embodiment, the pharmaceutical composition as described herein is administered by one or more intratumoral injections directly into a tumor. The injection may be applied to a single tumor or to more than one tumor.

[0118] The pharmaceutical compositions of the present invention can be administered in a manner appropriate to the disease being treated. Dosage amounts and frequency of administration will be determined by factors such as the route of administration, the patient's condition, and the type and severity of the patient's disease, although appropriate dosages may be determined based on clinical trials.

[0119] The following examples are offered for illustrative purposes only and not as a limitation. 6. EXAMPLES 6.1 Example 1: Xenogeneic compositions comprising porcine antigen-presenting cells 6.1.1 Isolation of Mononuclear Cells from Peripheral Blood

[0120] Pig blood (total volume of ~100 ml) collected in EDTA vacutainers (BD Biosciences, San Diego, USA) was diluted 1:1 with phosphate-buffered saline (PBS). 30 ml of the diluted blood were carefully layered over 20 ml of Ficoll Paque Plus (Cytiva, Uppsala, Sweden) in a 50 ml Falcon tube. Density gradient centrifugation was performed. Petition 870250092182, dated 08 / 10 / 2025, page 43 / 72 Centrifugation of 33 / 39 cells was performed at 600xg for 20 min with the centrifuge brakes off. After centrifugation, peripheral blood mononuclear cells (PBMCs) were collected from the interface and washed three times with PBS. Each wash consisted of a 5 min centrifugation step at 350xg followed by replacement of the supernatant with fresh PBS. After the final wash, the PBMCs were resuspended in 5 ml of ACK lysis buffer (Gibco, Thermo Fisher Scientific Inc., Waltham, USA) and incubated for 5 min. After red blood cell lysis, the PBMCs were centrifuged and resuspended in AIM V medium (Gibco, Thermo Fisher Scientific Inc., Waltham, USA) supplemented with 10% heat-inactivated fetal bovine serum (FBS; Gibco, Thermo Fisher Scientific Inc., Waltham, USA). Human PBMCs were similarly purified from leukocyte layers of anonymous healthy donors obtained from the Karolinska University Hospital Blood Bank (Stockholm, Sweden). 6.1.2 Isolation of porcine CD14+ cells

[0121] Porcine CD14+ cells were isolated by positive selection of magnetic beads using a cross-matched human antiCD14 MicroBeads isolation kit (Miltenyi, Bergisch Gladbach, Germany), according to the manufacturer's instructions. In two separate purifications from 1x10⁸ porcine PBMCs, the yield of positively selected CD14+ was 20% of the initial cell count. PBMC cell viability and CD14+ fraction after positive selection was >93% when analyzed using trypan blue exclusion (Gibco, Thermo Fisher Scientific Inc., Waltham, USA). 6.2 Example 2: In vivo experiment in mice using human PBMCs - murine melanoma model

[0122] Human peripheral blood mononuclear cells (PBMCs) were isolated from peripheral blood using the Ficoll density gradient centrifugation method.

[0123] PBMCs were then labeled with CFSE (carboxyfluorescein succinimidyl ester). CFSE-labeled PBMCs were injected intratumorally at a dose of approximately 1 x 106 cells. Petition 870250092182, dated 08 / 10 / 2025, page 44 / 72 34 / 39 in C57BL / 6J mice expressing the B16 melanoma cell line.

[0124] The comparison of tumor size between the treatment and control groups was performed by measuring the tumor volume on day 12. The results are shown in Table 1 below and in FIG 2. Table 1: Average tumor size Treatment (n = 12) 265.8 mm3 Control (n = 18) 538.2 mm3 6.3 Example 3: In vivo experiment in mice using human PBMCs - murine bladder cancer model

[0125] Human PBMCs were isolated from peripheral blood using the Ficoll density gradient centrifugation method.

[0126] PBMCs were then labeled with CFSE (carboxyfluorescein succinimidyl ester). CFSE-labeled PBMCs were injected intratumorally at a dose of approximately 1 x 106 cells into C57BL / 6J mice expressing the MB49 carcinogen-induced bladder cancer cell line.

[0127] The comparison of tumor size between the treatment and control groups was performed by measuring the tumor volume on day 7. The results are shown in the table below and in FIG. 3. Table 2: Average tumor size: Treatment (n = 10) 132.0 mm3; Control (n = 18) 310.6 mm3 6.4 Example 4: Mixed lymphocyte reaction

[0128] Human PBMCs were stained with Violet Proliferation Dye 450 (VPD450; BD Biosciences, San Diego, USA) following the manufacturer's instructions. In the first xenogeneic Mixed Lymphocyte Reaction (MLR) study, human PBMCs stained with VPD450 were mixed with non-porcine PBMCs. Petition 870250092182, dated 08 / 10 / 2025, p. 45 / 72 35 / 39 stained in various ratios. Specifically, starting with a 10:10 ratio of human to pig cells and proceeding with a fixed number of human cells combined with a gradual decrease in the number of pig cells to reach a 10:1 ratio of human to pig cells. The wells with a 10:10 ratio contained a combination of 2x10⁵ human PBMCs and 2x10⁵ pig PBMCs, while the wells with a 10:1 human to pig cell ratio contained a combination of 2x10⁵ human PBMCs and 2x10⁴ pig PBMCs. In the second xenogeneic study, VPD450-stained human PBMCs were mixed with unstained pig CD14+ cells in various ratios, as described above. All samples were resuspended in 96-well round-bottom cell culture plates in 200 µL of AIM V medium supplemented with 10% heat-inactivated FBS and incubated at 37°C and 5% CO2 for seven days. On day five, 100 µL of fresh culture medium was added to each well.Both xenogeneic MLR studies included two (n=2) PBMC donors matched separately with a single porcine donor. Cells from no donor / species were inactivated via irradiation / chemical treatment, resulting in a bidirectional xenogeneic MLR; however, only human cells were labeled with the proliferation dye. Control wells included allogeneic MLR consisting of 2x10⁴ human PBMCs matched with 2x10⁴ human PBMCs from an unrelated donor. Additional control wells received 2x10⁴ human PBMCs from a single donor.

[0129] On day seven of MLR, cells were washed twice in saline solution and incubated with Fc receptor binding inhibitor (Invitrogen; Thermo Fisher Scientific Inc., Waltham, USA), followed by anti-CD3 VioGreen staining (Miltenyi, Bergisch Gladbach, Germany; Clone REA613), anti-CD45RA BV650 staining (BD Biosciences, San Diego, USA; Clone HI100), anti-CD8 BB515 staining (BD Biosciences, San Diego, USA; Clone RPA-T8), anti-CCR7 PE staining (Miltenyi, Bergisch Gladbach, Germany; Clone REA546), anti-HLA-DR PerCPCy5.5 staining (BD Biosciences, San Diego, USA; Clone G46-6), and anti-CD4 APC-Vio770 staining. Petition 870250092182, dated 08 / 10 / 2025, page 46 / 72 36 / 39 (Miltenyi, Bergisch Gladbach, Germany; Clone REA623) and anti-CD56 APC (Miltenyi, Bergisch Gladbach, Germany; Clone REA196). Samples were stained while resuspended in staining buffer (FBS) (BD Biosciences, San Diego, USA), incubated in the dark at 4°C, and after two washing steps, acquired using a Celesta FACS flow cytometer (BD Biosciences, San Diego, USA). Post-acquisition data analysis was conducted using FlowJo 10.8.1 software (FlowJo LLC, Ashland, USA). 6.5 Example 5: In vitro study comparison: porcine PBMC vs human PBMC

[0130] Porcine PBMCs were plated with human PBMCs in varying human-to-porcine cell ratios (h:p), as described in the MLR study above. The human-to-porcine cell ratios (h:p) were h:p = 10:10, 10:9, 10:8, 10:7, 10:6, 10:5, 10:4, 10:3, 10:2, and 10:1. Separately, human PBMCs alone and allogeneic human PBMCs were evaluated as reference samples.

[0131] Human CD4 T cell counts and human CD8 T cell counts were assessed on day 7 for all samples, including reference samples. The results are shown in FIG. 4A and FIG. 4B, respectively.

[0132] Human NK cell counts were assessed on day 7 for all samples, including reference samples. Results are shown in FIG. 4C. 6.6 Example 6: In vitro study comparison: porcine monocytes (CD14+ cells) vs human PBMCs

[0133] Porcine monocytes (containing CD14+ cells) were plated with human PBMCs in varying human-to-porcine cell ratios (h:p), as described in the MLR study above. The human-to-porcine cell ratios (h:p) were h:p = 10:10, 10:9, 10:8, 10:7, 10:6, 10:5, 10:4, 10:3, 10:2, and 10:1. Separately, human PBMCs alone and allogeneic human PBMCs were evaluated as reference samples. Petition 870250092182, dated 08 / 10 / 2025, page 47 / 72 37 / 39

[0134] Human CD4 T cell counts and human CD8 T cell counts were assessed on day 7 for all samples, including reference samples. The results are shown in FIG. 5A and FIG. 5B, respectively.

[0135] Human proliferating CD4 T cell counts and human proliferating CD8 T cell counts were also evaluated on Day 7 for all samples, including reference samples. The results are shown in FIG. 5C and FIG. 5D, respectively.

[0136] HLA-DR in human CD4 T cells and human CD8 T cells, in terms of mean fluorescence intensity (MFI), was evaluated on day 7 for all samples, including reference samples. The results are shown in FIG. 5E and FIG. 5F, respectively.

[0137] Human NK cell counts were assessed on day 7 for all samples, including reference samples. Results are shown in FIG. 5G. 6.7 Example 7: In vivo experiment in mice using porcine PBMCs

[0138] Porcine PBMCs are injected intratumorally at a dose of 1 x 106 cells into C57BL / 6J mice expressing the B16 melanoma cell line. Tumor size comparison between the treatment and control groups is performed by measuring tumor volume on day 7 or later.

[0139] A similar experiment is conducted in C57BL / 6J mice expressing the carcinogen-induced bladder cancer cell line MB49. 6.8 Example 8: Intratumoral administration of porcine PBMCs to human patients

[0140] This example evaluates the safety and / or efficacy of a composition comprising porcine antigen-presenting cells administered intratumorally as a single agent or in combination with other anticancer therapy in cancer patients. Petition 870250092182, dated 08 / 10 / 2025, page 48 / 72 38 / 39

[0141] Patients are divided into two treatment groups. The first group of patients will receive the treatment composition as monotherapy, while the second group of patients will receive the treatment composition in combination with a checkpoint inhibitor or a tyrosine kinase inhibitor. Patients will be treated with the treatment composition at increasing dose and / or frequency, starting with a dose of 10 x 10⁶ cells per injection. Patients may or may not have been previously treated with anticancer therapy. Some outcome measures are provided below.

[0142] Adverse events will be recorded as a measure of safety and tolerability, which will include changes in baseline vital signs (e.g., heart rate, blood pressure, body temperature), changes in baseline laboratory parameters, etc.

[0143] The immune response will be assessed by measuring immunological markers in the blood. The size of the tumor(s) will be assessed after 3 and 6 months or as needed based on tumor progression.

[0144] The systemic inflammatory response will be assessed, including the potential systemic release of cytokines, chemokines, and other relevant inflammatory parameters in the blood (e.g., IL-1R, IL-2, IL-4, IL-5, IL-6, IL-7, IL-8, IL-10, IL-12p70, IL-13, IL-17A, G-CSF, GM-CSF, IFN-gamma, MCP-1, MIP1 beta, and TNF-alpha).

[0145] Tumor control will be assessed, which will include CT scans / MRI scans, measurement of the number of tumor-specific T cells, measurement of AFP (alpha-fetoprotein) levels in the blood and / or measurement of the level of circulating tumor cells.

[0146] Tumor-specific immune responses and systemic immune response will be evaluated.

[0147] Long-term changes in Eastern Cooperative Oncology Group (ECOG) scores and / or Karnofsky Performance Status (KPS) scores will be assessed. In addition, long-term changes in Petition 870250092182, dated 08 / 10 / 2025, page 49 / 72 39 / 39 Quality of Life scores.

[0148] The therapeutic effect will be evaluated in terms of partial response, complete response, progression-free survival and / or overall survival. 7. Equivalents

[0149] Although the disclosure is described in detail with reference to its specific embodiments, it should be understood that variations that are functionally equivalent are within the scope of this disclosure. In fact, several modifications of the disclosure beyond those shown and described herein will become apparent to those skilled in the art from the preceding description. Such modifications are intended to be within the scope of the appended claims. Those skilled in the art will recognize, or will be able to verify the use of, by no more than routine experimentation, many equivalents to the specific embodiments of the disclosure described herein. Such equivalents are intended to be covered by the following claims.

[0150] All publications, patents and patent applications mentioned in this report are incorporated herein by reference in the descriptive report to the same extent as if each individual publication, patent or patent application were specifically and individually indicated to be incorporated herein by reference in its entirety. Petition 870250092182, dated 08 / 10 / 2025, pp. 50 / 72

Claims

1 / 6 CLAIMS 1. A method for treating a tumor in a subject in need thereof, the method characterized in that it comprises: administering a composition comprising antigen-presenting cells to the subject, wherein the antigen-presenting cells are obtained from a species that is different from the subject.

2. Method according to claim 1, characterized in that the tumor is a solid tumor and the administration is into the subject's tumor via an intratumoral injection.

3. A method, according to any one of claims 1 to 2, characterized in that the antigen-presenting cells are obtained from a species that is a pig.

4. A method according to any one of claims 1 to 3, characterized in that the antigen-presenting cells are obtained from a species that is a miniature pig.

5. A method, according to any one of claims 1 to 4, characterized in that the subject is a human being.

6. Method according to claim 3, characterized in that the pig is a pig deficient in alpha-1,3 galactosyltransferase.

7. Method according to claim 6, characterized in that the alpha-1,3 galactosyltransferase-deficient pig is an inbred pig with porcine leukocyte antigen (SLA).

8. Method according to claim 4, characterized in that the miniature pig is a miniature pig deficient in alpha-1,3 galactosyltransferase.

9. Method, according to claim 8, characterized in that the alpha-1,3 galactosyltransferase deficient pig is an inbred pig with porcine leukocyte antigen (SLA).

10. Method, according to any one of claims 1 to 9, characterized in that the method triggers a specific immune response to the tumor. Petition 870250092182, dated 08 / 10 / 2025, page 51 / 72 2 / 6 11. A method according to any one of claims 1 to 10, characterized in that the method yields an abscopal effect.

12. Method, according to any one of claims 1 to 11, characterized in that the tumor is a solid cancerous tumor.

13. Method according to claim 12, characterized in that the tumor is selected from the group consisting of sarcomas, carcinomas, lymphomas, breast tumors, prostate tumors, head and neck tumors, glioblastomas, bladder tumors, pancreatic tumors, liver tumors, ovarian tumors, colorectal tumors, lung tumors, skin tumors, lymphoid tumors, gastrointestinal tumors, gastrointestinal stromal tumors, cervical tumors, hepatocellular carcinomas, renal cell carcinomas, melanomas, colorectal carcinomas, esophageal carcinomas, brain tumors, renal tumors, lung tumors (including non-small cell lung cancer), gastric tumors, bile duct tumors, uterine tumors, and infantile (pediatric) tumors.

14. A method, according to any one of claims 1 to 13, characterized in that the tumor is resistant to treatment with chemotherapy and / or treatment with immunotherapy.

15. A method according to any one of claims 1 to 14, characterized in that the antigen-presenting cells are derived from one or more pigs or miniature pigs using a leukapheresis procedure, wherein the leukapheresis procedure generates a leucopak containing peripheral blood mononuclear cells, and the leucopak is further fractionated by counterflow elutriation.

16. A method according to any one of claims 1 to 15, characterized in that the composition comprising antigen-presenting cells is substantially free of pathogens.

17. Method, according to any one of claims 1 to 16, characterized in that the antigen-presenting cells are obtained from pig or miniature pig of different genotypes.

18. A method, according to any one of claims 1 to 17, characterized in that the composition comprising antigen-presenting cells is administered in single or multiple doses.

19. A method according to any one of claims 1 to 18, characterized in that the composition comprising antigen-presenting cells is administered via an intratumoral injection of at least about 1 x 10⁶ antigen-presenting cells per dose.

20. Method, according to any one of claims 1 to 19, characterized in that the composition comprising antigen-presenting cells is administered via an intratumoral injection of approximately 1 x 10⁶, approximately 5 x 10⁶, approximately 10 x 10⁶, approximately 15 x 10⁶, approximately 20 x 10⁶, approximately 25 x 10⁶, approximately 30 x 10⁶, approximately 35 x 10⁶, approximately 40 x 10⁶, approximately 45 x 10⁶, approximately 50 x 10⁶, approximately 55 x 10⁶, approximately 60 x 10⁶, approximately 65 x 10⁶, approximately 70 x 10⁶, approximately 75 x 10⁶, approximately 80 x 10⁶, approximately 85 x 10⁶, approximately 90 x 106, approximately 95 x 106, approximately 10 x 107, approximately 15 x 107, approximately 20 x 107, approximately 25 x 107, approximately 30 x 107, approximately 35 x 107, approximately 40 x 107, approximately 45 x 107 or approximately 50 x 107 antigen-presenting cells per dose.

21. A method according to any one of claims 1 to 20, characterized in that the antigen-presenting cells are substantially mature antigen-presenting cells.

22. A method according to any one of claims 1 to 21, characterized in that the antigen-presenting cells are not activated or stimulated.

23. A method according to any one of claims 1 to 22, characterized in that the composition comprises peripheral blood mononuclear cells (PBMCs).

24. Method, according to any one of claims 1 to 23, characterized in that the composition comprises monocytes. Petition 870250092182, dated 08 / 10 / 2025, p. 53 / 72 4 / 6 25. A method, according to any one of claims 1 to 24, characterized in that the composition comprises dendritic cells, macrophages, granulocytes, T cells, B cells and / or NK cells.

26. A method according to any one of claims 1 to 25, characterized in that the composition comprises at least about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95% or more of about 95% of PBMCs, wherein the PBMCs are mature, immature or a combination of mature and immature PBMCs.

27. A method according to any one of claims 1 to 26, characterized in that the composition comprises at least about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95% or more of about 95% monocytes, wherein the monocytes are mature, immature or a combination of mature and immature monocytes.

28. Method according to claim 27, characterized in that the composition comprises a mixture of at least about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95% or more of about 95% of monocytes and dendritic cells, wherein the monocytes and / or dendritic cells in said mixture may be mature, immature or a combination of mature and immature monocytes and / or dendritic cells.

29. Method according to claim 28, characterized in that the mixture comprises 10% to 95%, 10% to 90%, 10% to 80%, 10% to 70%, 10% to 60%, 10% to 50%, 10% to 40%, 10% to 30%, 10% to 20%, 20% to 95%, 20% to 90%, 20% to 80%, 20% to 70%, 20% to 60%, 20% to 50%, 20% to 40%, 20% to 30%, 30% to 95%, 30% to 90%, 30% to 80%, 30% to 70%, 30% to 60%, 30% to 50%, 30% to 40%, 40% to 95%, 40% to 90%, 40% to 80%, 40% to 70%, 40% to 60%, 40% to 50%, 50% to 95%, 50% to 90%, 50% to 80%, 50% to 70%, 50% to 60%, 60% to 95%, 60% to 90%, 60% to 80%, 60% to 70%, 70% to 95%, 70% to 90%, 70% Petition 870250092182, dated 08 / 10 / 2025, page. 54 / 72 5 / 6 to 80%, 80% to 95%, 80% to 90%, 90% to 95% or more than 95% of monocytes.

30. Method according to claim 28, characterized in that the mixture comprises 10% to 30%, 35% to 55%, 60% to 80% or 85% to 95% monocytes.

31. Method according to claim 28, characterized in that the mixture comprises at least about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95% or more than about 95% of monocytes.

32. Method according to claim 28, characterized in that the mixture comprises 10% to 95%, 10% to 90%, 10% to 80%, 10% to 70%, 10% to 60%, 10% to 50%, 10% to 40%, 10% to 30%, 10% to 20%, 20% to 95%, 20% to 90%, 20% to 80%, 20% to 70%, 20% to 60%, 20% to 50%, 20% to 40%, 20% to 30%, 30% to 95%, 30% to 90%, 30% to 80%, 30% to 70%, 30% to 60%, 30% to 50%, 30% to 40%, 40% to 95%, 40% to 90%, 40% to 80%, 40% to 70%, 40% to 60%, 40% to 50%, 50% to 95%, 50% to 90%, 50% to 80%, 50% to 70%, 50% to 60%, 60% to 95%, 60% to 90%, 60% to 80%, 60% to 70%, 70% to 95%, 70% to 90%, 70% to 80%, 80% to 95%, 80% to 90%, 90% to 95% or more than 95% of dendritic cells.

33. Method according to claim 28, characterized in that the mixture comprises 10% to 30%, 35% to 55%, 60% to 80% or 85% to 95% of dendritic cells.

34. Method according to claim 28, characterized in that the mixture comprises at least about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95% or more than about 95% of dendritic cells.

35. Method, according to any of claims 1 to 34, Petition 870250092182, dated 08 / 10 / 2025, pp. 55 / 72 6 / 6 characterized by the fact that the subject is receiving another anticancer therapy.

36. Method according to claim 35, characterized in that the other anticancer therapy comprises treatment with one or more immune checkpoint inhibitors.

37. Method according to claim 35, characterized in that the other anticancer therapy is an anti-CTLA4 therapy, anti-PD1 therapy, anti-PDL1 therapy, anti-LAG-3 therapy, tumor treatment fields (TTFs), cell-based therapy, a tyrosine kinase inhibitor, a VEGF inhibitor, or any combination thereof.

38. A method according to claim 35, characterized in that the other anticancer therapy comprises treatment with imatinib, sunitinib, regorafenib, pazopanib, nilotinib, avapritinib, ripretinib, sorafenib, pimitespib, ipilimumab, tremelimumab, nivolumab, pembrolizumab, cemiplimab, atelizumab, avelumab, durvalumab, relatlimab, or any combination thereof.

39. A method, according to any one of claims 35 to 38, characterized in that the subject does not respond to other anticancer therapy in the absence of administration of the composition comprising antigen-presenting cells.

40. Suitable pharmaceutical composition for intratumoral injection, characterized in that the pharmaceutical composition comprises antigen-presenting cells obtained from one or more pigs. Petition 870250092182, dated 08 / 10 / 2025, pp. 56 / 72

Images