Treatment of solid tumors with implantable polymers with small molecules

Abstract

Disclosed are implantable polymers that release therapeutic agents in proximity to a tumor in a patient. In some embodiments, a polyethylene glycol hydrogel can release a small molecule, like a STING agonist in proximity to a phosphatase and tensin homolog (PTEN)-null triple negative breast cancer (TNBC) in the patient.

Description

TREATMENT OF SOLID TUMORS WITH IMPLANTABLE POLYMERSWITH SMALL MOLECULES

[0001] This application claims the benefit of and priority to U.S. Provisional Application No. 63 / 596,834, filed on November 7, 2023, the entire contents of which are incorporated herein by reference.

[0002] All patents, patent applications and publications cited herein are hereby incorporated by reference in their entirety. The disclosures of these publications in their entireties are hereby incorporated by reference into this application in order to more fully describe the state of the art as known to those skilled therein as of the date of the invention described and claimed herein.

[0003] This patent disclosure contains material that is subject to copyright protection. The copyright owner has no objection to the facsimile reproduction by anyone of the patent document or the patent disclosure as it appears in the U.S. Patent and Trademark Office patent file or records, but otherwise reserves any and all copyright rights.FIELD

[0004] Disclosed are implantable biocompatible polymers that continuously release therapeutic agents (e.g., small molecules, including STING agonists) in the body to treat cancers.BACKGROUND

[0005] Injectable delivery of certain therapeutic agents to tumors in a subject can be toxic to immune cells (e.g., T cells) that attack the tumors. Other modes of delivery in these instances are sought.SUMMARY

[0006] Disclosed herein are implantable, biocompatible polymers (e g., polyethylene glycol- based hydrogels) that can release therapeutic agents, including STING agonists in a subject to treat a cancer. In some embodiments, a biocompatible polymer that contains a STING agonist can be implanted in proximity to a solid tumor in a subject. In some embodiments, the cancer can be a breast cancer. In some embodiments, the cancer can be a phosphatase and tensin homolog (PTEN) null, triple negative breast cancer (TBNC).

[0007] Disclosed are methods of delivering a therapeutic agent to a subject to treat a solid tumor by implanting a biocompatible polymer that includes a therapeutic agent into the subject.The biocompatible polymer continuously releases the therapeutic agent in the subject over time. In some embodiments, the cancer treated can be a breast cancer, including TNBC, including PTEN-null TBNC. In some embodiments, the implanted biocompatible polymer can contain a STING agonist.

[0008] In some embodiments, the implantable biocompatible polymer can be a biopsy marker. In some embodiments, the biopsy marker can have a core of poly(dioxanone) (PDO), polylactide (PLA), polyglycollide (PGA), polycaprolactone (PCL), or copolymers thereof; and a sealant surrounding the core, comprising a polyethylene glycol (PEG)-based hydrogel. In some embodiments, the biopsy marker can be a HydroMARK™ biopsy marker.

[0009] In some embodiments, disclosed herein is a method for treating a breast cancer in a patient by hydrating a dehydrated biopsy marker with a STING agonist and implanting the hydrated biopsy marker into the patient in proximity to cells of the breast cancer. The dehydrated biopsy marker can include a polyethylene glycol (PEG)-based hydrogel.

[0010] In some embodiments, disclosed herein is a method for treating a PTEN-null, TNBC in a subject by, without resecting the breast cancer, implanting a hydrated HydroMARK™ biopsy marker having a STING agonist in proximity to the breast cancer.

[0011] Disclosed herein are HydroMARK™ biopsy markers that have a small molecule, including a STING agonist.BRIEF DESCRIPTION OF THE FIGURES

[0012] The patent or application file contains at least one drawing executed in color. Copies of this patent or patent application publication with color drawing(s) will be provided by the Office upon request and payment of the necessary fee.

[0013] Figure 1. Defective STING trafficking renders TNBC cells hypersensitive to STING agonism. Figure 1A shows images depicting STING staining of a TNBC tissue microarray (TMA) (n=63) with scoring criteria. 0 = no staining in tumor cells; 1+ = faint staining in >10% of tumor cells; 2+ = moderate staining in >10% of tumor cells; 3+ = strong staining in >10% of tumor cells. Figure IB depicts images from two cases showing core to core discrepancies in staining intensity. Case 24 with score of 3+ and 0; Case 89 with score of 2 and 0. Figure 1C shows representative images of PTEN staining of TNBCs with STING 3+ Figure ID shows an immunoblot of a panel of TNBC cell lines for PTEN and STING. Figure IE showsan immunoblot of MDAMB231 unaltered (Parent) or containing CRTSPR control vector (Scramble) or PTEN knockout vectors (Sgl, Sg2, Sg3) for PTEN and STING. Cells were treated with PBS as control or ADU at lOpM or 50pM for 48h. Supernatants were then analyzed for CXCL10 expression by ELISA (n=3). *, P < 0.05; **, P < 0.005; ***, P < 0.0002; ****, P < 0.0001 by two-way ANOVA with Tukey’s MCT. Figure IF shows an immunoblot of MDA- MB231 Parent or containing CRISPR control vector (Scramble) or Rab7 knockout vectors (Sgl, Sg2) for Rab7 and STING. Cells were treated with ADU at 50pM for 48h, and supernatants were then analyzed for CXCL10 expression by ELISA (n=2). *, P < 0.05; ***, P< 0.0002; ****, P < 0.0001 by ANOVA with Tukey’s MCT. Figure 1G shows immunofluorescence staining for STING (Green) and lysosome (Red) colocalization was performed after MDAMB231 cells were treated with 50pM ADU or lOOnM CID1067700 or in combination for Ih and 6h (representative of n = 3 independent experiments). Co-localization was quantified using ImageJ (n=4). **, P < 0.005; ***, P < 0.0002 by two-way ANOVA with Tukey’s MCT. Figure 1H shows a growth inhibition assay of PTEN null and PTEN WT cell lines over 6 weeks. MDA-MB468 and HCC70 were treated twice-weekly with lOpM or 50pM ADU. MDAMB231 and HCC1806 were treated twice-weekly with lOqM or 50pM ADU + / - lOOnM CID1067700. Cell growth was visualized with crystal violet staining and then quantified using absorbance of 590nm (n=3) (Figure II). Figure 1 J shows a growth inhibition assay where MDA-MB231 Scramble and PTEN KO cell lines (Sgl, Sg3) were treated similarly as in Figure 1H, stained for visualization with crystal violet and then quantified using absorbance of 590nm (n=3). *, P < 0.05; **, P < 0.005; ***, P < 0.0002; ****, P < 0.0001 by ANOVA with Tukey’s MCT (Figure IK). Figure IL shows an immunoblot of MDMAB468 containing CRISPR control vector (Scramble) or Rab7 knockout vectors (Sgl, Sg2) for Rab7 and STING. Figure IM shows immunofluorescence staining for STING (Green) and lysosome (Red) colocalization was performed after MDAMB231 cells were treated with 50pM ADU, lOOnM CID1067700, or in combination for Ih and 6h. Co-localization was quantified using ImageJ (n=4). **, P < 0.005; ***, P < 0.0002 by two-way ANOVA with Tukey’s MCT. Figure IN shows a growth inhibition assay. Figure IO shows effects of various treatments on CXCL10 production. Figure IP shows H-score analysis of STING levels in the PTEN negative (n=38) and PTEN positive (n=2) subsets. Six PTEN low positive TNBCs were included in the PTEN negative subset. Figure IQ shows quantitative analysis and frequencydistribution of H-score of STING expression in PTEN negative subset (n=38). H-score <30 indicates low to negative expression. H-score >75 indicates high expression.

[0014] Figure 2. Impaired STING turnover enhances STING agonist induced T cell migration. Figure 2A shows a Luminex cytokine profding performed for scramble and Rab7 knockout cells using PTEN WT cell lines MDAMB231 and HCC1806, and the PTEN null cell line MDAMB468. Log-2 -fold change cytokine / chemokine differences between Rab7 knockout, Scramble treated with ADU, and Rab7 knockout treated with ADU relative to untreated Scramble cells are shown (n=2-4). Figure 2B shows CXCL10, IL-8, and IL-6 expression measured by ELISA in culture media (CM) of samples from the experiment described in Figure 1A. *, P < 0.05; **, P < 0.005; ***, P < 0.0002; ****, P < 0.0001 by ANOVA with Tukey’s MCT. Figure 2C depicts a schematics of CD8+ T cell migration assay utilizing a 3D microfluidic device. Figure 2D shows images of T cell (yellow) migration towards MDAMB231 spheroids (DAPI). MDAMB231 Scramble and Rab7 knockout spheroids were pretreated with 50pM ADU for 18h prior to loading the 3D microfluidic device. Migrated CD8+ T cells were quantified after 48h (n=8- 9). ***, P < 0.0002; ****, P < 0.0001 by ANOVA with Tukey’s MCT. Figure 2E shows images of T cell migration towards HCC1806 spheroids treated similarly as in Figure ID. Migrated CD8+ T cells were quantified after 48h (n=7-10). *, P < 0.05; **, P < 0.005 by ANOVA with Tukey’s MCT. Figure 2F shows CXCL10 quantification in MDAMB23 1 and HCC1806 cells treated with 50pM or lOpM ADU in the presence or absence of increasing doses of CID1067700 for 48h. Supernatants were analyzed for CXCL10 by ELISA (n=3-5). *, P < 0.05; ***, P < 0.0002; ****, P < 0.0001 by ANOVA with Tukey’s MCT. Figure 2G depicts a schematic of a Jurkat cell migration assay utilizing a 3D microfluidic device. Representative images of Jurkat cells (pink) migrating towards MDA-MB231 spheroids (DAPI) after treatment with lOOnM CID 1067700 + / - 50pM ADU-S100. Migrated Jurkat cells were quantified after 48h (n=4). *, P < 0.05; ***, P < 0.0002; ****, P < 0.0001 by ANOVA with Tukey’s MCT. Figure 2H shows CCL5 expression assessment by ELISA in the culture media (CM) used in Figure 1A for HCC1806. ***, P < 0.0002; ****, P < 0.0001 by ANOVA with Tukey’s MCT. Figure 21, Representative images of Jurkat cells (pink) migrating towards MDAMB23 1 spheroids (DAPI) after treatment with lOOnM CID1067700 + / - 50pM ADU. Migrated Jurkat cells were quantified after 48h (n=4). **, P < 0.005; ***, P < 0.0002; ****, P < 0.0001 by ANOVA with Tukey’s MCT. Figure 2 J shows multiplexed cytokine profiling ofcontrol PTEN null cell line MDAMB468 and PTEN WT cell lines MDAMB231 and HCC1806, compared with Rab7 KO - / + ADU treatment. Heatmap shows control cell normalized log-2 fold change (L2FC) of key STING-IRF3 and STING-NFKB cytokine / chemokines. Figure 2K is immunoblot of HCC1806 Parent or containing CRISPR control vector (Acramble) or Rab7 knockout vectors (Sgl, Sg2).

[0015] Figure 3. Impregnation of STING agonist in a PEG hydrogel biopsy marker result in its local delivery into TNBC. Figure 3A shows an image of desiccated radiographic biopsy markers and their corresponding image after hydrating for 2 hrs. Figure 3B shows an image of implanted PEG marker in a mouse on Day 0 and 4, with magnification views of the marker on Day 8 and 25. Figure 3C depicts a schematic of THP-lassay. Known amount of ADU-S100 is added to THP1 cells followed by ELISA assay to determine corresponding CXCL10 levels. Figure 3D shows average values of triplicates from Figure 3C that were plotted to generate a standard curve. Figure 3E shows a schematic of the assay used to determine the amount of ADU released from a PEG marker with lOOpg of ADU over time. Figure 3F shows CXCL10 ELISA values corresponding to Figure 3E that were used to determine the amount of ADU released at each time point utilizing the standard curve generated in Figure 3D. Figure 3G shows the amount of ADU-S100 released from the various biopsy markers after each marker was soaked with lOOpg ADU over time. Average values of triplicates are shown. Figure 3H shows the size of tumors developed in mice after 4T1 cells (10k) were inoculated into the mammary fat pad of BALB / c mice followed by implantation of PEG marker with PBS or lOOpg ADU on Day 4. Individual tumor volume measurements are shown (n=5 mice per group). Figure 31 shows the size of tumors developed in mice after 4T1 cells (100k) were inoculated into the mammary fat pad of BALB / c mice. At tumor size of 150mm3 on Day 6, mice were implanted with PEG marker soaked with PBS or lOOpg ADU. Individual tumor volume measurements are shown (n=7 mice per group). Figure 3J shows images of a 4T1 mouse from Figure 31 treated with PEG+PBS on Day 25. Dissection of the tumor shows an intact PEG marker with a visible metal clip. Figure 3K (Left) shows the volume of the PEG marker at each time point after tumor implantation in the mouse. (Right) shows the amount of ADU released from the various biopsy markers embedded with lOOpg of ADU.

[0016] Figure 4. STING agonist monotherapy is toxic to T cells and does not result in T cell clonality. Figure 4A shows percentages of CD45+ leukocytes, lymphocytes, and T cells(CD3+, CD4+, CD8+) in primary human TNBC explants that were treated with 50pM ADU or PBS for 24h, followed by dissociation into single cell suspensions and analyzed by flow cytometry. Figure 4B shows percentages of cell viability in human CD8+ T cells that were treated with various dosages of ADU for 24h; cell viability was determined by CellTiter-Glo (CTG) assay at 24h (n=3). ****, P < 0.0001 by ANOVA with Tukey’s MCT. Figure 4C percentage of viability in CD3+, CD4+, and CD8+ T cells. Splenocytes were harvested from C57BL / 6 mice and treated with various dosages of ADU for 24h. Viability of CD3+, CD4+, and CD8+ T cells (n=2) was determined by flow cytometry. *, P < 0.05; **, P < 0.005; ***, P < 0.0002 by ANOVA with Tukey’s MCT. Figure 4D (Left) shows percentage of viable cells after human CD8+ T cells were treated with conditioned media and collected at various timepoints from PEG soaked with lOOpg ADU as in Figure 3E. Figure 4D (Right) shows CD8+ T cells that were treated with direct addition of PBS, direct addition of lOOpg ADU, PEG+PBS, or PEG+ADU embedded with lOOpg ADU. Cell viability was determined by CTG assay at 24h (n=3). The first panel shows treatment for 5 days with no media changes and cell viability was determined by CTG assay (N=3). The second panel shows cells treated the same way with surviving cells after ADU treatment replated at Day 5 and then stimulated with IL-2, IL-7, and IL-15, followed by CTG assay on Day 10 (n=3). Figure 4E shows the size of tumors after 4T1 cells (10k) were inoculated into the mammary fat pad of BALB / c mice, and at Day 4 a PEG marker with PBS or withlOOpg of ADU was implanted. On Day 30, another PEG marker with PBS or lOOpg of ADU was implanted. Individual tumor volume measurements are shown (PBS n=6, ADU n=10). * Denotes mice used for single-cell RNA-sequencing (scRNA-seq) and T-cell receptor- sequencing (TCR-seq). Figure 4F depicts a schematic of the experimental design used in Figure 4E. Figure 4G shows UMAP plots displaying unsupervised clustering of scRNA-seq data for PBMCs collected from PEG+PBS and PEG+ADU treated 4T1 tumors from E. Cell assignment was based on comparison to cluster gene expression signatures from established literature. Activated T cells was the main cluster to exhibit any associated clonotypes. Figure 4H shows UMAP plots displaying clusters with any associated TCR clonotypes. Figure 41, Pie charts demonstrating percentage of cells with unique TCR clonotypes of frequency=l, >=2, and >=5 in for PBS and ADU treatment groups. Figure 4J shows primary human TNBC explants treated with 50 pm ADU or PBS for 24h, followed by dissociation into single cell suspensions.Flow cytometry was performed to analyze percentages of live CD45+ leukocytes, CD3+ lymphocytes, and T cells (CD4+, CD8+) as depicted by the pie graph.

[0017] Figure 5A. Additional studies using STING agonist in PEG hydrogel biopsy markers. Figure 5A shows a study that did not use the PEG hydrogel. In established 4T1 tumors, on day 6, PBS or 100 pg ADU was injected intratum orally. Mean tumor volume measurements are shown (n=4-5 mice / group). Figure 5B shows representative IHC images of pSTATl and pTBKl staining of 4T1 tumors treated with PEG+PBS or PEG+100 pg ADU for 4 days. Scale bar, 100 pm. Figure 5C shows a schematic of orthotopic implantation of 100K 4T1 cells followed by PEG + PBS or 100 pg ADU implantation. ScRNA-seq and TCR-seq were performed with 4 pooled tumors / group 14 days later. Figure 5D (Left) UMAP plots displaying unsupervised clustering of scRNA-seq data for CD45+cells collected from Figure 5C. (Right) Volcano plot showing overview of differentially expressed genes (DEGs) in PEG+ADU versus PEG+PBS treated tumors in the cluster of APC + Macrophage in (K). Red dots indicate significantly upregulated genes. Blue dots indicate significantly downregulated genes. Figure 5E and 5F shows gene set enrichment analysis (GSEA) based on the expression fold-change values of ADU versus PBS treated tumors in the cluster of APC + Macrophage. GSEA plots for IFNy and IFNa response are shown in (E). Bar plot represents normalized enrichment scores (NES) for the top upregulated Hallmark gene sets (F). Quantitative data are represented as mean ± SEM; p values were calculated by two-way ANOVA followed by Sidak’s post hoc test (A), ns, not significant; ****, p < 0.0001.DETAILED DESCRIPTION

[0018] Disclosed herein are methods for administering small molecules, including STING agonists, to solid tumors (e.g., breast cancer) at doses that avoid negative effects to T cells, but that are still efficacious to treat the solid tumor (e g., immune-mediated clearance of STING- positive TNBCs). First, disclosed herein is that certain breast cancers, including phosphatase and tensin homolog (PTEN)-null, triple-negative breast cancers (TNBCs), can be hypersensitive to STING agonism, in some embodiments, due to defective Rab7 trafficking, and / or can result in enhanced T cell migration to the cancer. Second, we have discovered that certain biocompatible polymers containing anti-cancer therapeutics (e.g., small molecules, including STING agonists), implanted in proximity to a solid tumor in a subject, can continuously release the therapeuticagents over time. In the case of STING agonists, its release from the biocompatible polymers can avoid the harsh effects on T cells observed when STING agonists are injected intratumorally.

[0019] In some embodiments, the biocompatible polymer can be an expandable solid shape of polyethylene glycol (PEG) based hydrogel. The PEG can be dry / dessicated and expandable with fluid contact. In some embodiments, the polymers can be part of a biopsy site marker or biopsy clip. The biopsy site marker can include a metallic marker component. In some embodiments, a biopsy site marker can include a HydroMARK™ device.

[0020] In some embodiments, this platform provides for local delivery of STING agonists and other small molecules into a tumor microenvironment of solid cancers.

[0021] In some embodiments, the methods disclosed herein can be used to treat tumors in humans.Definitions

[0022] Detailed descriptions of one or more embodiments are provided herein. It is to be understood, however, that the present invention may be embodied in various forms. Therefore, specific details disclosed herein are not to be interpreted as limiting, but rather as a basis for the claims and as a representative basis for teaching one skilled in the art to employ the present invention in any appropriate manner.

[0023] The singular forms “a”, “an” and “the” include plural reference unless the context clearly dictates otherwise. The use of the word “a” or “an” when used in conjunction with the term “comprising” in the claims and / or the specification may mean “one,” but it is also consistent with the meaning of “one or more,” “at least one,” and “one or more than one.”

[0024] Wherever any of the phrases “for example,” “such as,” “including” and the like are used herein, the phrase “and without limitation” is understood to follow unless explicitly stated otherwise. Similarly, “an example,” “exemplary” and the like are understood to be nonlimiting.

[0025] The term “substantially” allows for deviations from the descriptor that do not negatively impact the intended purpose. Descriptive terms are understood to be modified by the term “substantially” even if the word “substantially” is not explicitly recited.

[0026] The terms “comprising” and “including” and “having” and “involving” (and similarly “comprises”, “includes,” “has,” and “involves”) and the like are used interchangeably and have the same meaning. Specifically, each of the terms is defined consistent with the common UnitedStates patent law definition of “comprising” and is therefore interpreted to be an open term meaning “at least the following,” and is also interpreted not to exclude additional features, limitations, aspects, etc. Thus, for example, “a process involving steps a, b, and c” means that the process includes at least steps a, b and c. Wherever the terms “a” or “an” are used, “one or more” is understood, unless such interpretation is nonsensical in context.

[0027] The term “about” is used herein to mean approximately, roughly, around, or in the region of. When the term “about” is used in conjunction with a numerical range, it modifies that range by extending the boundaries above and below the numerical values set forth. In general, the term “about” is used herein to modify a numerical value above and below the stated value by a variance of 20 percent up or down (higher or lower).Biocompatible Polymers

[0028] In some embodiments, the polymers disclosed herein are biocompatible. In some embodiments, the polymers are implantable. The polymers can retain a therapeutic agent, which therapeutic agent can be released from the polymer (e.g., diffuse out from the polymer) after the polymer is implanted in a subject. In some embodiments, the therapeutic agent can be released from the polymer continuously over a period of time in the subject (e.g., sustained release).

[0029] In some embodiments, the therapeutic agent can adsorb to the polymer. When implanted, the therapeutic agent can de-adsorb from the polymer. In some embodiments, the therapeutic agent can be absorbed by the polymer. For example, a therapeutic agent can be dissolved in a solvent (e.g., water), which solvent can be used to hydrate a dehydrated polymer (e.g, aqueous-expandable polymer)(absorption). When implanted, the therapeutic agent can move out of the hydrated polymer with the solvent as bodily fluids displace the solvent.

[0030] In some embodiments, a biocompatible polymer can be a biodegradable polymer, a non-biodegradable polymer or a natural polymer. In some embodiments, a biodegradable polymer can be a poly(dioxanone) (PDO), polyethylene glycol (PEG), poly(glycolic acid) (PGA), poly(lactic acid) (PLA), poly(lactic-co-glycolytic acid) (PLGA), poly(caprolactone) (PCL), or combinations thereof.

[0031] In some embodiments, a non-biodegradable polymer can be a poly(siloxane) (silicone) or poly(ethylene-vinyl acetate).

[0032] In some embodiments, a natural polymer can be an albumin, cellulose, chitosan, collagen elastin, gelatin, a milk protein, silk, and the like.

[0033] In some embodiments, the biocompatible polymer can be hydrophilic. In some embodiments, the biocompatible polymer can be a hydrogel. In some embodiments, the biocompatible polymer can be a PEG-containing hydrogel. A hydrogel can be chemically linked (e.g., covalent cross-linking bonds), physically-linked (e.g., non-covalent bonds), or a combination thereof.

[0034] In some embodiments, the biocompatible polymer can have poly(dioxanone) (PDO), polylactide (PLA), polyglycollide (PGA), poly caprolactone (PCL), copolymers thereof and the like.

[0035] In some embodiments, the biocompatible polymer can have a core that can be surrounded by a core-surrounding layer. In some embodiments, the core-surrounding layer can be a sealant. In some embodiments, the core can have poly(dioxanone) (PDO), polylactide (PLA), polyglycollide (PGA), polycaprolactone (PCL), and / or copolymers thereof. In some embodiments, the core can be surrounded by a sealant layer. The sealant layer can be a polyethylene glycol-based hydrogel.

[0036] In some embodiments, the biocompatible polymer (e.g., the core) can have a metal marker. In some embodiments, the marker can be titanium or stainless steel.

[0037] In some embodiments, the biocompatible polymer can have pores of pores of 10-500 pm in size. Spacing between pores in the polymer can be from about 5 to 100 nm.Biopsy Markers

[0038] In some embodiments, the biocompatible polymers described herein can be part of a biopsy marker or biopsy clip. The biopsy marker can have a metal marker, like a titanium or stainless-steel marker. Biopsy site markers can be used to mark areas of the body from which biopsies are taken. Implanted biopsy markers can remain indefinitely and be used for preoperative localization when the biopsy identifies cells that must be removed, or for future identification of the site when pathology indicates the biopsi ed cells are benign.

[0039] Herein, any biopsy marker can be used for delivery of therapeutic agents. In some embodiments, the biopsy markers can have a PEGylated substance, collagen, p-glucan or polylactic-co-glycolic acid. In some embodiments, the biopsy marker can be a HydroMARK™,MammoMARK®, MammoSTAR® or SecurMark® biopsy marker. In some embodiments, HydroMARK™ biopsy markers are marketd by Mammotome, Devicor Medical Products, Inc., which is part of Leica Biosystems, USA..

[0040] The HydroMARK™ device can be made of an expandable solid cylinder of polymerized and desiccated polyethylene glycol (PEG)-based hydrogel. The device can expand with fluid contact, and the implanted device can be visible under ultrasound for 6 weeks to 12 months. The device can contain an embedded metallic component, for example titanium or stainless steel, which can be permanently visible by X-ray and MR imaging.

[0041] In some embodiments, the biopsy markers can be dehydrated or dessicated (FIG. 3A). In some embodiments, the biopsy markers can be hydrated. Hydrated markers, that eventually are implanted into a subject, can be approximately 7, 9, 11, 13, 15, 17, or 19 mm in length.

[0042] In some embodiments, disclosed herein are biopsy markers that have been “loaded” with one or more therapeutic agents. In some embodiments, disclosed herein are HydroMARK™ biopsy markers that have been loaded with a STING agonist.Cancers

[0043] A wide variety of cancers can be treated using the compositions and methods disclosed herein. In some embodiments, the cancers can be solid tumors. In some embodiments, the cancers can be primary solid tumors or metastatic solid tumors. In some embodiments, the cancer can be a carcinoma, lymphoma, melanoma or sarcoma. In some embodiments, the cancer can be a bladder, bone, brain, breast, cervical, colorectal, gastrointestinal, kidney, liver, lung, ovarian, pancreatic, prostate, skin or thyroid cancer. In some embodiments, the cells of the cancer can have no or reduced amounts of functioning phosphatase and tensin homolog (PTEN). In some embodiments, PTEN can be inactivated or null.

[0044] In some embodiments, the cancer treated can be breast cancer. The breast cancer can include ductal carcinoma in situ (DCIS), HER2-positive breast cancer, invasive ductal carcinoma (IDC), invasive lobular carcinoma (ILC), mixed ductal and lobular carcinoma, inflammatory breast cancer or triple-negative breast cancer (TNBC). In some embodiments, the cells of the breast cancer can have no or reduced amounts of functioning phosphatase and tensin homolog (PTEN). In some embodiments, PTEN can be inactivated or null.

[0045] In some embodiments, the cancer can be one that overexpresses STING. STING (Stimulator of interferon genes) is a transmembrane protein encoded by TMEM173. STING is a regulator of the innate immune response to cytoplasmic double stranded DNA (dsDNA) in cancer. Following its activation, STING protein can undergo Ras-related protein Rab-7a (Rab7)- mediated lysosomal turnover. In some cells, including triple-negative breast cancer (TNBC) cells, this physiologic STING degradation can be disrupted by maintenance of Rab7 in a hyperphosphorylated inactive state.

[0046] We recently reported that tumor cell STING expression is elevated in TNBCs, whereas Luminal A and B subtypes are largely STING negative. In addition, when STING expression was examined in a tumor microarray (TMA) of 64 human TNBCs, there was a distinct trend of STINGhigh and STINGlow TNBCs, with approximately 61% of the TNBCs expressing STING.

[0047] In TNBC, phosphatase and tensin homolog (PTEN) inactivated or null cells maintain Rab7 in an inactive state. Trafficking of STING to lysosomes in impaired in these cells and the cells are hypersensitive to STING. Rab7 inhibitors exist and can be used to increase sensitivity to STING in cells that are not PTEN deficient.Therapeutic Agents

[0048] The therapeutic agents released by the implanted biocompatible polymers can be of a variety of types. In some embodiments, the therapeutic agents can be small molecules.

[0049] In some embodiments, the therapeutic agents can be STING agonists. A STING agonist can cause increase activity of STING. In some examples, STING agonists can cause increases in inflammatory cytokines that can remodel the tumor microenvironment and can increase anti-tumor T cell responses.

[0050] Herein, any STING agonist can be used. In some embodiments, the STING agonist can include ADU-S100, MK-1454, MK-2118, SB 11285, GSK3745417, BMS-986301, BISTING, E7766, TAK-676, SNX281, SYNB1891, and the like, or combinations thereof.

[0051] In some embodiments, a therapeutic can be a small molecule that is an estrogen receptor-target therapy. The estrogen receptor targeted therapy can be a selective estrogen receptor modulator (SERM). In some embodiments, the SERM can be tamoxifen, raloxifene, or the like.

[0052] In some embodiments, the therapeutic agent can be a Rab7 inhibitor.

[0053] In some embodiments, the biocompatible polymers can release two or more small molecules and / or STING agonists.Methods

[0054] In some embodiments, the biocompatible polymers can be “loaded” with a therapeutic agent. The therapeutic agents can be loaded into the biocompatible polymers before and / or after the polymers have been implanted into the body of a subject.

[0055] In some embodiments, prior to loading of a therapeutic agent, the biocompatible polymer can be dehydrated. A therapeutic agent can be loaded into the polymer by rehydrating the polymer with a solution that contains the therapeutic agent. The polymer can be hydrated using various volumes of solution containing the therapeutic agent. In some embodiments, the biocompatible polymer can be hydrated with a volume less than about 20 pl.

[0056] In some embodiments, the biocompatible polymer can be implanted into a body of a subject. The biocompatible polymer can be implanted in proximity to the tumor or cancer that is desired to be treated. The biocompatible polymer can be implanted within within 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 mm of the solid tumor. The biocompatible polymer can be implanted within the solid tumor. The biocompatible polymer can be implanted in the periphery of the solid tumor. The biocompatible polymer can remain in the body of the subject for various periods of time. In some embodiments, the polymer can remain in the body of the subject for days, weeks or months.

[0057] In some embodiments, the tumor is not resected prior to / during implantation of the biocompatible polymer. In some embodiments, the tumor can be resected prior to / during implantation of the biocompatible polymer.

[0058] Implanting the biocompatible polymer can include surgical implanting or implanting using a large-bore needle. In some embodiments, multiple biocompatible polymers (i.e., plugs) can be implanted.

[0059] In some embodiments, the biocompatible polymer can be loaded with a therapeutic agent after the polymer has been implanted. This can be done using a syringe containing the therapeutic agent to be loaded. A needle of the syringe can be inserted through the skin of the subject, and into the implanted polymer. The syringe can deliver the therapeutic agent (e.g., STING agonist) to the implanted polymer. In some embodiments, other substances can beinjected into the implanted biocompatible polymer. In some embodiments, a dehydrated biocompatible polymer (e.g., a biopsy marker) can be loaded with a therapeutic agent (e.g., STING agonist) and implanted into a subject. Subsequently, the same or another therapeutic agent and / or substance can be injected into the implanted biocompatible polymer.

[0060] In some embodiments, a large-bore needle can be used to insert a dehydrated biocompatible polymer (e.g., biopsy marker) through the skin and place it in proximity to a tumor in a subject. In some embodiments, another syringe containing a therapeutic agent can then inserted through the skin and into the placed biocompatible polymer. The therapeutic agent can be delivered to the placed biocompatible polymer using the syringe.

[0061] Implanted biocompatible polymers that provide a therapeutic agent to a solid tumor, as disclosed herein, can be used alone to treat a subject. In some embodiments, this treatment can be used in combination with other treatments. In some embodiments, the treatments disclosed herein can be used in combination with conventional therapy. In some embodiments, conventional therapy can include chemotherapy, endocrine therapy, and the like.Compositions

[0062] Disclosed herein are implantable biocompatible polymers that include a therapeutic agent. Embodiments of these biocompatible polymers plus therapeutic agents that can be combined with the biocompatible polymers have been described throughout this application.

[0063] In some embodiments, compositions of a biocompatible polymer that can include a metal marker or clip, and a therapeutic agent are disclosed. The biocompatible polymer that has the therapeutic agent can be hydrated.

[0064] In some embodiments, the biocompatible polymer can be a polymer core that includes the metal marker, surrounded by a polyethylene glycol (PEG)-based hydrogel. In some embodiments, the polymer core can be a poly(dioxanone) (PDO), polyethylene glycol (PEG), poly(glycolic acid) (PGA), poly(lactic acid) (PLA), poly(lactic-co-glycolytic acid) (PLGA), poly(caprolactone) (PCL), or combinations of these polymers. The PEG-based hydrogel surrounding the core can be called a “sealant”. In some embodiments, the metal marker or clip can be titanium, stainless-steel, and the like.

[0065] In some embodiments, the biocompatible polymer can be a biopsy marker. In some embodiments, the biopsy marker can be a combination biodegradable polymer encompassed by ahydrogel. In various embodiments, the biopsy marker can be of many different types, including HydroMARK™, MammoMARK®, MammoSTAR®, SecurMark®, and the like.

[0066] The therapeutic agents associated with the biocompatible polymer can be absorbed into and / or adsorbed onto the biocompatible polymer. In some embodiments, a dehydrated biocompatible polymer can be hydrated in a solution or suspension containing a therapeutic agent. The biocompatible polymer can have one or more associated therapeutic agents.

[0067] In some embodiments, the therapeutic agents can be small molecules. An example small molecule can be a STING agonist, as described herein. Example STING agonists can ADU-S100, MK-1454, MK-2118, SB11285, GSK3745417, BMS-986301, BI-STING, E7766, TAK-676, SNX281, SYNB1891, and other molecules that have the same or similar function.EXAMPLES

[0068] Examples are provided below to facilitate a more complete understanding of the invention. The following examples illustrate the exemplary modes of making and practicing the invention. However, the scope of the invention is not limited to specific embodiments disclosed in these Examples, which are for purposes of illustration only, since alternative methods can be utilized to obtain similar results.EXAMPLE 1Defective STING trafficking renders TNBC cells hypersensitive to STING agonism.

[0069] A STING immunohistochemistry study was performed on a TNBC tissue microarray (TMA) derived from treatment-naive patients (n=63) with clinicopathologic characteristics described in Table 1. A breast pathologist visually scored the samples using an intensity scale: 0 = no staining in tumor cells with intact expression in internal controls using endothelial and inflammatory cells; 1+ = faint staining in >10% of tumor cells; 2+ = moderate staining in >10% of tumor cells; 3+ = strong staining in >10% of tumor cells (Fig. 1A). More than half of the TNBCs showed STING expression, and four samples showed high variability ranging from 0 to 2+ / 3+ staining with similar histology in both the high and low STING expressing areas (Fig. IB). Among the STING 3+ TNBCs, 5 out of 6 specimens were available for PTEN staining and were either PTEN IHC negative or low. No association of heterogeneity with age, lymph node status, tumor size or grade was seen (Table 1).

[0070] Table 1. Clinicopathologic characteristics of triple negative breast cancer patients.1, 2

[0071] ’American Joint Committee on Cancer (8thedition) pT and pN categories

[0072] 2N / A is not applicable.

[0073] It was previously found that PTEN (phosphatase and tensin homolog; a tumor suppressor) null TNBCs maintain the lysosomal protein, Rab7 in an inactive state and, therefore, trafficking of STING to the lysosome can be impaired, resulting in baseline upregulation of STING protein and increased sensitivity to STING agonism (12). PTEN IHC was therefore performed on the TMA and confirmed this inverse relationship between PTEN and STING (Fig. 1C). Analysis of the larger TMA showed that PTEN null status had a tendency towards elevated tumor cell STING levels (Fig. IP and IQ). Similarly, using a panel of TNBC cell lines, PTEN null lines (MDAMB468, HCC1937, HCC70) showed higher STING protein expression than PTEN WT lines (HCC1143, HCC1806, MDAMB231) (Fig. ID). Furthermore, when PTEN was knocked out in MDAMB231, STING protein was upregulated and became sensitized to ADU- S100, producing higher levels of the interferon regulatory factor 3 (IRF3)-regulated chemokine CXCL10 (Fig. IE).

[0074] To broaden the use of STING agonists to include PTEN WT TNBCs, we targeted Rab7 directly through both protein knock out and the use of the Rab7 inhibitor (CID1067700), which acts as a competitor for the GTP and GDP binding site in Rab7, thus preventing nucleotide exchange and activation (24, 25). Rab7 knockout in MDAMAB231 resulted in a marked increase in STING protein and sensitized response to ADU-S100 with a marked increase in CXCL10 (Fig. IF). As a control, Rab7 knockout of PTEN WT MDAMB468 cells did notalter STING protein levels (Fig. IL).

[0075] To explore the immediate effects of Rab7 inhibition, MDAMB231 cells were treated with ADUS100, the Rab7 inhibitor CID1067700, and in combination. Immunofluorescence staining for STING and lysosome was then performed to monitor for colocalization. After ADU- S100 stimulation, there was distinct colocalization of STING to the lysosomes for degradation that was disrupted in the presence of Rab7 inhibition. (Fig. 1G; Fig IM). In a prior growth assay, chronic ADU- SI 00 treatment of the PTEN null cell lines MDAMB468 and HCC70 resulted in significant growth inhibition with low dose ADU-S100 at lOpM that was not seen with MDAMB231 and HCC1806. Addition of Rab7 inhibitor was able to sensitize both MDAMB23 1 and HCC1806 to ADU-S100 with a more significant effect in the latter (Fig. 1H) that is quantified by crystal violet absorbance (Fig.ll).

[0076] Since MDAMB231 had a less robust response to Rab7 inhibitor with ADUS100 at lOpM, MDAMB231 PTEN knockout cells were treated with ADU-S100 + / - Rab7 inhibitor and it was found that PTEN loss sensitized the cells to the lower concentration of ADU- SI 00 (lOpM) and that addition of Rab7 inhibitor further enhanced this effect (Fig. 1 J and Fig. IK; Fig. IN). Moreover, combination treatment also increased CXCL10 production in MDMAMB23 1 and HCC1806 but not in MDMAB468 in which Rab7 is already inactivated and a robust increase in CXCL10 has already been achieved with ADU-S100 treatment alone (Fig. IO).

[0077] These data show that PTEN null TNBCs have a dysregulation of STING that prevents its trafficking to the lysosomes for degradation, and in PTEN WT TNBCs this can be recapitulated with Rab7 knockout or addition of a Rab7 inhibitor.EXAMPLE 2Impaired STING turnover enhances STING agonist induced T cell migration.

[0078] Prior data showed upregulation of CXCL10 with Rab7 knockout. To better profile these resultant cytokines, a Luminex Assay was performed on MDAMB231 and HCC1806, using MDAMB468 as a control. In addition, scramble control cells and RAB7 knockout cells (Fig. 2K) for each cell line were treated with 50pM ADU-S100 to assess if cytokine levels could be further augmented. The key cytokines that were induced were TBK1-IRF3 regulated CXCL10 and CCL5 and the NF-KB- regulated cytokines IL-8 and IL-6 (Fig. 2A and 2J). Both CXCL10 and CCL5 have been implicated in T cell recruitment whereas IL-6 and IL-8 have beenassociated with both T-cell recruitment and immunosuppression (26). ELISA was then performed to confirm the upregulation of CXCL-10, IL-8, and IL-6 (Fig. 2B). The MDMAMB468 control did not show any further increases in cytokine production from Rab7 knockout as Rab7 is already inactivated in PTEN null cells, and the addition of ADU- SI 00 to scramble cells and Rab7 knockout cells, did not show any additive effects. However, MDA- MB231 cells with Rab7 knockout not only led to increased CXCL10 production but also increases in IL-8 and IL-6, both at baseline and in response to ADU-S100 stimulation. Similarly, HCC1806 Rab7 knockout cells showed an increased in CXCL10 level which was further enhanced with ADU-S100. There was also a significant increase in CCL5 (Fig. 2H), IL-8, and IL-6 with ADU-S100 stimulation of Rab7 knockout HCC1806 cells versus scramble.

[0079] Given that activating cytotoxic T lymphocytes depends on successful trafficking and infiltration to tumor tissue (27) and the cytokines that are upregulated by Rab7 knockout are largely involved in T cell trafficking, the potential in vivo implications for immune cell recruitment was modeled utilizing a previously described 3D microfluidic T-cell migration assay (Fig. 2C) (28). MDA-MB231 scramble and Rab7 knockout spheroids were pretreated with 50pM ADU-S100 and loaded into the migration zone of the 3D microfluidic device. Primary human CD8+ T cells were then introduced to a side channel, and T cell trafficking into the migration zone was evaluated after 48 hours. As shown in Fig. 2D, minimal T cells migrated towards the MDAMB231 scrambled spheroids even with ADU-S100 treatment. Since Rab7 knockout in PTEN WT TNBCs increases CXCL10 levels, significant T cell migration towards the MDAMB231 Rab7 knockout spheroids was observed, and ADU-S100 treatment further increased T cell trafficking into the migration zone. HCC 1806 showed a similar T cell migration pattern (Fig. 2E).

[0080] To assess whether these findings could be duplicated using Rab7 inhibitor CID1067700, it was first confirmed that addition of Rab7 inhibitor increases CXCL10 levels in both untreated and ADU-S100 treated MDAMB231 and HCC1806 cells. Next, the dosage of the Rab7 inhibitor with ADU- S100 was optimized to achieve the highest increase in CXCL10 levels for MDAMB231 and HCC 1806 (Fig. 2F). Using MDAMB231, spheroids were treated with lOOnM Rab7 inhibitor in the presence or absence of 50pM ADU-S100 in the 3D microfluidic device and then CXCR3-expressing Jurkat cells were introduced from a side channel. Significant migration of Jurkat cells was observed with ADU-S100 pretreatment that was additionallyenhanced by the Rab7 inhibitor (Fig. 2G; Fig 21).

[0081] Taken together, these findings identify PTEN null or PTEN WT with Rab7 knockout / inhibition as a genomic context in TNBCs for which STING agonists can be applied to not only induce cell intrinsic toxicity but to recruit T cells to the TME.EXAMPLE 3Impregnation of STING agonist in a PEG hydrogel biopsy marker results in its local delivery into TNBC.

[0082] In human clinical trials, intratumoral administration is seen as a barrier to therapeutic efficacy (14). In current clinical practice, after a patient has a breast biopsy, a metal marker is deployed to mark the area of biopsy. More recently, the metallic markers have been embedded in hydrogels that swell to optimize their visualization on ultrasound. These markers are FDA approved and can be made from polyethylene glycol, collagen, and beta-glucan. Once hydrated, these markers can fully expand within 6 hours, with the PEG hydrogel increasing proportionately the most in size (Fig. 3A). In an example, dehydrated PEG measures 5x1x1 mm and at maximum expansion it is 14x4x4 mm and can absorb at least 350 l of ddH20, which can be used to deliver an amount of desired drug.

[0083] We embedded ADU-S100 into PEG hydrogel markers in attempt to retain it in the TME. To ensure that the PEG will expand as expected, the PEG was implanted into the subcutaneous tissue of a Balb / c mouse. Significant expansion of the PEG was observed by Day 4, maximum expansion between Days 11-15, and by Day 25 there was some decrease in the size of the PEG (Fig. 3B, Fig. 3K).

[0084] In human breast tissue, the PEG is noted to be visible even after 6-12 months. To assess whether the PEG hydrogel can release ADU-S100 in a controlled fashion, a THP-1 cellbased reporter assay was developed. THP-1 cells were first treated with phorbol 12-myristate 13- acetate to differentiate into macrophages that are sensitive to STING agonist treatment and secrete CXCL10. We first established a standard curve for which the amount of ADU-S100 secreted from the PEG can be deduced from the corresponding amount of CXCL10 released. THP-1 cells were treated with a known amount of ADU-S100 and after 24 hours, supernatants were harvested for CXCL10 ELISA analysis (Fig. 3C) to generate a standard curve (Fig. 3D). Next, lOOpg of ADU-S100 was reconstituted into 20 pl of ddH20 and the PEG hydrogel was allowed to absorb the drug. The PEG+ADU-S100 was then covered by collagen to mimic the invivo tissue barrier with 1 ml of media placed on top and replaced with fresh media every 24 hours for a continuous 5-day collection. The harvested conditioned media at the 5 time points was then added to differentiated THP-1 cells for CXCL10 analysis (Fig. 3E).

[0085] The amount of ADU-S 100 released at the various time points was deduced (Fig. 3F) by using the standard curve previously generated in Fig. 3D. As expected, the PEG hydrogel was able to release ADU-S100 in a slow sustained fashion with release of 70pg of the embedded lOOpg by Day 5. Unlike the PEG hydrogel, the collagen and beta-glucan breast markers could not release ADU-S100 (Fig. 3G).

[0086] Next, the TNBC 4T1 model described above was treated with PEG hydrogel embedded with lOOpg of ADU-S100 versus PBS. Given the aggressiveness of the 4Tlmouse model, the PEG hydrogels were first implanted in a low tumor burden model at Day 4 after injecting 10K 4T1 cells into the 4th mammary fat pad. PEG+ADU-S100 prevented the 4T1 cells from establishing a tumor whereas PEG+PBS showed robust tumor growth (Fig. 3H). However, when PEG+ADU-S100 was implanted adjacent to an established 4T1 tumor, growth was inhibited initially, and by Day 20 there was exponential growth. (Fig. 31). At Day 25, the PEG marker could be easily identified adjacent to the tumor, and resection showed that the PEG marker was still intact (Fig. 3 J).

[0087] These data show that the PEG hydrogel marker is a new drug delivery method for ADU-S100.

[0088] In another experiment, we compared the results shown in Fig. 3H and 31 with direct intratumoral injection of ADU-S 100 and found minimal response in tumor growth compared to PEG delivery (Fig. 5A).EXAMPLE 4STING agonist monotherapy is toxic to T cells and does not result in T cell clonality.

[0089] T cells can become hyperresponsive to STING agonist, activating a distinct set of genes that results in apoptosis (34, 35). To investigate this potential toxicity, both a primary human TNBC explant and human CD8+ T cells were used. Explants were made from a patient’s TNBC as previously described (35) and were treated with PBS or 50pM ADU-S100 for 24 hours. The TNBC were examined by flow cytometry and showed a slight increase in CD45+ leukocytes and a small decrease in lymphocytes with ADU-S 100. A marked decrease in CD3+, CD4+ and CD8+ T cells was observed with ADU-S 100 treatment (Fig. 4A and 4J). Treatment ofprimary human CD8+ T cells with increasing concentrations of ADU-S100 showed a corresponding increase in T cell death (Fig. 4B).

[0090] Next, splenocytes were obtained from wild-type mice and the T population was sorted by flow cytometry into CD3+, CD4+, and CD8+ subpopulations. Each subpopulation was then treated with increasing concentrations of ADU-S100 for 24 hours and showed a corresponding increase in cell death, with CD8+ T cells being the most sensitive to treatment (Fig. 4C).

[0091] Given that PEG hydrogel can deliver STING agonist in a controlled sustained fashion (Fig. 3F), it was hypothesized that embedding ADU-S100 in the hydrogel can limit T cell toxicity. For example, if lOOpg of ADU-S100 is injected, the T cells are immediately exposed to the full dose of the drug which would likely result in immediate toxicity, whereas if the lOOpg is delivered over time in a controlled fashion that toxicity will likely be minimized. As such, human CD8+ T cells were plated in triplicate and then were treated with direct addition of PBS, direct addition of ADU- SI 00, PEG+PBS, or PEG+ADUS100. T cell toxicity was significantly higher with the direct ADU-S100 treatment versus embedding the drug in the PEG (Fig. 4D).EXAMPLE 5Additional studies of STING agonist delivery using a PEG hydrogel biopsy marker.

[0092] Based upon data shown in Example 3, we continued investigation of the potential of this PEG marker to deliver ADU-S100 in vivo using the well-established TNBC 4T1 syngeneic model (Aslakson, Cheryl J., and Fred R. Miller. "Selective events in the metastatic process defined by analysis of the sequential dissemination of subpopulations of a mouse mammary tumor." Cancer research 52.6, 1992, 1399-1405).

[0093] To determine whether the decrease in tumor size coincided with on-target tumor intrinsic effects of ADU-S100 being delivered by the PEG in this model, treated 4T1 mouse tumors were stained for pTBK l and pSTAT l at Day 4, revealing evidence of on target STING activation (Fig. 5B). Next, to better characterize the TME effects of ADU-S100 delivered by the PEG, we performed single cell RNA sequencing (scRNA-seq) on CD45+immune cells from 4T1 tumors 14 days after treatment (Fig. 5C). Uniform Manifold Approximation and Projection (UMAP) clustering revealed 5 different immune cell types (Fig. 5D), and we found that antigen presenting cell (APC) and macrophage clusters exhibed robust upregulation of IFN-induced genes by ADU-S100, includingwell assignificant enrichment for hallmark IFN gene signatures (Figs. 5E and 5F). Taken together, these findings confirmed effective PEG marker delivery of ADU-S100 into the TME.References

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[0141] Those skilled in the art will recognize, or be able to ascertain, using no more than routine experimentation, numerous equivalents to the specific substances and procedures described herein. Such equivalents are considered to be within the scope of this invention and are covered by the following claims.

Claims

CLAIMSWhat is claimed:

1. A method for delivering a therapeutic agent to a subject to treat a solid tumor, comprising implanting into the subject a biocompatible polymer including the therapeutic agent, wherein the biocompatible polymer (plug) continuously releases the therapeutic agent in the subject over time.

2. The method of claim 1, wherein the solid tumor is in a human.

3. The method of claim 2, wherein the solid tumor comprises a cancer.

4. The method of claim 3, wherein the cancer comprises a carcinoma, lymphoma, melanoma or sarcoma.

5. The method of claim 3, wherein the cancer comprises bladder, bone, brain, breast, cervical, colorectal, gastrointestinal, kidney, liver, lung, ovarian, pancreatic, prostate, skin or thyroid cancer.

6. The method of claim 5, wherein the breast cancer comprises ductal carcinoma in situ (DCIS), HER2-positive breast cancer, invasive ductal carcinoma (IDC), invasive lobular carcinoma (ILC), mixed ductal and lobular carcinoma, inflammatory breast cancer or triple-negative breast cancer (TNBC).

7. The method of claim 5, wherein the TNBC has a mutated or absent phosphatase and tensin homolog (PTEN).

8. The method of claim 2, wherein the solid tumor is a primary solid tumor or a metastatic solid tumor.

9. The method of claim 2, wherein the solid tumor is not resected prior to the implanting.

10. The method of claim 9, wherein the biocompatible polymer is implanted in proximity to the solid tumor.

11. The method of claim 10, wherein the biocompatible polymer is implanted within 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 mm of the solid tumor.

12. The method of claim 9, wherein the biocompatible polymer is implanted within the solid tumor.

13. The method of claim 9, wherein the biocompatible polymer is implanted in the periphery of the solid tumor.

14. The method of claim 2, wherein the solid tumor is resected prior to the implanting.

15. The method of claim 1, wherein the biocompatible polymer comprises a biodegradable polymer, a non-biodegradable polymer or a natural polymer.

16. The method of claim 15, wherein the biodegradable polymer comprises a poly(dioxanone) (PDO), polyethylene glycol (PEG), poly(glycolic acid) (PGA), poly(lactic acid) (PLA), poly(lactic-co-glycolytic acid) (PLGA), poly(caprolactone) (PCL), or combinations thereof.

17. The method of claim 15, wherein the non-biodegradable polymer comprises a poly(siloxane) (silicone) or poly(ethylene-vinyl acetate).

18. The method of claim 15, wherein the natural polymer comprises albumin, cellulose, chitosan, collagen elastin, gelatin, a milk protein or silk.

19. The method of claim 15, wherein the biocompatible polymer comprises a biodegradable polymer that remains in the subject for days, weeks, or months.

20. The method of claim 1, wherein the biocompatible polymer comprises a poly (urethane).

21. The method of claim 1, wherein the biocompatible polymer is hydrophilic.

22. The method of claim 1, wherein the biocompatible polymer comprises a hydrogel.

23. The method of claim 22, wherein the hydrogel comprises pores of 10-500 pm.

24. The method of claim 23, wherein spacing between pores in the hydrogel is from about 5 to about 100 nm.

25. The method of claim 22, wherein the hydrogel comprises a chemically-linked hydrogel or a physically-linked hydrogel.

26. The method of claim 22, wherein the biocompatible polymer comprises a PEG-based hydrogel.

27. The method of claim 1, wherein the biocompatible polymer comprises: a core comprising poly(dioxanone) (PDO), polylactide (PLA), polyglycollide (PGA), polycaprolactone (PCL), or copolymers thereof; and a sealant surrounding the core, comprising a polyethylene glycol (PEG)-based hydrogel.

28. The method of claim 27, wherein the core additionally comprises a titanium or stainless-steel marker.

29. The method of claim 1, wherein the biocompatible marker comprises a biopsy marker.

30. The method of claim 29, wherein the biopsy marker comprises a PEGylated substance, collagen, 0-glucan or polylactic-co-glycolic acid.

31. The method of claim 30, wherein the biopsy marker comprises an aqueous-expandable polymer.

32. The method of claim 29, wherein the biopsy marker comprises a HydroMARK™ biopsy marker.

33. The method of claim 29, wherein the biopsy marker is selected from the group consisting of a MammoMARK®, MammoSTAR® and SecurMark® biopsy marker.

34. The method of claim 1, wherein the therapeutic agent comprises a small molecule, large molecule, cell or combinations thereof.

35. The method of claim 34, wherein the therapeutic agent comprises a small molecule.

36. The method of claim 35, wherein the small molecule comprises a STING agonist.

37. The method of claim 36, wherein the STING agonist comprises ADU-S100, MK-1454, MK- 2118, SB11285, GSK3745417, BMS-986301, BI-STING, E7766, TAK-676, SNX281, SYNB1891, or combinations thereof.

38. The method of claim 35, wherein the small molecule comprises an estrogen receptor-targeted therapy.

39. The method of claim 38, wherein the estrogen-receptor targeted therapy comprises a selective estrogen receptor modulator (SERM).

40. The method of claim 39, wherein the SERM comprises tamoxifen or raloxifene.

41. The method of claim 1, wherein the biocompatible polymer is approximately 17, 15, 13, 11, 9 or 7 mm long when hydrated.

42. The method of claim 1, wherein the implanting comprises surgical implanting or implanting using a large-bore needle.

43. The method of claim 1, wherein multiple biocompatible polymer plugs are implanted.

44. The method of claim 2, wherein the method is used in combination with a conventional therapy.

45. The method of claim 44, wherein conventional therapy comprises chemotherapy or endocrine therapy.

46. A method for treating a solid tumor in a human subject, comprising: a. hydrating a dehydrated biopsy marker with a therapeutic agent; and b. implanting the hydrated biopsy marker in proximity to the solid tumor in the subject.

47. The method of claim 46, wherein the biopsy marker comprises a metal clip embedded in a biocompatible polymer.

48. The method of claim 46, wherein the biopsy marker comprises: a. a core comprising poly(dioxanone) (PDO), polylactide (PLA), polyglycollide (PGA), polycaprolactone (PCL), or copolymers thereof; and b. a sealant surrounding the core comprising a polyethylene glycol (PEG)-based hydrogel.

49. The method of claim 46, wherein the biopsy marker comprises a HydroMARK™ biopsy marker.

50. The method of claim 46, wherein the therapeutic agent comprises a small molecule.

51. The method of claim 50, wherein the small molecule comprises a STING agonist.

52. The method of claim 46, additionally comprising injecting a substance into the implanted biopsy marker subsequent to the implanting.

53. A method for treating a breast cancer in a human patient, comprising: hydrating a dehydrated biopsy marker with a STING agonist; wherein the dehydrated biopsy marker comprises a polyethylene glycol (PEG)- based hydrogel; and implanting the hydrated biopsy marker into the patient in proximity to the cells of the breast cancer.

54. The method of claim 53, wherein the dehydrated biopsy marker comprises a HydroMARK™ biopsy marker.

55. A method for loading a biopsy marker with a therapeutic agent, comprising hydrating the biopsy marker with an aqueous solution comprising the therapeutic agent to obtain a hydrated combination.

56. The method of claim 55, wherein the biopsy marker is hydrated with a volume of aqueous solution of less than about 20 pl.

57. The method of claim 55, additionally comprising implanting the hydrated combination into a human subject for sustained release of the therapeutic agent.

58. The method of claim 55, wherein the biopsy marker comprises a combination biodegradable polymer encompassed by a dehydrated hydrogel.

59. The method of claim 55, wherein the biopsy marker comprises a HydroMARK™ biopsy marker.

60. The method of claim 55, wherein the therapeutic agent comprises a small molecule.61 . The method of claim 60, wherein the small molecule comprises a STING agonist.

62. A method for treating a phosphatase and tensin homolog (PTEN)-null, triple negative breast cancer (TNBC), comprising without resecting the breast cancer, implanting a hydrated HydroMARK™ biopsy marker having a STING agonist in proximity to the breast cancer.

63. The method of claim 62, wherein the implanted HydroMARK™ biopsy marker additionally has a Rab7 inhibitor.

64. A HydroMARK™ biopsy marker comprising a STING agonist.

65. A composition, comprising: a biocompatible polymer that includes a metal marker; and a therapeutic agent absorbed into and / or adsorbed onto the biocompatible polymer.

66. The composition of claim 65, wherein the biocompatible polymer is hydrated.

67. The composition of claim 65, wherein the biocompatible polymer comprises a polymer core that includes the metal marker, surrounded by a polyethylene glycol (PEG)-based hydrogel.

68. The composition of claim 67, wherein the polymer core comprises a poly(dioxanone) (PDO), polyethylene glycol (PEG), poly(glycolic acid) (PGA), poly(lactic acid) (PLA), poly(lactic-co- glycolytic acid) (PLGA), poly(caprolactone) (PCL), or combinations thereof.

69. The composition of claim 65, wherein the metal marker comprises a titanium or stainless- steel marker.

70. The composition of claim 65, wherein the biocompatible polymer comprises a biopsy marker.71 . The composition of claim 70, wherein the biopsy marker comprises a HydroMARK™, MammoMARK®, MammoSTAR® or SecurMark® biopsy marker.

72. The composition of claim 70, wherein the biopsy marker comprises a combination biodegradable polymer encompassed by a hydrogel.

73. The composition of claim 72, wherein the biopsy marker comprises a HydroMARK™ biopsy marker.

74. The composition of claim 65, wherein the therapeutic agent comprises a small molecule.

75. The composition of claim 65, wherein the therapeutic agent comprises a STING agonist.

76. The composition of claim 75, wherein the STING agonist comprises ADU-S100, MK-1454, MK-2118, SB11285, GSK3745417, BMS-986301, BI-STING, E7766, TAK-676, SNX281, SYNB1891, or combinations thereof.

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