Nucleic Acid Compositions and Uses Thereof

Abstract

Compositions useful for the treatment and / or prevention of cancer and metastatic tumors are provided. For example, the compositions provided include biodegradable scaffolds or biomaterials that can act to promote or inhibit immune regulatory pathways and / or that can encode peptides that can promote or inhibit immune regulatory pathways. Methods for making the provided compositions, and kits containing materials for providing the same, are also provided.

Description

[Technical field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of U.S. Provisional Application No. 63 / 280,429, filed November 17, 2021, the contents of which are incorporated herein in their entirety. [Background technology]

[0002] Systemic administration of drugs, nutrients, or other substances into the circulatory system affects the entire body. Systemic administration routes include enteral administration (e.g., oral administration resulting in absorption of the drug via the digestive tract) and parenteral administration (e.g., intravenous, intramuscular, and subcutaneous injection). Administration of immunotherapeutic agents usually relies on these systemic administration routes, which may result in undesirable side effects. In some instances, certain promising therapeutic agents are extremely difficult to develop due to associated toxicities and limitations of current administration methods and systems. Surgery is often the first choice of treatment for solid tumor cancer, and is generally used in combination with systemic administration of anticancer therapy. However, surgery-induced immunosuppression is involved in the development of postoperative sepsis complications and tumor metastasis due to various metabolic and endocrine response changes, ultimately resulting in the death of many patients (Smyth, MJ et al. Nature Reviews Clinical Oncology, 2016, 13, 143-158). [Prior art documents] [Non-patent literature]

[0003] [Non-Patent Document 1] Smyth,MJet al.Nature Reviews Clinical Oncology,2016,13,143-158 Summary of the Invention [Means for solving the problem]

[0004] Systemic administration of immunotherapy may result in adverse side effects, e.g., inducing undesirable toxicity to non-cancerous cells and / or tissues, such as non-tumor-specific immune cells, and / or requiring high doses to achieve sufficient concentrations at the target site to induce a therapeutic response, and surgical removal of tumors may result in immunosuppression. Surgery may also induce cellular stress, which may involve activation of one or more physiological responses that promote wound healing after injury, for example. Such responses may include, for example, activation of neural, inflammatory, and / or pro-angiogenic signaling pathways, which may also promote cancer growth and / or metastatic spread. Inflammatory changes that may occur at the surgical site after tumor resection may include, for example, recruitment of immune and / or inflammatory cell type(s) and / or release of humoral factor(s). Local inflammatory wound response and systemic inflammatory processes together may activate dormant micrometastases or induce proliferation of residual cancer cells, thus increasing the risk of cancer recurrence.

[0005] The present disclosure provides insights, including identification of the causes of problems in certain prior art, including, inter alia, certain conventional approaches to cancer treatment. For example, the present disclosure understands that certain adverse events (e.g., skin rash, hepatitis, diarrhea, colitis, hypophysitis, thyroiditis, and adrenal insufficiency) that may be associated with systemic administration of immunotherapeutic agents may be immune-related and may be due, at least in part, to exposure of non-tumor specific immune cells to the systemically administered immunotherapeutic agents. In particular, the present disclosure understands that the high doses typically required for systemic administration to achieve sufficient concentrations in the tumor to induce a desired response may contribute to and / or be the cause of such undesirable effects. The present technology provides a system that solves such problems by, inter alia, providing localized delivery of immunotherapeutic agents that can improve efficacy by focusing the action of the drug as required. The present technology provides a system that solves such problems by providing localized delivery of immunotherapeutic agents that include nucleic acids or products encoded by nucleic acids.

[0006] Furthermore, the present disclosure provides insight that certain immunomodulatory agents traditionally used to treat autoimmune pathologies, when administered as described herein, may be useful in the treatment of cancer, despite otherwise expected off-target toxicity as opposed to that expected for anti-cancer immunomodulatory compounds. Furthermore, the present disclosure recognizes that certain immunomodulatory agents may be nucleic acids or products encoded by nucleic acids. Thus, the present disclosure teaches the utility of agents not previously believed to be useful for cancer therapy, and further teaches particularly effective and / or desirable delivery and dosing strategies for these and other agents.

[0007] In one aspect, the disclosure provides a method comprising administering to a target site in a subject for tumor resection a composition comprising: (i) a polymeric biomaterial; and (ii) a polynucleotide agent that encodes or modulates an immune-modulating polypeptide. In some embodiments, the polynucleotide agent encodes a cytokine that induces innate and / or adaptive immunity. In some, the polynucleotide agent activates a pattern recognition receptor that induces innate immunity. In some embodiments, the polynucleotide agent encodes a chemokine that induces immune cell recruitment. In some embodiments, the polynucleotide agent encodes an antibody that mediates immune checkpoint blockade or costimulation.

[0008] In some embodiments, the target site is a tumor resection site. In some embodiments, the target site is a site near the tumor resection site. In some embodiments, the target site is a sentinel lymph node. In some embodiments, the administering step is by injection.

[0009] In some embodiments, the composition is a liquid and the polymeric biomaterial is a viscous polymer solution. In some embodiments, the composition is a liquid and the polymeric biomaterial forms a polymeric network biomaterial in situ at the target site upon administration. In some embodiments, the polymeric network biomaterial comprises or is a crosslinked polymeric network biomaterial. In some embodiments, the polymeric network biomaterial comprises or is a non-crosslinked polymeric network biomaterial.

[0010] In some embodiments, the polymeric biomaterial is characterized in that when tested in vitro by placing the combination of the polymeric biomaterial and the polynucleotide agent in PBS (pH 7.4), less than 100% of the polynucleotide agent is released from the polymeric biomaterial within 3 hours. In some embodiments, the polymeric biomaterial is characterized in that when tested in vivo by administering the combination of the polymeric biomaterial and the polynucleotide agent to the mammary fat pad of a mouse subject, 50% or less of the polynucleotide agent is released in vivo 8 hours after administration. In some embodiments, the polymeric biomaterial is characterized in that when tested in vivo by administering the combination of the polymeric biomaterial and the polynucleotide agent to the mammary fat pad of a mouse subject, the polymeric biomaterial extends the release of the polynucleotide agent such that more polynucleotide agent is present in the mammary fat pad when evaluated 24 hours after administration than is observed when the polynucleotide agent is administered in solution. In some embodiments, the polymeric biomaterial is characterized in that the polynucleotide agent is released from the polymeric biomaterial and taken up by local cells such that (i) at least a subset of local immune cells express the immunomodulatory polypeptide encoded by the polynucleotide agent, (ii) at least a subset of local immune cells have increased expression of type 1 interferon in response to innate immune stimulation induced by the polynucleotide agent, and / or (iii) at least a subset of local immune cells have a change in the level and / or activity of the immunomodulatory polypeptide.

[0011] In some embodiments, the polymeric biomaterial is characterized by a storage modulus of about 10 Pa to about 5,000 Pa. In some embodiments, the polymeric biomaterial comprises or is a hydrogel. In some embodiments, the polymeric biomaterial comprises a positively charged polymer.

[0012] In some embodiments, the polymeric biomaterial further comprises a polynucleotide agent carrier. In some embodiments, the polynucleotide agent is complexed to the polynucleotide agent carrier. In some embodiments, the polynucleotide agent is supported on the polynucleotide agent carrier. In some embodiments, the polynucleotide agent carrier comprises a cationic agent and / or a lipid.

[0013] In some embodiments, the administering step does not involve administration of a tumor antigen to the subject of tumor resection. In some embodiments, the administering step does not involve administration of a microparticle to the subject of tumor resection. In some embodiments, the administering step does not involve adoptive transfer of immune cells to the subject of tumor resection.

[0014] In some embodiments, the composition further comprises an inhibitor of a proinflammatory immune response mediated by the p38 mitogen-activated protein kinase (MAPK) pathway. In some embodiments, the composition further comprises an activator of innate immunity. In some embodiments, the activator of innate immunity is or comprises a stimulator of interferon genes (STING) agonist. In some embodiments, the activator of innate immunity is or comprises a Toll-like receptor (TLR) 7 and / or TLR8 ("TLR7 / 8") agonist. In some embodiments, the composition further comprises an activator of adaptive immunity.

[0015] In some embodiments, the polymeric biomaterial forms a matrix or depot and the polynucleotide agent is within the polymeric biomaterial. In some embodiments, the polynucleotide agent is released by diffusion through the polymeric biomaterial. In some embodiments, the polymeric biomaterial is biodegradable in vivo.

[0016] In some embodiments, the tumor resection site is characterized by the absence of macroscopic residual tumor antigens. In some embodiments, the tumor resection subject is afflicted with metastatic cancer. In some embodiments, the method further comprises monitoring at least one metastatic site in the tumor resection subject after administration of a composition comprising (i) a polymeric biomaterial and (ii) a polynucleotide agent that encodes or modulates an immune-modulating polypeptide to a target site in the tumor resection subject.

[0017] In one aspect, the disclosure provides a method of producing a polymer network biomaterial composition, the method comprising: (a) providing a composition comprising one or more precursor components of a polymer network biomaterial and a polynucleotide agent that encodes or modulates an immunomodulatory polypeptide; and (b) allowing the precursor components to form a polymer network biomaterial in less than 10 minutes, wherein the polymer network biomaterial has a storage modulus of less than 5,000 Pa, and / or when tested in vitro by placing the combination of the polymer network biomaterial and the polynucleotide agent in PBS (pH 7.4), less than 100% of the polynucleotide agent is released from the polymer network biomaterial within 3 hours, and when tested in vivo by administering the combination of the polymer network biomaterial and the polynucleotide agent to the mammary fat pad of a mouse subject, 50% or less of the polynucleotide agent is released in vivo 8 hours after administration, and the polymer network biomaterial is capable of being biocompatible with the polymer network biomaterial. and allowing the polynucleotide agent released from the polymer network biomaterial to form a polymeric network biomaterial characterized in that, when tested in vivo by administering a combination of the polymer network biomaterial and polynucleotide agent to the mammary fat pad of a mouse subject, the polymer network biomaterial extends the release of the polynucleotide agent such that, when evaluated 24 hours after administration, more polynucleotide agent is present in the mammary fat pad than observed when the polynucleotide agent is administered in solution, and / or the polynucleotide agent released from the polymer network biomaterial forms a polymeric network biomaterial characterized in that (i) at least a subset of local immune cells express an immunomodulatory polypeptide encoded by the polynucleotide agent, (ii) at least a subset of local immune cells have increased expression of type 1 interferon in response to innate immune stimulation induced by the polynucleotide agent, and / or (iii) at least a subset of local immune cells have a change in the level and / or activity of the immunomodulatory polypeptide.

[0018] In some embodiments, the polymer network biomaterial comprises or is a crosslinked polymer network biomaterial. In some embodiments, the polymer network biomaterial comprises or is a non-crosslinked polymer network biomaterial. In some embodiments, the polymer network biomaterial comprises or is a hydrogel. In some embodiments, the polymer network biomaterial comprises a polynucleotide agent carrier. In some embodiments, the polynucleotide agent is complexed to the polynucleotide agent carrier. In some embodiments, the polynucleotide agent is supported on the polynucleotide agent carrier. In some embodiments, the polynucleotide agent carrier comprises a cationic agent and / or a lipid.

[0019] In one aspect, the disclosure provides a method of characterizing a polymer network biomaterial composition or a component(s) thereof, the method comprising: (a) providing a polymer network biomaterial composition in a buffer solution in vitro, the polymer network biomaterial composition comprising a candidate polymer biomaterial and a polynucleotide agent encoding or modulating an immunomodulatory polypeptide; and (b) determining whether less than 100% of the polynucleotide agent is released from the polymer biomaterial within 3 hours. In some embodiments, the buffer solution comprises or is PBS (pH 7.4). In some embodiments, the determining step further comprises measuring the amount of polynucleotide agent released from the polymer biomaterial at a predetermined time point over a period of at least 3 hours. In some embodiments, the determining step further comprises determining the release profile kinetics of the polynucleotide agent from the polymer biomaterial. In some embodiments, the method further comprises selecting a polymer network biomaterial composition characterized in that less than 100% of the polynucleotide agent is released from the polymer biomaterial within 3 hours.

[0020] In some embodiments, the method further comprises contacting the selected polymeric network biomaterial composition with a cell population and determining (a) whether the polynucleotide agent released from the candidate polymeric biomaterial is taken up by an immune cell, and / or (b) whether the immune cell taking up the polynucleotide agent exhibits at least one of the following biological activities: (i) expressing an immunomodulatory polypeptide encoded by the polynucleotide agent, (ii) exhibiting increased expression of type 1 interferon in response to innate immune stimulation induced by the polynucleotide agent, and / or (iii) exhibiting a change in the level and / or activity of the immunomodulatory polypeptide. In some embodiments, the immune cell comprises a bone marrow cell and / or a plasmacytoid dendritic cell. In some embodiments, the cell further comprises a non-immune cell. In some embodiments, the non-immune cell comprises a fibroblast and / or an endothelial cell. In some embodiments, the method further comprises determining (a) whether the polynucleotide agent released from the candidate polymeric biomaterial is taken up by the non-immune cell, and / or (b) whether the polynucleotide agent adversely affects the non-immune cell.

[0021] In one aspect, the disclosure provides a method of characterizing a polymer network biomaterial composition or a component(s) thereof, the method comprising: (a) administering a polymer network composition to a target site in a mouse subject in vivo, the polymer network biomaterial composition comprising a candidate polymer biomaterial and a polynucleotide agent that encodes or modulates an immunomodulatory polypeptide, and (b) determining whether (i) the candidate polymer biomaterial extends release of the polynucleotide agent at the target site for at least 24 hours upon administration relative to administration of the same polynucleotide agent in a solution without the polymer biomaterial, and / or (ii) whether the release profile of the polynucleotide agent from the candidate polymer biomaterial is characterized by no more than 50% of the polynucleotide agent being released in vivo 8 hours after administration. In some embodiments, the target site is a mammary fat pad. In some embodiments, the method further comprises selecting a polymer network biomaterial composition characterized in that (i) the candidate polymeric biomaterial, upon administration, extends the release of a polynucleotide agent at a target site for at least 24 hours relative to administration of the same polynucleotide agent in a solution without the polymeric biomaterial, and / or (ii) no more than 50% of the polynucleotide agent is released from the candidate polymeric biomaterial in vivo 8 hours after administration.

[0022] In some embodiments, the method further comprises determining (a) whether the polynucleotide agent released from the candidate polymeric biomaterial is taken up by an immune cell, and / or (b) whether the immune cell that takes up the polynucleotide agent exhibits at least one of the following biological activities: (i) expressing an immunomodulatory polypeptide encoded by the polynucleotide agent, (ii) exhibiting increased expression of type 1 interferon in response to innate immune stimulation induced by the polynucleotide agent, and / or (ii) exhibiting a change in the level and / or activity of the immunomodulatory polypeptide. In some embodiments, the method further comprises determining (c) whether cellular uptake of the polynucleotide agent released from the candidate polymeric biomaterial is delayed compared to when the cell is contacted with the polynucleotide agent in the absence of the candidate polymeric biomaterial, and / or (d) whether at least one of the biological activities induced by the polynucleotide agent is delayed compared to when the cell is contacted with the polynucleotide agent in the absence of the candidate polymeric biomaterial. In some embodiments, the immune cell comprises a myeloid cell and / or a plasmacytoid dendritic cell. In some embodiments, the method further comprises determining (e) whether the polynucleotide agent released from the candidate polymeric biomaterial is taken up by a non-immune cell, and / or (f) whether the polynucleotide agent adversely affects a non-immune cell. In some embodiments, the non-immune cell comprises a fibroblast and / or an endothelial cell.

[0023] In one aspect, the disclosure provides a method comprising administering to a target site in a subject undergoing tumor resection a composition comprising (i) a polynucleotide agent carrier and (ii) a polynucleotide agent that encodes or modulates an immune modulating polypeptide. In some embodiments, the polynucleotide agent carrier comprises a cationic agent and / or a lipid. [Brief description of the drawings]

[0024] [Figure 1]1 is a graphical representation of a survival analysis. Data are presented as Kaplan-Meier survival curves showing the survival of female BALB / cJ mice orthotopically inoculated with 4T1-Luc2 cells. These cells were allowed to generate tumors, which were then surgically resected. Immediately after tumor resection, an exemplary composition comprising a polymer network (crosslinked hyaluronic acid hydrogel) and an innate immune system agonist (RIG-1 agonist) was implanted at the tumor resection site. Administration of the localized composition promoted sustained release of the active agent (RIG-1 agonist) and resulted in a significantly increased lifespan of the animals when compared to control implants lacking the active agent. [Diagram 2] 1 is a graphical representation of a survival analysis. Data are presented as Kaplan-Meier survival curves showing the survival of female BALB / cJ mice orthotopically inoculated with 4T1-Luc2 cells. The cells were allowed to generate tumors, which were then surgically resected. Immediately after tumor resection, an exemplary composition comprising a polymer network (poloxamer and hyaluronic acid hydrogel) and an innate immune system agonist (RIG-1 agonist) was injected into the tumor resection site. Localized administration of the composition promoted sustained release of the active agent (RIG-1 agonist) and resulted in increased longevity of the animals when compared to control implants lacking the active agent. [Diagram 3] 1 is a graphical representation of a survival analysis. Data are presented as Kaplan-Meier survival curves showing the survival of female BALB / cJ mice orthotopically inoculated with 4T1-Luc2 cells. The cells were allowed to generate tumors, which were then surgically resected. Immediately after tumor resection, an exemplary composition comprising a polymer network (crosslinked hyaluronic acid hydrogel) and a nucleic acid encoding an innate immune system agonist (interleukin-15 (IL-15) mRNA) was implanted at the tumor resection site. Administration of the localized composition promoted sustained release of the active agent (IL-15 mRNA) and resulted in increased longevity of the animals when compared to control implants lacking the active agent. [Figure 4]1 is a graphical representation of a survival analysis. Data are presented as Kaplan-Meier survival curves showing the survival of female BALB / cJ mice orthotopically inoculated with 4T1-Luc2 cells. The cells were allowed to generate tumors, which were then surgically resected. Immediately after tumor resection, an exemplary composition comprising a polymer network (poloxamer hydrogel or poloxamer and hyaluronic acid hydrogel) and a nucleic acid encoding an innate immune system agonist (interleukin-15 (IL-15) mRNA) was injected into the tumor resection site. Localized administration of a composition containing hyaluronic acid resulted in an increase in the lifespan of the animals when compared to a composition lacking hyaluronic acid. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0025] A specific definition It should be noted that the concentrations of the individual polymeric components in the polymeric biomaterial(s) preparations described herein are each expressed as % (w / w) or weight %. As used herein, the concentration, % (by weight), of a polymeric component in a polymeric biomaterial(s) preparation is determined based on the mass or weight of the polymeric component relative to the sum of (i) the total mass or weight of all individual polymeric components present in the polymeric biomaterial(s) preparation, and (ii) the total mass or weight of solvent used in the polymeric biomaterial(s) preparation.

[0026] Activator of adaptive immune response: The term "activator of adaptive immune response" refers to an agent that activates (e.g., increases the activity of) the adaptive immune system (and / or one or more features of the adaptive immune system) in a subject (e.g., a subject to which it is administered and / or a subject otherwise in need thereof) compared to the absence of the agent. Such activation may restore or enhance anti-tumor function, for example, by neutralizing inhibitory immune checkpoints and / or triggering costimulatory receptors, ultimately generating helper and / or effector T cell responses against immunogenic antigens expressed by cancer cells, and generating memory B and / or T cell populations. In certain embodiments, the activator of adaptive immune response involves modulation of adaptive immune responses and / or leukocyte trafficking. Examples of activators of adaptive immune response include, for example, those described in WO2018 / 045058, the contents of which are incorporated herein by reference in their entirety for purposes described herein.

[0027] Activator of the innate immune response: The term "activator of the innate immune response" refers to an agent that activates (e.g., increases the activity of) the innate immune system (and / or one or more features of the innate immune system) in a subject (e.g., a subject to which it is administered and / or a subject otherwise in need thereof) compared to the absence of the agent. Such activation may stimulate (e.g., increase the expression level and / or activity of) one or more agents that initiate an inflammatory response (e.g., an immunostimulatory inflammatory response) and / or help induce an adaptive immune response, e.g., resulting in the development of antigen-specific acquired immunity. In some embodiments, activation of the innate immune system may result in the recruitment of relevant immune cells, including, but not limited to, neutrophils, basophils, eosinophils, natural killer cells, dendritic cells, monocytes, and macrophages, cytokine production, proliferation and / or survival of leukocytes, and improved T cell priming, e.g., by presentation of antigens by antigen presenting cells and / or increased expression levels and / or activity of costimulatory molecules. Examples of activators of the innate immune response include, for example, those described in WO2018 / 045058, the contents of which are incorporated by reference in their entirety for the purposes described herein.

[0028] Administration: As used herein, the terms "administer", "administering", or "administration" typically refer to administering a composition to a subject to achieve delivery of an agent or payload, which is or is contained in the composition, to a target site or site to be treated. Those skilled in the art will be aware of the various routes that may be utilized for administration of different agents to a subject, e.g., a human, in the appropriate context. For example, the terms "administer", "administering", or "administration", in the context of administering a composition, including a composition described herein, may refer to implantation, absorption, ingestion, injection, inhalation, parenteral administration, or other introduction of a composition described herein, although administering may refer in some embodiments to implantation, or in some embodiments to injection.

[0029] Agent: As used herein, the term "agent" may refer to a physical entity or phenomenon. In some embodiments, an agent may be characterized by a particular feature and / or effect. In some embodiments, an agent may be a compound, molecule, or entity of any chemical class, including, for example, a small molecule, a polypeptide, a nucleic acid, a sugar, a lipid, a metal, or a combination or complex thereof. In some embodiments, the term "agent" may refer to a compound, molecule, or entity that includes a polymer. In some embodiments, the term may refer to a compound or entity that includes one or more polymeric moieties. In some embodiments, the term "agent" may refer to a compound, molecule, or entity that is substantially free of a particular polymer or polymeric moiety. In some embodiments, the term may refer to a compound, molecule, or entity that lacks or is substantially free of any polymer or polymeric moiety.

[0030] Agonist: Those skilled in the art will understand that the term "agonist" can be used to refer to an agent, condition, or event whose presence, level, degree, type, or form correlates with an increase in the level and / or activity of another agent (i.e., an agonized agent) and / or an increase or induction of one or more biological events. In general, agonists can be or include agents of various chemical classes, including, for example, small molecules, polypeptides, nucleic acids, carbohydrates, lipids, metals, inorganic crystals, and / or any other entity that exhibits related activation activity. In some embodiments, an agonist can be direct (where the agonist exerts its effect directly on its target), and in some embodiments, an agonist can be indirect (where the agonist exerts its effect other than by binding to its target, for example, by interacting with a regulator of the target such that the level or activity of the target is altered). A partial agonist competes with a full agonist to interact with its target and / or its modulators and may therefore act as a competitive antagonist in the presence of a full agonist, thereby resulting in (i) a decrease in one or more effects of another agent and / or (ii) a decrease in one or more biological events compared to that observed with the full agonist alone.

[0031] Antagonist: Those skilled in the art will understand that the term "antagonist" may refer to an agent, condition, or event whose presence, level, extent, type, or form is associated with a decrease in the level and / or activity of another agent (i.e., an antagonized agent) and / or a decrease or suppression of one or more biological events. In general, antagonists may include agents from various chemical classes, including, for example, small molecules, polypeptides, nucleic acids, carbohydrates, lipids, metals, and / or any other entity that exhibits relevant inhibitory activity. In some embodiments, an antagonist may be a "direct antagonist" in that it directly binds to its target, and in some embodiments, an antagonist may be an "indirect antagonist" in that it exerts its effect by means other than directly binding to its target, for example, by interacting with a regulator of the target such that the level or activity of the target is altered.

[0032] Antibody: As used herein, the term "antibody" refers to a polypeptide that contains standard immunoglobulin sequence elements sufficient to confer specific binding to a particular target antigen. As is known in the art, naturally produced intact antibodies are approximately 150 kD tetrameric agents composed of two identical heavy chain polypeptides (about 50 kD each) and two identical light chain polypeptides (about 25 kD each) that associate with each other into what is commonly referred to as a "Y-shaped" structure. Each heavy chain is composed of at least four domains (each about 110 amino acids long): an amino-terminal variable (VH) domain (located at the tip of the Y structure), followed by three constant domains: CH1, CH2, and a carboxy-terminal CH3 (located at the base of the stem of the Y). A short region known as a "switch" connects the heavy chain variable and constant regions. A "hinge" connects the CH2 and CH3 domains to the rest of the antibody. Two disulfide bonds in this hinge region connect the two heavy chain polypeptides to each other in intact antibodies. Each light chain is composed of two domains, an amino-terminal variable (VL) domain separated from each other by another "switch", followed by a carboxy-terminal constant (CL) domain. An intact antibody tetramer is composed of two heavy-light chain dimers, with one disulfide bond linking the heavy and light chains together and two other disulfide bonds linking the heavy chain hinge regions together so that the dimers are connected to form a tetramer. Naturally produced antibodies are also typically glycosylated on the CH2 domain. Each domain in a natural antibody has a structure characterized by an "immunoglobulin fold" formed from two beta sheets (e.g., three-, four-, or five-stranded sheets) packaged together in a compressed antiparallel beta barrel. Each variable domain contains three hypervariable loops known as "complementarity determining regions" (CDR1, CDR2 and CDR3) and four somewhat invariant "framework" regions (FR1, FR2, FR3 and FR4).When a natural antibody folds, the FR regions form a beta sheet that provides the structural framework of the domain, and the CDR loop regions from both the heavy and light chains come together in three-dimensional space to create a single hypervariable antigen-binding site located at the tip of a Y-structure. The Fc region of a naturally occurring antibody binds to elements of the complement system and also to receptors on effector cells, including, for example, effector cells that mediate cytotoxicity. As is known in the art, the affinity and / or other binding properties of the Fc region for the Fc receptor can be modulated through glycosylation or other modifications. In some embodiments, the antibodies produced and / or utilized according to the present disclosure comprise a glycosylated Fc domain, including Fc domains with modified or engineered such glycosylation. For purposes of this disclosure, in certain embodiments, any polypeptide or polypeptide complex that includes a sufficient immunoglobulin domain sequence found in a natural antibody may be referred to and / or used as an "antibody," regardless of whether such polypeptide is produced naturally (e.g., generated by an organism in response to an antigen) or produced by recombinant engineering, chemical synthesis, or other artificial systems or methodologies. In some embodiments, an antibody is polyclonal, and in some embodiments, an antibody is monoclonal. In some embodiments, an antibody has constant region sequences that are characteristic of mouse, rabbit, primate, or human antibodies. In some embodiments, antibody sequence elements are humanized, primatized, chimeric, etc., as known in the art. Furthermore, the term "antibody," as used herein, may, in appropriate embodiments (unless otherwise specified or clear from the context), refer to any of the constructs or formats known or developed in the art for utilizing the structural and functional characteristics of antibodies in alternative presentations.For example, in some embodiments, the antibody utilized in accordance with the present disclosure is in a format selected from, but not limited to, an intact IgA, IgG, IgE or IgM antibody; a bispecific or multispecific antibody (e.g., Zybodies®, etc.); an antibody fragment such as a Fab fragment, a Fab' fragment, a F(ab')2 fragment, a Fd' fragment, a Fd fragment, and an isolated CDR or set thereof; a single chain Fv; a polypeptide-Fc fusion; a single domain antibody, an alternative scaffold or an antibody mimic (e.g., anti-cullins, FN3 monobodies, DARPins, Affibodies, Affilins, Affimers, Affitins, Alphabodies, Avimers, Fynomers, Im7, VLR, VNAR, Trimab, CrossMab, Trident); a nanobody, a binanobody, a F(ab')2, a Fab', a di-sdFv, a single domain antibody, a trifunctional antibody, a diabody, and a minibody, etc. In some embodiments, the relevant formats may be or include Adnectins®; Affibodies®; Affilins®; Anticalins®; Avimers®; BiTEs®; cameloid antibodies; Centyrins®; Ankyrin repeat proteins or DARPINs®; Dual Affinity Retargeting (DART) agents; Fynomers®; Shark single domain antibodies such as IgNAR; Immune Mobilizing Monoclonal T Cell Receptors against Cancer (ImmTAC); KALBITOR®; MicroProteins; Nanobodies® minibodies; masked antibodies (e.g., Probodies®); Small Modular ImmunoPharmaceuticals ("SMIPs™"); single chain or tandem diabodies (TandAb®); TCR-like antibodies; Trans-bodies®; TrimerX®; VHHs. In some embodiments, the antibody may lack covalent modifications (eg, glycan attachment) that it would have if produced naturally.In some embodiments, the antibody may include a covalent modification (e.g., the attachment of a glycan, a payload (e.g., a detectable moiety, a therapeutic moiety, a catalytic moiety, etc.), or other pendant group (e.g., polyethylene glycol, etc.).

[0033] Bioadhesive: The term "bioadhesive" refers to a biocompatible agent that can adhere to a target surface, e.g., a tissue surface. In some embodiments, the bioadhesive can adhere to a target surface, e.g., a tissue surface, and can remain on the target surface for a period of time. In some embodiments, the bioadhesive can be biodegradable. In some embodiments, the bioadhesive can be a natural agent that can be prepared or obtained, e.g., by isolation or synthesis, and in some embodiments, the bioadhesive can be a non-natural agent that can be designed and / or manufactured by the hand of man (e.g., by processing, synthesis, and / or recombinant production depending on the agent), as will be understood by those of skill in the art. In some particular embodiments, the bioadhesive can be or include a polymeric material that can be composed of or contain multiple monomers, such as, for example, sugars. Certain exemplary bioadhesives include cyanoacrylates (Dermabond, 2-octyl cyanoacrylate; Indermil, n-butyl-2-cyanoacrylate; Histoacryl and Histoacryl Blue, n-butyl-2-cyanoacrylate), albumin and glutaraldehyde (BioGlue™, bovine serum albumin and 10% glutaraldehyde), fibrin glues (Tisseel™, pooled human plasma fibrinogen and thrombin; Evicel™, pooled human plasma fibrinogen and thrombin; Vitagel™, autologous plasma fibrinogen and thrombin; Cryoseal™ system, autologous plasma fibrinogen and thrombin), gelatin and / or resorcinol crosslinked with formaldehyde and / or glutaraldehyde, polysaccharide-based adhesives (e.g., alginate, chitosan, collagen, dextran, and / or gelatin), PEG, acrylate, polyamine, or urethane variants (isocyanate terminated prepolymers), and / or combinations thereof.Other examples of bioadhesives known in the art, for example those described in Mehdizadeh and Yang “Design Strategies and Applications of Tissue Bioadhesives” Macromol Biosci 13:271-288 (2013), may be used for the purposes of the methods described herein. In some embodiments, the bioadhesive may be a degradable bioadhesive. Examples of such degradable bioadhesives include, but are not limited to, fibrin glue, gelatin-resorcinol-formaldehyde / glutaraldehyde adhesive, hydrogel adhesives based on poly(ethylene glycol) (PEG), polysaccharide adhesives, polypeptide adhesives, polymer adhesives, biomimetic bioadhesives, and those described in Bhagat and Becker “Degradable Adhesives for Surgery and Tissue Engineering” Biomacromolecules 18:3009-3039 (2017).

[0034] Biocompatible: The term "biocompatible" as used herein refers to a material that, for example, when placed in contact with living tissue in vivo, does not cause significant harm to such tissue. The biocompatibility of a material may be evaluated by the ability of such material to pass the biocompatibility tests set forth in International Standards Organization (ISO) Standard No. 10993, and / or US Pharmacopeia (USP) 23, and / or US Food and Drug Administration (FDA) Blue Book Memorandum No. G95-1 "Use of International Standard ISO-10993, Biological Evaluation of Medical Devices Part-1: Evaluation and Testing." Typically, these tests measure the toxicity, infectivity, pyrogenicity, irritation potential, reactivity, hemolytic activity, carcinogenicity, and / or immunogenicity of the material. In certain embodiments, a material is "biocompatible" if it is not itself toxic to cells in the in vivo environment of its intended use. In certain embodiments, materials are "biocompatible" if their addition to cells in vitro results in 20% or less cell death and / or if their administration in vivo does not induce significant inflammation or other such adverse effects that are clinically undesirable for the purposes described herein. As will be understood by those of skill in the art, such significant inflammation is distinguishable from the transient low-grade inflammation that typically accompanies surgery or the introduction of a foreign body into the body. Furthermore, those of skill in the art will understand upon reading this disclosure that, in some embodiments, the polymeric biomaterial(s) preparations described herein and / or their individual polymeric components are biocompatible if a degree of immune modulation (e.g., innate immune agonism) over a defined period of time is clinically beneficial and / or desirable, for example, to provide anti-tumor immunity.

[0035] Biologics: The terms "biologic," "biological drug," and "biological product" refer to a broad range of products, such as vaccines, blood and blood components, allergens, somatic cells, gene therapy, tissues, nucleic acids, and proteins. Biologics may contain carbohydrates, proteins, or nucleic acids, or complex combinations of these substances, or may be living entities such as cells and tissues. Biologics may be isolated from a variety of natural sources (e.g., human, animal, microbial) and / or produced by biotechnological methods and / or other techniques.

[0036] Biological sample: "Biological sample" refers to a primary sample obtained from a biological source and / or, in some embodiments, a sample derived therefrom (e.g., by processing). Those skilled in the art will understand that a biological sample may include or be selected from, for example, tissue samples (such as tissue sections and needle biopsies of tissues), cell samples (such as cytological smears (such as Pap or blood smears) or samples of cells obtained by microdissection), whole organism samples (such as yeast or bacterial samples), or cell fractions, fragments, or organelles (such as those obtained by lysing cells and separating their components by centrifugation or other methods). Other examples of biological samples include blood, serum, urine, semen, feces, cerebrospinal fluid, interstitial fluid, mucus, tears, sweat, pus, biopsy tissue (e.g., obtained by surgical or needle biopsy), nipple aspirate, milk, vaginal fluid, saliva, swabs (such as oral swabs), or any material containing biomolecules derived from a first biological sample.

[0037] Cancer: The term "cancer" refers to a malignant tumor (Stedman's Medical Dictionary, 25th ed.; Hensyl ed.; Williams & Wilkins: Philadelphia, 1990). Of particular interest in the context of some embodiments of the present disclosure are cancers that are treated by cell killing and / or ablative therapy (e.g., surgical resection and / or certain chemotherapeutic modalities such as cytotoxic therapy). In some embodiments, the cancers treated according to the present disclosure are those that have been surgically resected (i.e., at least one tumor has been surgically removed). In some embodiments, the cancers treated according to the present disclosure are those for which resection is the standard of care. In some embodiments, the cancers treated according to the present disclosure are those that have metastasized. In certain embodiments, exemplary cancers include acoustic neuroma, adenocarcinoma, adrenal carcinoma, anal carcinoma, angiosarcoma (e.g., lymphangioendotheliosarcoma, angiosarcoma), appendix cancer, benign monoclonal gammopathy, biliary tract cancer (e.g., cholangiocarcinoma), bile duct cancer, bladder cancer, bone cancer, breast cancer (e.g., adenocarcinoma of the breast, papillary carcinoma of the breast, mammary carcinoma, thyroid ... cancer), medullary carcinoma of the breast), brain cancer (e.g., meningioma, glioblastoma, glioma (e.g., astrocytoma, oligodendroglioma), medulloblastoma), bronchial carcinoma, carcinoid tumor, cardiac tumor, cervical cancer (e.g., cervical adenocarcinoma), choriocarcinoma, chordoma, craniopharyngioma, colorectal cancer (e.g., colon carcinoma, rectal carcinoma, colorectal adenocarcinoma), connective tissue carcinoma, epithelial carcinoma, ductal carcinoma in situ, ependymoma, endotheliosarcoma (e.g., Kaposi's sarcoma, multiple idiopathic hemorrhagic sarcoma), endometrial cancer (e.g., uterine carcinoma, uterine sarcoma), esophageal cancer (e.g., esophageal adenocarcinoma, Barrett's adenocarcinoma), Ewing's sarcoma, eye cancer (e.g., intraocular melanoma, retinoblastoma), familial hypereosinophilia hypereosinophilia), gallbladder cancer, stomach cancer (e.g., gastric adenocarcinoma), gastrointestinal stromal tumor (GIST), germ cell cancer, head and neck cancer (e.g., head and neck squamous cell carcinoma), oral cancer (e.g., oral squamous cell carcinoma), throat cancer (e.g., laryngeal cancer, pharyngeal cancer, nasopharyngeal cancer,oropharyngeal cancer), hematopoietic cancers (e.g., leukemias such as acute lymphocytic leukemia (ALL) (e.g., B-cell ALL, T-cell ALL), acute myeloid leukemia (AML) (e.g., B-cell AML, T-cell AML), chronic myeloid leukemia (CML) (e.g., B-cell CML, T-cell CML), and chronic lymphocytic leukemia (CLL) (e.g., B-cell CLL, T-cell CLL)), Hodgkin's lymphoma (HL) (e.g., B-cell HL, T-cell HL) and non-Hodgkin's lymphoma (NHL) (B-cell NHL, e.g., diffuse large cell lymphoma (DLCL) (e.g., diffuse large cell type B-cell lymphoma), follicular lymphoma, chronic lymphocytic leukemia / small lymphocytic lymphoma (CLL / SLL), mantle cell lymphoma (MCL), marginal zone B-cell lymphoma (e.g., mucosa-associated lymphoid tissue (MALT) lymphoma, nodal marginal zone B-cell lymphoma, splenic marginal zone B-cell lymphoma), primary mediastinal B-cell lymphoma, Burkitt lymphoma, lymphoplasmacytic lymphoma (i.e., Waldenström's macroglobulinemia), hairy cell leukemia (HCL), immunoblastic large cell lymphoma, precursor B-lymphoblastic lymphoma, and primary central nervous system (CNS) lymphoma lymphomas such as precursor T-lymphoblastic lymphoma / leukemia, peripheral T-cell lymphomas (PTCL) (e.g., T-cell NHL such as cutaneous T-cell lymphoma (CTCL) (e.g., mycosis fungoides, Sézary syndrome), angioimmunoblastic T-cell lymphoma, extranodal natural killer T-cell lymphoma, enteropathy-type T-cell lymphoma, subcutaneous panniculitis-like T-cell lymphoma, and anaplastic large cell lymphoma), mixtures of one or more of the above leukemias / lymphomas, multiple myeloma, heavy chain disease (e.g., alpha chain disease, gamma chain disease, mu chain disease), hemangioblastoma, histiocytosis, subcutaneous leukemia, bronchitis, and bronchitis. Pharyngeal cancer, inflammatory myofibroblastic tumor, immune cell amyloidosis, kidney cancer (e.g., nephroblastoma, also known as Wilms' tumor, renal cell carcinoma), liver cancer (e.g., hepatocellular carcinoma (HCC), malignant hepatoma), lung cancer (e.g., bronchogenic carcinoma, small cell lung cancer (SCLC), non-small cell lung cancer (NSCLC), adenocarcinoma of the lung), leiomyosarcoma (LMS), mastocytosis (e.g., systemic mastocytosis), melanoma, midline carcinoma, multiple endocrine neoplasia syndrome, muscle cancer, myelodysplastic syndrome (MDS), mesothelioma, myeloproliferative disorder (MPD) (e.g., polycythemia vera (PV), essential thrombocytosis (ET),Primary myelofibrosis (AMM), also known as myelofibrosis (MF), chronic idiopathic myelofibrosis, chronic myelogenous leukemia (CML), chronic neutrophilic leukemia (CNL), hypereosinophilic syndrome (HES)), nasopharyngeal carcinoma, neuroblastoma, neurofibroma (e.g., neurofibromatosis (NF) type 1 or 2, schwannomatosis), neuroendocrine carcinoma (e.g., gastroenteropancreatic neuroendocrine tumor (GEP-NET), carcinoid tumor), osteosarcoma (e.g., bone cancer), ovarian cancer (e.g., cystadenocarcinoma, ovarian embryonal carcinoma, ovarian adenocarcinoma), papillary adenocarcinoma, pancreatic cancer (e.g., pancreatic adenocarcinoma, intraductal papillary mucinous neoplasm (IPMN), islet cell tumor), parathyroid carcinoma, papillary adenocarcinoma, penile cancer (e.g., Paget's disease of the penis and scrotum), pharyngeal carcinoma, pinealoma, pituitary carcinoma, pleuropulmonary blastoma, primitive neuroectodermal tumor ( PNT), plasma cell neoplasms, paraneoplastic syndromes, intraepithelial neoplasia, prostate cancer (e.g., prostatic adenocarcinoma), rectal cancer, rhabdomyosarcoma, retinoblastoma, salivary gland cancer, skin cancer (e.g., squamous cell carcinoma (SCC), keratoacanthoma (KA), melanoma, basal cell carcinoma (BCC)), small intestine cancer (e.g., appendix ulcer), soft tissue sarcoma (e.g., malignant fibrous histiocytoma (MFH), liposarcoma, malignant peripheral nerve sheath tumor (MPNST), chondrosarcoma, fibrosarcoma, myxosarcoma), sebaceous gland carcinoma, gastric cancer, small intestine cancer, sweat gland carcinoma, synovium, testicular cancer (e.g., seminoma, testicular embryonal carcinoma), thymic cancer, thyroid cancer (e.g., papillary thyroid carcinoma, papillary thyroid carcinoma (PTC), medullary thyroid carcinoma), urethral cancer, uterine cancer, vaginal cancer, and vulvar cancer (e.g., Paget's disease of the vulva).

[0038] Carbohydrate polymer: The term "carbohydrate polymer" refers to a polymer that is or includes, for example, one or more carbohydrates having a carbohydrate backbone. For example, in some embodiments, a carbohydrate polymer refers to a polysaccharide or oligosaccharide, or a polymer containing multiple monosaccharide units covalently linked. The monosaccharide units may all be identical, or in some cases, there may be more than one type of monosaccharide unit present in the carbohydrate polymer. In certain embodiments, the carbohydrate polymer is naturally occurring. In certain embodiments, the carbohydrate polymer is synthetic (i.e., not naturally occurring). In some embodiments, the carbohydrate polymer may include chemical modifications. In some embodiments, the carbohydrate polymer is a linear polymer. In some embodiments, the carbohydrate polymer is a branched polymer.

[0039] Chemotherapeutic Agent: The term "chemotherapeutic agent" refers to a therapeutic agent known to be used in chemotherapy for cancer. For example, in some embodiments, a chemotherapeutic agent can inhibit the proliferation of rapidly growing cancer cells and / or kill cancer cells. Examples of such chemotherapeutic agents include, but are not limited to, alkylating agents, antimetabolites, topoisomerase inhibitors, and / or mitotic inhibitors.

[0040] Combination therapy: As used herein, the term "combination therapy" refers to those situations in which a subject is exposed to two or more treatment regimens (e.g., two or more therapeutic agents) simultaneously. In some embodiments, the two or more regimens may be administered simultaneously, in some embodiments, such regimens may be administered sequentially (e.g., all "doses" of a first regimen are administered prior to administration of any dose of a second regimen), and in some embodiments, such agents are administered in overlapping dosing regimens. In some embodiments, "administration" of a combination therapy may include administering one or more agent(s) or modality(s) in combination to a subject receiving the other agent(s) or modality(s). For clarity, combination therapy does not require that the individual agents be administered together (or necessarily simultaneously) in a single composition, although in some embodiments, two or more agents or active portions thereof may be administered together in a combination composition or even in a combination compound (e.g., as part of a single chemical complex or covalent conjugate).

[0041] Colloid: As used herein, the term "colloid" refers to a homogenous solution or suspension of particles (e.g., polymer particles) dispersed throughout a continuous medium (e.g., an aqueous buffer system). In some embodiments, a colloid is an emulsion. In some embodiments, a colloid is a sol. In some embodiments, a colloid is a gel.

[0042] Equivalent: As used herein, the term "equivalent" refers to two or more agents, entities, situations, sets of conditions, etc. that may not be identical to each other, but are sufficiently similar to allow a comparison between them so that a person skilled in the art would understand that a conclusion can be reasonably drawn based on the observed differences or similarities. In some embodiments, an equivalent set of conditions, situations, individuals, or populations is characterized by a number of substantially identical characteristics and one or a few different characteristics. A person skilled in the art will understand what degree of identity is required for two or more such agents, entities, situations, sets of conditions, etc. to be considered equivalent in any given situation in context. For example, a person skilled in the art will understand that a set of situations, individuals, or populations are equivalent to each other when they are characterized by a sufficient number and type of substantially identical characteristics to warrant a reasonable conclusion that differences in results obtained or phenomena observed under or with different sets of situations, individuals, or populations are caused by or indicate variations in those characteristics. Those skilled in the art will also understand that when the term "equivalent" is used in the context of a comparison of two or more values, such values ​​are equivalent to each other such that the difference in the values ​​does not result in a significant difference in the therapeutic outcome, e.g., induction of anti-tumor immunity and / or incidence of tumor regrowth and / or metastasis. For example, in some embodiments, equivalent release rates refer to such release rate values ​​within 15% over a 48-hour period. In some embodiments, equivalent release rates refer to such release rate values ​​within 20% over a 48-hour period. In some embodiments, equivalent release rates refer to such release rate values ​​within 15% over a 24-hour period.

[0043] Condition, Disease, or Disorder: The terms "condition," "disease," and "disorder" are used interchangeably.

[0044] Critical gelation temperature: As used herein, "critical gelation temperature," abbreviated as "CGT," refers to the threshold temperature at or above which the precursor state of a biomaterial preparation (e.g., as described herein) transitions to a polymeric network state (e.g., a hydrogel state) as described herein. In some embodiments, the critical gelation temperature may correspond to a sol-gel transition temperature. In some embodiments, the critical gelation temperature may correspond to a lower critical solution temperature. For a general description of thermoresponsive gels, see Taylor et al., "Thermoresponsive Gels," Gels (2017) 3:4, the contents of which are incorporated herein by reference for purposes described herein. As described herein, certain embodiments of the biomaterial preparations described herein have been shown to form a polymeric network state when they are exposed to a temperature of about 35-40°C. One of skill in the art will understand upon reading this disclosure that such biomaterial preparations do not necessarily have a CGT of about 35-40°C, but rather may have a CGT lower than 35-40°C. For example, in some embodiments, a provided biomaterial preparation may have a CGT of about 20-28°C.

[0045] Crosslinking: As used herein, the term "crosslinking" refers to interactions and / or linkages between one entity and another to form a network. For example, in some embodiments, the crosslinks present in a polymer network can be or include intramolecular crosslinks, intermolecular crosslinks, or both. In some embodiments, crosslinking can include interactions and / or linkages between one polymer chain(s) and another polymer chain(s) to form a polymer network. In some embodiments, crosslinking can be achieved using one or more physical crosslinking approaches, including, for example, one or more environmental triggers and / or physicochemical interactions. Examples of environmental triggers include, but are not limited to, pH, temperature, and / or ionic strength. Non-limiting examples of physicochemical interactions include hydrophobic interactions, charge interactions, hydrogen bonding interactions, steric complexation, and / or supramolecular chemistry. In some embodiments, crosslinking can be achieved using one or more covalent crosslinking approaches (e.g., the link between two entities is or includes a covalent bond) based on chemical reactions that can include, for example, in some embodiments, the reaction of aldehydes and amines to form Schiff bases, the reaction of aldehydes and hydrazides to form hydrazines, and / or the Michael reaction of acrylates with either primary amines or thiols to form secondary amines or sulfides. Examples of such covalent crosslinking approaches include, but are not limited to, small molecule crosslinking and polymer-polymer crosslinking. Various methods for physical and covalent crosslinking of polymer chains are described, for example, in Hoare and Kohane, "Hydrogels in drug delivery: Progress and challenges" Polymer (2008) 49: 1993-2007, the entire contents of which are incorporated herein by reference for the purposes disclosed herein.

[0046] Crosslinker: The term "crosslinker" or "crosslinking agent", as used interchangeably herein, refers to an agent that links one entity (e.g., one polymer chain) to another entity (e.g., another polymer chain). In some embodiments, the link between the two entities (i.e., "crosslink") is or includes a covalent bond. In some embodiments, the link between the two entities is or includes an ionic bond or ionic interaction. In some embodiments, the crosslinker is a chemical crosslinker that may be or include, for example, in some embodiments, a small molecule (e.g., dialdehyde or genipin) to induce the formation of a covalent bond between an aldehyde and an amino group. In some embodiments, the crosslinker includes a photosensitive functional group. In some embodiments, the crosslinker includes a pH-sensitive functional group. In some embodiments, the crosslinker includes a heat-sensitive functional group.

[0047] Disease: As used herein, the term "disease" refers to a disorder or condition that typically impairs the normal function of a tissue or system of a subject (e.g., a human subject) and is typically manifested by characteristic signs and / or symptoms. Examples of diseases suitable for the techniques provided herein include, but are not limited to, autoimmune diseases, inflammatory diseases, bone diseases, metabolic diseases, neurological and neurodegenerative diseases, cancer, cardiovascular diseases, allergies and asthma, Alzheimer's disease, and hormone-related diseases. In some embodiments, the disease suitable for the techniques provided herein is cancer.

[0048] Effective amount: An "effective amount" is an amount sufficient to induce a desired biological response, e.g., treatment of a condition that the subject may suffer from. As will be understood by one of skill in the art, the effective amount of a composition or an agent contained in a composition may vary depending on such factors as the desired biological endpoint, the physical, chemical, and / or biological properties of the agent in the composition (e.g., pharmacokinetics and / or degradation), the condition being treated, and the age and health of the subject. In some embodiments, the amount may be effective for therapeutic treatment, and alternatively or additionally, in some embodiments, the amount may be effective for prophylactic treatment. For example, in the treatment of cancer, an effective amount may prevent tumor regrowth, reduce tumor burden, or halt tumor growth or spread. One of skill in the art will understand that an effective amount need not be contained in a single dosage form. Rather, administration of an effective amount may involve administration of multiple doses (e.g., according to a dosing regimen), potentially over time. For example, in some embodiments, an effective amount may be an amount administered in a dosing regimen that has been established to achieve a particular result with statistical significance when administered to a relevant population.

[0049] Hydrate: The term "hydrate" as used herein has its art-recognized meaning and refers to an aggregate of a compound (e.g., which may be a salt form of the compound) with one or more water molecules. Typically, the number of water molecules contained in a hydrate of a compound is in a definite ratio to the number of compound molecules in the hydrate. Thus, a hydrate of a compound can be represented, for example, by the general formula R×xH2O, where R is the compound and x is a number greater than 0. A given compound can form two or more types of hydrates, including, for example, monohydrates (x is 1), hypohydrates (x is a number greater than 0 and less than 1, e.g., hemihydrates (R×0.5H2O)), and polyhydrates (x is a number greater than 1, e.g., dihydrates (R×2H2O) and hexahydrates (R×6H2O)).

[0050] Hydrogel: The term "hydrogel" has its art-recognized meaning and refers to a material formed from a network of polymer chains that are hydrophilic and may be found as a colloidal gel in which an aqueous phase is the dispersion medium. In some embodiments, hydrogels are natural or synthetic polymer networks that are highly absorbent (e.g., capable of absorbing and / or retaining more than 90% water). In some embodiments, hydrogels have a degree of flexibility comparable to that of natural tissues, for example, due to their significant water content.

[0051] Immunotherapy: The term "immunotherapy" refers to therapeutic agents that promote the treatment of disease by inducing, enhancing, or suppressing immune responses. Immunotherapies designed to induce or amplify immune responses are classified as activating immunotherapies, while immunotherapies that reduce or suppress immune responses are classified as suppressing immunotherapies. Immunotherapies are usually, but not necessarily, biological drugs. Numerous immunotherapies are used to treat cancer. These include, but are not limited to, monoclonal antibodies, adoptive cell transfer, cytokines, chemokines, vaccines, nucleic acids, small molecule inhibitors, and small molecule agonists. For example, useful immunotherapies may include, but are not limited to, inducers of type I interferon, interferon, stimulator of interferon genes (STING) agonists, TLR7 / 8 agonists, IL-15 superagonists, COX inhibitors (e.g., COX-1 inhibitors and / or COX-2 inhibitors), anti-PD-1 antibodies, anti-CD137 antibodies, and anti-CTLA-4 antibodies. In some embodiments, certain polymeric biomaterial(s) preparations provided herein are themselves immunomodulatory (e.g., sufficient to induce anti-tumor immunity) in the absence of immunotherapy and thus do not include administration of such immunotherapy as described herein.

[0052] Immunomodulatory payload: As used herein, the term "immunomodulatory payload" refers to a separate immunomodulatory agent (e.g., small molecule, polypeptide (e.g., including cytokines), nucleic acid, etc.) that can be carried by or distributed within a polymeric biomaterial(s) preparation such as those provided and / or utilized herein, where the immunomodulatory agent provides a therapeutic effect that modulates or alters (e.g., induces, enhances, or suppresses, etc.) one or more aspects of the immune response in a subject. Examples of immunomodulatory payloads include, but are not limited to, activators of adaptive immune responses, activators of innate immune responses, inhibitors of proinflammatory pathways, immunomodulatory cytokines, or immunomodulatory therapeutics, as well as those described in WO2018 / 045058 and WO2019 / 183216, and any combination thereof. The contents of the aforementioned patent applications are incorporated herein by reference for the purposes described herein. In some embodiments, the immunomodulatory payload is or includes an innate immune modulating payload (e.g., an immunomodulatory payload that induces or stimulates innate immunity and / or one or more features of innate immunity). In some embodiments, the innate immune modulating payload is or includes an activator of the innate immune response. In some embodiments, the immune modulating payload is or includes an adaptive immune modulating payload, e.g., an activator of the adaptive immune response. In some embodiments, the immune modulating payload is or includes an inhibitor of a proinflammatory pathway, e.g., an inhibitor of a proinflammatory immune response mediated by the p38 mitogen-activated protein kinase (MAPK) pathway. In some embodiments, the immune modulating payload is or includes an immunomodulating cytokine. In some embodiments, the immune modulating payload is or includes an immunomodulating therapeutic. As will be appreciated by one of skill in the art, the immune modulating payload does not include components (e.g., precursor components) and / or by-products of the polymeric biomaterial(s) preparation (e.g., as described and / or utilized herein) that are generated, e.g., by chemical, enzymatic, and / or biological reactions, e.g., degradation.

[0053] Implantation: The terms "implantable," "implantation," "implanting," and "implant" refer to the placement of a subject composition at a specific location in a subject, such as within a tumor resection site or within a sentinel lymph node, typically by conventional surgical methods.

[0054] Increase, induce, or reduce: As used herein, these terms or grammatically equivalent comparative terms refer to a value relative to an equivalent reference measurement. For example, the assessed value achieved in a subject can be "increased" relative to that obtained in the same subject under different conditions (e.g., before or after an event, or in the presence or absence of an event, such as administration of a composition or preparation described and / or utilized herein), or in a different, equivalent subject (e.g., in an equivalent subject different from the subject of interest in the absence of previous exposure to a condition, e.g., administration of a composition or preparation described and / or utilized herein). In some embodiments, the comparative term refers to a statistically relevant difference (e.g., of sufficient incidence and / or magnitude to achieve statistical relevance). A person skilled in the art will know or be able to easily determine the degree and / or incidence of difference necessary or sufficient to achieve such statistical significance in a given context.

[0055] Inhibit: The term "inhibit" or "inhibition" is not limited to complete inhibition only. Thus, in some embodiments, partial inhibition or relative reduction is included within the scope of the term "inhibition". For example, in the context of regulating the level (e.g., expression and / or activity) of a target, the term refers to a reduction in the level (e.g., expression and / or activity) of the target to a level that is reproducibly and / or statistically significantly lower than an initial or other suitable reference level, which in some embodiments may be, for example, the baseline level of the target. In some embodiments, the term refers to a reduction in the level (e.g., expression and / or activity) of the target to a level that is less than 75%, less than 50%, less than 40%, less than 30%, less than 25%, less than 20%, less than 10%, less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2%, less than 1%, less than 0.5%, less than 0.1%, less than 0.01%, less than 0.001%, or less than 0.0001% of the initial level, which may be, for example, the baseline level of the target. In the context of risk and / or incidence of tumor recurrence and / or metastasis, the term, in some embodiments, refers to a reduction in the risk or incidence of tumor recurrence and / or metastasis to a level that is reproducibly and / or statistically significantly lower than an initial or other suitable reference level, which can be, for example, a baseline level of risk or incidence of tumor recurrence and / or metastasis in the absence of or prior to administration of a composition described herein. In some embodiments, the term refers to a reduction in the risk or incidence of tumor recurrence and / or metastasis to a level that is less than 75%, less than 50%, less than 40%, less than 30%, less than 25%, less than 20%, less than 10%, less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2%, less than 1%, less than 0.5%, less than 0.1%, less than 0.01%, less than 0.001%, or less than 0.0001% of an initial level, which may be, for example, a baseline level of risk or incidence of tumor recurrence and / or metastasis in the absence of or prior to administration of a composition described herein.In the context of modulating immune cell function (e.g., by inhibiting activity and / or expression of a target), the term, in some embodiments, refers to a reduction in activity and / or expression of a target to a level that is reproducibly and / or statistically significantly lower than an initial or other suitable reference level, which can be, for example, a baseline level of activity and / or expression of the target in the absence or prior to administration of a composition described herein.

[0056] Inhibitor: As used herein, the term "inhibitor" refers to an agent whose presence or level correlates with a decrease in the level or activity of a modulated target. In some embodiments, an inhibitor may act directly (where it exerts an effect on a target directly, e.g., by binding to the target), and in some embodiments, an inhibitor may act indirectly (where it exerts an effect by interacting with and / or otherwise altering a regulator of the target such that the level and / or activity of the target is reduced). In some embodiments, an inhibitor is one whose presence or level correlates with a target level or activity that is reduced relative to a particular reference level or activity (e.g., observed under appropriate reference conditions such as the presence of a known inhibitor or the absence of an inhibitor disclosed herein). In some embodiments, an inhibitor may be a small molecule, a polynucleotide, an oligonucleotide, a polysaccharide, a polypeptide, a protein, an antibody, and / or a functional portion thereof.

[0057] Inhibitors of proinflammatory pathways: As used herein, the term "inhibitors of proinflammatory pathways" refers, in some embodiments, to agents that inhibit or reduce inflammation associated with immunosuppression. In some embodiments, such inhibitors of proinflammatory pathways refer to agents that prevent the recruitment of immunosuppressive cells or prevent acute inflammation. Such acute inflammation and / or recruitment of immunosuppressive cells may occur following local trauma, including that caused by surgery. In some embodiments, inhibitors of proinflammatory pathways may inhibit, for example, immune responses that induce inflammation, including, for example, the production of inflammatory cytokines (e.g., including but not limited to, TGF-β and IL-10), increased activity and / or proliferation of M2-like macrophages, and recruitment of associated immune cells, including but not limited to, myeloid cells, neutrophils, and mast cells. Examples of inhibitors of proinflammatory pathways include, for example, those described in International Application No. 2019 / 183216, the contents of which are incorporated herein by reference in their entirety for purposes described herein.

[0058] Lymph node: As known in the art, the term "lymph node" refers to a component of the lymphatic system, which is a small structure located throughout the body through which lymphatic fluid flows. It is understood that lymph nodes filter certain substances from lymphatic fluid. Lymph nodes may also contain immune cells that may be involved in immune responses throughout the body, for example. In some embodiments, lymph nodes may be or include sentinel lymph nodes (i.e., lymph nodes through which cancer cells are most likely to spread from a primary tumor).

[0059] Marker: Marker, as used herein, refers to an entity or moiety whose presence or level is characteristic of a particular condition or event. In some embodiments, the presence or level of a particular marker may be characteristic of the presence or stage of a disease, disorder, or condition. To give one example, in some embodiments, the term refers to a gene expression product that is characteristic of a particular tumor, tumor subclass, tumor stage, etc. Alternatively or additionally, in some embodiments, the presence or level of a particular marker correlates with the activity (or activity level) of a particular signaling pathway, which may be characteristic of, for example, a particular class of tumor. The statistical significance of the presence or absence of a marker may vary depending on the particular marker. In some embodiments, the detection of a marker is highly specific in that it reflects a high probability that the tumor is of a particular subclass. Such specificity may come at the expense of sensitivity (i.e., a negative result may occur even when the tumor is one that is expected to express the marker). Conversely, a marker with a high sensitivity may be less specific than one with a lower sensitivity. Those skilled in the art will understand that in many embodiments, a useful marker need not discriminate with 100% accuracy.

[0060] Isomers: It should also be understood that compounds that have the same molecular formula but differ in the nature or sequence of bonding of their atoms or the arrangement of their atoms in space are termed "isomers." Isomers that differ in the arrangement of their atoms in space are termed "stereoisomers."

[0061] Metastasis: The terms "metastasis," "metastatic," or "metastasizing" refer to the spread or migration of cancer cells from a primary or original tumor to another organ or tissue, typically identifiable by the presence of a "secondary tumor" or "secondary cell mass" of the tissue type of the primary or original tumor and not of the organ or tissue in which the secondary (metastatic) tumor is located. For example, prostate cancer that has migrated to bone is said to be metastatic prostate cancer, and includes cancerous prostate cancer cells growing in the bone tissue.

[0062] Microparticles: As used herein, the term "microparticles" refers to particles having a longest dimension (e.g., diameter) between 1 micrometer and 1,000 micrometers (μm). In some embodiments, a microparticle may be characterized by a longest dimension (e.g., diameter) between 1 μm and 500 μm. In some embodiments, a microparticle may be characterized by a longest dimension (e.g., diameter) between 1 μm and 100 μm. In many embodiments, a population of microparticles is characterized by an average size (e.g., longest dimension) of less than about 1,000 μm, about 500 μm, about 100 μm, about 50 μm, about 40 μm, about 30 μm, about 20 μm, or about 10 μm, and often greater than about 1 μm. In many embodiments, a microparticle may be substantially spherical (e.g., such that its longest dimension may be its diameter).

[0063] Monosaccharide: As used herein, the term "monosaccharide" is given its ordinary meaning as used in the art and refers to a simple form of sugar consisting of a single sugar unit that cannot be further broken down into smaller sugar building blocks or moieties. Common examples of monosaccharides include, for example, glucose (dextrose), fructose, galactose, mannose, ribose, etc. Monosaccharides can be classified according to the number of carbon atoms of the carbohydrate, for example, into trioses having three carbon atoms, such as glyceraldehyde and / or dihydroxyacetone; tetroses having four carbon atoms, such as erythrose, threose, and / or erythrulose; pentoses having five carbon atoms, such as arabinose, lyxose, ribose, xylose, ribulose, and / or xylulose; hexoses having six carbon atoms, such as allose, altrose, galactose, glucose, gulose, idose, mannose, talose, fructose, psicose, sorbose, and / or tagatose; heptoses having seven carbon atoms, such as mannoheptulose and / or sedoheptulose; octose having eight carbon atoms, such as 2-keto-3-deoxy-manno-octonate; nonoses having nine carbon atoms, such as sialose; and decoses having ten carbon atoms. The above monosaccharides include both D-monosaccharides and L-monosaccharides. Alternatively, the monosaccharide can be a monosaccharide variant in which the sugar unit includes one or more substituents other than hydroxyl, such as deoxy, H substituents, heteroatom substituents (e.g., S, Cl, F, etc.). Such variants can be, but are not limited to, ethers, esters, amides, acids, phosphates, and amines. Amine variants (i.e., amino sugars) include, for example, glucosamine, galactosamine, fructosamine, and / or mannosamine. Amide variants include, for example, N-acetylated amine variants of sugars (e.g., N-acetylglucosamine and / or N-acetylgalactosamine).

[0064] Modulator: As used herein, the term "modulator" can be or include an entity whose presence or level in a system in which an activity of interest is observed correlates with a change in the level and / or nature of that activity compared to that observed under otherwise equivalent conditions in the absence of the modulator. In some embodiments, a modulator is an activator or agonist in that the activity of interest is increased in its presence compared to that observed under otherwise equivalent conditions in the absence of the modulator. In some embodiments, a modulator is an antagonist or inhibitor in that the activity of interest is reduced in its presence compared to that observed under otherwise equivalent conditions in the absence of the modulator. In some embodiments, a modulator interacts directly with a target entity whose activity is of interest. In some embodiments, a modulator interacts indirectly with a target entity whose activity is of interest (e.g., interacts with one or more entities that interact with and / or associate with the target entity). In some embodiments, the modulator affects the level of the target entity of interest; alternatively or additionally, in some embodiments, the modulator affects the activity of the target entity of interest without affecting the level of the target entity. In some embodiments, the modulator affects both the level and activity of the target entity of interest such that differences in observed activity are not entirely explained or disproportionate to the differences in observed levels. In some embodiments, the modulator can be a small molecule, a polynucleotide, an oligonucleotide, a polysaccharide, a polypeptide, a protein, an antibody, and / or functional portions thereof.

[0065] Modulators of macrophage effector function: The term "modulators of macrophage effector function" refers to agents that activate macrophage effector function or deplete immunosuppressive macrophages or macrophage-derived suppressor cells. Such enhancement may recruit macrophages and myeloid components to destroy tumors and their stroma, including tumor vasculature. Macrophages may be induced to secrete anti-tumor cytokines and / or perform phagocytosis, including antibody-dependent cellular phagocytosis.

[0066] Neutrophil function regulator: As used interchangeably herein, the terms "neutrophil regulator" and "neutrophil function regulator" refer to a regulator of one or more biological functions and / or phenotypes of neutrophils. For example, in some embodiments, a neutrophil function regulator may inhibit neutrophil recruitment, survival, and / or proliferation. Additionally or alternatively, in some embodiments, a neutrophil function regulator may regulate neutrophil-associated effector functions, which may include, but are not limited to, regulating the production and / or secretion of one or more immunomodulatory molecules (e.g., immunomodulatory cytokines and / or chemokines) and / or alter the extracellular matrix modification capabilities of neutrophils. In some embodiments, a neutrophil function regulator (e.g., those described herein) may act or target only neutrophils. In some embodiments, a regulator of neutrophil function (e.g., as described herein) may act on neutrophils as well as at least one additional type of immune cell, such as other subsets of myeloid-derived suppressor cells (MDSCs), macrophages, and / or monocytes. Those skilled in the art will appreciate that at least one subset of neutrophils may exhibit similar immune activity to one or more certain subsets of MDSCs and thus may be considered polymorphonuclear and / or granulocytic MDSCs (e.g., as described in Mehmeti-Ajradini et al., "Human G-MDSCs are neutrophils at distinct maturation stages promoting tumor growth in breast cancer" Life Science Alliance, September 21, 2020, and Brandau et al., "A subset of mature neutrophils contains the strongest PMN-MDSC activity in blood and tissue of patients with head and neck cancer" The Journal of Immunology, May 1, 2020, the contents of each of which are incorporated herein by reference for purposes described herein).

[0067] Nanoparticles: As used herein, the term "nanoparticles" refers to particles having a longest dimension (e.g., diameter) of less than 1000 nanometers (nm). In some embodiments, a nanoparticle may be characterized by a longest dimension (e.g., diameter) of less than 300 nm. In some embodiments, a nanoparticle may be characterized by a longest dimension (e.g., diameter) of less than 100 nm. In many embodiments, a nanoparticle may be characterized by a longest dimension of about 1 nm to about 100 nm, or about 1 nm to about 500 nm, or about 1 nm to 1,000 nm. In many embodiments, a population of nanoparticles is characterized by an average size (e.g., longest dimension) of less than about 1,000 nm, about 500 nm, about 100 nm, about 50 nm, about 40 nm, about 30 nm, about 20 nm, or about 10 nm, and often greater than about 1 nm. In many embodiments, a nanoparticle may be substantially spherical, such that its longest dimension may be its diameter. In some embodiments, a nanoparticle has a diameter of less than 100 nm, as defined by the National Institutes of Health.

[0068] Neoplasms and Tumors: The terms "neoplasm" and "tumor" are used interchangeably herein and refer to an abnormal mass of tissue whose proliferation exceeds and is not coordinated with the growth of normal tissue. A neoplasm or tumor can be "benign" or "malignant" depending on the following characteristics: degree of cellular differentiation (including morphology and function), rate of growth, local invasion, and metastasis. "Benign neoplasms" are generally well differentiated, have characteristically slower growth than malignant neoplasms, and remain localized at the site of origin. In addition, benign neoplasms do not have the ability to invade, invade, or metastasize to distant sites. Exemplary benign neoplasms include, but are not limited to, lipomas, chondromas, adenomas, acrochordons, senile hemangiomas, seborrheic keratosis, lentigines, and sebaceous hyperplasia. In some cases, certain "benign" tumors may subsequently give rise to malignant neoplasms, which may arise as a result of further genetic alterations in a subpopulation of the tumor's tumor cells; these tumors are referred to as "pre-malignant neoplasms." One example of a pre-malignant neoplasm is a teratoma. In contrast, "malignant neoplasms" are generally poorly differentiated (anaplastic) and have characteristically rapid growth accompanied by progressive infiltration, invasion, and destruction of surrounding tissue. Moreover, malignant neoplasms generally have the ability to metastasize to distant sites.

[0069] Nucleic Acid: The term "nucleic acid" as used herein is consistent with its ordinary meaning and may be represented as a polynucleotide. However, when used as "nucleic acid" or "at least one nucleic acid", this does not refer to a single molecule, but rather to a single species of nucleic acid.

[0070] Payload: In general, the term "payload" as used herein refers to an agent that may be incorporated into the polymeric biomaterial(s) preparation described herein. In some embodiments, the payload may be any chemical class of compound, molecule, or entity, including, for example, a small molecule, peptide, polypeptide, nucleic acid, sugar (e.g., polysaccharide), lipid, metal, or combination or complex thereof. In some embodiments, the payload may be or include a biological modifier, a detectable agent (e.g., dye, fluorophore, radioactive label, etc.), a detection agent, a nutrient, a therapeutic agent, a mineral, a growth factor, a cytokine, an antibody, a hormone, an extracellular matrix protein (collagen, vitronectin, fibrin, etc.), an extracellular matrix sugar, a chemoattractant, a polynucleotide (e.g., DNA, RNA, antisense molecule, plasmid, etc.), a microorganism (e.g., virus), etc., or a combination thereof. In some embodiments, the payload is or includes a therapeutic agent. Examples of therapeutic agents include, but are not limited to, analgesics, antibiotics, antibodies, anticoagulants, antiemetics, cells, clotting agents, cytokines, growth factors, hormones, immunomodulators, polynucleotides (e.g., DNA, RNA, antisense molecules, plasmids, etc.), and combinations thereof. In some embodiments, the payload may be or may include a cell or organism, or a fraction, extract, or component thereof. Alternatively or additionally, in some embodiments, the payload may be or may include a natural product, in that it is found in nature and / or obtained from nature. Alternatively or additionally, in some embodiments, the term may be used to refer to one or more entities that are artificial, in that it is designed, engineered, and / or produced through the action of the hand of man, and / or is not found in nature. In some embodiments, the payload may be or may include an agent in isolated or pure form, and in some embodiments, such an agent may be in crude form.

[0071] Pharmaceutically acceptable salts: The term "pharmaceutical acceptable salts" refers to salts that are suitable for use, for example, in contact with human and / or animal tissues, without undue toxicity, irritation, allergic reaction, etc., within the scope of sound medical judgment, and are commensurate with a reasonable benefit / risk ratio. Pharmaceutically acceptable salts are well known in the art. For example, Berge et al. describe pharmaceutical acceptable salts in detail in J. Pharmaceutical Sciences, 1977, 66, 1-19, the contents of which are incorporated herein by reference for the purposes set forth herein. Pharmaceutically acceptable salts that can be utilized according to certain embodiments of the present disclosure include, for example, those derived from suitable inorganic and organic acids and bases. Examples of pharma- ceutically acceptable non-toxic acid addition salts are salts of amino groups formed with inorganic acids such as hydrochloric, hydrobromic, phosphoric, sulfuric, and perchloric acids, or with organic acids such as acetic, oxalic, maleic, tartaric, citric, succinic, or malonic acids, or by using other methods known in the art, such as ion exchange. Other pharma- ceutically acceptable salts include adipate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, formate, fumarate, glucoheptonate, glycerophosphate, gluconate, hemisulfate, heptanoate, hexanoate, hydroiodide, 2-hydroxy-ethanesulfonate, Examples of suitable salts include, but are not limited to, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, pamoate, pectinate, persulfate, 3-phenylpropionate, phosphate, picrate, pivalate, propionate, stearate, succinate, sulfate, tartrate, thiocyanate, p-toluenesulfonate, undecanoate, and valerate.Salts derived from appropriate bases include alkali metal, alkaline earth metal, ammonium, and N+(C1-C4 alkyl)4 salts. Representative alkali metal or alkaline earth metal salts include sodium, lithium, potassium, calcium, magnesium salts, and the like. Further pharma-ceutically acceptable salts include non-toxic ammonium, quaternary ammonium, and amine cations formed, where appropriate, using counterions such as halides, hydroxides, carboxylates, sulfates, phosphates, nitrates, lower alkylsulfonates, and arylsulfonates.

[0072] Polynucleotide Carrier: The term "polynucleotide carrier," "polynucleotide agent carrier," or "nucleic acid carrier" refers to any agent that facilitates endocytosis of a nucleic acid by a eukaryotic cell. These may include, but are not limited to, lipids, proteins, and small molecules.

[0073] Poloxamer: As used herein, the term "poloxamer" refers to a polymer preparation of one or more poloxamers or a polymer preparation containing one or more poloxamers. In some embodiments, the poloxamer in the polymer preparation may be unconjugated or unmodified, for example, a typical triblock copolymer that contains a hydrophobic chain of polyoxypropylene (polypropylene glycol, PPG) adjacent to two hydrophilic chains of polyoxyethylene (polyethylene glycol, PEG). In some embodiments, the polymer preparation of one or more poloxamers or the polymer preparation containing one or more poloxamers may not be filtered (e.g., such a polymer preparation may contain impurities and / or relatively low molecular weight polymer molecules compared to a comparable polymer preparation that is filtered). Examples of poloxamers include, but are not limited to, poloxamer 124 (P124, also known as Pluronic® L44 NF), poloxamer 188 (P188, also known as Pluronic F68NF), poloxamer 237 (P237, also known as Pluronic F87 NF), poloxamer 338 (P338, also known as Pluronic F108 NF), poloxamer 407 (P407, also known as Pluronic F127 NF), and combinations thereof.

[0074] Polymer: The term "polymer" is given its ordinary meaning as used in the art, i.e., a molecular structure that includes one or more repeating units (monomers) linked by covalent bonds. The repeating units may all be identical, or in some cases, there may be more than one type of repeating unit present within a polymer (e.g., a copolymer). In certain embodiments, the polymer is naturally occurring. In certain embodiments, the polymer is synthetic (i.e., not naturally occurring). In some embodiments, the polymer is a linear polymer. In some embodiments, the polymer is a branched polymer. In some embodiments, a polymer for use according to the present disclosure is not a polypeptide. In some embodiments, a polymer for use according to the present disclosure is not a nucleic acid. In some embodiments, a polymer for use according to the present disclosure is not a polypeptide.

[0075] Polymer combination preparation: As used herein, the term "polymer combination preparation" refers to a polymer biomaterial that comprises at least two different polymer components. For example, in many embodiments, the polymer combination preparation described herein is a polymer biomaterial that comprises a first polymer component and a second first polymer component, where the first polymer component is or comprises at least one poloxamer, and the second polymer component is or comprises a non-poloxamer polymer. In some embodiments, the polymer combination preparation described herein is a polymer biomaterial in a precursor state, which may be useful, for example, for administration to a subject. In some embodiments, the polymer combination preparation described herein is a polymer biomaterial in a polymer network state.

[0076] Polymeric biomaterial: As used herein, a "polymeric biomaterial" is a material that is or comprises at least one polymer or at least one polymer moiety and is biocompatible. In many embodiments, the polymeric biomaterial is or comprises at least one polymer, and in some embodiments, the polymer can be or comprises a copolymer. In some embodiments, the polymeric biomaterial is or comprises a preparation of at least two different polymeric components (e.g., a preparation containing a poloxamer and a second polymeric component that is not a poloxamer). Those skilled in the art will recognize that a particular polymer may exist and / or be available in a variety of forms (e.g., length, molecular weight, charge, topography, surface chemistry, degree and / or type of modification such as alkylation, acylation, quaternization, hydroxyalkylation, carboxyalkylation, thiolation, phosphorylation, glycosylation, etc.), and in some embodiments, such a preparation of a polymer may include a particular level and / or distribution of such form(s). Additionally or alternatively, it will be appreciated that in some embodiments, one or more immune modulating properties of the polymeric biomaterial may be tuned by its biomaterial property(ies), including, for example, the polymeric biomaterial (e.g., modulated by the hydrophobic and / or hydrophilic moieties, chemical moieties, and / or charge properties of the polymeric biomaterial) and / or the topography of the polymeric biomaterial (e.g., modulated by the size, shape, and / or surface structure), as described, for example, in Mariani et al. “Biomaterials: Foreign Bodies or Tuners for the Immune Response?” International Journal of Molecular Sciences, 2019, 20, 636, the contents of which are incorporated herein by reference in their entirety for the purposes set forth herein. In some embodiments, the polymeric biomaterial may be in a polymer network state. In some embodiments, the polymeric biomaterial may be in an injectable form, for example, a precursor state (e.g., a viscous solution).For example, a polymeric biomaterial may include its precursor components to be formed in situ (e.g., upon administration to a subject). In some embodiments, the polymeric biomaterial may be a liquid. In some embodiments, the polymeric biomaterial is a viscous solution. In some embodiments, the polymeric biomaterial is a colloid. In some embodiments, the polymeric biomaterial may be a solid. In some embodiments, the polymeric biomaterial may be a crystal (e.g., an inorganic crystal).

[0077] Polymer Network: The term "polymer network" is used herein to describe a collection of polymer chains that interact with each other. In some embodiments, the polymer network forms a three-dimensional structural material. In some embodiments, the polymer network can be formed by linking polymer chains ("crosslinked polymer network") using a crosslinker (e.g., as described herein). In some embodiments, the polymer network transitions from a precursor state when it is exposed to a temperature at or above a critical gelation temperature, and the polymer network state has a viscosity that significantly exceeds (e.g., at least 50% or more) that of the precursor state, and the polymer network state includes crosslinks that are not present in the precursor state. In some embodiments, the polymer network can be formed by non-covalent or non-ionic intermolecular association of the polymer chains, for example, via hydrogen bonding. In some embodiments, the polymer network can be formed by a combination of chemically crosslinking the polymer chains and non-covalent or non-ionic intermolecular association of the polymer chains.

[0078] Proinflammatory cytokine: As used herein, the term "proinflammatory cytokine" refers to a protein or glycoprotein molecule secreted by cells (e.g., cells of the immune system) that induces an inflammatory response. As will be understood by those skilled in the art, inflammation can be immunostimulatory or immunosuppressive depending on the biological context.

[0079] Proinflammatory immune response: As used herein, the term "proinflammatory immune response" refers to an immune response that induces inflammation, including, for example, the production of inflammatory cytokines (e.g., including but not limited to, CXCL10, IFN-α, IFN-β, IL-1β, IL-6, IL-18, and / or TNF-α), increased activity and / or proliferation of Th1 cells, recruitment of myeloid cells, etc. In some embodiments, a proinflammatory immune response can be or can include one or both of acute and chronic inflammation.

[0080] Proliferative disease: "Proliferative disease" refers to a disease resulting from abnormal growth or expansion by proliferation of cells (Walker, Cambridge Dictionary of Biology; Cambridge University Press: Cambridge, UK, 1990). Proliferative diseases may be associated with 1) pathological proliferation of normally quiescent cells, 2) pathological migration of cells from their normal location (e.g., metastasis of tumor cells), 3) pathological expression of proteolytic enzymes such as matrix metalloproteinases (e.g., collagenase, gelatinase, and elastase), or 4) pathological angiogenesis, such as in proliferative retinopathies and tumor metastasis. Exemplary proliferative diseases include cancer (i.e., "malignant neoplasms"), benign neoplasms, angiogenesis or diseases associated with angiogenesis, inflammatory diseases, autoinflammatory diseases, and autoimmune diseases.

[0081] Prophylactically effective amount: A "prophylactically effective amount" is an amount sufficient to prevent a condition (e.g., significantly delay the onset or recurrence of one or more symptoms or characteristics of the condition such that it / they are not detected at a time that would be expected in the absence of administration of that amount). A prophylactically effective amount of a composition refers to an amount of a therapeutic agent(s) alone or in combination with other agents that provides a prophylactic benefit in preventing a condition. The term "prophylactically effective amount" can encompass an amount that improves overall prophylaxis or enhances the prophylactic efficacy of another prophylactic agent. One of skill in the art will appreciate that a prophylactically effective amount need not be contained in a single dosage form. Rather, administration of an effective amount can involve administration of multiple doses, potentially over time (e.g., according to a dosing regimen).

[0082] Risk: As will be understood from the context, "risk" of a disease, disorder, and / or condition refers to the likelihood that a particular individual will develop the disease, disorder, and / or condition. In some embodiments, the risk is expressed as a percentage. In some embodiments, the risk is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90-100%. In some embodiments, the risk is expressed as a relative risk to a risk associated with a reference sample or group of reference samples. In some embodiments, the reference sample or group of reference samples has a known risk of the disease, disorder, condition, and / or event. In some embodiments, the reference sample or group of reference samples is from an individual comparable to the particular individual. In some embodiments, the relative risk is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more. In some embodiments, risk may reflect, for example, one or more genetic attributes that may predispose an individual to developing (or not developing) a particular disease, disorder, and / or condition, hi some embodiments, risk may reflect one or more epigenetic events or attributes and / or one or more lifestyle or environmental events or attributes.

[0083] Salts: As used herein, the term "salts" refers to any and all salts, including pharma- ceutically acceptable salts.

[0084] Sample: As used herein, the term "sample" typically refers to an aliquot of material obtained or derived from a source of interest as described herein. In some embodiments, the source of interest is a biological or environmental source. In some embodiments, the source of interest may be or include a cell or an organism such as a microorganism, a plant, or an animal (e.g., a human). In some embodiments, the source of interest is or includes a biological tissue or fluid. In some embodiments, the biological tissue or fluid may be or include amniotic fluid, aqueous humor, peritoneal fluid, bile, bone marrow, blood, breast milk, cerebrospinal fluid, earwax, chyle, chime, ejaculate, endolymph, exudate, feces, gastric acid, gastric juice, lymph, mucus, pericardial fluid, peritoneal fluid, pleural fluid, pus, mucosal secretions, saliva, sebum, semen, serum, smegma, sputum, synovial fluid, sweat, tears, urine, vaginal secretions, vitreous humor, vomit, and / or combinations or component(s) thereof. In some embodiments, the biological fluid may be or include intracellular fluid, extracellular fluid, intravascular fluid (plasma), interstitial fluid, lymph, and / or cellular permeate fluid. In some embodiments, the biological fluid may be or include phytoexudates. In some embodiments, the biological tissue or sample may be obtained, for example, by aspiration, biopsy (e.g., fine needle or tissue biopsy), swab (e.g., oral, nasal, skin, or vaginal swab), scraping, surgery, washing or lavage (e.g., bronchoalveolar, ductal, nasal, ocular, oral, uterine, vaginal, or other washing or lavage). In some embodiments, the biological sample is or includes cells obtained from an individual. In some embodiments, the sample is a "primary sample" obtained directly from the source of interest by any suitable means. In some embodiments, as will be clear from the context, the term "sample" refers to a preparation obtained by processing the primary sample (e.g., by removing one or more components of the primary sample and / or by adding one or more agents). For example, filtration using a semipermeable membrane.Such a "processed sample" may include, for example, nucleic acids or proteins extracted from a sample or obtained by subjecting the primary sample to one or more techniques, such as amplification or reverse transcription of nucleic acids, isolation and / or purification of certain components.

[0085] Small molecule: The term "small molecule" or "small molecule therapeutic" refers to a molecule having a relatively low molecular weight, whether naturally occurring or artificially created (e.g., by chemical synthesis). Typically, a small molecule is an organic compound (i.e., it contains carbon). A small molecule may contain multiple carbon-carbon bonds, stereocenters, and other functional groups (e.g., amines, hydroxyls, carbonyls, heterocycles, and the like). In certain embodiments, the molecular weight of a small molecule is about 1,000 g / mol or less, about 900 g / mol or less, about 800 g / mol or less, about 700 g / mol or less, about 600 g / mol or less, about 500 g / mol or less, about 400 g / mol or less, about 300 g / mol or less, about 200 g / mol, or about 100 g / mol or less. In certain embodiments, the molecular weight of the small molecule is at least about 100 g / mol, at least about 200 g / mol, at least about 300 g / mol, at least about 400 g / mol, at least about 500 g / mol, at least about 600 g / mol, at least about 700 g / mol, at least about 800 g / mol, or at least about 900 g / mol, or at least about 1,000 g / mol. Combinations of the above ranges (e.g., at least about 200 g / mol and up to about 500 g / mol) are also possible. In certain embodiments, the small molecule is a therapeutically active agent such as a drug (e.g., a molecule approved by the US Food and Drug Administration as provided in the Code of Federal Regulations (CFR)). The small molecule may be complexed with one or more metal atoms and / or metal ions. In this example, the small molecule is also referred to as a "small organometallic molecule." Preferred small molecules are biologically active in that they produce a biological effect in animals, preferably mammals, and more preferably humans. Small molecules include, but are not limited to, radionuclides and imaging agents. In certain embodiments, the small molecule is a drug. Preferably, but not necessarily, the drug is one that has already been deemed safe and effective for use in humans or animals by the appropriate government or regulatory agency.For example, drugs approved for human use are listed by the FDA under 21 C.FR §§ 330.5, 331-361, and 440-460, which are incorporated herein by reference, and drugs for veterinary use are listed by the FDA under 21 C.FR §§ 500-589, the contents of each of which are incorporated herein by reference for purposes described herein, and such listed drugs are typically considered acceptable for use in accordance with the present disclosure.

[0086] Solvate: The term "solvate" as used herein has its art-understood meaning and refers to an aggregate of a compound (e.g., which may be a salt form of a compound) with one or more solvent atoms or molecules. In some embodiments, the solvate is a liquid. In some embodiments, the solvate is in a solid form (e.g., a crystalline form). In some embodiments, the solid form of the solvate is suitable for isolation. In some embodiments, the association between the solvent atom(s) and the compound in the solvate is a non-covalent association. In some embodiments, such association is or includes hydrogen bonding, van der Waals interactions, or a combination thereof. In some embodiments, the solvent in which the atom(s) is included in the solvate may be or include one or more of water, methanol, ethanol, acetic acid, DMSO, THF, diethyl ether, and the like. A suitable solvate may be a pharmaceutically acceptable solvate, and in some particular embodiments, the solvate is a hydrate, ethanolate, or methanolate. In some embodiments, the solvate may be a stoichiometric or non-stoichiometric solvate.

[0087] Subject: A "subject" to which administration is contemplated includes, but is not limited to, humans (i.e., male or female of any age group, e.g., a pediatric subject (e.g., infant, child, adolescent) or an adult subject (e.g., young adult, middle-aged adult, or elderly adult)) and / or non-human animals, e.g., mammals (e.g., domestic animals such as primates (e.g., cynomolgus monkeys, rhesus monkeys), cows, pigs, horses, sheep, goats, cats, and / or dogs, and / or birds (e.g., chickens, ducks, geese, and / or turkeys). In certain embodiments, the animal is a mammal (e.g., at any stage of development). In some embodiments, the animal (e.g., a non-human animal) may be a transgenic or genetically engineered animal. In some embodiments, the subject is a tumor resection subject, e.g., a subject who has recently undergone tumor resection. In some embodiments, the tumor resection subject is a subject who has undergone tumor resection less than 72 hours (e.g., less than 48 hours, less than 24 hours, less than 12 hours, less than 6 hours, or less) prior to receiving the compositions described herein. In some embodiments, the tumor resection subject is a subject who has undergone tumor resection less than 48 hours prior to receiving the compositions described herein. In some embodiments, the tumor resection subject is a subject who has undergone tumor resection less than 24 hours prior to receiving the compositions described herein. In some embodiments, the tumor resection subject is a subject who has undergone tumor resection less than 12 hours prior to receiving the compositions described herein.

[0088] Substantially: As used herein, the term "substantially" refers to a qualitative condition that indicates the entire or nearly entire extent or degree of a characteristic or property of interest. Those skilled in the art will understand that the agent of interest will achieve or avoid, if at all, an absolute result, e.g., the agent of interest will have no actual effect on the immune response, e.g., inflammation. Thus, the term "substantially" is used herein to express the potential lack of absoluteness inherent in many biological and chemical effects.

[0089] Sustained: As used interchangeably herein, the terms "sustained" or "extended" generally refer to prolonging an effect and / or process over a desired period of time. For example, in the context of sustained immunomodulation (e.g., in the presence of a composition or preparation described herein and / or utilized herein), such immunomodulatory effects may be observed for a longer period of time following administration of a particular immunomodulatory payload compared to that observed in the context of a composition comprising a biomaterial preparation, and administration of the same payload without such a biomaterial preparation otherwise described herein. In the context of sustained release of one or more agents of interest from a composition described herein (e.g., one or more polynucleotide agents encoding or modulating an immunomodulatory polypeptide incorporated into a biomaterial preparation described herein), such release may occur on a time scale ranging from about 30 minutes to several weeks or more. In some embodiments, the extent of sustained or extended release may be characterized in vitro or in vivo. For example, in some embodiments, release kinetics may be tested in vitro by placing the preparations and / or compositions described herein in a buffered aqueous solution (e.g., PBS at pH 7.4). In some embodiments, when a composition described herein is placed in a buffered aqueous solution (e.g., PBS at pH 7.4), less than 100% or less (including, for example, 90% or less, 80% or less, 70% or less, 50% or less or less) of one or more agents of interest (e.g., one or more polynucleotide agents encoding or modulating an immunomodulatory polypeptide incorporated into a biomaterial preparation described herein) is released from the biomaterial within 3 hours. In some embodiments, the release kinetics can be tested in vivo, for example, by administering (e.g., implanting) the composition to a target site (e.g., mammary fat pad) of an animal subject (e.g., a murine subject).In some embodiments, when the composition is administered (e.g., implanted) into a target site (e.g., mammary fat pad) of an animal subject (e.g., a murine subject), 70% or less or less (including, e.g., 60% or less, 50% or less, 40% or less, 30% or less or less) of one or more agents of interest (e.g., one or more polynucleotide agents encoding or modulating an immunomodulatory polypeptide incorporated into a biomaterial preparation described herein) is released in vivo after 8 hours.

[0090] Targeted Agents: The term "targeted agents," when used in reference to anticancer agents, means those that block the growth and spread of cancer by interfering with specific molecules ("molecular targets") involved in the growth, progression, and / or spread of cancer. Targeted agents may also be referred to as "targeted cancer therapies," "molecularly targeted drugs," "molecularly targeted therapies," or "precision medicines." Targeted agents differ from conventional chemotherapy in that they typically act on specific molecular targets specifically associated with cancer and / or a particular tumor or tumor type, stage, etc., whereas many chemotherapeutic agents act on all rapidly dividing cells (e.g., whether the cells are cancerous or not). Targeted agents are purposefully selected or designed to interact with their targets, whereas many standard chemotherapeutics are specified to kill cells.

[0091] Tautomers: The term "tautomers" or "tautomeric" refers to two or more interconvertible compounds resulting from at least one formal migration of a hydrogen atom and at least one change in valence (e.g., from a single bond to a double bond, a triple bond to a single bond, or vice versa). The exact ratio of tautomers depends on several factors, including temperature, solvent, and pH. Tautomerizations (i.e., reactions that provide tautomeric pairs) can be catalyzed by acids or bases. Exemplary tautomerizations include keto-enol, amide-imide, lactam-lactim, enamine-imine, and enamine-(different enamine) tautomerizations.

[0092] Test subject: As used herein, the term "test subject" refers to a subject to which the techniques provided herein are applied for experimental studies, e.g., to evaluate the effectiveness of compositions and / or preparations described herein in biomaterial degradation and / or anti-tumor immunity. In some embodiments, the test subject may be a human test subject or a population of human test subjects. For example, in some embodiments, the human test subject may be a normal healthy subject. In some embodiments, the human test subject may be a tumor resection subject. In some embodiments, the test subject may be a mammalian non-human animal or a population of mammalian non-human animals. Non-limiting examples of such mammalian non-human animals include mice, rats, dogs, pigs, rabbits, etc., which in some embodiments may be normal healthy subjects, while in some embodiments may be tumor resection subjects. In some embodiments, the mammalian non-human animal may be a transgenic or genetically engineered animal.

[0093] Therapeutic Agent: The term "therapeutic agent" refers to an agent that has one or more properties that result in a desired, usually beneficial, physiological effect. For example, a therapeutic agent may treat, ameliorate, and / or prevent a disease. A person of skill in the art reading this disclosure will understand that the term "therapeutic agent" as used herein does not require a particular level or type of therapeutic activity, as may be required for a regulatory agency to deem an agent "therapeutic" for regulatory purposes. As will be understood by a person of skill in the art, upon reading this disclosure, in some embodiments, certain polymeric biomaterial(s) preparations described herein (in the absence of an immunomodulatory payload) may have one or more properties that contribute to and / or achieve a desired physiological effect, and thus may be considered to be a "therapeutic agent" as that term is used herein (regardless of whether such biomaterials are deemed pharmacologic active by any particular regulatory agency). In some embodiments, a therapeutic agent that may be utilized in the preparations, compositions, and / or methods described herein (e.g., including the polymeric biomaterial(s) preparations described herein) may be or may include an immunomodulatory payload. In some embodiments, a therapeutic agent that may be utilized in the preparations, compositions, and / or methods described herein (including, for example, the polymeric biomaterial(s) preparations described herein) may be or include a non-immunomodulatory payload including, for example, a biologic, a small molecule, a nucleic acid, a polypeptide, or a combination thereof. In some embodiments, a therapeutic agent that may be utilized in the preparations, compositions, and / or methods described herein (including, for example, the polymeric biomaterial(s) preparations described herein) may be or include a chemotherapeutic agent, and in some embodiments, may be or include a cytotoxic agent.

[0094] Therapeutically effective amount: A "therapeutically effective amount" is an amount sufficient to provide a therapeutic effect in the treatment of a condition, which may be or include, for example, a reduction in frequency and / or severity and / or a delay in the onset of one or more features or symptoms associated with the condition. A therapeutically effective amount refers to an amount of a therapeutic agent(s) alone or in combination with other therapies that provides a therapeutic effect in the treatment of a condition. A "therapeutically effective amount" may include an amount that improves overall therapy, reduces or avoids the symptoms or causes of a condition, or enhances the therapeutic effectiveness of another therapeutic agent. Those skilled in the art will understand that a therapeutically effective amount need not be contained in a single dosage form. Rather, administration of an effective amount may involve administration of multiple doses, potentially over time (e.g., according to a dosing regimen, particularly one that has been established to provide an appropriate effect with a desired statistical confidence when applied to a relevant population).

[0095] Temperature-responsive: As used herein, the term "temperature-responsive," in the context of a temperature-responsive polymer or biomaterial (e.g., a polymeric biomaterial), refers to a polymer or biomaterial (e.g., a polymeric biomaterial) that exhibits an instantaneous or discontinuous change in one or more of its properties at a critical temperature (e.g., a critical gelation temperature). For example, in some embodiments, one or more of such properties is or includes the solubility of the polymer or biomaterial in a particular solvent. By way of example only, in some embodiments, a temperature-responsive polymer or biomaterial (e.g., a polymeric biomaterial) is characterized as being a homogenous polymer solution or colloid that is stable below a critical temperature (e.g., a critical gelation temperature) and instantaneously forms a polymer network (e.g., a hydrogel) when the critical temperature (e.g., a critical gelation temperature) is reached or exceeded. In some embodiments, a temperature-responsive polymer or biomaterial (e.g., a polymeric biomaterial) may be temperature reversible, e.g., in some embodiments where a polymer solution may instantaneously form a polymer network at a temperature above a critical gelation temperature, such resulting polymer network may instantaneously revert to a homogeneous polymer solution when the temperature is reduced below the critical gelation temperature.

[0096] Treatment: The terms "treatment," "treat," and "treating" refer to reversing, alleviating, delaying the onset of, or inhibiting the progression of a "pathological condition" as described herein (e.g., a disease, disorder, or condition including one or more signs or symptoms thereof), e.g., a cancer or tumor. In some embodiments, treatment may be administered after one or more signs or symptoms have developed or been observed. Treatment may be continued after symptoms have resolved, e.g., to delay or prevent recurrence and / or spread.

[0097] Tumor: The terms "tumor" and "neoplasm" are used interchangeably herein and refer to an abnormal mass of tissue whose proliferation exceeds and is not coordinated with the growth of normal tissue. Neoplasms or tumors can be "benign" or "malignant" depending on the following characteristics: degree of cellular differentiation (including morphology and function), rate of growth, local invasion, and metastasis. "Benign neoplasms" are generally well differentiated, have characteristically slower growth than malignant neoplasms, and remain localized at the site of origin. In addition, benign neoplasms do not have the ability to invade, invade, or metastasize to distant sites. Exemplary benign neoplasms include, but are not limited to, lipomas, chondromas, adenomas, acrochordons, senile hemangiomas, seborrheic keratosis, lentigines, and sebaceous hyperplasia. In some cases, certain "benign" tumors may subsequently give rise to malignant neoplasms, which may arise as a result of further genetic alterations in a subpopulation of the tumor's tumor cells; these tumors are referred to as "pre-malignant neoplasms." One example of a pre-malignant neoplasm is a teratoma. In contrast, "malignant neoplasms" are generally poorly differentiated (anaplastic) and have characteristically rapid growth accompanied by progressive infiltration, invasion, and destruction of surrounding tissue. Moreover, malignant neoplasms generally have the ability to metastasize to distant sites.

[0098] Tumor removal: As used herein, the term "tumor removal" encompasses partial or complete removal of a tumor, which may result from cancer treatment, e.g., surgical resection. In some embodiments, tumor removal refers to the physical removal of part or all of a tumor by surgery (i.e., "tumor resection"). In some embodiments, tumor removal may result from surgical tumor resection and adjuvant therapy (e.g., chemotherapy, immunotherapy, and / or radiation therapy). In some embodiments, adjuvant therapy may be administered after surgical tumor resection, e.g., at least 24 hours or more after surgical tumor resection.

[0099] Tumor resection subjects: As used herein, the term "tumor resection subjects" refers to subjects who have undergone or have recently undergone tumor resection. In some embodiments, a tumor resection subject is a subject in which at least 70% or more of the total tumor mass (including at least 80%, at least 90%, at least 95%, at least 98%, at least 99% or more (including 100%)) has been removed by surgical resection. In some cases, even when gross visual examination shows that all of the total tumor mass has apparently been removed, those skilled in the art will understand that there may be some residual cancer cells present microscopically at the visible margins of the resection. In some embodiments, a tumor resection subject may be determined to have negative margins (i.e., based on, for example, histological evaluation of the tissue surrounding the tumor resection site, there are no cancer cells present microscopically at the margins of the resection). In some embodiments, a tumor resection subject may be determined to have positive margins (i.e., based on, for example, histological evaluation of the tissue surrounding the tumor resection site, there are cancer cells present microscopically at the margins of the resection). In some embodiments, a tumor resection subject may have micrometastases and / or dormant disseminated cancer cells that may be encouraged to progress / grow due to a physiological response to surgery. In some embodiments, a tumor resection subject is administered a composition (e.g., as described and / or utilized herein) immediately after tumor resection is performed (e.g., administered during surgery). In some embodiments, a tumor resection subject receives a composition (e.g., as described and / or utilized herein) within 24 hours after surgery, including, for example, within 18 hours, within 12 hours, within 6 hours, within 3 hours, within 2 hours, within 1 hour, within 30 minutes, or less.

[0100] Tumor resection site: The term "tumor resection site" generally refers to a site where a portion or all of a tumor has been or is being removed via tumor resection. In some embodiments, a tumor resection site refers to a site where at least 70% or more of the total tumor mass has been removed by surgical resection, including at least 80%, at least 90%, at least 95%, at least 98%, at least 99%, or more (including 100%). In some cases, even when gross visual examination shows that all of the total tumor mass has apparently been removed, those skilled in the art will understand that there may be some residual cancer cells present microscopically at the visible resection margin. In some embodiments, a tumor resection site may be determined to have negative margins (i.e., no cancer cells are present microscopically at the resection margin, for example, based on histological evaluation of the tissue surrounding the tumor resection site). In some embodiments, a tumor resection site may be determined to have positive margins (i.e., cancer cells are present microscopically at the resection margin, for example, based on histological evaluation of the tissue surrounding the tumor resection site).

[0101] Variant: As used herein, the term "variant" refers to an entity that exhibits significant structural identity with a reference entity, but is structurally different from the reference entity in the presence or level of one or more chemical moieties compared to the reference entity. In many embodiments, a variant also differs functionally from the reference entity. In general, whether a particular entity is properly considered to be a "variant" of a reference entity is based on the degree of structural identity with the reference entity. As will be understood by those skilled in the art, any biological or chemical reference entity has certain characteristic structural elements. A variant is, by definition, a different chemical entity that shares one or more such characteristic structural elements. To give some examples, small molecules may have a distinctive core structural element (e.g., a macrocyclic core) and / or one or more distinctive pendant moieties, such that variants of small molecules share a core structural element and distinctive pendant moieties but differ in other pendant moieties and / or the type of linkages present within the core (single vs. double, E vs. Z, etc.); polypeptides may have distinctive sequence elements comprised of multiple amino acids that have designated positions relative to one another in linear or three-dimensional space and / or that contribute to a particular biological function; and nucleic acids may have distinctive sequence elements comprised of multiple nucleotide residues that have designated positions relative to other nucleotide residues in linear or three-dimensional space. For example, a variant biomaterial (e.g., a variant polymer or a polymeric biomaterial comprising a variant polymer) may differ from a reference biomaterial (e.g., a reference polymer or polymeric biomaterial) as a result of one or more structural modifications (e.g., addition, deletion, and / or modification, and / or grafting of chemical moieties), provided that the variant biomaterial (e.g., a variant polymer or a polymeric biomaterial comprising such a variant polymer) may retain the desired property(s) and / or function(s) (e.g., immunomodulation and / or temperature responsiveness) of the reference biomaterial.For example, a variant of an immunomodulatory biomaterial may differ from a reference immunomodulatory biomaterial (e.g., a reference polymer or polymeric biomaterial) as a result of one or more structural modifications (e.g., addition, deletion, and / or modification, and / or grafting of chemical moieties), provided that the variant biomaterial (e.g., a variant polymer or a polymeric biomaterial comprising such a variant polymer), e.g., when used in the methods described herein, may affect the immune system (e.g., by stimulating innate immunity). In some embodiments, a variant immunomodulatory biomaterial (e.g., a variant polymer or a polymeric biomaterial comprising a variant polymer) is characterized in that, when assessed 24 hours after administration of such variant immunomodulatory biomaterial (e.g., a variant polymer or a polymeric biomaterial comprising a variant polymer) to a target site in a subject, the amount of one or more proinflammatory cytokines (e.g., without limitation, CXCL10, IFN-α, IFN-β, IL-1β, IL-6, IL-18, and / or TNF-α) observed at the target site and / or in the systemic circulation of the subject is at least 60% or more (including, for example, at least 70%, at least 80%, at least 90%, at least 95%, at least 98%, or up to 100%) of the amount observed when a reference biomaterial (e.g., a reference polymer or polymeric biomaterial) is administered to the target site. In some embodiments, the variant immunomodulatory biomaterial (e.g., a variant polymer or a polymeric biomaterial comprising a variant polymer) is characterized in that the amount of one or more proinflammatory cytokines (e.g., without limitation, CXCL10, IFN-α, IFN-β, IL-1β, IL-6, IL-18, and / or TNF-α) observed at the target site and / or in the systemic circulation of a subject, when assessed 24 hours after administration of such variant biomaterial (e.g., a variant polymer or a polymeric biomaterial comprising a variant polymer) to a target site in a subject, is at least 1.1-fold or more (including, for example, at least 1.5-fold, at least 2-fold, at least 3-fold, at least 4-fold, at least 5-fold or more) the amount observed when a reference biomaterial (e.g., a reference polymer or polymeric biomaterial) is administered to the target site. In some embodiments, the variant biomaterial (e.g., variant polymeric biomaterial) exhibits at least one physical property that differs from that of the reference biomaterial (e.g., reference polymeric biomaterial). For example, in some embodiments, the variant biomaterial (e.g., variant polymeric biomaterial) may exhibit increased water solubility (e.g., at physiological pH) compared to the water solubility of the reference biomaterial (e.g., reference polymeric biomaterial). In some embodiments, the variant has 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 structural modifications compared to the reference. In some embodiments, the variant has a small number (e.g., less than 5, 4, 3, 2, or 1) structural modifications (e.g., alkylation, acylation, quaternization, hydroxyalkylation, carboxyalkylation, thiolation, phosphorylation, glycosylation, etc.). In some embodiments, the variant has no more than 5, 4, 3, 2, or 1 addition or deletion of chemical moieties compared to the reference, and in some embodiments, no additions or deletions. In some embodiments, a variant is a substance that can be produced from a reference by chemical manipulation, hi some embodiments, a variant is an entity that can be produced through the performance of a synthetic process that is substantially similar (e.g., shares multiple steps) with the process that produces the reference.

[0102] DETAILED DESCRIPTION OF CERTAIN EMBODIMENTS Provided herein is a composition comprising at least one or more polymeric biomaterials and one or more nucleic acids. In some embodiments, the provided compositions can localize the delivery of one or more nucleic acids that can code for an active agent (e.g., an immunomodulatory agent) or function directly as an active agent to a target site (e.g., a site where a tumor has been removed and / or where tumor cells have been treated or killed, e.g., by chemotherapy or radiation, and / or in / near an untreated tumor), thereby concentrating the action of the active agent at the target site in need thereof. Such compositions can be particularly useful in the treatment of cancer.

[0103] In some embodiments, the compositions may include a polymeric biomaterial(s), a nucleic acid(s), and optionally one or more additional immunomodulatory molecules. In some embodiments, the compositions provided may include a polymeric biomaterial(s) and a nucleic acid(s) that function as a regulator of an adaptive immune response and / or a regulator of an innate immune response. In some embodiments, the additional immunomodulatory molecule may include an inhibitor(s) of a proinflammatory immune response mediated by the p38 mitogen-activated protein kinase (MAPK) pathway (e.g., a p38 MAPK inhibitor). In some embodiments, the compositions may include an activator of an innate immune response, a cytokine, and / or a chemokine. In some embodiments, the compositions may include one or more activators of an adaptive immune response. In some embodiments, the compositions provided may further include an additional therapeutic agent (e.g., an additional inhibitor of a proinflammatory pathway, a regulator of macrophage effector function, and / or a chemotherapeutic agent, etc.).

[0104] In some embodiments, the compositions may include therapeutic agents (e.g., acting as immunomodulators or nucleic acids encoding immunomodulators) that may mediate or inhibit inflammation (e.g., chronic inflammation) induced by trauma or microtrauma (e.g., by surgery such as surgical tumor resection, keyhole surgery, or injections), thus providing a unique tool for the treatment of cancer, particularly solid tumors. In some embodiments, the therapeutic agents provided may modify and / or reduce or inhibit the activity of myeloid-derived suppressor cells (MDSCs). In some embodiments, the therapeutic agents provided may modify and / or reduce or inhibit the activity of neutrophils. In some embodiments, the therapeutic agents provided may modify and / or reduce or inhibit the activity of macrophages. In some embodiments, the therapeutic agents provided may modify and / or reduce or inhibit the recruitment of immunosuppressive cells. In some embodiments, the therapeutic agents provided may modify and / or reduce or inhibit acute inflammation. In some embodiments, the therapeutic agents provided may activate the innate immune response system and / or the adaptive immune response system. The compositions, methods, and kits provided herein are also advantageous over existing methods in that they do not require administration of cells (e.g., adoptive cell transfer) or the incorporation or presence of additional components such as nanoparticles, microparticles, certain peptides, or tumor antigens.

[0105] In some embodiments, the compositions described herein are useful for treating cancer (e.g., solid tumors or metastases) in the perioperative setting. In some embodiments, the compositions may deliver immunotherapy in a subject in need thereof by implantation or injection of the composition at a therapeutically necessary site. In some embodiments, the compositions provided herein are particularly advantageous over existing immunotherapies, at least because they may, in some embodiments, release an immunomodulator (e.g., a nucleic acid that acts as an immunomodulator or encodes an immunomodulator) directly to the tumor resection site, avoiding systemic administration. Thus, some embodiments of the compositions described herein provide a vehicle for drug delivery at the tumor resection site that avoids potential toxicity that may be associated with conventional systemic administration of immunotherapy. Concentrating the immunotherapy at the tumor resection site may improve efficacy as well. In certain embodiments, the compositions provided are useful for delaying and / or inhibiting tumor growth, preventing cancer recurrence, preventing tumor metastasis, and / or preventing primary tumor regrowth.

[0106] In particular, in some embodiments, the present disclosure provides techniques for the suppression of immune responses which may themselves develop additional immune suppression (e.g., the activity of certain macrophages, neutrophils, and / or MDSCs).

[0107] Without wishing to be bound by any particular theory, the present disclosure notes that in some embodiments, the techniques provided herein may reduce a type of inflammation that is commonly observed in the context of chronic inflammation (e.g., often associated with autoimmune disease), but may be activated in an acute setting (i.e., post-surgery) as described herein. The present disclosure provides insight that inflammation-targeting therapy by harnessing the power of localized delivery of nucleic acids as described herein may be uniquely useful in the post-tumor resection setting. Those skilled in the art will appreciate that many therapeutic strategies designed to ameliorate or stimulate autoimmune responses and / or effective at doing so may be devastating in the setting of tumor resection, as they may result in the exacerbation of tumor progression phenotypes. The present disclosure teaches that, notwithstanding this general principle, targeting certain immune-modulating pathways via targeted and / or localized administration of nucleic acids as described herein is surprisingly useful in the context of cancer treatment.

[0108] In some embodiments, the described compositions comprising polymeric biomaterials and nucleic acids may be combined with therapies including, for example, other immunomodulatory strategies, such as activation / agonism of the innate immune system via small molecule or peptide administration (e.g., via administration of agents such as STING agonists or TLR agonists).

[0109] composition Polymer Biomaterials The compositions described herein generally include at least one biomaterial, referred to throughout as polymeric biomaterial(s). In some embodiments, the polymeric biomaterial may form a polymer network that may act as a scaffold or depot for the dispersion of at least some of the additional components (e.g., nucleic acids) contained within the composition. In some embodiments, the scaffold or depot includes any synthetic or naturally occurring material that is suitable for containing and promoting the sustained or extended release of any therapeutic agent in the compositions described herein. Thus, the polymeric biomaterial may have physical properties that provide some of the advantageous properties of the compositions described herein (e.g., storage modulus, biodegradation, and / or release profile of the therapeutic agent).

[0110] In certain embodiments, the composition comprises a polymeric biomaterial(s) that can extend the release of a therapeutic agent (e.g., a nucleic acid) when delivered to a target site (e.g., a tumor resection site) relative to administration of the same therapeutic agent in solution. In certain embodiments, the polymeric biomaterial(s) extends the release of the therapeutic agent at the tumor resection site by at least 5 minutes, 10 minutes, 20 minutes, 30 minutes, 40 minutes, 50 minutes, 60 minutes, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 7 hours, 8 hours, 9 hours, 10 hours, 11 hours, 12 hours, 18 hours, 24 hours, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 2 weeks, 3 weeks, or 4 weeks relative to administration of the same therapeutic agent in solution. In some embodiments, the polymeric biomaterial(s) prolong the release of at least one therapeutic agent (e.g., a nucleic acid encoding or acting as an immunomodulatory agent) such that, when evaluated at a particular time point after administration, more of the therapeutic agent is present at the tumor resection site relative to the level observed when the therapeutic agent is administered in solution. For example, in some embodiments, when evaluated 24 hours after administration, the amount of the therapeutic agent released and present at the tumor resection site is at least 30% greater (e.g., at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90% or more) than the amount observed when the therapeutic agent is administered in solution. In some embodiments, when evaluated 48 hours after administration, the amount of the therapeutic agent released and present at the tumor resection site is at least 30% greater (e.g., at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90% or more) than the amount observed when the therapeutic agent is administered in solution. In some embodiments, when assessed 3 days after administration, the amount of therapeutic agent released and present at the tumor resection site is at least 30% greater (including, for example, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90% or more) than the amount observed when the therapeutic agent is administered in solution.In some embodiments, when assessed 5 days after administration, the amount of therapeutic agent released and present at the tumor resection site is at least 30% greater (including, for example, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90% or more) than the amount observed when the therapeutic agent is administered in solution.

[0111] In some embodiments, compositions comprising the polymeric biomaterial(s) preparation described herein (e.g., in a precursor state or in a polymer network state) have a viscosity of, for example, 9000 mPa·s or less, 8000 mPa·s or less, 7000 mPa·s or less, 6000 mPa·s or less, 5000 mPa·s or less, 4000 mPa·s or less, 3500 mPa·s or less, 3000 mPa·s or less, 2500 mPa·s or less, 2000 mPa·s or less, 3500 mPa·s or less, 3000 mPa·s or less, 2500 mPa·s or less, 3000 mPa·s or less, 4000 mPa·s or less, 5000 mPa·s or less, 5000 mPa·s or less, 6000 mPa·s or less, 7000 mPa·s or less, 8000 mPa·s or less, 9000 mPa·s or less, 9000 mPa·s or less, 9000 mPa·s or less, 1 ... The viscosity may be characterized by a viscosity of 10,000 mPa·s or less, including a·s or less, 1500 mPa·s or less, 1000 mPa·s or less, 500 mPa·s or less, 250 mPa·s or less, 200 mPa·s or less, 150 mPa·s or less, 100 mPa·s or less, 75 mPa·s or less, 50 mPa·s or less, 25 mPa·s or less, 20 mPa·s or less, 15 mPa·s or less, 10 mPa·s or less, or less. In some embodiments, a composition comprising a polymeric biomaterial(s) preparation described herein (e.g., in a precursor state or in a polymer network state) has a viscosity of, e.g., at least 10 mPa·s, at least 20 mPa·s, at least 30 mPa·s, at least 40 mPa·s, at least 50 mPa·s, at least 60 mPa·s, at least 70 mPa·s, at least 80 mPa·s, at least 90 mPa·s, at least 100 mPa·s, at least 125 mPa·s, at least 150 mPa·s, at least The viscosity may be characterized by a viscosity of at least 5 mPa·s or greater, including at least 175 mPa·s, at least 250 mPa·s, at least 500 mPa·s, at least 1000 mPa·s, at least 1500 mPa·s, at least 2000 mPa·s, at least 2500 mPa·s, at least 3000 mPa·s, at least 4000 mPa·s, at least 5000 mPa·s, at least 6000 mPa·s, at least 7000 mPa·s, at least 8000 mPa·s, at least 9000 mPa·s or greater. Combinations of the above ranges are also possible.For example, in some embodiments, a composition comprising a polymeric biomaterial(s) preparation described herein (e.g., in a precursor state or polymeric network state) may be characterized by a viscosity of 5 mPa·s to 10,000 mPa·s, or 10 mPa·s to 5000 mPa·s, or 5 mPa·s to 200 mPa·s, or 20 mPa·s to 100 mPa·s, or 5 mPa·s to 20 mPa·s. One of skill in the art reading this disclosure will understand that, in some cases, the viscosity of a composition comprising a polymeric biomaterial(s) preparation described herein may be selected or adjusted based on, for example, the route of administration (e.g., injection vs. implantation), the injection volume and / or time, and / or the duration of the effect of the stimulation of innate immunity. As will also be understood by one of skill in the art, the viscosity of a polymer depends, for example, on the temperature and the concentration of the polymer in the test sample. In some embodiments, the viscosity of a composition comprising a polymeric biomaterial(s) preparation described herein may be characterized by a viscosity of 1000 s at 20° C., for example, at 1000 s. -1 The shear rate may be measured at 0.05 to 1.05 mm.

[0112] In some embodiments where the composition comprising the polymeric biomaterial(s) preparation described herein is in a polymeric network state, such a polymeric network state can be at least 100 Pa, at least 200 Pa, at least 300 Pa, at least 400 Pa, at least 500 Pa, at least 600 Pa, at least 700 Pa, at least 800 Pa, at least 900 Pa, at least 1000 Pa, at least 1100 Pa, at least 1200 Pa, at least 1300 Pa, at least 1400 Pa, at least 1500 Pa, at least 1600 Pa, at least 1700 Pa, at least 1800 Pa, at least 1900 Pa, at least 2000 Pa, at least 2100 Pa, at least 2200 Pa, at least 2300 Pa, at least 2400 Pa, at least 2500 Pa, at least 2600 Pa, at least 2700 Pa, at least 2800 Pa, at least 2900 Pa, at least 3000 Pa, at least 3100 Pa, at least 3200 Pa, at least 3300 Pa, at least 3400 Pa, at least 3500 Pa, at least 3600 Pa, at least 3700 Pa, at least 3800 Pa, at least 3900 Pa, at least 4000 Pa, at least 4100 Pa, at least 4200 Pa, at least 4300 Pa, at least 4400 Pa, at least 4500 Pa, at least 4600 Pa, at least 4700 Pa, at least 4800 Pa, at least 4900 Pa, at least 5000 Pa, at least 5100 Pa, at least 5200 Pa, at least 5300 Pa, at least 5400 Pa, The polymer network state of the composition comprising the polymeric biomaterial(s) preparation described herein may be characterized by a storage modulus of 10 kPa or less, 9 kPa or less, 8 kPa or less, 7 kPa or less, 6 kPa or less, or less than 9 kPa. Combinations of the above ranges are also possible. For example, in some embodiments, such polymer network states of compositions comprising the polymeric biomaterial(s) preparations described herein may be characterized by a storage modulus of 100 Pa to 10 kPa, or 200 Pa to 5000 Pa, or 300 Pa to 2500 Pa, or 500 Pa to 2500 Pa, or 100 Pa to 500 Pa.Those skilled in the art will appreciate that various rheological characterization methods (e.g., those described in Weng et al., “Rheological Characterization of in situ Crosslinkable Hydrogels Formulated from Oxidized Dextran and N-Carboxyethyl Chitosan” Biomacromolecules, 8:1109-1115 (2007)) can be used to measure the storage modulus of a material, and in some cases, the storage modulus of a material can be measured by a rheometer and / or dynamic mechanical analysis (DMA). Those skilled in the art will also appreciate that rheological properties can vary with ambient conditions, e.g., temperature and / or pH. Thus, in some embodiments, the provided polymer combination preparations are characterized by a storage modulus (e.g., as described herein) measured at a subject's body temperature (e.g., 37° C. for a human subject), at a pH of, e.g., 5-8, or at physiological pH (e.g., pH 7). As will be apparent to those skilled in the art upon reading the disclosure provided herein, the storage modulus of a polymer combination preparation, e.g., provided in the form of particles, refers to the bulk storage modulus of the particles in the population.

[0113] In certain embodiments, the composition comprising the polymeric biomaterial is composed of a positively charged polymer. In certain embodiments, the polymeric biomaterial is composed of a negatively charged polymer(s). In certain embodiments, the polymeric biomaterial is composed of a neutral polymer(s). In certain embodiments, the polymeric biomaterial is selected from the group consisting of hyaluronic acid, alginate, chitosan, chitin, chondroitin sulfate, dextran, gelatin, collagen, starch, cellulose, polysaccharides, fibrin, poly-L-lysine, methylcellulose, ethylene-vinyl acetate (EVA), poly(lactic-co-glycolic) acid (PLGA), polylactic acid (PLA), polyglycolic acid (PGA), polyethylene glycol (PEG), PEG diacrylate (PEGDA), disulfide-containing PEGDA (PEGSSDA), PEG dimethacrylate (PEGDMA), polydioxanone (PDO), polyhydroxybutyrate (PHB), poly(2-hydroxyethyl methacrylate) (pHEMA), polycaprolactone (PCL), poly(beta-amino ester) (PBAE), poly(ester amide), poly(propylene glycol) (PPG ), poly(aspartic acid), poly(glutamic acid), poly(propylene fumarate) (PPF), poly(sebacic anhydride) (PSA), poly(trimethylene carbonate) (PTMC), poly(desaminotyrosyl tyrosine alkyl ester carbonate) (PDTE), poly[bis(trifluoroethoxy)phosphazene], polyoxymethylene, single-walled carbon nanotubes, polyphosphazenes, polyanhydrides, poly(N-vinyl-2-pyrrolidone) (PVP), poly(vinyl alcohol) (PVA), poly(acrylic acid) (PAA), poly(methacrylic acid) (PMA), polyacetals, poly(alpha esters), poly(ortho esters), polyphosphates, polyurethanes, polycarbonates, polyamides, polyhydroxyalkanoates, polyglycerols, polyglucuronic acid, and / or combinations and / or derivatives thereof.

[0114] In certain embodiments, the composition comprising at least one polymeric biomaterial is comprised of a hydrophobic polymeric biomaterial(s). In certain embodiments, the hydrophobic polymeric biomaterials include ethylene-vinyl acetate (EVA), poly(lactic-co-glycolic) acid (PLGA), polylactic acid (PLA), polyglycolic acid (PGA), polydioxanone (PDO), polyhydroxybutyrate (PHB), polycaprolactone (PCL), poly(ester amide), poly(propylene fumarate) (PPF), poly(sebacic anhydride) (PSA), poly(trimethylene carbonate) (PTMC), poly(desaminotyrosyl tyrosine) (PSA ... Examples of suitable hydrophobic polymeric biomaterials include poly[bis(trifluoroethoxy)phosphazene], polyoxymethylene, single-walled carbon nanotubes, polyphosphazenes, polyanhydrides, poly(N-vinyl-2-pyrrolidone) (PVP), poly(acrylic acid) (PAA), poly(methacrylic acid) (PMA), poly(alpha esters), poly(ortho esters), polyphosphate esters, polyurethanes, polycarbonates, polyamides, and / or polyhydroxyalkanoates. In some embodiments, the use of hydrophobic polymeric biomaterials may be particularly useful when the therapeutic agent(s) in the composition are hydrophilic. In some embodiments where the polymeric biomaterial is hydrophobic, the hydrophobic therapeutic agent is expected to be released over a longer period of time (e.g., days / weeks) rather than a relatively short period of time (hours / days). Thus, in certain embodiments, when the polymeric biomaterial is a hydrophobic polymer, the therapeutic agent(s) of the composition are therefore hydrophilic molecules. Additionally, in certain embodiments, when the biomaterial is a hydrophobic polymer, the therapeutic agent(s) of the composition are hydrophobic and / or neutral molecules.

[0115] In certain embodiments, the provided compositions comprising at least one polymeric biomaterial include a crosslinked biologic. In certain embodiments, the biologic is crosslinked by the self-immolative crosslinker dithiobis(ethyl 1H-imidazole-1-carboxylate) (DIC). In certain embodiments, the resulting polymer network or hydrogel is loaded with a small molecule. In certain embodiments, the small molecule comprises a nucleic acid. In certain embodiments, the provided compositions may include a polynucleotide and a polynucleotide agent carrier.

[0116] The polymeric biomaterials useful in the compositions described herein are biocompatible. In some embodiments, the polymeric biomaterials are biodegradable in vivo. In some embodiments, the compositions provided herein can be chemically and / or biologically degraded within a physiological environment, such as within the body. Degradation of the provided compositions can occur at various rates, depending on the components and polymeric biomaterials used. For example, in some embodiments, the half-life of the provided compositions (the time at which 50% of the composition is degraded into monomers and / or other non-polymeric moieties) can be on the order of days, weeks, months, or years. In some embodiments, the compositions can be biologically degraded, for example, by enzymatic activity or cellular mechanisms, in some cases, for example, by exposure to lysozyme (e.g., having a relatively low pH), or by simple hydrolysis. In some embodiments, the provided compositions can be degraded into monomers and / or other non-polymeric moieties that cells can either recycle or dispose of without significant toxic effects on the cells. In some embodiments, the provided compositions are stable in vivo to deliver the drug to the intended target within a suitable length of time.

[0117] In certain embodiments, the composition comprises a polymeric biomaterial(s) capable of forming a polymeric network with or without the addition of a crosslinking agent. In some embodiments, the polymeric biomaterial(s) can form a polymeric network biomaterial in less than 10 minutes. The embodiments of the compositions described herein can be prepared by any method known in the art of pharmacology. In certain embodiments, such a preparation method includes the steps of adding thiol-modified hyaluronic acid to a mold, adding a nucleic acid (e.g., a nucleic acid encoding or acting as a regulator of an immune response), optionally adding an activator of an adaptive immune response to the mold, optionally adding a chemokine or cytokine to the mold, optionally adding an activator of an innate immune response to the mold, adding a crosslinker to the mold (e.g., a thiol-reactive PEGDA crosslinker), and allowing the mixture to solidify for at least 10 minutes, at least 15 minutes, at least 20 minutes, at least 25 minutes, at least 30 minutes, at least 35 minutes, at least 40 minutes, at least 45 minutes, at least 50 minutes, at least 55 minutes, at least 1 hour, at least 90 minutes, at least 2 hours, at least 3 hours, at least 4 hours, at least 5 hours, or at least 6 hours.

[0118] In certain embodiments, the composition comprises at least one biomaterial, and the biomaterial(s) may form a polymer network. In certain embodiments, the polymer network is a hydrogel. In certain embodiments, the polymer network is crosslinked. The polymer network or hydrogel may be crosslinked using any method known in the art, such as chemical crosslinking methods (e.g., by using small molecule crosslinkers that may be derived from natural sources or may be synthesized), polyelectrolyte crosslinking (e.g., mixing a polymer with a second polymer that contains an opposite charge), thermally induced crosslinking, photoinduced crosslinking (e.g., using vinylsulfone, methacrylate, acrylate), pH induced crosslinking, and enzyme-catalyzed crosslinking. In some embodiments, one or more crosslinking methods described in Parhi, Adv Pharm Bull., Review 7(4): 515-530 (2017) (herein incorporated by reference for purposes described herein) may be used in forming the polymer network or hydrogel. In some embodiments, the polymer network or hydrogel is a non-crosslinked polymer network or hydrogel.

[0119] In certain embodiments, compositions comprising polymeric biomaterials (e.g., cross-linked or non-cross-linked) include hyaluronic acid, alginate, chitosan, chondroitin sulfate, dextran, gelatin, collagen, starch, cellulose, polysaccharides, fibrin, poly-L-lysine, methylcellulose, polyethylene glycol (PEG), PEG diacrylate (PEGDA), disulfide-containing PEGDA (PEGSSDA), PEG dimethacrylate (PEGDMA), poly(2-hydroxyethyl methacrylate) (pHEMA), poly(beta-amino ester) (PBAE), poly(aspartic acid), poly(glutamic acid), poly(propylene glycol) (PPG), poly(vinyl alcohol) (PVA), polyacetal, polyglycerol, polyglucuronic acid, and / or combinations thereof. In certain embodiments, when the polymeric biomaterial is a polymer network or hydrogel, the therapeutic agent(s) of the composition are hydrophilic molecules. In certain embodiments, when the polymeric biomaterial is a polymer network or hydrogel, the therapeutic agent(s) of the composition are hydrophobic molecules. In certain embodiments, when the polymeric biomaterial is a polymer network or hydrogel, the therapeutic agent(s) of the composition are hydrophobic or hydrophilic molecules. In certain embodiments, when the polymeric biomaterial is a polymer network or hydrogel, the therapeutic agent(s) of the composition are amphiphilic molecules (e.g., both hydrophobic and hydrophilic molecules).

[0120] In certain embodiments, the composition comprising the polymeric biomaterial is composed of hyaluronic acid, chitosan, and / or alginate. In certain embodiments, the polymeric biomaterial is cross-linked hyaluronic acid, cross-linked chitosan, and / or cross-linked alginate. In certain embodiments, the polymeric biomaterial comprises cross-linked hyaluronic acid, cross-linked chitosan, or cross-linked alginate. In certain embodiments, the polymeric network or hydrogel is hyaluronic acid, chitosan, or alginate. In certain embodiments, the polymeric biomaterial comprises hyaluronic acid. In certain embodiments, the polymeric biomaterial comprises cross-linked hyaluronic acid. In certain embodiments, the polymeric biomaterial is hyaluronic acid. In certain embodiments, the polymeric biomaterial is cross-linked hyaluronic acid. In certain embodiments, the polymeric network is composed of two or more different polymeric biomaterials. In certain embodiments, the polymeric network or hydrogel comprises hyaluronic acid and poloxamer.

[0121] Exemplary Poloxamers and Their Variants In some embodiments, the polymer included in the composition comprising the polymeric biomaterial(s) preparation described herein may be or may include a poloxamer. Poloxamers are typically block copolymers that contain a hydrophobic chain of polyoxypropylene (e.g., polypropylene glycol, PEG, and / or poly(ethylene oxide), PEO) adjacent to two hydrophilic chains of polyoxyethylene (e.g., polyethylene glycol, PEG, and / or poly(ethylene oxide), PEO). Poloxamers are known under the trade names Synperonic, Pluronic, and / or Kolliphor. Generally, poloxamers are non-ionic surfactants that, in some embodiments, may have good solubilizing ability, low toxicity, and / or high compatibility with cells, body fluids, and a wide range of chemicals.

[0122] In some embodiments, the poloxamer for use according to the present disclosure may be any poloxamer known in the art. For example, as will be understood by those skilled in the art, poloxamers are generally named with the letter P (for poloxamer) followed by three digits. The first two digits are multiplied by 100 to obtain the approximate molecular weight of the polyoxypropylene chain, and the last digit is multiplied by 10 to obtain the percentage of polyoxyethylene content. By way of example only, P407 refers to a poloxamer with a polyoxypropylene molecular weight of 4000 g / mole and a polyoxyethylene content of 70%. Those skilled in the art will also understand that for the trade names Pluronic and Synperonic, the coding of such poloxamers begins with a letter to define its physical form at room temperature (e.g., L=liquid, P=paste, F=flake (solid)), followed by two or three digits, where the first digit of the numerical designation (two of the three digits) is multiplied by 300 to indicate the approximate molecular weight of the polyoxypropylene chain, and the last digit is multiplied by 10 to obtain the percentage of polyoxyethylene content. By way of example only, L61 refers to a liquid preparation of poloxamer with a polyoxypropylene molecular weight of 1800 g / mole and a polyoxyethylene content of 10%. In addition, as will be apparent to those skilled in the art, poloxamer 181 (P181) is equivalent to Pluronic L61 and Synperonic PE / L61.

[0123] In some embodiments, poloxamers that may be included in compositions comprising the polymeric biomaterial(s) preparations described herein may include poloxamer 124 (e.g., Pluronic L44 NF), poloxamer 188 (e.g., Pluronic F68NF), poloxamer 181 (e.g., Pluronic L61), poloxamer 182 (e.g., Pluronic L62), poloxamer 184 (e.g., Pluronic L64), poloxamer 237 (e.g., Pluronic F87 NF), poloxamer 338 (e.g., Pluronic F108 NF), poloxamer 331 (e.g., Pluronic L101), poloxamer 407 (e.g., Pluronic F127 NF), or combinations thereof. In some embodiments, the provided polymeric biomaterial(s) preparations may include at least two or more different poloxamers. Additional poloxamers listed in Table 1 of Russo and Villa “Poloxamer Hydrogels for Biomedical Applications” Pharmaceutics (2019) 11(12):671, the contents of which are incorporated herein by reference for purposes described herein, may also be useful in preparing the polymeric biomaterial(s) described herein.

[0124] In some embodiments, the poloxamer that may be included in the composition comprising the polymeric biomaterial(s) preparation described herein may be or may include poloxamer 407 (P407). In some embodiments, P407 is a triblock poloxamer copolymer with a hydrophobic PPO block adjacent to two hydrophilic PEO blocks. The approximate length of the two PEO blocks is typically 101 repeat units, while the approximate length of the PPO block is 56 repeat units. In some embodiments, P407 has an average molecular weight of about 12,600 Da, of which about 70% corresponds to PEO. In some embodiments, P407 may easily self-assemble to form micelles, depending on the concentration and ambient temperature. Without wishing to be bound by a particular theory, dehydration of the hydrophobic PPO block in combination with hydration of the PEO block may result in the formation of spherical micelles, and the subsequent packaging of the micellar structure results in a 3D cubic lattice that constitutes the main structure of the poloxamer hydrogel. They are also non-toxic and stable, and therefore suitable for use as controlled release therapeutics. As will be appreciated by those skilled in the art, P407 concentrations in hydrogel formulations based on binary poloxamer / water mixtures typically range from 16-20 w / v%, with values ​​of about 18 w / v% being most frequently used. See, for example, Pereia et al. "Formulation and Characterization of Poloxamer 407®: Thermoreversible Gel Containing Polymeric Microparticles and Hyaluronic Acid" Quim. Nova, Vol. 36, No. 8, 1121-1125 (2013), the contents of which are incorporated herein by reference in their entirety for the purposes set forth herein.

[0125] In some embodiments, the poloxamer that may be included in the compositions comprising the polymeric biomaterial(s) preparations described herein may be or may include a poloxamer described in International Patent Application No. PCT / US21 / 42110, filed July 17, 2021, the entire contents of which are incorporated herein by reference for purposes described herein.

[0126] In some embodiments, the provided temperature-responsive polymeric biomaterial(s) preparations include a first polymer component (e.g., a poloxamer as described herein) and a second polymer component that is not a poloxamer. In some embodiments, the second polymer component may be present in the provided polymeric biomaterial(s) preparations at a concentration of 15% (w / w) or less. In some embodiments, the second polymer component may be present in the provided polymeric biomaterial(s) preparations at a concentration of 10% (w / w) or less, including, for example, at a concentration of 10% (w / w), 9% (w / w), 8% (w / w), 7% (w / w), 6% (w / w), 5% (w / w), 4% (w / w), 3% (w / w), 2% (w / w), 1% (w / w), 0.5% (w / w) or less. In some embodiments, the second polymer component is present in a provided polymeric biomaterial(s) preparation in an amount of, for example, at least 0.2% (w / w), at least 0.3% (w / w), at least 0.4% (w / w), at least 0.5% (w / w), at least 0.6% (w / w), at least 0.7% (w / w), at least 0.8% (w / w), at least 0.9% (w / w), at least 1% (w / w), at least 1.5 ... The second polymer component may be present at a concentration of at least 0.1% (w / w), including at least 2% (w / w), at least 2.5% (w / w), at least 3% (w / w), at least 3.5% (w / w), at least 4% (w / w), at least 4.5% (w / w), at least 5% (w / w), at least 6% (w / w), at least 7% (w / w), at least 8% (w / w), at least 9% (w / w), at least 10% (w / w) or more. In some embodiments, the second polymer component in the provided polymeric biomaterial(s) preparations may be present at a concentration of 0.1% (w / w) to 10% (w / w), or 0.1% (w / w) to 8% (w / w), or 0.1% (w / w) to 5% (w / w), or 1% (w / w) to 5% (w / w).In some embodiments, the second polymer component may be present in a provided polymeric biomaterial(s) preparation at a concentration of 0.5% (w / w) to 10% (w / w), or 0.5% (w / w) to 5% (w / w), or 1% (w / w) to 10% (w / w), or 1% (w / w) to 5% (w / w), or 2% to 10% (w / w).

[0127] In some embodiments, the second polymer component included in the provided polymeric biomaterial(s) preparations can be or can include at least one biocompatible and / or biodegradable polymer component, including, for example, at least two, at least three, at least four or more.Examples of such biocompatible and / or biodegradable polymer components include immunomodulatory polymers, carbohydrate polymers (e.g., polymers that are or include a carbohydrate backbone, including, for example, carbohydrates, e.g., chitosan, alginate, hyaluronic acid, and / or variants thereof), polyacrylic acid, silica gel, polyethyleneimine (PEI), polyphosphazene, and / or variants thereof), cellulose, chitin, chondroitin sulfate, collagen, dextran, gelatin, ethylene-vinyl chloride ... elastomer acetate (EVA), fibrin, poly(lactic-co-glycolic) acid (PLGA), polylactic acid (PLA), polyglycolic acid (PGA), polyethylene glycol (PEG), PEG diacrylate (PEGDA), disulfide-containing PEGDA (PEGSSDA), PEG dimethacrylate (PEGDMA), polydioxanone (PDO), polyhydroxybutyrate (PHB), poly(2-hydroxyethyl methacrylate) (pHEMA), polycarboxybetaine (PCB), polysulfobetaine (PS B), polycaprolactone (PCL), poly(beta-amino ester) (PBAE), poly(ester amide), poly(propylene glycol) (PPG), poly(aspartic acid), poly(glutamic acid), poly(propylene fumarate) (PPF), poly(sebacic anhydride) (PSA), poly(trimethylene carbonate) (PTMC), poly(desaminotyrosyltyrosine alkyl ester carbonate) (PDTE), poly[bis(trifluoroethoxy)phosphazene], polyoxymethylene, monolayer carbon Examples of suitable polyether ether esters include, but are not limited to, nanotubes, polyanhydrides, poly(N-vinyl-2-pyrrolidone) (PVP), poly(vinyl alcohol) (PVA), poly(acrylic acid) (PAA), poly(methacrylic acid) (PMA), polyacetals, poly(alpha esters), poly(ortho esters), polyphosphates, polyurethanes, polycarbonates, polyamides, polyhydroxyalkanoates, polyglycerol, polyglucuronic acid, starch, variants thereof, and / or combinations thereof.

[0128] In some embodiments, the second polymer component included in the provided polymeric biomaterial(s) preparation is or includes an immunomodulatory polymer, e.g., a polymer that modulates one or more aspects of the immune response (e.g., a polymer that induces innate immune agonism). In some embodiments, the immunomodulatory polymer can be or include a polymeric agonist of innate immunity as described in International Patent Application No. PCT / US20 / 31169, filed May 1, 2020 (published as WO2020 / 223698A1), the entire contents of which are incorporated herein by reference for purposes described herein.

[0129] In some embodiments, the second polymer component included in the provided polymeric biomaterial(s) preparation can be or comprises a carbohydrate polymer, e.g., a polymer that is or comprises a carbohydrate backbone, including but not limited to carbohydrates, e.g., hyaluronic acid, chitosan, and / or variants thereof.

[0130] Exemplary Hyaluronic Acid and Its Variants In some embodiments, the polymer included in the composition comprising the polymeric biomaterial(s) preparation described herein may be or may include an anionic polymer. In some embodiments, the anionic polymer may be or may include hyaluronic acid or a variant thereof. Hyaluronic acid (HA), also known as hyaluronan or hyaluronate, is a non-sulfated member of a class of polymers known as glycosaminoglycans (GAGs) that are widely distributed in body tissues. HA is found as an extracellular matrix component of tissues forming a pericellular coat on the cell surface. In some embodiments, HA is a cytosolic acid (CAA)-derived hyaluronan ... 14 H 21 NO 11 ) n (which in some embodiments may be present as a salt, e.g., sodium, potassium, and / or calcium salt), where n may vary depending on the source, isolation procedure, and / or determination method.

[0131] In some embodiments, HA that may be useful according to the present disclosure may be isolated or derived from a number of natural sources. For example, in some embodiments, HA may be isolated or derived from, including, for example, human umbilical cord, rooster comb, and / or vertebrate connective matrix. In some embodiments, HA may be isolated or derived from the capsule components of bacteria such as Streptococci. See, for example, Kendall et al, (1937), Biochem. Biophys. Acta, 279, 401-405. In some embodiments, HA and / or variants thereof may be produced via microbial fermentation. In some embodiments, HA and / or variants thereof may be recombinant HA or variants thereof, produced using gram-positive and / or gram-negative bacteria as hosts, including, but not limited to, Bacillus sp., Lactococcos lactis, Agrobacterium sp., and / or Escherichia coli.

[0132] As discussed in International Patent Application No. PCT / US20 / 31169, filed May 1, 2020, the biological activity of HA varies depending on its molecular weight; for example, high molecular weight HA (high MWHA) can have anti-inflammatory or immunosuppressive activity, and low molecular weight HA (low MWHA) can exhibit pro-inflammatory or immunostimulatory behavior. For example, Gao et al. “A low molecular weight hyaluronic acid derivative accelerates excisional wound healing by modulating pro-inflammation, promoting epithelialization and neovascularization, and remodeling collagen” IntJ. Mol Sci (2019) 20:3722, Cyphert et al. “Size Matters: Molecular Weight Specificity of Hyaluronan Effects in Cell Biology.” Int. J. Cell Biol.(2015)2015:563818, Dicker et al. “Hyaluronan:A simple polysaccharide with diverse biological functions”Acta Biomater.(2014)10:1558-1570, Aya and Stern“Hyaluronan in wound healing:Rediscovering a major player.”Wound Repair Regen.(2014)22:579-593 and Frenkel “The role of hyaluronan in wound See, “Methods of healing” Int. Wound J. (2014) 11:159-163, the entire contents of each of which are incorporated by reference in their entirety for purposes described herein.Thus, in some embodiments, HA or variants thereof that may be included in compositions comprising the polymeric biomaterial(s) preparations described herein may have a low molecular weight, e.g., an average molecular weight of 500 kDa or less, including 450 kDa, 400 kDa, 350 kDa, 300 kDa, 250 kDa, 200 kDa, 150 kDa, 100 kDa, 50 kDa or less. In some embodiments, HA or variants thereof that may be included in compositions comprising the polymeric biomaterial(s) preparations described herein may have an average molecular weight of about 100 kDa to about 150 kDa. In some embodiments, HA or variants thereof that may be included in compositions comprising the polymeric biomaterial(s) preparations described herein may have an average molecular weight of about 300 kDa to about 400 kDa. In some embodiments, compositions comprising the polymeric biomaterial(s) preparations described herein may include poloxamers (e.g., as described herein) and low molecular weight HA or variants thereof in the absence of an immunomodulatory payload, which may be useful for inducing innate immune agonism.

[0133] In some embodiments, HA or variants thereof that may be included in compositions comprising the polymeric biomaterial(s) preparations described herein may have a high molecular weight, e.g., an average molecular weight of greater than 500 kDa or more, including 550 kDa, 600 kDa, 650 kDa, 700 kDa, 750 kDa, 800 kDa, 850 kDa, 900 kDa, 950 kDa, 1 Mda, 1.1 Mda, 1.2 Mda, 1.3 Mda, 1.4 Mda, 1.5 Mda, 1.6 Mda, 1.7 Mda, 1.8 Mda, 1.9 Mda, 2 Mda, 2.5 Mda, 3 Mda, 3.5 Mda, 4 Mda, 4.5 MDa or more. In some embodiments, HA or variants thereof that may be useful according to the present disclosure may have an average molecular weight of about 600 kDa to about 900 kDa. In some embodiments, HA or variants thereof that may be useful according to the present disclosure may have an average molecular weight of about 1 MDa to about 3 MDa. In some embodiments, compositions comprising the polymeric biomaterial(s) preparations described herein may include poloxamers (e.g., those described in U.S. Provisional Patent Application No. 63 / 053488, filed July 17, 2020, which is incorporated by reference in its entirety for all purposes) and high molecular weight HA or variants thereof, in the absence of an immunomodulatory payload, may be useful in resolving inflammation (e.g., immunosuppressive inflammation).

[0134] In some embodiments, the compositions comprising the polymeric biomaterial(s) preparations described herein comprise hyaluronic acid variants. In some embodiments, the hyaluronic acid variants are water-soluble. In some embodiments, the hyaluronic acid variants can be chemically modified hyaluronic acid, for example, in some embodiments, the hyaluronic acid is esterified. Examples of chemical modifications to hyaluronic acid include, but are not limited to, the addition of thiol, haloacetate, butanediol, diglycidyl, ether, dihydrazide, aldehyde, glycan, and / or tyramine functional groups. Additional hyaluronic acid modifications and variants are known in the art. See, e.g., Highley et al., "Recent advances in hyaluronic acid hydrogels for biomedical applications" Curr Opin Biotechnol (2016) Aug 40:35-40; Burdick & Prestwich, "Hyaluronic acid hydrogels for biomedical applications" Advanced Materials (2011); Prestwhich, "Hyaluronic acid-based clinical biomaterials derived for cell and molecule delivery in regenerative medicine" J. Control Release (2011) Oct 30;155(2):193-199, each of which is incorporated by reference in their entirety for purposes described herein.

[0135] In some embodiments, compositions comprising the polymeric biomaterial(s) preparations described herein that include HA also include at least one poloxamer. As discussed in U.S. Provisional Patent Application No. 63 / 053488, filed July 17, 2020, which is incorporated by reference in its entirety for all purposes, the combination of HA and poloxamer can be particularly effective for the purposes described herein.

[0136] In certain embodiments, hyaluronic acid can be crosslinked by conjugating thiols (EXTRACEL®, HYSTEM®), methacrylates, hexadecylamides (HYMOVIS®), and tyramines (CORGEL®). Hyaluronic acid can also be directly crosslinked with formaldehyde (HYLAN-A®) or divinyl sulfone (HYLAN-B®).

[0137] In certain embodiments, the hyaluronic acid comprises a thiol-modified hyaluronic acid and a crosslinker. In certain embodiments, the polymer network or hydrogel comprises a thiol-modified hyaluronic acid (e.g., GLYCOSIL®) and a thiol-reactive PEGDA crosslinker (e.g., EXTRALINK®). In certain embodiments, the thiol-modified hyaluronic acid and the thiol-reactive PEGDA crosslinker are combined to form a crosslinked polymer network or hydrogel useful in the compositions described herein.

[0138] In certain embodiments, the amounts and concentrations of thiol-modified hyaluronic acid, thiol-reactive hyaluronic acid, and crosslinker can be adjusted to provide a composition with desired physical properties, such as having a storage modulus of about 500 Pa to about 3000 Pa.

[0139] Exemplary Alginates and Variants Thereof In certain embodiments, the composition comprising at least one polymeric biomaterial described herein comprises alginate. Alginate in its non-salt form is known as alginic acid and / or algin. Alginate is a polysaccharide widely distributed in the cell walls of brown algae, which is hydrophilic and forms a viscous gum when hydrated. Metals such as sodium and calcium are utilized to create alginate. Alginic acid has a linear structure of a heteropolysaccharide. Generally, alginic acid is composed of d-mannuronic acid and l-guluronic acid. Alginic acid is present in many markets as a salt, sodium alginate. Alginates have the ability to form gels in the presence of dipolar ions (such as calcium ions) due to the carboxyl groups on their polymer chains. Due to the porous structure and high water absorption capacity of calcium alginate gel, its use in wound dressing and / or cell immobilization has been previously proposed.

[0140] In certain embodiments, the composition comprising at least one polymeric biomaterial is comprised of the polymeric biomaterial alginate. In certain embodiments, the polymeric biomaterial comprises cross-linked alginate. In certain embodiments, the polymeric biomaterial is an alginate. In certain embodiments, the polymeric biomaterial is cross-linked alginate. In certain embodiments, the polymeric biomaterial may form a polymeric network or hydrogel, the polymeric biomaterial comprises alginate. In certain embodiments, the polymeric network or hydrogel comprises cross-linked alginate. In certain embodiments, the polymeric network or hydrogel is an alginate. In certain embodiments, the polymeric network or hydrogel is cross-linked alginate. In certain embodiments, the polymeric biomaterial does not comprise alginate. In certain embodiments, the polymeric biomaterial is not an alginate. In certain embodiments, the polymeric network or hydrogel is not an alginate. In certain embodiments, the polymeric network or hydrogel does not comprise alginate.

[0141] In certain embodiments, the alginate can be ionically crosslinked by adding a salt (eg, calcium chloride) that promotes crosslinking.

[0142] In certain embodiments, the alginate comprises an alginate and a cross-linking agent (e.g., calcium chloride). In certain embodiments, the polymer network or hydrogel comprises an alginate and a cross-linking agent (e.g., calcium chloride). In certain embodiments, the alginate and calcium chloride (e.g., an ionic cross-linking agent) are combined to form a cross-linked polymer network or hydrogel useful in the compositions described herein.

[0143] In certain embodiments, the amounts and concentrations of alginate and calcium chloride may be adjusted to provide a composition with desired physical properties, such as having a storage modulus of about 500 Pa to about 3000 Pa.

[0144] Exemplary Chitosan and Its Variants In some embodiments, the compositions comprising the polymeric biomaterial(s) preparations described herein may be or may include a cationic polymer, hi some embodiments, the cationic polymer may be or may include chitosan or a variant thereof. Examples of chitosan and / or variants thereof that may be included in the polymer combination preparations described herein include, but are not limited to, chitosan, chitosan salts (e.g., chitosan HCl, chitosan chloride, chitosan lactate, chitosan acetate, chitosan glutamate), alkyl chitosan, aromatic chitosan, carboxyalkyl chitosan (e.g., carboxymethyl chitosan), hydroxyalkyl chitosan (e.g., hydroxypropyl chitosan, hydroxyethyl chitosan), aminoalkyl chitosan, acylated chitosan, phosphorylated chitosan, thiolated chitosan, quaternary ammonium chitosan (e.g., N-(2-hydroxyl)propyl-3-trimethylammonium chitosan chloride), guanidinyl chitosan, chitosan oligosaccharides, glycated chitosan (e.g., N-dihydrogalactosyl), and variants or combinations thereof. In some embodiments, compositions comprising the polymeric biomaterial(s) preparations described herein include a carboalkylchitosan (e.g., carboxymethylchitosan).

[0145] One of skill in the art will appreciate that in some cases, chitosan and / or variants thereof may be produced by deacetylation of chitin. In some embodiments, chitosan or variants thereof included in a poloxamer-containing polymer combination preparation (e.g., as described herein) are characterized by a degree of deacetylation (i.e., percent of acetyl groups removed) of at least 70% or more, including, for example, at least 75%, at least 80%, at least 85%, at least 90%, at least 95% or more (including up to 100%). In some embodiments, chitosan or variants thereof are characterized by a degree of deacetylation of 99% or less, 95% or less, 90% or less, 85% or less, 80% or less, 75% or less, or less. Combinations of the above ranges are also possible. For example, chitosan or variants thereof may be characterized by a degree of deacetylation of 80%-95%, 70%-95%, or 75%-90%. As will be appreciated by one of skill in the art, the degree of deacetylation (DA%) can be determined by a variety of methods known in the art, such as, in some cases, by NMR spectroscopy.

[0146] In some embodiments, the chitosan or variant thereof included in the composition comprising the polymeric biomaterial(s) preparation described herein has a molecular weight of, for example, at least 20 kDa, at least 30 kDa, at least 40 kDa, at least 50 kDa, at least 60 kDa, at least 70 kDa, at least 80 kDa, at least 90 kDa, at least 100 kDa, at least 110 kDa, at least 120 kDa, at least 130 kDa, at least 140 kDa, at least 150 kDa, at least 160 kDa, at least 170 kDa, at least 180 kDa, at least 200 kDa, at least 250 kDa, at least 260 kDa, at least 270 kDa, at least 280 kDa, at least 300 kDa, at least 350 kDa, at least 360 kDa, at least 370 kDa, at least 380 kDa, at least 390 kDa, at least 400 kDa, at least 400 kDa, at least 45 ... Da, at least 190 kDa, at least 200 kDa, at least 210 kDa, at least 220 kDa, at least 230 kDa, at least 240 kDa, at least 250 kDa, at least 260 kDa, at least 270 kDa, at least 280 kDa, at least 290 kDa, at least 300 kDa, at least 350 kDa, at least 400 kDa, at least 500 kDa, at least 600 kDa, at least 700 kDa, or more, e.g., at least 10 kDa or more. In some embodiments, the chitosan or variant thereof included in the composition comprising the polymeric biomaterial(s) preparation described herein may have an average molecular weight of 750 kDa or less, including, for example, 700 kDa or less, 600 kDa or less, 500 kDa or less, 400 kDa or less, 300 kDa or less, 200 kDa or less, 100 kDa or less, 50 kDa or less, or less. Combinations of the above ranges are also possible. For example, in some embodiments, the chitosan or variant thereof included in the composition comprising the polymeric biomaterial(s) preparation described herein is characterized by an average molecular weight of 10 kDa to 700 kDa, or 20 kDa to 700 kDa, or 30 kDa to 500 kDa, or 150 kDa to 600 kDa, or 150 kDa to 400 kDa, or 50 kDa to 150 kDa, or 10 kDa to 50 kDa.In some embodiments, the chitosan or variant thereof included in the composition comprising the polymeric biomaterial(s) preparation described herein is characterized by an average molecular weight of 20 kDa to 700 kDa, or 30 kDa to 500 kDa. As described herein, the average molecular weight can be a number average molecular weight, a weight average molecular weight, or a peak average molecular weight.

[0147] In some embodiments, the chitosan or variant thereof included in the composition comprising the polymeric biomaterial(s) preparation described herein is characterized by a molecular weight distribution in the range of 10 kDa to 700 kDa, or 20 kDa or 700 kDa, or 30 kDa to 500 kDa, or 150 kDa to 600 kDa, or 150 kDa to 400 kDa, or 50 kDa to 150 kDa, or 10 kDa to 50 kDa. In some embodiments, the chitosan or variant thereof included in the composition comprising the polymeric biomaterial(s) preparation described herein is characterized by a molecular weight distribution in the range of 20 kDa to 700 kDa, or 30 kDa to 500 kDa.

[0148] In some embodiments, the chitosan or variant thereof included in the composition comprising the polymeric biomaterial(s) preparation described herein may be characterized by a viscosity of 3500 mPa·s or less, including, for example, 3000 mPa·s or less, 2500 mPa·s or less, 2000 mPa·s or less, 1500 mPa·s or less, 1000 mPa·s or less, 500 mPa·s or less, 250 mPa·s or less, 200 mPa·s or less, 150 mPa·s or less, 100 mPa·s or less, 75 mPa·s or less, 50 mPa·s or less, 25 mPa·s or less, 20 mPa·s or less, 15 mPa·s or less, 10 mPa·s or less, or less. In some embodiments, the chitosan or variant thereof may be characterized by a viscosity of at least 5 mPa·s or greater, including, for example, at least 10 mPa·s, at least 20 mPa·s, at least 30 mPa·s, at least 40 mPa·s, at least 50 mPa·s, at least 60 mPa·s, at least 70 mPa·s, at least 80 mPa·s, at least 90 mPa·s, at least 100 mPa·s, at least 125 mPa·s, at least 150 mPa·s, at least 175 mPa·s, at least 250 mPa·s, at least 500 mPa·s, at least 1000 mPa·s, at least 1500 mPa·s, at least 2000 mPa·s, at least 2500 mPa·s or greater. Combinations of the above ranges are also possible. For example, in some embodiments, such viscous polymer solutions of or including chitosan or variants thereof may be characterized by a viscosity of 5 mPa·s to 3000 mPa·s, or 5 mPa·s to 300 mPa·s, or 5 mPa·s to 200 mPa·s, or 20 mPa·s to 200 mPa·s, or 5 mPa·s to 20 mPa·s. In some embodiments, the viscosity of chitosan or variants thereof described herein is measured at 1% in 1% acetic acid at 20° C.

[0149] In some embodiments, compositions comprising the polymeric biomaterial(s) preparations described herein comprise at least one or more (e.g., 1, 2, 3 or more) chitosan and / or variants thereof (e.g., including modified chitosan and / or a salt of chitosan or modified chitosan, such as a chloride salt or a glutamate salt). For example, in some embodiments, the chitosan and / or variants thereof (e.g., including modified chitosan and / or a salt of chitosan or modified chitosan, such as a chloride salt or a glutamate salt) may be characterized by a degree of deacetylation of 70%-95%, or 75%-90%, or 80%-95%, or greater than 90%. In some embodiments, chitosan and / or variants thereof (including, e.g., modified chitosan and / or a salt of chitosan or modified chitosan, such as the chloride salt or glutamate salt) may be characterized by an average molecular weight (e.g., measured as chitosan or a chitosan salt, e.g., chitosan acetate) of 10 kDa to 700 kDa, 20 kDa to 600 kDa, 30 kDa to 500 kDa, 150 kDa to 400 kDa, or 200 kDa to 600 kDa. In some embodiments, chitosan and / or variants thereof (including, e.g., modified chitosan and / or a salt of chitosan or modified chitosan such as the chloride salt or glutamate salt) may be characterized by a molecular weight distribution (e.g., measured as chitosan or a chitosan salt, e.g., chitosan acetate) in the range of 10 kDa to 700 kDa, 20 kDa to 600 kDa, 30 kDa to 500 kDa, 150 kDa to 400 kDa, or 200 kDa to 600 kDa. In some embodiments, chitosan and / or variants thereof (including, e.g., a salt thereof, such as the chloride salt or glutamate salt) may be characterized by a viscosity in the range of 5 to 3000 mPa·s, or 5 to 300 mPa·s, or 20 to 200 mPa·s.In some embodiments, such chitosan and / or variants thereof (e.g., including salts thereof such as chloride or glutamate) may be or include PROTASAN™ UltraPure chitosan chloride and / or chitosan glutamate (e.g., available from NovoMatrix®, a business unit of FMC Health and Nutrition (now part of Du Pont, product numbers: CL113, CL114, CL213, CL214, G113, G213, G214)). In some embodiments, such chitosan and / or variants thereof (including, for example, salts thereof such as the chloride salt or glutamate salt) may be or include chitosan, chitosan oligomers, and / or variants thereof (including, for example, chitosan HCl, carboxymethyl chitosan, chitosan lactate, chitosan acetate), obtained, for example, from Heppe Medical Chitosan GMBH (e.g., Chitoceuticals® or Chitoscience®).

[0150] In some embodiments, the chitosan or variant thereof included in the composition comprising the polymeric biomaterial(s) preparation described herein is or includes a carboxyalkylchitosan (e.g., carboxymethylchitosan) characterized by at least one or all of the following characteristics: (1) a degree of deacetylation of 80%-95%, (ii) an average molecular weight of 30 kDa-500 kDa, or a molecular weight distribution of 30 kDa-500 kDa, and (iii) a viscosity in the range of 5-300 mPa·s.

[0151] In some embodiments, the chitosan or variant thereof included in the composition comprising the polymeric biomaterial(s) preparation described herein is or comprises a variant of chitosan (e.g., as described herein). In some embodiments, such a variant of chitosan may comprise chemical modification(s) of one or more chemical moieties of the chitosan chain, e.g., hydroxyl and / or amino groups. In some embodiments, such a variant of chitosan is or comprises a modified chitosan (e.g., a chitosan modified by the addition of one or more mono- or oligosaccharide side chains to one or more of its free amino groups), such as, but not limited to, a glycated chitosan. Exemplary saccharified chitosans that are useful herein include, but are not limited to, those described in, for example, US5,747,475, US6,756,363, WO2013 / 109732, US2018 / 0312611, and US2019 / 0002594, the contents of each of which are incorporated herein by reference for purposes described herein.

[0152] In some embodiments, the chitosan or variant thereof included in the composition comprising the polymeric biomaterial(s) preparation described herein is or includes chitosan conjugated to a polymer (e.g., a hydrophilic polymer such as polyethylene glycol) that increases its solubility in an aqueous environment.

[0153] In some embodiments, the chitosan or variant thereof included in the composition comprising the polymeric biomaterial(s) preparation described herein is or includes a thiolated chitosan. Various modifications to chitosan, including but not limited to carboxylation, PEGylation, galactosylation (or other glycosylation), and / or thiolation, are known in the art, for example, as described in Ahmadi et al. Res Pharm Sci., 10(1):1-16 (2015), the contents of which are incorporated herein by reference for purposes described herein. Those skilled in the art reading this disclosure will understand that other modified chitosans may be useful for the particular application in which the method is performed.

[0154] In some embodiments, compositions comprising the polymeric biomaterial(s) preparation described herein that includes chitosan or a variant thereof also include at least one poloxamer. As discussed in U.S. Provisional Patent Application No. 63 / 053488, filed July 17, 2020, which is incorporated by reference in its entirety for all purposes, a combination of chitosan and poloxamer may be particularly effective for the purposes described herein.

[0155] Polynucleotide agent(s) encoding or modulating an immunomodulatory polypeptide The compositions provided include at least one nucleic acid or polynucleotide agent. In certain embodiments, the nucleic acid(s) provided may be deoxyribonucleic acid(s) and / or ribonucleic acid(s). In certain embodiments, the nucleic acid(s) provided may be chemically modified in a manner that may alter stability, function, or other characteristics. In certain embodiments, the compositions include at least one nucleic acid or polynucleotide agent that acts to directly regulate and / or modify the immune system and / or encodes a molecule that regulates the immune system or regulates an immunomodulatory peptide. In some embodiments, the nucleic acid encodes or regulates an immunomodulatory peptide, including a cytokine, a chemokine, an antibody, an innate immune system regulator, an adaptive immune system regulator, a proinflammatory peptide, an anti-inflammatory peptide, a growth factor, a hormone, and / or a domain, combination, or hybrid construct of any of these.

[0156] In certain embodiments, the compositions provided include at least one polynucleotide agent that modulates or encodes a molecule that induces innate immunity by activating a pattern recognition receptor. In certain embodiments, the activator of the innate immune response is a ligand of a pattern recognition receptor (PRR). In certain embodiments, the activator of the innate immune response is an agonist of a pattern recognition receptor (PRR). In certain embodiments, the activator of the innate immune response can be recognized by a Toll-like receptor (TLR), a nucleotide-binding oligomerization domain-like receptor (NLR), a C-type lectin receptor (CLR), and / or a RIG-1-like receptor (RLR).

[0157] In certain embodiments, the compositions provided may comprise at least one nucleic acid comprising a sequence encoding a cytokine. Alternatively, in some embodiments, the compositions comprising at least one nucleic acid may further comprise a cytokine.

[0158] Cytokines are a broad category of small proteins (approximately 5-20 kDa) that are important in cell signaling. Their release affects the behavior of surrounding cells. Cytokines can induce innate and / or adaptive immunity. Cytokines participate in autocrine, paracrine, and endocrine signaling as immunomodulatory agents. Cytokines include chemokines, interferons, interleukins, lymphokines, and tumor necrosis factors. Cytokines are produced by a wide range of cells, including immune cells such as macrophages, B lymphocytes, T lymphocytes, and mast cells, as well as endothelial cells, fibroblasts, and various stromal cells. They act through receptors and play a key role in the immune system. Cytokines regulate the balance between humoral and cell-based immune responses and modulate the maturation, proliferation, and responsiveness of specific cell populations. Some cytokines enhance or inhibit the action of other cytokines in a complex manner. Cytokines are important in host responses to infection, immune responses, inflammation, trauma, sepsis, cancer, and reproduction.

[0159] Furthermore, it is currently known in the art that delivery, dosing, and scheduling methods, as well as toxicity-related issues, need to be addressed to enable the immune stimulatory functions of many cytokines and chemokines to be fully utilized.

[0160] In certain embodiments, the compositions provided include at least one nucleic acid encoding a cytokine and / or at least one cytokine, including cytokines such as IL-1, IL-1α, IL-1β, IL-2, IL-2 superkine, IL-6, IL-7, IL-9, AM0010, IL-12, IL-15, IL-15 superagonist, ALT-803, NIZ985, IL-16, IL-18, IL-19, IL-20, IL-21, IL-22, IL-23, IL-24, IL-25, IL-26, IL-27, IL-28, IL-29, IL-30, IL-31, IL-32, IL-33, IL-34, IL-35, IL-36, IL-37, IL-38, IL-39, IL-40, IL-41, IL-42, IL-43, IL-44, IL-45, IL-46, IL-47, IL-48, IL-49 ...9, IL-49, IL-49, IL-49, IL-49, IL-49, , IL-21, an IL-21 superagonist, denenicoquine, an IL-21 superagonist antibody, IFN-α, IFN-β, IFN-γ, TNF-α, GM-CSF, a cytokine fusion, RG7461, RG7813, M9241, NKTR-214, NKTR-255, BMS-982470, BG-00001, Flt3L, or CDX-301, and / or the nucleotide sequences encoding them.

[0161] In certain embodiments, the compositions provided comprise at least one nucleic acid encoding a cytokine and / or at least one cytokine, including ALT-803, NIZ985, denenicoquine, RG7461, RG7813, M9241, IFN-α, IFN-β, or IFN-γ, and / or the nucleotide sequences encoding them.

[0162] In certain embodiments, the compositions provided comprise at least one nucleic acid encoding a cytokine, and / or at least one cytokine, including an IL-15 superagonist and / or IL-21, and / or the nucleotide sequences encoding them.

[0163] In certain embodiments, compositions provided include at least one nucleic acid encoding a cytokine and / or at least one cytokine, including an IL-15 superagonist, IL-21, IFN-α, IFN-β, IFN-γ, CCL4, CCL5, CXCL9, or CXCL10, and / or nucleotide sequences encoding same, in certain embodiments, the encoded sequence and / or cytokine is an IL-15 superagonist, IFN-α, IFN-β, or IFN-γ.

[0164] IL-15 (Interleukin 15) is a cytokine structurally similar to IL-2 and is secreted by mononuclear phagocytes following infection with a virus(es). IL-15 induces cell proliferation of natural killer cells, cells whose primary role is to kill virus-infected cells. The combination of IL-15 with soluble IL-15Rα produces a complex called IL-15 superagonist (IL-15sa), which has greater biological activity than IL-15 alone. IL-15sa is an antitumor and antiviral agent due to its ability to selectively proliferate NK and memory CD8+T (mCD8+T) lymphocytes.

[0165] In certain embodiments, the compositions provided include at least one nucleic acid encoding a cytokine and / or at least one cytokine, including an IL-15 superagonist known as ALT-803, and / or a nucleotide sequence encoding the same. ALT-803 is believed to induce memory CD8+ T cell proliferation, upregulate receptors involved in innate immunity, secrete interferon gamma, and acquire the ability to kill malignant cells in the absence of antigenic stimulation. Thus, ALT-803 may promote the proliferation and activation of memory CD8+ T cells while converting them into innate immune effector cells that exhibit robust anti-malignant tumor activity. ALT-803 is a fusion protein of an IL-15 variant and an IL-15Rα / Fc complex that has recently entered clinical trials as a direct immunomodulator. ALT-803 exhibits a greater than 25-fold enhancement in biological activity compared to IL-15.

[0166] In certain embodiments, the compositions provided include at least one nucleic acid encoding a cytokine and / or at least one cytokine, the cytokine included being NIZ985 (hetIL-15) and / or the nucleotide sequence encoding same. Studies have shown that hetIL-15 administration can promote increased tumor infiltration and persistence of CD8+ T cells, including tumor-specific T cells, resulting in an increased CD8+ / Treg ratio. Tumor-resident CD8+ T cells exhibit characteristics of effector cells, characterized by increased proliferation (Ki67+) and high cytotoxicity (Granzyme B+). In the absence of hetIL-15, a smaller population of tumor-infiltrating T cells exhibits high levels of the exhaustion marker PD-1, potentially limiting their anti-cancer efficacy. Provision of hetIL-15 can result in a significant decrease in lymphocyte expression of PD-1, mitigating one potential mechanism of the exhaustion phenotype. Preclinical cancer studies support the use of hetIL-15 in tumor immunotherapy approaches to promote the development of antitumor responses by prioritizing effector over regulatory cells.

[0167] In certain embodiments, the compositions provided include at least one nucleic acid encoding a cytokine and / or at least one cytokine, including interferon alpha (IFN-α) and / or a nucleotide sequence encoding same. IFN-α proteins are produced by white blood cells. They are primarily involved in the innate immune response to viral infections.

[0168] In certain embodiments, the compositions provided include at least one nucleic acid encoding a cytokine and / or at least one cytokine, including interferon beta (IFN-β), and / or a nucleotide sequence encoding the same. IFN-β includes a protein produced by fibroblasts and is involved in the innate immune response. IFN-β stimulates both macrophages and NK cells to induce antiviral responses and is also active against tumors. In mice, IFN-β inhibits immune cells to produce growth factors, thereby slowing tumor growth, and inhibits other cells from producing angiogenic growth factors, thereby blocking tumor angiogenesis and preventing tumors from connecting to the vasculature.

[0169] In certain embodiments, the compositions provided include at least one nucleic acid encoding a cytokine and / or at least one cytokine, the cytokine included being interferon gamma (IFN-γ) and / or the nucleotide sequence encoding same. IFN-γ, or type II interferon, is a cytokine useful in innate and adaptive immunity. IFN-γ is a key activator of macrophages and an inducer of class II major histocompatibility complex (MHC) molecule expression. In vitro studies of IFN-γ in cancer cells are extensive, and results indicate the anti-proliferative activity of IFN-γ, which results in growth inhibition or cell death, generally induced by apoptosis, but sometimes by autophagy. Clinical administration of IFN-γ has led to improved survival of patients with ovarian, bladder, and melanoma cancers.

[0170] In certain embodiments, the compositions provided include at least one nucleic acid encoding a chemokine and / or at least one chemokine. Chemokines are a family of small cytokines. The primary role of chemokines is to act as chemoattractants that induce cell migration. Some chemokines control cells of the immune system during the process of immune surveillance, such as directing lymphocytes to lymph nodes, where they may screen for invading pathogens by interacting with antigen-presenting cells present in these tissues. These are known as homeostatic chemokines, and are produced and secreted without the need to stimulate their source cell(s). Some chemokines play a role in development, directing cells to tissues where they promote angiogenesis (the growth of new blood vessels) or provide specific signals important for the maturation of the cells. Other chemokines are inflammatory and are released from a wide variety of cells in response to bacteria, viruses, and agents that cause physical damage, such as silica or uric acid crystals that occur in gout. Their release is often stimulated by proinflammatory cytokines, such as interleukin-1. Inflammatory chemokines function primarily as chemoattractants for leukocytes, recruiting monocytes, neutrophils, and other effector cells from the blood to sites of infection or tissue injury. Certain inflammatory chemokines activate cells to initiate an immune response or promote wound healing. They are released by many different cell types and serve to guide cells of both the innate and adaptive immune systems.

[0171] Moreover, it is currently known in the art that delivery, dosing, and scheduling methods, as well as toxicity-related issues, need to be addressed to allow the immune stimulatory functions of many chemokines to be fully utilized. Thus, in some embodiments, provided compositions may overcome many of the known barriers to utilizing chemokines for immunotherapy.

[0172] In certain embodiments, compositions provided include at least one nucleic acid encoding a chemokine and / or at least one chemokine, including CCL1, CCL2, CCL3, CCL4, CCL5, CCL17, CCL19, CCL21, CCL22, CXCL9, CXCL10, CXCL11, CXCL13, CXCL16, or CX3CL1, and / or the nucleotide sequences encoding them.

[0173] In certain embodiments, the provided compositions comprising at least one nucleic acid may include a nucleic acid encoding a regulator (e.g., an inhibitor) of a proinflammatory pathway. In some embodiments, the nucleic acid-encoded inhibitor of a proinflammatory pathway comprises an antibody. In some embodiments, the provided compositions may include two or more inhibitors of a proinflammatory pathway. In some embodiments, the inhibitor of a proinflammatory pathway may prevent recruitment of immunosuppressive cells. In some embodiments, the inhibitor of a proinflammatory pathway may prevent acute inflammation. In some embodiments, the inhibitor of a proinflammatory pathway may inhibit an immune response that induces inflammation, including, for example, production of one or more proinflammatory cytokines (e.g., TNF-alpha, IL-1 beta, and / or IL-6), increased activity and / or proliferation of Th1 cells, recruitment of myeloid cells, and the like. For example, in some embodiments, the inhibitor of a proinflammatory pathway may be an inhibitor of IL-1 beta. In some embodiments, the inhibitor of a proinflammatory pathway may be an inhibitor of IL-6.

[0174] In certain embodiments, the provided compositions may act as or contain at least one nucleic acid encoding an inhibitor of a proinflammatory pathway. In some embodiments, the inhibitor of a proinflammatory pathway is an inhibitor of the proinflammatory immune response mediated by the p38 mitogen-activated protein kinase (MAPK) pathway as described herein.

[0175] In certain embodiments, the provided composition may act as an inhibitor of proinflammatory pathway that prevents the recruitment of immunosuppressive cells or may contain at least one nucleic acid that codes for the inhibitor of the proinflammatory pathway.In certain embodiments, the inhibitor of the proinflammatory pathway is an inhibitor, antagonist, or partial agonist of CCR2, CCR5, CXCR2, CXCR4, CXCL12, or CCL2.In certain embodiments, the inhibitor of the proinflammatory pathway is an inhibitor, antagonist, or partial agonist of CCR5, CXCR2, CXCL12, or CCL2.

[0176] In certain embodiments, the provided compositions may include at least one nucleic acid that acts as an inhibitor of a proinflammatory pathway or encodes an inhibitor of a proinflammatory pathway, and an optional further inhibitor of the proinflammatory pathway that is an inhibitor, antagonist, or partial agonist of CCR2. In certain embodiments, CCR2 is associated with the p38 MAPK pathway (e.g., as described in Montague, et al., J. Inflammation 2018, 15:101, and Xu, et al., Am. J. Transl. Res. 2017, 9, 2878-2890, each of which is incorporated herein by reference for the purposes described herein). In certain embodiments, the optional inhibitor, antagonist, or partial agonist of CCR2 is PF-04136309, CCX872-B, or prosalizumab. In certain embodiments, the inhibitor of proinflammatory pathway is PF-04136309, CCX872-B or prosalizumab.In certain embodiments, the inhibitor of proinflammatory pathway is not an inhibitor, antagonist or partial agonist of CCR2.In certain embodiments, the inhibitor of proinflammatory pathway is not PF-04136309.

[0177] In certain embodiments, the provided compositions may comprise at least one nucleic acid that acts as an inhibitor of a proinflammatory pathway or encodes an inhibitor of a proinflammatory pathway, and the inhibitor of a proinflammatory pathway is an inhibitor, antagonist, or partial agonist of CCR5. In certain embodiments, CCR5 is associated with the p38 MAPK pathway (e.g., as described in Lei, et al., Biochem. Biophys. Res. Commun. 2005, 329, 610-615, and Manes, et al., J. Exp. Med. 2003, 198, 1381-1389, each of which is incorporated herein by reference for the purposes described herein). In certain embodiments, the nucleic acid encoded inhibitor, antagonist, or partial agonist of CCR5 is optionally coupled to maraviroc, DAPTA, GSK706769, INCB009471, GW873140, vicriviroc, or PRO140.

[0178] In certain embodiments, the provided compositions may include at least one nucleic acid that acts as or encodes an inhibitor of a proinflammatory pathway, the inhibitor of a proinflammatory pathway being an inhibitor, antagonist, or partial agonist of CCR2 and / or CCR5. In certain embodiments, the nucleic acid encoded inhibitor, antagonist, or partial agonist of CCR2 and CCR5 is optionally coupled with PF-04634817, cenicriviroc, or BMS-813160.

[0179] In certain embodiments, the provided composition may comprise at least one nucleic acid that acts as an inhibitor of proinflammatory pathway or encodes an inhibitor of proinflammatory pathway, and the inhibitor of proinflammatory pathway is an inhibitor, antagonist, or partial agonist of CXCR2.In certain embodiments, the inhibitor, antagonist, or partial agonist encoded by the nucleic acid of CXCR2 is optionally coupled with danilixin, QBM076, SX-682, or SB225002.

[0180] In certain embodiments, the provided compositions may comprise at least one nucleic acid that acts as an inhibitor of a proinflammatory pathway or encodes an inhibitor of a proinflammatory pathway, and the inhibitor of a proinflammatory pathway is an inhibitor, antagonist, or partial agonist of CXCR4. In certain embodiments, CXCR4 is associated with the p38 MAPK pathway (e.g., as described in Lei, et al., Biochem. Biophys. Res. Commun. 2005, 329, 610-615, and Trushin, et al., J. Immunol. 2007, 178, 4846-4853, each of which is incorporated herein by reference for the purposes described herein). In certain embodiments, the nucleic acid-encoded inhibitor, antagonist, or partial agonist of CXCR4 is optionally coupled with plerixafor, AMD070, AMD3465, AMD11070, LY2510924, MSX-122, TG-0054, CX-01, X4P-001, BL-8040, USL311, or SP01A. In certain embodiments, the inhibitor of the proinflammatory pathway is not an inhibitor, antagonist, or partial agonist of CXCR4.

[0181] In certain embodiments, the provided compositions may comprise at least one nucleic acid that acts as an inhibitor of proinflammatory pathway or encodes an inhibitor of proinflammatory pathway, and the inhibitor of proinflammatory pathway is an inhibitor, antagonist, or partial agonist of CXCL12. In certain embodiments, CXCL12 is related to p38MAPK pathway (e.g., as described in Gao, et al., Int. J. Clin. Exp. Pathol. 2018, 11, 3119-3125, which is incorporated herein by reference for the purposes described herein).

[0182] In certain embodiments, the compositions provided may include at least one nucleic acid that acts as an inhibitor of a proinflammatory pathway or encodes an inhibitor of a proinflammatory pathway, the inhibitor of the proinflammatory pathway being an inhibitor, antagonist, or partial agonist of CCL2. In certain embodiments, CCL2 is associated with the p38 MAPK pathway (e.g., as described in Cho, et al., J. Neuroimmunol. 2008, 199, 94-103, and Marra, et al., Am. J. Physiol. Gastrointest. Liver Physiol. 2004, 287, G18-26, each of which is incorporated herein by reference for purposes described herein). In certain embodiments, the inhibitor, antagonist, or partial agonist encoded by the nucleic acid of CCL2 is optionally coupled with a bindarit.

[0183] In certain embodiments, the provided compositions may comprise at least one nucleic acid that acts as or encodes an inhibitor of a proinflammatory pathway, and the inhibitor of a proinflammatory pathway is optionally selected from the group consisting of PF-04136309, CCX872-B, prosalizumab, maraviroc, DAPTA, GSK706769, INCB009471, GW873140, vicrivirine, and the like. and coupled with , PRO140, PF-04634817, cenicriviroc, BMS-813160, danilixin, QBM076, SX-682, SB225002, plerixafor, AMD070, AMD3465, AMD11070, LY2510924, MSX-122, TG-0054, CX-01, X4P-001, BL-8040, USL311, or SP01A.

[0184] In certain embodiments, provided compositions may comprise at least one nucleic acid that acts as or encodes an inhibitor of a proinflammatory pathway, optionally coupled with CCX872-B, prosalizumab, maraviroc, DAPTA, GSK706769, INCB009471, GW873140, vicriviroc, PRO140, PF-04634817, cenicriviroc, BMS-813160, danilixin, QBM076, SX-682, SB225002, plerixafor, AMD070, AMD3465, AMD11070, LY2510924, MSX-122, TG-0054, CX-01, X4P-001, BL-8040, USL311, or SP01A.

[0185] In certain embodiments, the provided compositions may include at least one nucleic acid that acts as or encodes an inhibitor of a proinflammatory pathway, which prevents acute inflammation. In certain embodiments, the inhibitor of a proinflammatory pathway is an anti-IL-1α antibody, an anti-IL-1β antibody, an anti-IL-1R antibody, an IL-1 inhibitor, an anti-IL-6 antibody, an anti-IL-6R antibody, an anti-IL17 antibody, an anti-IL-17A antibody, an anti-IL-17RA antibody, an anti-IL-23 / IL-12 antibody, or an anti-IL-23 antibody.

[0186] In certain embodiments, the provided compositions may include at least one nucleic acid encoding an inhibitor of a proinflammatory pathway, and the inhibitor of the proinflammatory pathway is an anti-IL-1α antibody. In certain embodiments, the anti-IL-1α antibody is MABp1. In certain embodiments, the inhibitor of the proinflammatory pathway is MABp1.

[0187] In certain embodiments, the provided compositions may include at least one nucleic acid encoding an inhibitor of a proinflammatory pathway, and the inhibitor of the proinflammatory pathway is an anti-IL-1β antibody. In certain embodiments, IL-1β is associated with the p38 MAPK pathway (e.g., Kulawik, et al., J.Biol.Chem. 2017, 292, 6291-6302; Rovin, et al., Cytokine 1999, 11, 118-126; Laporte, et al., Am. J.Physiol.Lung Cell Mol.Physiol. 2000, 279, L932-L941; Baldassare, et al., J.Immunol. 1999, 162, 5367-5373; and Weber, et al. Sci.Signal. 2010, 3, cm1, each of which is incorporated herein by reference for purposes described herein). In certain embodiments, the anti-IL-1β antibody is canakinumab. In certain embodiments, the inhibitor of the proinflammatory pathway is canakinumab.

[0188] In certain embodiments, the provided compositions may include at least one nucleic acid encoding an inhibitor of a proinflammatory pathway, and the inhibitor of the proinflammatory pathway is an anti-IL-1R antibody. In certain embodiments, IL-1R is associated with the p38 MAPK pathway (e.g., as described in Weber, et al., Sci. Signal. 2010, 3, cm1, and Jain, et al., Nat. Commun. 2018, 9:3185, each of which is incorporated herein by reference for purposes described herein). In certain embodiments, the anti-IL-1R antibody is anakinra. In certain embodiments, the inhibitor of the proinflammatory pathway is anakinra.

[0189] In certain embodiments, the provided compositions may comprise at least one nucleic acid encoding an inhibitor of proinflammatory pathway, and the inhibitor of proinflammatory pathway is an IL-1 inhibitor.In certain embodiments, the IL-1 inhibitor is rilonacept.In certain embodiments, the inhibitor of proinflammatory pathway is rilonacept.

[0190] In certain embodiments, the provided compositions may include at least one nucleic acid encoding an inhibitor of a proinflammatory pathway, and the inhibitor of the proinflammatory pathway is an anti-IL-6 antibody. In certain embodiments, IL-6 is associated with the p38 MAPK pathway (see, for example, Sinfield, et al., Biochem. Biophys. Res. Commun. 2013, 430, 419-424; Suzuki, et al., FEBS Lett. 2000, 465, 23-27; and Nishikai-Yan Shen, et al., PLoS One 2017, 12, 1-17, each of which is incorporated herein by reference for purposes described herein). In certain embodiments, the anti-IL-6 antibody is olokizumab, clazakizumab, OPR-003, sirukumab, ARGX-109, FE301, or FM101. In certain embodiments, the inhibitor of the proinflammatory pathway is olokizumab, clazakizumab, OPR-003, sirukumab, ARGX-109, FE301, or FM101.

[0191] In certain embodiments, the provided composition may comprise at least one nucleic acid encoding an inhibitor of proinflammatory pathway, and the inhibitor of proinflammatory pathway is an anti-IL-6R antibody.In certain embodiments, the anti-IL-6R antibody is tocilizumab, sarilumab, or bovalizumab.In certain embodiments, the inhibitor of proinflammatory pathway is tocilizumab, sarilumab, or bovalizumab.

[0192] In certain embodiments, the provided compositions may include at least one nucleic acid encoding an inhibitor of a proinflammatory pathway, the inhibitor of the proinflammatory pathway being an anti-IL-17 antibody. In certain embodiments, IL-17 is associated with the p38 MAPK pathway (e.g., as described in Noubade, et al., Blood 2011, 118, 3290-3300; Roussel, et al., J. Immunol. 2010, 184, 4531-4537; and Mai, et al., J. Biol. Chem. 2016, 291, 4939-4954, each of which is incorporated herein by reference for purposes described herein). In certain embodiments, the anti-IL-17 antibody is ixekizumab, bimekizumab, ALX-0761, CJM112, CNTO6785, LY3074828, SCH-900117, or MSB0010841. In certain embodiments, the inhibitor of the proinflammatory pathway is ixekizumab, bimekizumab, ALX-0761, CJM112, CNTO6785, LY3074828, SCH-900117, or MSB0010841.

[0193] In certain embodiments, the provided compositions may include at least one nucleic acid encoding an inhibitor of a proinflammatory pathway, and the inhibitor of the proinflammatory pathway is an anti-IL-17A antibody. In certain embodiments, the anti-IL17A antibody is secukinumab. In certain embodiments, the inhibitor of the proinflammatory pathway is secukinumab.

[0194] In certain embodiments, the provided compositions may include at least one nucleic acid encoding an inhibitor of a proinflammatory pathway, and the inhibitor of the proinflammatory pathway is an anti-IL-17RA antibody. In certain embodiments, the anti-IL17RA antibody is brodalumab. In certain embodiments, the inhibitor of the proinflammatory pathway is brodalumab.

[0195] In certain embodiments, the provided compositions may include at least one nucleic acid encoding an inhibitor of a proinflammatory pathway, and the inhibitor of the proinflammatory pathway is an anti-IL-23 / IL-12 antibody. In certain embodiments, the anti-IL-23 / IL-12 antibody is ustekinumab or briakinumab. In certain embodiments, the inhibitor of the proinflammatory pathway is ustekinumab or briakinumab.

[0196] In certain embodiments, the compositions provided may include at least one nucleic acid encoding an inhibitor of a proinflammatory pathway, and the inhibitor of the proinflammatory pathway is an anti-IL-23 antibody. In certain embodiments, IL-23 is associated with the p38 MAPK pathway (e.g., as described in Tang, et al., Immunology 2012, 135, 112-124, and Canavese, et al., J. Clin. Exp. Dermatol. Res. 2011, S2:002. doi:10.4172 / 2155-9554, each of which is incorporated herein by reference for purposes described herein). In certain embodiments, the anti-IL-23 antibody is tildrakizumab, BI655066, or guselkumab. In certain embodiments, the inhibitor of the proinflammatory pathway is tildrakizumab, BI655066, or guselkumab.

[0197] In certain embodiments, provided compositions may comprise at least one nucleic acid encoding an inhibitor of a proinflammatory pathway, the inhibitor of a proinflammatory pathway being MABp1, canakinumab, anakinra, rilonacept, olokizumab, clazakizumab, OPR-003, sirukumab, ARGX-109, FE301, tocilizumab, sarilumab, bovalilizumab, ixekizumab, bimekizumab, sonelokimab (ALX-0761, MSB0010841), CJM112, CNTO6785, mirikizumab (LY3074828), SCH-900117, secukinumab, brodalumab, ustekinumab, briakinumab, tildrakizumab, risankizumab (BI655066), or guselkumab.

[0198] In certain embodiments, the compositions provided may comprise at least one nucleic acid that acts as an inhibitor of a proinflammatory pathway or encodes an inhibitor of a proinflammatory pathway, and the inhibitor of a proinflammatory pathway is a TGFβR inhibitor. In certain embodiments, TGFβR is associated with the p38 MAPK pathway (e.g., Yu, et al., EMBOJ. 2002, 21, 3749-3759; Sato, et al., J. Invest. Dermatol. 2002, 118, 704-711; and Hanafusa, et al., J. Biol. Chem. 1999, 274, 27161-27167, each of which is incorporated herein by reference for the purposes described herein). In certain embodiments, the TGFβR inhibitor encoded by the nucleic acid is optionally coupled with galunisertib. In certain embodiments, the inhibitor of a proinflammatory pathway is galunisertib.

[0199] In certain embodiments, the compositions provided may comprise at least one nucleic acid comprising a sequence encoding a growth factor. Alternatively, in some embodiments, the compositions comprising at least one nucleic acid may further comprise a growth factor.

[0200] Growth factors are a broad category of molecules that contribute to cell proliferation, wound healing, and sometimes stimulating cell differentiation. Growth factors can include proteins and steroid hormones. Growth factors are involved in signaling between cells and play an important role in the immune system.

[0201] In certain embodiments, compositions provided include at least one nucleic acid encoding a growth factor, and / or at least one growth factor, wherein the nucleotide sequence is an amino acid sequence encoding an autocrine motility factor, an adrenomedullin, an angiopoietin (e.g., angiopoietin-1, angiopoietin-2), an autocrine motility factor, a bone morphogenetic protein (BMP) (e.g., BMP1, BMP2, BMP3, BMP4, BMP5, BMP6, BMP7, BMP8a, BMP8b, BMP10, BMP11, or BMP15), a ciliary neurotrophic factor family protein (e.g., ciliary neurotrophic factor family protein, trophic factor (CNTF), leukemia inhibitory factor (LIF), or interleukin-6 (IL-6)), colony-stimulating factors (e.g., macrophage colony-stimulating factor (M-CSF), granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin-3 (IL-3), or promegapoietin), epidermal growth factor (EGF), ephrin ligands (e.g., ephrin A1, A2, A3, A4, A5, B1, B2, and B3), erythropoietin, fibroblast growth factor (FGF) (e.g., FGF1, FGF2, FGF3, FGF4, FGF5, FGF6, FGF7, FGF8, FGF9, FGF10, FGF11, FGF12, FGF13, FGF14, FGF15, FGF16, FGF17, FGF18, FGF19, FGF20, FGF21, FGF22, or FGF23), bovine growth hormone (BGH), glial cell line-derived neurotrophic factor (GDNF) family ligand (e.g., GDNF, neuturin, artemin, or persephin), growth differentiation factor 9 (GDF9), hepatocyte growth factor (HGF), hepatoma-derived growth factor (HDGF) , insulin, insulin-like growth factor (IGF) (e.g., IGF-1 or IGF-2), interleukin (IL) (e.g., IL-1, IL-2, IL-3, IL-4, IL-5, IL-6, or IL-7), keratinocyte growth factor (KGF), mitogen stimulating factor (MSF), macrophage stimulating protein (MSP), myostatin (GDF-8), neuregulin (NRG) (e.g., NRG1, NRG2, NRG3, or NRG4), neurotrophins (e.g., brain-derived neurotrophic factor (BDNF), nerve growth factor (NGF),Growth factors that code for and / or include neurotrophin-3 (NT-3), or neurotrophin-4 (NT-4), placental growth factor (PGF), platelet-derived growth factor (PDGF), renalase (RNLS), T cell growth factor (TCGF), transforming factor (TGF) (e.g., TGF-α, and TGF-β1, TGF-β2, or TGF-β3), tumor necrosis factor alpha (TNF-α), vascular endothelial growth factor (VEGF), or a WNT family protein (e.g., WNT1, WNT2, WNT3, WNT4, WNT5, WNT6, WNT7A, WNT7B, WNT8A, WNT8B, WNT9A, WNT9B, WNT10A, WNT10B, WNT11, or WNT16).

[0202] In certain embodiments, the compositions provided may comprise at least one nucleic acid comprising a sequence encoding a hormone. Alternatively, in some embodiments, the compositions comprising at least one nucleic acid may further comprise a hormone.

[0203] Hormones are a class of signaling molecules that can function in multicellular organisms and in signal transduction between distant cells. Hormones are a broad category of molecules that include peptide hormones, amino acid derivative hormones, steroid hormones, and eicosanoid hormones. Hormones play important immunoregulatory roles, including regulating lymphocyte trafficking, promoting differentiation of T cell populations, and regulating T cell and antigen-presenting cell activity, see, for example, Luhder et al., "Short but powerful: short peptide hormones and their role in autoimmune inflammation" J Neuroimmunol. (2009) 271(1-2):1-7, the contents of which are incorporated herein by reference for purposes described herein. In certain embodiments, the hormones can be naturally occurring hormones or synthetic analogs of naturally occurring hormones.

[0204] In certain embodiments, compositions provided include at least one nucleic acid encoding a hormone, and / or at least one hormone, wherein the nucleotide sequence encodes and / or the hormone is adrenocorticotropic hormone (ACTH), amylin, angiotensin, atrial natriuretic peptide (ANP), bradykinin, calcitonin, cholecystokinin (CCK), gastrin, ghrelin, glucagon, glucagon-like peptide-1 (GLP-1), glucagon-like peptide-2 (GLP-2), gonadotropin-releasing hormone (GnRH), growth hormone, follicle-stimulating hormone (FSH), insulin, kallidin, leptin, luteinizing hormone (LH), melanocyte-stimulating hormone (MSH), oxytocin, parathyroid hormone (PTH), prolactin, renin, somatostatin, thyroid-stimulating hormone (TSH), thyrotropin-releasing hormone (TRH), vasoprepin, or vasoactive intestinal peptide (VIP). In certain embodiments, provided compositions comprise at least one nucleic acid encoding a hormone and / or at least one hormone, including abaloparatide, abarelix, afamelanotide, albiglutide, aviptadil, buserelin, carbetocin, cargutocin, carperitide, cetrorelix, degarelix, deslorelin, desmopressin, dulaglutide, elcatonin, exenatide, felypressin, ganirelix, and / or the nucleotide sequences encoding them are oxalate, ghrelin, gonadorelin, goserelin, histrelin, icatibant, lanreotide, leuprorelin, liraglutide, lixisenatide, nafarelin, nesiritide, octreotide, ornipressin, pasireotide, pramlintide, saralasin, taltirelin, teduglutide, teriparatide, terlipressin, tesamorelin, tetracosactide, thymopentin, or triptorelin.

[0205] In certain embodiments, the at least one nucleic acid encoding a hormone and / or the compositions provided comprising at least one hormone are useful in the treatment of cancer, hi certain embodiments, the at least one nucleic acid encoding a hormone and / or the compositions provided comprising at least one hormone are useful in the treatment of non-proliferative diseases. In certain embodiments, the provided compositions comprising at least one nucleic acid encoding a hormone and / or at least one hormone are useful for treating acromegaly, acute decompensated heart failure, bleeding esophageal varices, breast cancer, Cushing's disease, diabetes insipidus, endometriosis, erectile dysfunction, erythropoietic protoporphyria, hereditary angioedema, HIV lipodystrophy, hypercalcemia, hypoglycemia, neuroendocrine tumors, nocturia, osteoporosis, Paget's disease, postpartum hemorrhage, precocious puberty, primary amenorrhea, prostate cancer, renovascular hypertension, short bowel syndrome, spinocerebellar degeneration, type 1 diabetes, type 2 diabetes, various autoimmune diseases, various infectious diseases, various inflammatory diseases, vasodilatory shock, or West syndrome, as described, for example, in Lau and Dunn. "Therapeutic peptides: historical perspectives, current development trends, and future directions" Bioorg Med. See Chem (2018) 26(10):2700-2707, the contents of which are incorporated by reference for purposes herein.

[0206] In certain embodiments, the compositions provided may include a single therapeutic nucleic acid encoding a single immunomodulator. In certain embodiments, the compositions provided may include multiple nucleic acids encoding multiple different immunomodulators. In certain embodiments, the compositions provided may include at least one nucleic acid encoding multiple different immunomodulators operably linked on the same nucleic acid strand. In certain embodiments, the multiple nucleic acids may each encode a portion or part of the final protein or RNA product, which is generated as a result of homologous recombination, splicing, and / or tertiary or quaternary amino acid sequencing.

[0207] Polynucleotide Agent Carrier(s) In certain embodiments, the provided compositions may include at least one polymeric biomaterial, at least one nucleic acid, and optionally one or more polynucleotide agent carriers. In some embodiments, the provided compositions may include at least one nucleic acid and one or more polynucleotide agent carriers. In some embodiments, at least one nucleic acid of the provided compositions is complexed to a polynucleotide agent carrier. In some embodiments, the provided nucleic acid is supported on a polynucleotide agent carrier. In some embodiments, the provided compositions include at least one polynucleotide agent carrier, which may be a lipid, lipoplex, liposome, lipid nanoparticle (LNP), stable nucleic acid lipid particle (SNALP), SNALP based on ionizable lipid, lipidoid, DOTAP, DOTMA, DOPE, DSPE, DODMA, DODAP, DOGS, DC-Chol, PC, Chol, cationic polymer, amphiphilic copolymer(s), PEG, polyamino acid, polylactic or glycolic acid, polycaprolactone, poly(beta-amino ester), protamine, polyethyleneimine, gold, silver, and combinations and / or derivatives thereof, see, e.g., Ni et al. Life (Basel) 2019; 9 (3): 59, the contents of which are incorporated herein in their entirety for purposes described herein. In certain embodiments, the provided compositions do not contain one or more polynucleotide agent carriers.

[0208] In some embodiments, the nucleic acid of the exemplary compositions may function without being endocytosed. In some embodiments, the nucleic acid provided may function after endocytosis. In some embodiments, the nucleic acid of the exemplary compositions provided does not require a polynucleotide agent carrier for endocytosis. In some embodiments, the endocytosis of the nucleic acid of the exemplary compositions is assisted by a polynucleotide agent carrier. In embodiments in which the nucleic acid of the exemplary drug delivery composition is endocytosed, the endocytic cell may be said to be transfected. Transfection may occur with or without the assistance of a nucleic acid carrier.

[0209] Polynucleotides are generally macromolecules that can be highly hydrophilic / anionic while exhibiting relatively complex supramolecular assemblies. To successfully deliver intact polynucleotides to cells, important intracellular and extracellular barriers must be overcome. First, the polynucleotide must be protected from biodegradation by extracellular (e.g., serum and tissue-specific) nucleases. Once the nucleic acid reaches the cell, if it encodes an active agent, it must penetrate the anionic plasma membrane bilayer, a dynamic structure composed of phospholipids, membrane proteins, and cholesterol that are held together primarily through hydrophobic interactions. The desired outcome of transfection can then be observed only if the nucleic acid escapes intracellular degradation and performs its function (e.g., activate the innate immune system directly or through the creation of the encoded product). A method for delivering polynucleotides to all target cells while inducing minimal or negligible toxicity is desirable. Those skilled in the art will recognize that while there are numerous approaches developed for transfection, there are no universally effective materials and / or methods that work optimally in all cells.

[0210] In certain embodiments, to aid in endocytosis and / or transfection, the nucleic acids of the provided compositions may be modified through various base, backbone, or sugar modifications that may act to promote nucleic acid stability and transfection efficiency. In certain embodiments, chemical modifications may be used on one, two, or all three of the nucleic acid components, the sugar-phosphate backbone, the sugar moiety, and the heterocyclic base moiety. In certain embodiments, the nucleic acids of the exemplary compositions may include any, all, or none of the following changes. Sugar phosphate backbone modifications include, but are not limited to, phosphorothioate (PS), boranophosphate (BP), phosphonoacetate (PACE), morpholino (PMO), and peptide nucleic acid (PNA). Modifications of the sugar moiety include, but are not limited to, 2'-O-methyl (2'-O-ME), 2'-O-methoxyethyl (2'-MOE), 2'-fluoro (2'-F), 2'-deoxy-2'-fluoro-β-d-arabinonucleic acid (2'-FANA), 4'-thio (4'-S), 4'-C-aminomethyl-2'-O-Me, locked nucleic acid (LNA), tricycloDNA (tcDNA), unlocked nucleic acid (UNA). Modifications of the heterocyclic base include, but are not limited to, 5-bromo-uracil, 5-iodo-uracil, 2,6-diamino-purine, and 5-methylcytosine.

[0211] In certain embodiments, the provided compositions may include at least one nucleic acid and at least one polynucleotide chemical carrier. Chemical carriers are often rationally designed to overcome extracellular and intracellular obstacles that prevent nucleic acids from entering cells. Polynucleotide carriers may take advantage of the anionic charge of nucleic acids to either electrostatically condense them to a relatively small size suitable for internalization, or encapsulate them behind physical barriers, and / or in some cases, adsorb them to a surface. These effects may mask the negative charge on the nucleic acid. In summary, these effects may protect the nucleic acid from endonucleases, promote internationalization, and in some cases promote transcription and / or translation.

[0212] In certain embodiments, the compositions provided may include at least one nucleic acid and at least one cell membrane penetrating peptide (CPP). CPPs are usually relatively short (5-30 amino acids long) peptides of diverse properties that can carry various cargos, including polynucleotides, across cell membranes. CPPs can be classified into cationic peptides rich in basic amino acids such as lysine and arginine, as well as amphipathic peptides that contain both hydrophilic and hydrophobic amino acids. Amphipathic peptides may acquire different secondary structures depending on the spatial arrangement of hydrophobic and hydrophilic residues under different ambient conditions. Those skilled in the art will recognize that the charge number and density, size, hydrogen bonds, and secondary structure of CPPs may play an important role in determining their interaction with different components of the cell membrane and therefore their internalization. As carriers, CPPs can be either directly conjugated to polynucleotides or ionically complexed with nucleic acids. Covalent conjugation approaches allow the generation of conjugates with well-defined structures and stoichiometries. To further improve in vivo stability, CPPs can be modified with fatty acids or cholesterol. Examples of CPPs that can be included in the provided compositions include, but are not limited to, oligoarginine (R9), penetratin, lactoferrin, PepFect6 peptide, pepR, pepM, arginine, cysteine, gelatin, and peptides derived from the HIV-1 protein Tat.

[0213] In certain embodiments, the compositions provided may include hydrophobic substances or lipids. Cationic hydrophobic molecules are commonly used vehicles for nucleic acid delivery because they can be relatively easily synthesized and chemically modified for different applications. In certain embodiments, the polynucleotide agent carrier included in the exemplary composition may include, but is not limited to, commercially available cationic lipids: N-[1-(2,3-dioleoyloxy)-proper]-N,N,N-trimethylammonium (DOTMA, Lipofectin), 2,3-dioleyloxy-N-[2-sperminecarboxamido]ethyl-N,N-dimethyl-1-propane ammonium trifluoroacetate (DOSPA), 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP), N-[1-(2,3-dimyristyloxy)propyl]-N,N-dimethyl-N-(2-hydroxyethyl)ammonium bromide (DMRIE), 3b-[N-(N,N-dimethylaminoethane)carbamoyl]cholesterol (DC-Chol), dioctadecylamidoglyceryl spermine (DOGS, Transfectam), and / or dimethyldioctadecylammonium bromide (DDAB). Cationic lipids can be formulated with nucleic acids as stable nucleic acid lipid particles (SNALPs).

[0214] In certain embodiments, the provided compositions may include polymeric carriers of nucleic acids. In certain embodiments, the polymeric carriers may be chemically conjugated or complexed through electrostatic interactions or wrapped around nucleic acids to facilitate their delivery to target cells. In some embodiments, the polymeric carriers of nucleic acids may include, but are not limited to, polyethyleneimine (PEI), chitosan, PEG-PPL=PLLeu, PEI-CG-PEI, PgP, and / or any combination thereof.

[0215] Regulators of innate immune responses In certain embodiments, the compositions provided comprise at least one nucleic acid that acts as a regulator of the innate immune response or encodes a regulator of the innate immune response. In certain embodiments, the compositions provided may comprise two or more nucleic acids that act as regulators of the innate immune response or encode regulators of the innate immune response. The primary functions of the innate immune response include recruiting immune cells to the site of infection through the production of chemical factors, including specialized chemical mediators (e.g., cytokines), activating the complement cascade to identify bacteria, activate cells, and promote clearance of antibody complexes or dead cells, identifying and removing foreign bodies present in organs, tissues, blood, and lymph by specialized white blood cells, activating the adaptive immune system through a process known as antigen presentation, and acting as a physical and chemical barrier (e.g., epithelial surfaces, gastrointestinal tract) against infectious agents. Typically, white blood cells are white blood cells that carry out the functions of the innate immune system. These cells include natural killer cells, mast cells, eosinophils, basophils, macrophages, neutrophils, and dendritic cells. These cells function within the immune system by identifying and eliminating pathogens that can cause infection.

[0216] Inhibitors of innate immune responses The present disclosure recognizes that, among other things, by inhibiting inflammatory response, the risk of cancer recurrence can be reduced and thus survival can be extended. The present disclosure recognizes that, among other things, by inhibiting the proinflammatory immune response mediated by the p38 mitogen-activated protein kinase (MAPK) pathway (e.g., by acting as a p38 MAPK inhibitor or administering a nucleic acid encoding a p38 MAPK inhibitor) at the target site (e.g., tumor resection site), the risk of cancer recurrence can be reduced and thus survival can be extended. It is unexpected that MAPK inhibition can promote anti-tumor immunity, since it has been reported that MAPK targeting therapy (e.g., inhibition of BRAF / MEK / ERK module) induces a transcriptional signature associated with resistance to anti-PD-1 immune checkpoint blockade therapy, which in turn can negatively affect responsiveness to anti-PD-1 / L1 cancer therapy (see, e.g., Hugo, et al., Cell 2016,165,35-44, which is incorporated herein by reference for the purposes described herein).

[0217] Thus, in some embodiments, compositions are provided herein that act as or contain at least one nucleic acid encoding an inhibitor of a proinflammatory immune response mediated by the p38 mitogen-activated protein kinase (MAPK) pathway. In some embodiments, compositions provided herein may contain two or more nucleic acids that act as or contain an inhibitor of a proinflammatory immune response mediated by the p38 mitogen-activated protein kinase (MAPK) pathway. In some embodiments, where applicable, inhibitors of a proinflammatory immune response mediated by the p38 mitogen-activated protein kinase (MAPK) pathway may be delivered as nucleic acids encoding specific inhibitors.

[0218] The p38 family of MAPKs includes the p38α, p38β, p38γ, and p38δ isoforms. p38 MAPK is activated by multiple immune receptors, and thus inhibition of signaling modules or regulatory targets that function either upstream or downstream of p38 may provide an effective and selective method of inhibiting the molecular pathways and proinflammatory immune responses that it mediates.

[0219] For example, p38MAPK can be activated by mitogen-activated protein kinase kinase 3 (MAP2K3), mitogen-activated protein kinase kinase 6 (MAP2K6), mitogen-activated protein kinase kinase kinase 1 (MAP3K1), and / or mitogen-activated protein kinase kinase kinase 4 (MAP3K4). Thus, inhibition of upstream targets of p38MAPK can be effective in inhibiting the p38MAPK pathway.

[0220] Inhibition of downstream targets of p38MAPK may also be an effective means of inhibiting the p38MAPK pathway. Downstream of p38MAPK, for example, mitogen-activated protein kinase interacting protein kinase 1 and 2 (MNK1 and MNK2) are activated by the p38MAPK pathway. MNK kinases play an important role in regulating mRNA translation and are therefore major mediators of carcinogenic progression, drug resistance, proinflammatory cytokine production, and cytokine signaling. Mitogen and stress-activated kinases 1 and 2 (MSK1 and MSK2) are also downstream targets of p38MAPK and affect inflammatory responses. MAP kinase-activated protein kinases 2, 3, and 5 (MK2, MK3, MK5) are activated by p38MAPK and are involved in cellular stress and inflammatory responses.

[0221] In view of the above, inhibition of p38MAPK pathway through at least one nucleic acid acting as an inhibitor of upstream or downstream components of p38MAPK pathway or encoding the inhibitor may provide a therapeutic strategy for the treatment of cancer. In particular, local inflammatory wound response and systemic inflammatory process may together activate dormant micrometastasis or induce the proliferation of residual cancer cells, thus increasing the risk of cancer recurrence. Therefore, inhibiting the proinflammatory immune response mediated by p38MAPK pathway at tumor resection site may reduce the risk of cancer recurrence and prolong the survival of subjects.

[0222] In certain embodiments, the inhibitor of the proinflammatory immune response mediated by the p38MAPK pathway is a p38MAP kinase inhibitor. In certain embodiments, the provided compositions may act as or contain at least one nucleic acid encoding an inhibitor of p38α, p38β, p38γ, and / or p38δMAP kinase. In certain embodiments, the provided compositions may act as or contain at least one nucleic acid encoding a p38MAP kinase inhibitor, and include semapimod, pexmetinib, BMS-582949, losmapimod, pamapimod, ralimetinib, doramapimod, VX-702, VX-745, TAK-715, SB239063, SB202190, SB203580, SCIO469, PH-797804, AZD7624, ARRY-797, ARRY-614, AVE- 9940, LY3007113, skepinone-L, UM-164, SCIO323, SX-011, SK-F860002, SB706504, SB681323, CHF-6297, RWJ-67657, Org48762-0, ML3403, JX-401, EO-1428, DBM1285, AMG-548, AL-8697, PD-169316, PF-03715455, PH-797804, selonsertib, sorafenib, dilmapimod, and / or any combination thereof. In certain embodiments, provided compositions may include at least one nucleic acid that acts as or encodes a p38 MAP kinase inhibitor and may further include a quinazolinone, pyrimidopyrimidone, pyridopyrimidone, pyrazole, quinolinone, naphthyridinone core structure, and / or any combination thereof. In certain embodiments, provided compositions may further include the p38 MAP kinase inhibitor losmapimod.

[0223] In certain embodiments, the p38 MAP kinase inhibitor binds to the ATP binding site of p38 MAP kinase, hi certain embodiments, the p38 MAP kinase inhibitor is an allosteric inhibitor of p38 MAP kinase.

[0224] In certain embodiments, the compositions provided may comprise at least one nucleic acid that acts as an inhibitor of a proinflammatory immune response mediated by the p38MAPK pathway or that encodes an inhibitor of the proinflammatory immune response, the inhibitor acting on an upstream effector of p38MAPK. In certain embodiments, the at least one nucleic acid that acts as an inhibitor of a proinflammatory immune response mediated by the p38MAPK pathway or that encodes an inhibitor of the proinflammatory immune response is an inhibitor of RIPK1, RIPK2, RIPK3, RIPK4, RAC1, CDC42, MTK1, TAK1, MEKK1, MEKK2, MEKK3, MEKK4, DLK, MLK2, TAO1, TAO2, TLP2, TPL2, ASK1, MKK3, MKK4, and / or MKK6. In certain embodiments, the at least one nucleic acid that acts as an inhibitor of a proinflammatory immune response mediated by the p38MAPK pathway or that encodes an inhibitor of the proinflammatory immune response is an inhibitor of a downstream effector of p38MAPK. In certain embodiments, at least one nucleic acid that acts as or encodes an inhibitor of a proinflammatory immune response mediated by the p38 MAPK pathway is an inhibitor of MK2, MK3, MNK1, MNK2, MSK1, MSK2, MSK3, RSK, PP2A, and / or cPLA2.

[0225] Activators of the innate immune response In some embodiments, the compositions provided may include an activator of the innate immune response. In some embodiments, the compositions provided may include two or more activators of the innate immune response. In some embodiments, where applicable, the activator of the innate immune response may act as a specific activator or be delivered as a nucleic acid encoding the specific activator. The primary functions of the innate immune response include recruiting immune cells to the site of infection through the production of chemical factors, including specialized chemical mediators (e.g., cytokines), activating the complement cascade to identify bacteria, activate cells, and promote clearance of antibody complexes or dead cells, identifying and removing foreign bodies present in organs, tissues, blood, and lymph by specialized white blood cells, activating the adaptive immune system through a process known as antigen presentation, and acting as a physical and chemical barrier (e.g., epithelial surfaces, gastrointestinal tract) against infectious agents. Typically, white blood cells are the white blood cells that carry out the functions of the innate immune system. These cells include natural killer cells, mast cells, eosinophils, basophils, macrophages, neutrophils, and dendritic cells. These cells function within the immune system by identifying and eliminating pathogens that can cause infection.

[0226] In certain embodiments, the compositions provided may further comprise activation of a pro-inflammatory innate immune response mediated by p38 mitogen-activated protein.

[0227] In certain embodiments, provided compositions may further comprise at least one activator of the innate immune response.

[0228] In certain embodiments, the compositions provided include at least one nucleic acid that acts as an activator of the innate immune response or that encodes an activator of the innate immune response that is a ligand of a pattern recognition receptor (PRR). In certain embodiments, the at least one nucleic acid that acts as an activator of the innate immune response or that encodes an activator of the innate immune response is an agonist of the pattern recognition receptor (PRR).

[0229] In certain embodiments, provided compositions comprise at least one nucleic acid that acts as or encodes an activator of the innate immune response, and the activator is an inducer of type I interferon. In certain embodiments, the activator of the innate immune response is a recombinant interferon.

[0230] In certain embodiments, provided compositions comprise at least one nucleic acid that acts as or encodes an activator of the innate immune response, where the activator is an effective inducer of NK cell activation and / or proliferation. In certain embodiments, "effective inducer" refers to an activator of the innate immune response that directly induces NK cell activation and / or proliferation.

[0231] In certain embodiments, provided compositions comprise at least one nucleic acid that acts as or encodes an activator of the innate immune response, where the activator is an effective inducer of dendritic cell activation and / or maturation. In certain embodiments, "effective inducer" refers to an activator of the innate immune response that directly induces dendritic cell activation and / or maturation.

[0232] In certain embodiments, provided compositions comprise at least one nucleic acid that acts as or encodes an activator of the innate immune response, and the activator is an effective inducer of type I interferon by dendritic cells. In certain embodiments, "effective inducer" refers to an activator of the innate immune response that directly induces type I interferon by dendritic cells.

[0233] In certain embodiments, the compositions provided comprise at least one nucleic acid that acts as or codes for an activator of innate immunity. In some embodiments, the compositions provided comprise at least one nucleic acid that acts as or codes for an activator of innate immunity and an optional additional activator of innate immune response, where the additional activator is a small molecule or a biologic. In certain embodiments, the activator of innate immune response is a small molecule. In certain embodiments, the activator of innate immune response is a biologic.

[0234] In certain embodiments, the activator of the innate immune response is a stimulator of interferon genes (STING) agonist, a cytoplasmic DNA sensor (CDS) agonist, a Toll-like receptor (TLR) agonist, a C-type lectin receptor (CLR) agonist, a NOD-like receptor (NLR) agonist, a RIG type I receptor (RLR) agonist, and / or an inflammasome inducer.

[0235] In certain embodiments, the activator of the innate immune response is a stimulator of interferon genes (STING) agonist, a Toll-like receptor (TLR) agonist, and / or a NOD-like receptor (NLR) agonist. In certain embodiments, the activator of the innate immune response is a stimulator of interferon genes (STING) agonist and / or a Toll-like receptor (TLR) agonist. In certain embodiments, the activator of the innate immune response is a stimulator of interferon genes (STING) agonist, a TLR7 agonist, and / or a TLR8 agonist.

[0236] In certain embodiments, the activator of the innate immune response is 3'3'-cGAMP, 2'3'-cGAMP, 2'3'-cGAM(PS)2(Rp / Rp), 2'3'-cGAM(PS)2(Rp / Sp), 2'2'-cGAMP, c-di-AMP, 2'3'-c-di-AMP, 2'3'-c-di-AMP(PS)2(Rp / Rp), 2'3'-c-di-AMP(PS)2(Rp / Sp), c-di-GMP, c-di-IMP, HSV-60, ISD, VACV-70, poly(dA:dT), poly(dG:dC), heat-killed bacteria, lipoglycan ... Polysaccharides (LPS), lipoteichoic acid, peptidoglycans (PGNs), synthetic lipoproteins, poly(A:U), poly(I:C), monophosphoryl lipid A (MPLA), GSK1795091, G100, SD-101, MGN1703, CMP-001, flagellin (FLA), polyU, poly(dT), gardiquimod, imiquimod (R837), base analogues, adenine analogues, guanosine analogues, purine derivatives, benoazepine analogues, imidazoquinolines, thiazoquinolines, loxoribine, resiquimod (R848), dactolisib, sumanirole, N1-guanidine Lysinyl[4-((6-amino-2-(butylamino)-8-hydroxy-9H-purin-9-yl)methyl)benzoyl]spermine (CL307), CL264, CL097, CL075, CL347, CL401, CL413, CL419, CL531, CL553, CL572, MEDI9197, MEDI5083, hypoxanthine, TL8-506, PF-4878691, isatoribine, SM-324405, SM-324406, AZ12441970, AZ12443988, CpG oligonucleotides, bacterial DNA, beta-glucan , beta-glucans from fungal and bacterial cell walls, γ-D-Glu-mDAP (iE-DAP), iE-DAP derivatives, muramyl dipeptide (MDP), MDP derivatives, 5' triphosphate double-stranded RNA, poly(dA:dT), ATP, chitosan, potassium aluminum sulfate, calcium pyrophosphate dihydrate, silica dioxide, MurNAc-L-Ala-γ-D-Glu-mDAP (M-TriDAP), xanthenone analogs (e.g., DMXAA, vadimezan), TREX1 inhibitors, cyclic dinucleotides, LHC165, GSK-2245035, RG7854,GS-9620, GS-9688, EMD1201081, PF-3512676, BO-112, RGT-100, MK-1454, SB-11285, NKTR-262, CDX-301, 2'3'-c-di-GMP, cAIMP, cAIM(PS)2(Rp / Sp), derivatives thereof, and / or pharma- ceutically acceptable salts thereof.

[0237] In certain embodiments, the activator of the innate immune response is a fluorinated derivative of any of the innate immune response activators described herein. In certain embodiments, the activator of the innate immune response is a difluorinated cAIMP (c-(2'FdAMP-2'FdIMP)). In certain embodiments, the activator of the innate immune response is a difluorinated cAIM(PS)2(Rp / Sp). In certain embodiments, the activator of the innate immune response is an O-methylated derivative of any of the above activators.

[0238] In certain embodiments, the activator of the innate immune response is 3'3'-cGAMP, 2'3'-cGAMP, 2'3'-cGAM(PS)2(Rp,Rp), 2'3'-cGAM(PS)2(Rp,Sp), 2'2'-cGAMP, c-di-AMP, 2'3'-c-di-AMP, 2'3'-c-di-AM(PS)2(Rp,Rp), 2'3'-c-di-AM(PS)2(Rp,Sp), c-di-GMP, 2'3'-c-di-GMP, 2'3'-c-di-GM(PS)2(Rp,Rp), 2'3'-c-di -GM(PS)2(Rp,Sp), c-di-IMP, resiquimod, CpG oligonucleotides, polyinosinic acid:polycytidylic acid, LHC165, GSK-2245035, RG7854, GS-9620, GS-9688, EMD1201081, PF-3512676, BO-112, RGT-100, MK-1454, SB-11285, NKTR-262, CDX-301, 2'3'-c-di-GMP, cAIMP, cAIM(PS)2(Rp / Sp), and / or pharma- ceutically acceptable salts thereof.

[0239] In certain embodiments, the activator of the innate immune response is 3'3'-cGAMP, 2'3'-cGAMP, 2'3'-cGAM(PS)2(Rp,Rp), 2'3'-cGAM(PS)2(Rp,Sp), 2'2'-cGAMP, c-di-AMP, 2'3'-c-di-AMP, 2'3'-c-di-AM(PS)2(Rp,Rp), 2'3'-c-di-AM Fluorinated derivatives of (PS)2(Rp,Sp), c-di-GMP, 2'3'-c-di-GMP, 2'3'-c-di-GM(PS)2(Rp,Rp), 2'3'-c-di-GM(PS)2(Rp,Sp), c-di-IMP, 2'3'-c-di-GMP, cAIMP, cAIM(PS)2(Rp / Sp), and / or pharma- ceutically acceptable salts thereof.

[0240] In certain embodiments, the activator of the innate immune response is an O-methylated derivative of 3'3'-cGAMP, 2'3'-cGAMP, 2'3'-cGAM(PS)2(Rp,Rp), 2'3'-cGAM(PS)2(Rp,Sp), 2'2'-cGAMP, c-di-AMP, 2'3'-c-di-AMP, 2'3'-c-di-AM(PS)2(Rp,Rp), 2'3'-c-di-AM(PS)2(Rp,Sp), c-di-GMP, 2'3'-c-di-GMP, 2'3'-c-di-GM(PS)2(Rp,Rp), 2'3'-c-di-GM(PS)2(Rp,Sp), c-di-IMP, and / or a pharma- ceutically acceptable salt thereof.

[0241] In certain embodiments, the activator of the innate immune response is 2'3'-cGAMP, 2'3'-c-di-AM (PS) 2 (Rp, Rp), MurNAc-L-Ala-γ-D-Glu-mDAP (M-TriDAP), c-di-GMP, and / or resiquimod. In certain embodiments, the activator of the innate immune response is 2'3'-cGAMP, 2'3'-c-di-AM (PS) 2 (Rp, Rp), MurNAc-L-Ala-γ-D-Glu-mDAP (M-TriDAP), and / or resiquimod. In certain embodiments, the activator of the innate immune response is 2'3'-cGAMP, 2'3'-c-di-AM (PS) 2 (Rp, Rp), and / or resiquimod. In certain embodiments, the activator of the innate immune response is 2'3'-c-di-AM(PS)2(Rp,Rp) and / or resiquimod. In certain embodiments, the activator of the innate immune response is cAIMP and / or a fluorinated derivative thereof. In certain embodiments, the activator of the innate immune response is difluorinated cAIMP.

[0242] In certain embodiments, the activator of innate immune response is 2'3'-cGAMP or a pharma- ceutically acceptable salt thereof. In particular, 2'3'-cGAMP (cyclic [G(2',5')pA(3',5')p]) has been described to function as an endogenous second messenger and induce STING-dependent type I interferon response. 2'3'-cGAMP has also been shown to be an effective adjuvant to promote the production of antigen-specific antibodies and T cell responses in mice. 2'3'-cGAMP exerts antiviral functions in the cells in which it is produced, but can cross cell membranes by passive diffusion and exert effects on neighboring cells. [ka]

[0243] In certain embodiments, the activator of the innate immune response is 2'3'-c-di-AM(PS)2(Rp,Rp) or a pharma- ceutically acceptable salt thereof. 2'3'-c-di-AM(PS)2(Rp,Rp) is the Rp,Rp isomer of the 2'3' bisphosphorothioate analog of 3'3'-cyclic adenosine monophosphate (c-di-AMP). It is also a STING agonist. [ka]

[0244] In certain embodiments, the activator of the innate immune response is cAIMP, its difluorinated derivative, its difluorinated bisphosphorothioate derivative (cAIM(PS)2(Rp / Sp)), and / or its pharma- ceutically acceptable salt. cAIMP and its derivatives are also STING agonists. [ka] [ka] [ka]

[0245] In certain embodiments, the activator of innate immune response is a STING agonist, and the STING agonist is a cyclic dinucleotide.In certain embodiments, the cyclic dinucleotide is any cyclic dinucleotide disclosed in U.S. Patent No. 10,106,574, granted on October 23, 2018, the entire contents of which are incorporated herein by reference.In certain embodiments, the cyclic dinucleotide is any cyclic dinucleotide disclosed in U.S. Patent No. 10,131,686, granted on November 20, 2018, the entire contents of which are incorporated herein by reference.

[0246] In certain embodiments, the activator of the innate immune response is MK-1454.

[0247] In certain embodiments, the activator of the innate immune response is a cytoplasmic DNA sensor (CDS) agonist. In certain embodiments, the CDS agonist is a cyclic GMP-AMP synthase (cGAS) agonist.

[0248] In certain embodiments, the activator of the innate immune response is any STING agonist or cGAS agonist disclosed in U.S. Patent No. 10,336,786, granted July 2, 2019, the entire contents of which are incorporated herein by reference. In certain embodiments, the activator of the innate immune response is any STING agonist or cGAS agonist disclosed in U.S. Patent Application No. 14 / 268,967, filed May 2, 2014, the entire contents of which are incorporated herein by reference. In certain embodiments, the activator of the innate immune response is any STING agonist or cGAS agonist disclosed in U.S. Patent Application No. 9,840,533, granted December 12, 2017, the entire contents of which are incorporated herein by reference. In certain embodiments, the activator of the innate immune response is any STING agonist or cGAS agonist disclosed in U.S. Patent No. 10,176,292, granted January 8, 2019, the entire contents of which are incorporated herein by reference.

[0249] In certain embodiments, the activator of the innate immune response is any STING agonist disclosed in U.S. Patent Application No. 13 / 057,662, filed June 14, 2011, the entire contents of which are incorporated herein by reference. In certain embodiments, the activator of the innate immune response is any STING agonist disclosed in U.S. Patent Application No. 9,695,212, granted July 4, 2017, the entire contents of which are incorporated herein by reference. In certain embodiments, the activator of the innate immune response is any STING agonist disclosed in U.S. Patent Application No. 15 / 035,432, filed May 19, 2016, the entire contents of which are incorporated herein by reference. In certain embodiments, the activator of the innate immune response is any STING agonist disclosed in U.S. Patent Application No. 16 / 069,201, filed January 11, 2018, the entire contents of which are incorporated herein by reference. In certain embodiments, the activator of the innate immune response is any STING agonist disclosed in U.S. Patent No. 10,604,542, granted March 31, 2020, the entire contents of which are incorporated herein by reference. In certain embodiments, the activator of the innate immune response is any STING agonist disclosed in International Patent Application No. PCT / US2014 / 038525, filed May 18, 2014, the entire contents of which are incorporated herein by reference. In certain embodiments, the activator of the innate immune response is any STING agonist disclosed in U.S. Patent No. 9,770,467, granted September 26, 2017, the entire contents of which are incorporated herein by reference. In certain embodiments, the activator of the innate immune response is any STING agonist disclosed in International Patent Application No. PCT / IB2016 / 057265, filed January 12, 2016, the entire contents of which are incorporated herein by reference.

[0250] In certain embodiments, the activator of the innate immune response is MurNAc-L-Ala-γ-D-Glu-mDAP (M-TriDAP) or a pharma- ceutically acceptable salt thereof. M-TriDAP is a peptidoglycan (PGN) degradation product found primarily in gram-negative bacteria. M-TriDAP is recognized by the intracellular sensors NOD1 (CARD4) and, to a lesser extent, NOD2 (CARD15). Recognition of M-TriDAP by NOD1 / NOD2 induces a signaling cascade involving the serine / threonine RIP2 (RICK, CARDIAK) kinase interacting with IKK, which results in the activation of NF-κB and the production of inflammatory cytokines such as TNF-α and IL-6. M-TriDAP induces the activation of NF-κB at a level similar to that of Tri-DAP.

[0251] In certain embodiments, the activator of the innate immune response is a TLR7 agonist. In certain embodiments, the activator of the innate immune response is a TLR8 agonist. In certain embodiments, the activator of the innate immune response is a TLR7 agonist and a TLR8 agonist.

[0252] In certain embodiments, the activator of the innate immune response is an immune response modifier (IRM).

[0253] In certain embodiments, the activator of the innate immune response is any IRM disclosed in U.S. Patent No. 5,714,608, granted February 3, 1998, the entire contents of which are incorporated herein by reference. In certain embodiments, the activator of the innate immune response is any IRM disclosed in U.S. Patent No. 6,039,969, granted March 21, 2000, the entire contents of which are incorporated herein by reference. In certain embodiments, the activator of the innate immune response is any IRM disclosed in U.S. Patent No. 6,200,592, granted March 13, 2000, the entire contents of which are incorporated herein by reference. In certain embodiments, the activator of the innate immune response is any IRM disclosed in U.S. Patent No. 4,689,338, granted August 25, 1987, the entire contents of which are incorporated herein by reference. In certain embodiments, the activator of the innate immune response is any IRM disclosed in U.S. Patent No. 5,446,153, granted on August 29, 1995, the entire contents of which are incorporated herein by reference. In certain embodiments, the activator of the innate immune response is any IRM disclosed in U.S. Patent No. 6,194,425, granted on February 2, 2001, the entire contents of which are incorporated herein by reference. In certain embodiments, the activator of the innate immune response is any IRM disclosed in U.S. Patent No. 6,110,929, granted on August 29, 2000, the entire contents of which are incorporated herein by reference. In certain embodiments, the activator of the innate immune response is any IRM disclosed in U.S. Patent No. 7,544,697, granted on June 9, 2009, the entire contents of which are incorporated herein by reference.

[0254] In certain embodiments, the activator of the innate immune response is CL307 (N1-glycinyl[4-((6-amino-2-(butylamino)-8-hydroxy-9H-purin-9-yl)methyl)benzoyl]spermine) or a pharma- ceutically acceptable salt thereof. CL307 is a highly potent TLR7 agonist. Titration experiments have shown that CL307 induces robust NF-κB activation even at concentrations as low as 20 nM (10 ng / ml). [ka]

[0255] In certain embodiments, the activator of the innate immune response is CL264 or a pharma- ceutically acceptable salt thereof. CL264 induces activation of NF-κB and secretion of IFN-α in TLR7-expressing cells. CL264 is a TLR7-specific ligand and does not stimulate TLR8 even at high concentrations (>10 μg / ml). In HEK293 cells transfected with TLR7, CL264 induces NF-κB activation at a concentration of 0.1 μM, 5-10 times lower than imiquimod. [ka]

[0256] In certain embodiments, the activator of the innate immune response is loxoribine or a pharma- ceutically acceptable salt thereof. 7 Rank and C 8 Loxoribine is a guanosine analogue derivatized at the 5'-position. This nucleoside is a highly potent stimulator of the immune system. Loxoribine activates the innate immune system via TLR7, and this activation requires endosomal maturation. Loxoribine recognition is restricted to TLR7. [ka]

[0257] In certain embodiments, the activator of the innate immune response is hypoxanthine or a pharma- ceutically acceptable salt thereof. Hypoxanthine is a naturally occurring purine derivative. [ka]

[0258] In certain embodiments, the activator of innate immune response is TL8-506 or a pharma- ceutically acceptable salt thereof. TL8-506 is a benzazepine compound that is an analog of the Toll-like receptor 8 (TLR8) agonist VTX-2337. TL8-506 activates TLR8 more potently than R848 and CL075. TL8-506 is about 50-fold and about 25-fold more potent than R848 and CL075, respectively, in inducing NF-κB activation in HEK293 cells transfected with TLR8. TL8-506 is a selective agonist of TLR8. [ka]

[0259] In certain embodiments, the activator of the innate immune response is PF-4878691, isatoribine, SM-324405, SM-324406, AZ12441970, AZ12443988, GSK-2245035, RG7854, GS-9620, LHC165, NKTR-262, GS-9688, VTX-2337, or a pharma- ceutically acceptable salt thereof. PF-4878691, isatoribine, SM-324405, SM-324406, AZ12441970, AZ12443988, GSK-2245035, RG7854, and GS-9620 are TLR7 agonists. LHC165 and NKTR-262 are both TLR7 and TLR8 agonists. GS-9688 and VTX-2337 are TLR8 agonists.

[0260] In certain embodiments, the activator of the innate immune response is an imidazoquinoline derivative, including dactolisib, imiquimod, gardiquimod, resiquimod, sumanirole, and pharma- ceutically acceptable salts thereof.

[0261] In certain embodiments, the activator of the innate immune response is CL097 or a pharma- ceutically acceptable salt thereof. CL097 is a highly water-soluble derivative of resiquimod (≧20 mg / ml). CL097 is a TLR7 and TLR8 ligand. It induces NF-κB activation at 0.4 μM (0.1 μg / ml) in TLR7-transfected HEK293 cells and at 4 μM (1 μg / ml) in TLR8-transfected HEK293 cells. [ka]

[0262] In certain embodiments, the activator of the innate immune response is CL075 or a pharma- ceutically acceptable salt thereof. CL075 (3M002) is a thiazoloquinolone derivative that stimulates TLR8 in human peripheral blood mononuclear cells. It activates NF-kB and preferentially induces the production of TNF-α and IL-12. CL075 also induces the secretion of IFN-α through TLR7, but to a lesser extent. In TLR8-transfected HEK293 cells, 0.4 μM (0.1 μg / ml) induces the activation of NF-kB, and in TLR7-transfected HEK293 cells, approximately 10-fold more CL075 is required to activate NF-kB. [ka]

[0263] In certain embodiments, the activator of the innate immune response is MEDI9197 or a pharma- ceutically acceptable salt thereof. MEDI9197 (3M052) is an injectable TLR7 and TLR8 agonist. It is an imidazoquinoline immune response modifier (IRM) with a C18 lipid moiety and is designed to be slow to disperse from the application site. [ka]

[0264] In certain embodiments, the activator of the innate immune response is resiquimod (R848) or a pharma- ceutically acceptable salt thereof. In particular, resiquimod is an agent that acts as an immune response modifier and has antiviral and antitumor activity. It is used as a topical gel to treat skin lesions such as those caused by herpes simplex virus and cutaneous T-cell lymphoma. It is also used as an adjuvant to increase the efficacy of vaccines. It has several mechanisms of action, being an agonist of Toll-like receptors 7 (TLR7) and 8 (TLR8) and an upregulator of opioid growth factor receptors. [ka]

[0265] In certain embodiments, the activator of innate immune response is a TLR7 selective antedrug. In certain embodiments, the activator of innate immune response is SM-324405, AZ12441970, or a pharmaceutically acceptable salt thereof.

[0266] In certain embodiments, the activator of innate immune response is GS-9620. In certain embodiments, the activator of innate immune response is PF-4878691. In certain embodiments, the activator of innate immune response is NKTR-262. In certain embodiments, the activator of innate immune response is LHC165.

[0267] In certain embodiments, the activator of the innate immune response is an inflammasome inducer. Inflammasomes are multimeric protein complexes that are essential for host defense against infection and endogenous danger signals. They promote the secretion of the proinflammatory cytokines interleukin (IL)-1β and IL-18, and trigger a rapid, proinflammatory form of cell death called pyroptosis.

[0268] In certain embodiments, the activator of the innate immune response is an inducer of the NLRP3, AIM2, NLRC4, or NLRP1 inflammasome.

[0269] In certain embodiments, the activator of the innate immune response is [ka] or a pharma- ceutically acceptable salt thereof, wherein R 1 is H and R 2 is H or R 1 is a butyl group, and R 2 is H or R 1 is H and R 2 But -CO 2 CH 3 or R 1 is a butyl group, and R 2 But -CO 2 CH 3 It is.

[0270] In certain embodiments, the activator of the innate immune response is an imadazoquinoline, an imidazonaphthyridine, a pyrazolopyridine, an aryl substituted imidazoquinoline, a compound having a 1-alkoxy 1H-imidazo ring system, an oxazolo[4,5-c]-quinolin-4-amine, a thiazolo[4,5-c]-quinolin-4-amine, a selenazolo[4,5-c]-quinolin-4-amine, an imidazonaphthyridine ... 1-Substituted, 2-Substituted 1H-Imidazo[4,5-C]quinolin-4-amines, Condensed Cycloalkylimidazopyridines, 1H-Imidazo[4,5-c]quinolin-4-amines, 1-Substituted 1H-Imidazo-[4,5-c]quinolin-4-amines, Imidazo-[4,5-C]quinolin-4-amines, 2-Ethyl 1H-Imidazo[4,5-C]quinolin-4-amines, Oligomeric 1H-Imidazo[4,5-C]quinolin-4-amines dazo[4,5-c]quinolin-4-amine, 6,7-dihydro-8-(imidazol-1-yl)-5-methyl-1-oxo-1H,5H-benzo[ij]quinolizine-2-carboxylic acid, pyridoquinoxaline-6-carboxylic acid, 6,7-dihydro-8-(imidazol-1-yl)-5-methyl-1-oxo-1H,5H-benzo[ij]quinolizine-2-carboxylic acid, substituted naphtho[ij]quinolizine, substituted pyridoquinoxaline-6-carboxylic acid, 7-hydroxy-benzo[ij]quinolizine-2-carboxylic acid derivatives, substituted benzo[ij]quinolizine-2-carboxylic acid, 7-hydroxy-benzo[ij]quinolizine-2-carboxylic acid, substituted pyrido[1,2,3,-de]-1,4-benzoxazine, N-methylenemalonate of tetrahydroquinoline, and / or pharma-ceutically acceptable salts thereof.

[0271] In certain embodiments, the activator of the innate immune response is any NLRP3 agonist disclosed in U.S. Patent Application No. 15 / 253,215, filed August 31, 2016, the entire contents of which are incorporated herein by reference.

[0272] In certain embodiments, the activator of the innate immune response is a RORy agonist. A RORy agonist is an agent that promotes RORy activity, such as by binding to and activating RORy or increasing the expression of RORy in a patient or cell population. A RORy agonist can be, for example, a small organic molecule, a polypeptide, or a nucleic acid. Various RORy agonists have been reported in the literature, such as U.S. Pat. No. 9,394,315, Zhang et al. in Mol.Pharmacol. (2012) vol.82, pages 583-590, and Wang et al. in ACSChem.Biol. (2010), vol.5, pages1029-1034, each of which is incorporated herein by reference for the purposes described herein.

[0273] In certain embodiments, the activator of the innate immune response is [ka] and pharma- ceutically acceptable salts thereof.

[0274] In certain embodiments, the activator of the innate immune response is a general or specific compound described in U.S. Pat. No. 9,394,315, granted July 19, 2016, the entire contents of which are incorporated herein by reference, such as a compound of formula (I) as follows: [ka] or a pharma- ceutically acceptable salt thereof, A is aryl, aralkyl, heteroaryl, cycloalkyl, or heterocycloalkyl, each of which is selected from the group consisting of halogen, hydroxyl, C 1-6 Alkyl, C 1-6 Haloalkyl, C 1-6 Hydroxyalkyl, C 1-6 Alkoxy, C 1-6 Haloalkoxy, -N(R 4 )(R 5 ), -CO 2R 6 , -C(O)R 6 , -CN, -C 1-4 Alkylene-C 1-4 Alkoxy, -C 1-4 Alkylene-N(R 4 )(R 5 ), -C 1-4 Alkylene-CO 2 R 6 , -OC 1-6 Alkylene-N(R 4 )(R 5 ), -N(R 4 )C(O)-C 1-6 Alkylene-N(R 4 )(R 5 ), -S(O) p C 1-6 Alkyl, -SO 2 N(R 4 )(R 5 ), -N(R 4 )SO 2 (C 1-6 alkyl), -C(O)N(R 4 )(R 5 ), and -N(R 4 )C(O)N(R 4 )(R 5 and optionally substituted with 1, 2, or 3 substituents independently selected from the group consisting of: X is -O-[C(R 6 )(R 7 )]-[C(R 6 ) 2 ] m -Ψ, -OC(R 6 ) 2- C(R 6 )(R 7 )-C(R 6 ) 2 -Ψ, -OC(R 6 ) 2- C(R 6 )(R 7 )-Ψ, -C(R 6 ) 2- [C(R 6 )(R 7 )]-[C(R 6 ) 2 ]m-Ψ, -C(O)-[C(R 6 )(R7 )]-[C(R 6 ) 2 ]m-Ψ, -C(R 6 ) 2- N(R 8 )-[C(R 6 )(R 7 )]-[C(R 6 ) 2 ]m-Ψ, -C(R 6 )=N-Ψ, -C(R 6 ) 2 C(R 6 )=N-Ψ, -N=C(R 6 )-Ψ, or -N=C(R 6 )C(R 6 ) 2 -Ψ, where ψ is the bond to the sulfonamide ring nitrogen atom in formula I; Y is -N(R 2 )(R 3 ) or -O-aralkyl, wherein the aralkyl is selected from halogen, hydroxyl, C 1-6 Alkoxy, C 1-6 Haloalkoxy, C 1-6 Alkyl, C 1-6 Haloalkyl, -N(R 4 )(R 5 ), -CN, -CO 2- C 1-6 Alkyl, -C(O)-C 1-6 Alkyl, -C(O)N(R 4 )(R 5 ), -S(O) p C 1-6 Alkyl, -SO 2 N(R 4 )(R 5 ), and -N(R 4 )SO 2 (C 1-6 alkyl), R 1 is, independently for each occurrence, hydrogen, halogen, or C 1-6 represents an alkyl group, R 2 is -C(O)-aryl, -C(O)-aralkyl, -C(O)-[C(R 6 )2 ] m -cycloalkyl, -C(O)-[C(R 6 ) 2 ] m -heterocyclyl, -C(O)-C 1-6 Alkyl, -C(O)-C 1-6 Alkylene-C 1-6 Alkoxyl, -C(O)-C 1-6 Alkylene-cycloalkyl, or -C(O)-C 1-6 alkylene-heterocycloalkyl, each of which is selected from halogen, hydroxyl, C 1-6 Alkoxy, C 1-6 Haloalkoxy, C 1-6 Alkyl, C 1-6 Haloalkyl, -N(R 4 )(R 5 ), -CN, -CO 2- C 1-6 Alkyl, -C(O)-C 1-6 Alkyl, -C(O)N(R 4 )(R 5 ), -S(O) p C 1-6 Alkyl, -SO 2 N(R 4 )(R 5 ), and -N(R 4 )SO 2 (C 1-6 alkyl), R 3 is hydrogen or C 1-6 is alkyl, R 4 and R 5 each independently for each occurrence is hydrogen or C 1-6 Represents alkyl or R 4 and R 5 form, together with the nitrogen atom to which they are attached, a 3- to 7-membered heterocycle; R 6 is, independently for each occurrence, hydrogen or C 1-6 represents an alkyl group, R 7 But hydrogen, hydroxyl, C 1-6Hydroxyalkyl, C 1-6 Alkyl, C 1-6 Haloalkyl, -CO 2 R 6 , C 1-6 Alkylene-CO 2 R 6 , C 1-4 Hydroxyalkylene-CO 2 R 6 , -N(R 4 )(R 5 ), C 1-6 Alkylene-N(R 4 )(R 5 ), C 1-6 Hydroxyalkylene-N(R 4 )(R 5 ), -N(R 4 )C(O)R 9 , C 1-6 Alkylene-N(R 4 )C(O)R 9 , C 1-6 Alkylene-C(O)N(R 4 )(R 5 ), -N(R 4 )CO 2- C 1-6 Alkyl or C 1-6 Alkylene-N(R 4 )(C(O)N(R 4 )(R 5 ) or R 7 is heterocycloalkyl or C 1-4 Alkylene-heterocycloalkyl, wherein the heterocycloalkyl is oxo, halogen, hydroxyl, C 1-6 Alkyl, C 1-6 Haloalkyl, C 1-6 Hydroxyalkyl, C 1-6 Alkoxy and C 1-6 optionally substituted with 1, 2, or 3 substituents independently selected from the group consisting of haloalkoxy; R 8 But hydrogen, C 1-6 Alkyl, or -C(O)-C 1-6 is alkyl, R 9 But hydrogen, C 1-6Alkyl, C 1-6 Hydroxyalkyl, C 1-6 Alkylene-N(R 4 )(R 5 ), or C 1-6 Alkylene-N(R 4 )C(O)-C 1-6 is alkyl, n is 1 or 2; m and p are each independently represented by 0, 1, or 2 for each occurrence.

[0275] In certain embodiments, the activator of the innate immune response is any RORy agonist disclosed in U.S. Patent No. 9,394,315, granted July 19, 2016, the entire contents of which are incorporated herein by reference. In certain embodiments, the activator of the innate immune response is any RORy agonist disclosed in U.S. Patent No. 10,532,088, granted January 14, 2020, the entire contents of which are incorporated herein by reference.

[0276] In certain embodiments, the activator of innate immune response is a RIG-I-like receptor (RLR) agonist. In certain embodiments, the activator of innate immune response is RGT-100.

[0277] Regulators of adaptive immunity In certain embodiments, the compositions described herein comprise a nucleic acid that encodes or acts as a regulator(s) of an adaptive immune response.

[0278] The adaptive immune response system, also known as the acquired immune system, is a subsystem of the overall immune system that includes highly specialized systemic cells and processes that eliminate or prevent pathogen growth. The adaptive immune system is one of two major immune strategies found in vertebrates (the other is the innate immune system). Adaptive immunity generates immunological memory after an initial response to a specific pathogen, resulting in an enhanced response to subsequent encounters with that pathogen. This process of acquired immunity is the basis of vaccination. Like the innate system, the adaptive system contains both humoral and cell-mediated immune components. Unlike the innate immune system, the adaptive immune system is highly specific to a particular pathogen.

[0279] The adaptive immune response system is triggered in vertebrates when a pathogen evades the innate immune response system, produces a threshold level of antigen, and generates a "stranger" or "danger" signal that activates dendritic cells. The main functions of the adaptive immune system include the recognition of specific "non-self" antigens in the presence of "self" during the antigen presentation process, the generation of responses tailored to eliminate specific pathogens or pathogen-infected cells, and the development of immunological memory in which pathogens are "remembered" via memory B cells and memory T cells.

[0280] Activators of the adaptive immune response In some embodiments, compositions provided herein can include one or more nucleic acids that encode or act as activators of the adaptive immune response, hi some embodiments, an activator of the adaptive immune response is or includes a nucleic acid that can directly activate the adaptive immune system and / or encodes an activator of the adaptive immune system.

[0281] Useful approaches to activate the adaptive immune response system (e.g., to activate therapeutic antitumor immunity) include immune checkpoint blockade. Immune checkpoints refer to the numerous inhibitory pathways built into the immune system that are essential for regulating the duration and magnitude of physiological immune responses in peripheral tissues to maintain self-tolerance and minimize collateral tissue damage. Tumors employ certain immune checkpoint pathways as a primary mechanism of immune tolerance, particularly to T cells specific for tumor antigens. Because many of the immune checkpoints are initiated by ligand-receptor interactions, they can be easily blocked by antibodies or regulated by recombinant forms of the ligand or receptor. Cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) antibodies were the first of this class of immunotherapeutic agents to receive FDA approval (ipilimumab). Preliminary clinical findings with blockade of additional immune checkpoint proteins, such as programmed cell death protein 1 (PD-1), indicate broad and diverse opportunities to enhance antitumor immunity with the potential to generate sustained clinical responses.

[0282] PD-1, which functions as an immune checkpoint, plays a key role in downregulating the immune system by preventing the activation of T cells, thereby reducing autoimmunity and promoting self-tolerance. The inhibitory effect of PD-1 is achieved through a dual mechanism that promotes apoptosis (programmed cell death) in antigen-specific T cells in lymph nodes while simultaneously reducing apoptosis in regulatory T cells (suppressor T cells). A new class of therapeutic agents that block PD-1, PD-1 inhibitors (e.g., anti-PD-1 antibodies), activate the immune system to attack tumors and are therefore used to treat several types of cancer. In addition, antibodies to programmed death ligand 1 (PD-L1) provide similar effects on the activation of adaptive immune responses as antibodies that target PD-1. Thus, compositions containing anti-PD-L1 antibodies are expected to provide similar therapeutic effects as compositions containing anti-PD-1 antibodies.

[0283] In certain embodiments, the activator of the adaptive immune response is a small molecule. In certain embodiments, the activator of the adaptive immune response is a biologic. In certain embodiments, the biologic and / or small molecule is a nucleic acid, or an RNA or protein product encoded by a nucleic acid. In certain embodiments, the biologic is a protein. In certain embodiments, the biologic is an antibody or a fragment thereof. In certain embodiments, the biologic is a nucleic acid that encodes a protein or RNA that can act as an activator of the adaptive immune response.

[0284] In certain embodiments, the compositions provided include at least one nucleic acid encoding an antibody. In certain embodiments, the compositions provided include at least one nucleic acid, and optionally an antibody. In certain embodiments, the nucleic acid encodes an activator of the adaptive immune response, or the antibody is an activator of the adaptive immune response, and the encoded activator is an antibody. In certain embodiments, the activator of adaptive immunity is an anti-PD-1 antibody, an anti-PD-L1 antibody, an anti-CTLA-4 antibody, an anti-TIM3 antibody, an anti-OX40 antibody, an anti-GITR antibody, an anti-LAG-3 antibody, an anti-CD137 antibody, an anti-CD3 antibody, an anti-CD27 antibody, an anti-CD28 antibody, an anti-CD28H antibody, an anti-CD30 antibody, an anti-CD39 antibody, an anti-CD40 antibody, an anti-CD43 antibody, an anti-CD47 antibody, an anti-CD48 antibody, an anti-CD70 antibody, an anti-CD43 antibody, an anti-CD47 antibody, an anti-CD48 antibody, an anti-CD43 antibody, an anti-CD47 antibody, an anti-CD43 ... Antibodies, anti-CD73 antibody, anti-CD96 antibody, anti-CD123 antibody, anti-CD155 antibody, anti-CD160 antibody, anti-CD200 antibody, anti-CD244 antibody, anti-ICOS antibody, anti-TNFRSF25 antibody, anti-TMIG D2 antibody, anti-DNAM1 antibody, anti-BTLA antibody, anti-LIGHT antibody, anti-TIGIT antibody, anti-VISTA antibody, anti-HVEM antibody, anti-Siglec antibody, anti-GAL1 antibody, anti-GAL3 antibody, anti-GAL9 antibody , anti-BTNL2 (butrophilin) ​​antibody, anti-B7-H3 antibody, anti-B7-H4 antibody, anti-B7-H5 antibody, anti-B7-H6 antibody, anti-KIR antibody, anti-LIR antibody, anti-ILT antibody, anti-CEACAM1 antibody, anti-C EACAM5 antibody, anti-CEACAM6 antibody, anti-MICA antibody, anti-MICB antibody, anti-NKG2D antibody, anti-NKG2A antibody, anti-A2AR antibody, anti-C5aR antibody, anti-TGFβ antibody, anti-TGFβR antibody, anti-CX and nucleic acids encoding anti-CR4 antibodies, anti-CXCL12 antibodies, anti-CCL2 antibodies, anti-IL-10 antibodies, anti-IL-13 antibodies, anti-IL-23 antibodies, anti-phosphatidylserine antibodies, anti-neuropilin antibodies, anti-GalCer antibodies, anti-HER2 antibodies, anti-VEGFA antibodies, anti-VEGFR antibodies, anti-EGFR antibodies, anti-Tie2 antibodies, anti-CCR4 antibodies, anti-TRAIL-DR5 antibodies, and / or any combination thereof.

[0285] In certain embodiments, the activator of the adaptive immune response comprises at least one nucleic acid encoding a fragment of any of the antibodies listed herein and / or a fragment of any of the antibodies listed herein. Certain embodiments. In certain embodiments, the activator of the adaptive immune response is a nucleic acid encoding a humanized form of any of the antibodies listed herein. In certain embodiments, the activator of the adaptive immune response is a nucleic acid encoding a single chain of any of the antibodies listed herein. In certain embodiments, the activator of the immune response is a nucleic acid encoding a multimeric form (e.g., a dimeric IgA molecule, a pentavalent IgM molecule) of any of the antibodies listed herein.

[0286] In certain embodiments, the activator of the adaptive immune response is or is a nucleic acid encoding an anti-PD-1 antibody, an agonistic anti-CD137 antibody, an agonistic anti-CD40 antibody, an anti-CTLA-4 antibody, an anti-LAG-3 antibody, an anti-TIM3, or a combination thereof. In certain embodiments, the activator of the adaptive immune response is a nucleic acid encoding an anti-PD-1 antibody or an anti-CTLA-4 antibody. In certain embodiments, the activator of the adaptive immune response is a nucleic acid encoding an anti-PD-1 antibody. In certain embodiments, the activator of the adaptive immune response is a nucleic acid encoding an anti-CTLA-4 antibody. In certain embodiments, the activator of the adaptive immune response is a nucleic acid encoding an agonistic anti-CD137 antibody. In certain embodiments, the activator of the adaptive immune response is a nucleic acid encoding an anti-LAG-3 antibody. In certain embodiments, the activator of the adaptive immune response is a nucleic acid encoding an anti-TIM3 antibody.

[0287] In certain embodiments, the activator of the adaptive immune response is a nucleic acid encoding an antibody and / or is an antibody. In certain embodiments, the nucleic acid is selected from the group consisting of pembrolizumab, nivolumab, pidilizumab, ipilimumab, tremelimumab, durvalumab, atezolizumab, avelumab, PF-06801591, utomilumab, PDR001, PBF-509, MGB453, LAG525, AMP-224, INCSHR1210, INCAGN1876, INCAGN1949, samalizumab, PF-05082566, urelumab, lirilumab, lurizumab, BMS-936559, BMS-936561, BMS-986004, BMS-986012, BMS-986016, BMS-986178, IMP32 1, IPH2101, IPH2201, IPH5401, IPH4102, IPH4301, IPH52, IPH53, Valilumab, Ulocuplumab, Monalizumab, MEDI0562, MEDI0680, MEDI1873, MEDI6383, MEDI6469, MEDI9447, AMG228, AMG820, CC-90002, CDX-1127, CGEN15001T, CGEN15022, CGEN15029, CGEN15049, CGEN15027, CGEN15052, CGEN15092, CX-072, CX-2009, CP-870893, Lucatumumab, Dacetuzumab, Chi Lob7 / 4, RG6058, RG7686, RG7876, RG7888, TRX518, MK-4166, IMC-CS4, emactuzumab, trastuzumab, pertuzumab, obinutuzumab, cabilalizumab, margetuximab, enoblitzumab, mogamulizumab, panitumumab, carlumab, ramucirumab, bevacizumab, rituximab, cetuximab, fresolimumab, denosumab, MGA012, AGEN1884, AGEN2034, LY3300054, JTX-4014, teplizumab, FP The antibody may be coated with or be an antibody selected from the group consisting of A150, PF-04136309, PF-06747143, AZD5069, GSK3359609, FAZ053, TSR022, MBG453, REGN2810, REGN3767, MOXR0916, PF-04518600, RO7009789, BMS986156, GWN323, JTX-2011, NKTR-214, GSK3174998, DS-8273a, NIS793, BGB-A317, and / or any combination thereof.

[0288] In certain embodiments, the activator of the adaptive immune response is an antibody-encoding nucleic acid and / or is an antibody. In certain embodiments, the nucleic acid encodes or the antibody is pembrolizumab, nivolumab, pidilizumab, ipilimumab, tremelimumab, durvalumab, atezolizumab, REGN2810, MGA012, AGEN1884, AGEN2034, LY3300054, JTX-4014, avelumab, and / or any combination thereof.

[0289] In certain embodiments, the activator of the adaptive immune response is an antibody mimetic or antibody fusion, or a nucleic acid encoding either the antibody mimetic or the antibody fusion.

[0290] In certain embodiments, the activator of the adaptive immune response is a nucleic acid encoding a bispecific antibody and / or is a bispecific antibody. In certain embodiments, the nucleic acid is selected from the group consisting of RG7802 (an antibody targeting carcinoembryonic antigen (CEA) and the CD3 receptor), RG7828 (a bispecific monoclonal antibody targeting CD20 on B cells and CD3 on T cells), RG7221 (a bispecific monoclonal antibody targeting VEGF and angiopoietin 2), RG7386 (a bispecific monoclonal antibody targeting FAP and DR5), ERY974 (a bispecific monoclonal antibody targeting CD3 and glypican 3), MGD012 (a bispecific monoclonal antibody targeting PD-1 and LAG-3), AMG211 (a bispecific T cell engager targeting CD3 and CEA), MEDI573 (a bispecific monoclonal antibody targeting IGF1 and IGF2), MEDI565 (a bispecific monoclonal antibody targeting CD3 and CEA), FS1 7 (undisclosed target), FS18 (bispecific monoclonal antibody targeting LAG3 and an undisclosed target), FS20 (undisclosed target), FS22 (undisclosed target), FS101 (bispecific monoclonal antibody targeting EGFR and HGF), FS117 (undisclosed target), FS118 (bispecific monoclonal antibody targeting LAG3 and PD-L1), RO6958688 (bispecific monoclonal antibody targeting CD3 and CEA), MCLA-128 (bispecific monoclonal antibody targeting HER2 and HER3), M7824 (bifunctional fusion protein targeting PD-L1 and TGFβ), MGD009 (humanized antibody recognizing both B7-H3 and CD3), MGD013 (bispecific PD-1 and LAG-3 antibody), and / or any combination thereof, or the bispecific antibody is one of them.

[0291] In certain embodiments, the activator of adaptive immune response is an antibody-drug conjugate.In certain embodiments, the antibody-drug conjugate is trastuzumab emtansine, inotuzumab ozogamicin, PF-06647020, PF-06647263, PF-06650808, RG7596, RG7841, RG7882, RG7986, DS-8201, ABBV-399, glembatumumab vedotin, inotuzumab ozogamicin, MEDI4276, or a pharmaceutically acceptable salt thereof.

[0292] In certain embodiments, the activator of adaptive immune response is a small molecule.In certain embodiments, the small molecule is IDO inhibitor, TGFβR inhibitor, BRAF inhibitor, KIT inhibitor, A2aR inhibitor, Tie2 inhibitor, arginase inhibitor, iNOS inhibitor, HIF1α inhibitor, STAT3 inhibitor, PGE2 inhibitor, PDE5 inhibitor, RON inhibitor, mTOR inhibitor, JAK2 inhibitor, HSP90 inhibitor, PI3K-AKT inhibitor, β-catenin inhibitor, GSK3β inhibitor, IAP inhibitor, HDAC inhibitor, DNMT inhibitor, BET inhibitor, COX2 inhibitor, PDGFR inhibitor, VEGFR inhibitor, BCR-ABL inhibitor, proteasome inhibitor, angiogenesis inhibitor, MEK inhibitor, BRAF+MEK inhibitor, pan-RAF inhibitor, EGFR inhibitor, PARP inhibitor, glutaminase inhibitor, WNT inhibitor, FAK inhibitor, ALK inhibitor, CDK4 / 6 inhibitor or FGFR3 inhibitor.

[0293] In certain embodiments, the small molecule is celecoxib, sunitinib, imatinib, vemurafenib, dabrafenib, bortezomib, vorinostat, pomalidomide, thalidomide, lenalidomide, epacadostat, indoximide, GDC0919, BMS986205, AZD8055, AZD4635, CPI-444, PBF509, LCL161, CB-839, CB-1158, FPA008, BLZ945, IPI-549, pexidartinib, galunisertib, virinapant, trametinib, cobimetinib, binimetinib, ensartib, gefitinib, pazopanib, sorafenib, nisartin ... ntedanib, SYM004, veliparib, olaparib, BGB-290, everolimus, LXH254, azacitidine, decitabine, guadecitabine, RRX001, CC486, romidepsin, entinostat, panobinostat, tamoxifen, ibrutinib, idelalisib, capmatinib, selumetinib, abemaciclib, palbociclib, glasdegib, enzalutamide, AZD9150, PF-06840003, SRF231, Hu5F9-G4, CC-900002, TTI-621, WNT974, BGJ398, LY2874455, or a pharma- ceutically acceptable salt thereof.

[0294] Additional medications In certain embodiments, the compositions provided may act as or contain nucleic acids encoding therapeutic agents not specifically disclosed in the "Modulators of Innate Immune Responses" or "Modulators of Adaptive Immunity" sections.

[0295] In certain embodiments, the compositions provided may comprise at least one nucleic acid that acts as or encodes a regulator of macrophage effector function. Macrophages are immune cells derived from circulating monocytes, present in all tissues, and involved in many conditions of pathology. Macrophages play a dichotomous role in cancer, where they can promote tumor growth but also serve as important immune effectors of therapeutic antibodies. Macrophages express all classes of Fcγ receptors and have the potential to destroy tumors through the process of antibody-dependent cellular phagocytosis. Many studies have shown that macrophage phagocytosis is the primary mechanism of action of many antibodies approved to treat cancer. Thus, several approaches to enhance macrophage responses to therapeutic antibodies are under investigation, including the search for new targets and the development of antibodies with enhanced functions. Macrophage responses to antibody therapy may be enhanced with engineered Fc variants, bispecific antibodies, or antibody-drug conjugates. Macrophages have shown success as effectors of cancer immunotherapy.

[0296] In certain embodiments, the provided compositions may include at least one nucleic acid that acts as or encodes a regulator of macrophage effector function, and the regulator of macrophage effector function is a regulator of suppressive myeloid cells, including myeloid-derived suppressor cells (MDSCs). In certain embodiments, the regulator of macrophage effector function may kill, deplete, or enhance macrophages and / or MDSCs. In certain embodiments, the regulator of macrophage effector function is an anti-CD40 antibody, an anti-CD47 antibody, an anti-CSF1 antibody, or an anti-CSF1R antibody. In certain embodiments, the regulator of macrophage effector function is SRF231, Hu5F9-G4, CC-900002, or TTI-621 (anti-CD47 antibody). In certain embodiments, the regulator of macrophage effector function is MCS-110 (anti-CSF1 antibody). In certain embodiments, the regulator of macrophage effector function is FPA008, RG7155, IMC-CS4, AMG820, or UCB6352 (anti-CSF1R antibody).

[0297] In some embodiments, the provided compositions may include at least one nucleic acid that acts as or encodes an effector function regulator, and optionally additional small molecules. In certain embodiments, the optional regulator of macrophage effector function is a small molecule inhibitor of CSF1R. In certain embodiments, the regulator of macrophage effector function is BLZ945, GW2580, or PLX3397 (a small molecule inhibitor of CSF1R).

[0298] In some embodiments, provided compositions may include at least a biomaterial and at least one nucleic acid that acts as or encodes a modulator of effector function, where the modulator is a BTK inhibitor, an ITK inhibitor, a PI3K gamma inhibitor, or a PI3K delta inhibitor.

[0299] In certain embodiments, the provided compositions comprising at least a biological material and a nucleic acid may further comprise an oncolytic virus, including, but not limited to, herpes simplex virus (e.g., HSV1716, OncoVex GM-CSF), adenovirus (e.g., H101, Onyx-15), poliovirus (e.g., PV1 (RIPO)), reovirus (e.g., reolysin), Seneca virus (e.g., NTX-010, SVV-001), Rigville virus, Maraba virus, measles, Newcastle disease virus, vaccinia, or ECHO virus.

[0300] In certain embodiments, provided compositions comprising at least biological material and nucleic acid can further comprise a radioisotope (e.g., as part of a molecule or on a bead). In certain embodiments, the radioisotope is yttrium-90, palladium-103, iodine-125, cesium-131, or iridium-192.

[0301] In certain embodiments, the compositions provided include at least a biomaterial and a nucleic acid, which may be a chemotherapeutic agent. In certain embodiments, the compositions provided include at least a biomaterial, a nucleic acid, and an optional additional chemotherapeutic agent. In certain embodiments, the chemotherapeutic agent may include antiestrogen (e.g., tamoxifen, raloxifene, and megestrol), LHRH agonists (e.g., goscrclin and leuprolide), antiandrogens (e.g., flutamide and bicalutamide), photodynamic therapy (e.g., vertoporfin (BPD-MA), phthalocyanines, photosensitizer Pc4, and demethoxy-hypocrelin A (2BA-2-DMHA)), nitrogen mustards (e.g., cyclophosphamide, ifostat, cyclophosphamide, cyclophosphamide, ifostat, etc.). sufamide, trofosfamide, chlorambucil, estramustine, and melphalan), nitrosoureas (e.g., carmustine (BCNU) and lomustine (CCNU)), alkylsulfonates (e.g., busulfan and treosulfan), triazenes (e.g., dacarbazine and temozolomide), platinum-containing compounds (e.g., cisplatin, carboplatin, and oxaliplatin), vinca alkaloids (e.g., vincristine, vinblastine, vindesine, and vinorelbine). , taxoids (e.g., paclitaxel or nanoparticle albumin-bound paclitaxel (ABRAXANE), docosahexaenoic acid-bound paclitaxel (DHA-paclitaxel, taxoplexin), polyglutamic acid-bound paclitaxel (PG-paclitaxel, paclitaxel poliglumex, CT-2103, XYOTAX), tumor-activated prodrug (TAP) ANG1005 (angiopep-2 bound to three molecules of paclitaxel), paclitaxel-EC-1 (erbB2-recognizing paclitaxel linked to peptide EC-1), and glucose-conjugated paclitaxel, e.g., 2'-paclitaxel 2-glucopyranosyl methyl succinate, docetaxel, taxol), epipodophyllins (e.g., etoposide, etoposide phosphate, teniposide, topotecan, 9-aminocamptothecin, camptoirinotecan, irinotecan, crisnatol, and mitomycin C), antimetabolites, DHFR inhibitors (e.g., methotrexate, dichloromethotrexate,trimetrexate, and edatrexate), IMP dehydrogenase inhibitors (e.g., mycophenolic acid, tiazofurin, ribavirin, EICAR), ribonucleoside reductase inhibitors (e.g., hydroxyurea and deferoxamine), uracil analogs (e.g., 5-fluorouracil (5-FU), floxuridine, doxifluridine, latitrexed, tegafur uracil, and capecitabine), cytosine analogs (e.g., cytarabine (arabinose), C), cytosine arabinoside, and fludarabine), purine analogs (e.g., mercaptopurine and thioguanine), vitamin D3 analogs (e.g., EB1089, CB1093, and KH1060), isoprenylation inhibitors (e.g., lovastatin), dopaminergic neurotoxins (e.g., 1-methyl-4-phenylpyridinium ion), cell cycle inhibitors (e.g., staurosporine), actinomycins (e.g., actinomycin D, dactinomycin), bleomycins (e.g., bleomycin A2, bleomycin B2, and peplomycin), anthracyclines (e.g., daunorubicin, doxorubicin, PEGylated liposomal doxorubicin, idarubicin, epirubicin, pirarubicin, zorubicin, and mitoxantrone), MDR inhibitors (e.g., verapamil), Ca, 2+ These include, but are not limited to, ATPase inhibitors (e.g., thapsigargin), oblimersen, gemcitabine, carminomycin, leucovorin, pemetrexed, cyclophosphamide, dacarbazine, procarbidine, prednisolone, dexamethasone, campatecin, plicamycin, asparaginase, aminopterin, methopterin, porfiromycin, melphalan, leurocidin, leurosine, chlorambucil, trabectedin, procarbazine, discodermolide, carminomycin, aminopterin, hexamethylmelamine, and / or pharmaceutically acceptable salts thereof.

[0302] In certain embodiments, the chemotherapeutic agent is an immunomodulatory chemotherapeutic agent. In certain embodiments, the chemotherapeutic agent has a known immunomodulatory function (e.g., induction of immunogenic cell death or depletion of immunosuppressive regulatory immune cells). In certain embodiments, the chemotherapeutic agent is included in the drug delivery composition due to its immunotherapeutic properties, rather than its use as a traditional cancer cell-specific cytotoxic chemotherapy. In certain embodiments, the drug delivery composition does not include a chemotherapeutic agent. In certain embodiments, the drug delivery composition does not include a cytotoxic agent.

[0303] In some embodiments, the provided composition may include at least a biomaterial and at least one nucleic acid acting as a targeted agent or encoding a targeted agent. In certain embodiments, the targeted agent includes, but is not limited to, an IDO inhibitor, a TGFβR inhibitor, an arginase inhibitor, an iNOS inhibitor, a HIF1α inhibitor, a STAT3 inhibitor, a CSF1R inhibitor, a PGE2 inhibitor, a PDE5 inhibitor, a RON inhibitor, an mTOR inhibitor, a JAK2 inhibitor, an HSP90 inhibitor, a PI3K-AKT inhibitor, a β-catenin inhibitor, a GSK3β inhibitor, an IAP inhibitor, an HDAC inhibitor, a DNMT inhibitor, a BET inhibitor, an A2AR inhibitor, a BRAF+MEK inhibitor, a pan-RAF inhibitor, a PI3Kγ inhibitor, a PI3Kδ inhibitor, an EGFR inhibitor, a VEGF inhibitor, a PARP inhibitor, a glutaminase inhibitor, a BTK inhibitor, an ITK inhibitor, a WNT inhibitor, a FAK inhibitor, an ALK inhibitor, a CDK4 / 6 inhibitor, or an FGFR3 inhibitor.

[0304] In some embodiments, the compositions provided may include at least a biomaterial and at least one nucleic acid acting as a targeted agent or encoding a targeted agent. In some embodiments, the compositions provided may include at least a biomaterial and at least one nucleic acid acting as a targeted agent or encoding a targeted agent, and optionally additional targeted agents. In some embodiments, the targeted agents include imatinib, thalidomide, lenalidomide, tyrosine kinase inhibitors (e.g., axitinib (AG013736), bosutinib (SKI-606), cediranib (RECENTIN™, AZD2171), dasatinib (SPRYCEL®, BMS-354825), erlotinib (TARCEVA®), gefitinib (IRESSA®), imatinib (Gl eevec®, CGP57148B, STI-571), lapatinib (TYKERB®, TYVERB®), lestaurtinib (CEP-701), neratinib (HKI-272), nilotinib (TASIGNA®), semaxanib (semakinib, SU5416), sunitinib (SUTENT®, SU11248), toceranib (PALLADIA®), vandetanib (ZACTIM A (registered trademark), ZD6474), vatalanib (PTK787, PTK / ZK), trastuzumab (HERCEPTIN (registered trademark)), bevacizumab (AVASTIN (registered trademark)), rituximab (RITUXAN (registered trademark)), cetuximab (ERBITUX (registered trademark)), panitumumab (VECTIBIX (registered trademark)), ranibizumab (Lucentis (registered trademark)), nilotinib (TASIGNA (registered trademark)), sorafenib ( NEXAVAR®), everolimus (AFINITOR®), alemtuzumab (CAMPATH®), gemtuzumab ozogamicin (MYLOTARG®), temsirolimus (TORISEL®), ENMD-2076, PCI-32765, AC220, dovitinib lactate (TKI258, CHIR-258), BIBW2992 (TOVOKTM), SGX523, PF-04217903,PF-02341066, PF-299804, BMS-777607, ABT-869, MP470, BIBF1120 (VARGATEF®), AP24534, JNJ-26483327, MGCD265, DCC-2036, BMS-690154, CEP-11981, tivozanib (AV-951), OSI-930, MM-121, XL-184, XL-647, and / or X L228), proteasome inhibitors (e.g., bortezomib (VELCADE)), mTOR inhibitors (e.g., rapamycin, temsirolimus (CCI-779), everolimus (RAD-001), ridaforolimus, AP23573 (Ariad), AZD8055 (AstraZeneca), BEZ235 (Novartis), BGT226 (Norvartis), XL765 (Sanofi Aventis), PF-4691502 (Pfizer), GDC0980 (Genetech), SF1126 (Semafoe) and OSI-027 (OSI)), epacadostat, indoximide, GDC0919, BMS986205, AZD4635, CPI-444, PBF509, LCL161, CB-839, CB-1158, FPA008, BLZ945, IPI-549, pexidartinib, galunisertib, birinapant, trametinib, dabrafenib, vemurafenib, cobimetinib, binimetinib, ensartib, pazopanib, nintedanib, SYM004, veliparib, olapa The present invention includes, but is not limited to, rib, BGB-290, LXH254, azacitidine, decitabine, guadecitabine, RRX001, CC486, romidepsin, entinostat, vorinostat, panobinostat, tamoxifen, ibrutinib, idelalisib, capmatinib, selumetinib, abemaciclib, palbociclib, glasdigib, enzalutamide, AZD9150, PF-06840003, SRF231, Hu5F9-G4, CC-900002, TTI-621, WNT974, BGJ398, LY2874455, anti-Tie2 antibodies, or pharma- ceutically acceptable salts thereof.

[0305] Drug Delivery Composition Embodiments In some embodiments, the compositions provided comprise at least one polymeric biomaterial and at least one nucleic acid. In certain embodiments, the compositions provided comprise a multi-polymeric biomaterial and at least one nucleic acid.

[0306] In certain embodiments, provided compositions comprise a polymeric biomaterial(s) and two or more nucleic acids.

[0307] In certain embodiments, compositions provided include polymeric biomaterial(s), one or more nucleic acids, and one or more polynucleotide carriers.

[0308] In certain embodiments, compositions provided include at least one polymeric biomaterial and at least one nucleic acid encoding a peptide.

[0309] In certain embodiments, compositions provided include polymeric biomaterial(s), at least one nucleic acid encoding a peptide, and at least one polynucleotide carrier.

[0310] In certain embodiments, compositions provided include a polymeric biomaterial(s), one or more nucleic acids, a polynucleotide carrier, and an inhibitor of a proinflammatory pathway.

[0311] In certain embodiments, provided compositions comprise a polymeric biomaterial(s), one or more nucleic acids, a polynucleotide carrier, an inhibitor of a proinflammatory pathway, and an activator of the innate immune response.

[0312] In certain embo...

Claims

1. (i) a polymeric biomaterial; and (ii) a polynucleotide agent that encodes or regulates an immunomodulatory polypeptide, the composition being for use in a method comprising administering the composition to a target site in a subject undergoing tumor resection.

2. (a) the polynucleotide agent (i) encodes a cytokine that induces innate and / or adaptive immunity, or (ii) activates a pattern recognition receptor that induces innate immunity, or (iii) encodes a chemokine that induces immune cell recruitment, or (iv) encodes an antibody that mediates immune checkpoint blockade or costimulation, and / or (b) the target site is (i) the site of tumor resection, or (ii) a site near the site of tumor resection, or (iii) a sentinel lymph node; and / or (c) the administering step is by injection, and optionally (i) the composition is a liquid and the polymeric biomaterial is a viscous polymer solution, or (ii) the composition is a liquid and the polymeric biomaterial forms a polymeric network biomaterial in situ at the target site upon administration, and optionally the polymeric network biomaterial comprises or is a crosslinked or non-crosslinked polymeric network biomaterial. A composition for use according to claim 1.

3. The polymeric biomaterial is characterized in that (a) when tested in vitro by placing the combination of the polymeric biomaterial and the polynucleotide agent in PBS (pH 7.4), less than 100% of the polynucleotide agent is released from the polymeric biomaterial within 3 hours, or (b) when tested in vivo by administering the combination of the polymeric biomaterial and the polynucleotide agent to the mammary fat pad of a mouse subject, 50% or less of the polynucleotide agent is released in vivo 8 hours after said administration, or (c) when tested in vivo by administering the combination of the polymeric biomaterial and the polynucleotide agent to the mammary fat pad of a mouse subject, the polymeric biomaterial is characterized in that the polynucleotide agent is released when evaluated 24 hours after administration. or (d) prolonging the release of the polynucleotide agent such that more polynucleotide agent is present in the mammary fat pad than would be observed if the polynucleotide agent were administered in solution; or (d) the polynucleotide agent is released from the polymeric biomaterial and taken up by local cells such that (i) at least a subset of local immune cells express the immunomodulatory polypeptide encoded by the polynucleotide agent, (ii) at least a subset of local immune cells have increased expression of type 1 interferon in response to innate immune stimulation induced by the polynucleotide agent, and / or (iii) at least a subset of local immune cells have a change in the level and / or activity of the immunomodulatory polypeptide. A composition for use according to claim 1. (a) the polymeric biomaterial (i) is characterized by a storage modulus of 10 Pa to 5,000 Pa, or (ii) comprises or is a hydrogel, or (iii) comprises a positively charged polymer, and / or (b) the polymeric biomaterial further comprises a polynucleotide agent carrier, optionally comprising a cationic agent and / or a lipid, and the polynucleotide agent is, for example, (i) complexed to the polynucleotide agent carrier or (ii) supported on the polynucleotide agent carrier; and / or (c) the administering step does not involve administration of a tumor antigen to the subject of the tumor resection; and / or (d) the administering step does not involve administration of microparticles to the subject of the tumor resection; and / or (e) the administering step does not involve adoptive transfer of immune cells to the subject of the tumor resection; and / or (f) the composition further comprises an inhibitor of a pro-inflammatory immune response mediated by the p38 mitogen-activated protein kinase (MAPK) pathway; and / or (g) the composition further comprises an activator of innate immunity, optionally (i) the activator of innate immunity is or comprises a stimulator of interferon genes (STING) agonist, or (ii) the activator of innate immunity is or comprises a Toll-like receptor (TLR) 7 and / or TLR8 ("TLR7 / 8") agonist, and / or (h) the composition further comprises an activator of adaptive immunity; and / or (i) the polymeric biomaterial forms a matrix or depot, the polynucleotide agent is within the polymeric biomaterial, and optionally the polynucleotide agent is released by diffusion through the polymeric biomaterial, e.g., the polymeric biomaterial is biodegradable in vivo; and / or (j) the tumor resection site is characterized by the absence of macroscopic residual tumor antigens; and / or (k) the tumor resection subject is afflicted with metastatic cancer, and optionally the method includes monitoring at least one metastatic site in the tumor resection subject after said administering. A composition for use according to any one of claims 1 to 3.

5. 1. A method for producing a polymer network biomaterial composition, said method comprising: (a) providing a composition comprising one or more precursor components of a polymer network biomaterial and a polynucleotide agent that encodes or modulates an immunomodulatory polypeptide; (b) allowing the precursor components to form a polymer network biomaterial in less than 10 minutes, wherein the polymer network biomaterial comprises: the polymer network biomaterial has a storage modulus of less than 5,000 Pa; and / or when tested in vitro by placing the combination of the polymer network biomaterial and the polynucleotide agent in PBS (pH 7.4), less than 100% of the polynucleotide agent is released from the polymer network biomaterial within 3 hours; when the combination of the polymer network biomaterial and the polynucleotide agent is tested in vivo by administering it to the mammary fat pad of a mouse subject, 50% or less of the polynucleotide agent is released in vivo 8 hours after said administration; and / or when tested in vivo by administering a combination of the polymer network biomaterial and the polynucleotide agent to the mammary fat pad of a mouse subject, the polymer network biomaterial prolongs the release of the polynucleotide agent such that, when evaluated 24 hours after administration, more polynucleotide agent is present in the mammary fat pad than is observed when the polynucleotide agent is administered in solution; and allowing the polynucleotide agent released from the polymer network biomaterial to form characterized in that (i) at least a subset of local immune cells express the immunomodulatory polypeptide encoded by the polynucleotide agent, (ii) at least a subset of local immune cells have increased expression of type 1 interferon in response to innate immune stimulation induced by the polynucleotide agent, and / or (iii) at least a subset of local immune cells have a change in the level and / or activity of the immunomodulatory polypeptide.

6. The method of claim 5, wherein (a) the polymer network biomaterial comprises or is a crosslinked or non-crosslinked polymer network biomaterial, and / or (b) the polymer network biomaterial comprises or is a hydrogel, and / or (c) the polymer network biomaterial comprises a polynucleotide agent carrier, optionally the polynucleotide agent carrier comprising a cationic agent and / or a lipid, for example, (i) the polynucleotide agent is complexed to the polynucleotide agent carrier, or (ii) the polynucleotide agent is supported on the polynucleotide agent carrier.

7. 1. A method for characterizing a polymer network biomaterial composition or its component(s), said method comprising: (a) providing in vitro a polymer network biomaterial composition in a buffer solution, said polymer network biomaterial composition comprising a candidate polymer biomaterial and a polynucleotide agent that encodes or modulates an immunomodulatory polypeptide; (b) determining whether less than 100% of the polynucleotide agent is released from the polymeric biomaterial within 3 hours. (a) the buffer solution comprises or is PBS (pH 7.4); and / or (b) the determining step further comprises measuring the amount of the polynucleotide agent released from the polymeric biomaterial at predetermined time points over a period of at least 3 hours, and optionally the determining step further comprises determining a release profile kinetics of the polynucleotide agent from the polymeric biomaterial; and / or (c) the method further comprises selecting a polymer network biomaterial composition characterized in that less than 100% of the polynucleotide agent is released from the polymer biomaterial within 3 hours. The method of claim 7.

9. contacting the selected polymer network biomaterial composition with a population of cells; (a) whether the polynucleotide agent released from the candidate polymeric biomaterial is taken up by immune cells; and / or (b) the immune cells that take up the polynucleotide agent exhibit the following biological activity: (i) expressing said immunomodulatory polypeptide encoded by said polynucleotide agent; (ii) exhibiting increased expression of type 1 interferon in response to innate immune stimulation induced by the polynucleotide agent; and / or (iii) determining whether the immunomodulatory polypeptide exhibits at least one of: Optionally, (A) the immune cells comprise myeloid cells and / or plasmacytoid dendritic cells, or (B) the cells further comprise non-immune cells, e.g., the non-immune cells comprise fibroblasts and / or endothelial cells, e.g., the method further comprises determining whether the polynucleotide agent released from the candidate polymeric biomaterial is taken up by non-immune cells and / or whether the polynucleotide agent adversely affects the non-immune cells.

8. The method of claim 8(c).

10. 1. A method for characterizing a polymer network biomaterial composition or its component(s), said method comprising: (a) administering a polymer network composition to a target site in a mouse subject in vivo, wherein the polymer network biomaterial composition comprises a candidate polymer biomaterial and a polynucleotide agent that encodes or modulates an immunomodulatory polypeptide; (b) (i) whether the candidate polymeric biomaterial, upon administration, extends the release of the polynucleotide agent at the target site for at least 24 hours relative to administration of the same polynucleotide agent in a solution without the polymeric biomaterial; and / or (ii) determining whether the release profile of the polynucleotide agent from the candidate polymeric biomaterial is characterized by 50% or less of the polynucleotide agent being released in vivo 8 hours after administration. (a) the target site is a mammary fat pad; and / or (b) the method further comprises selecting a polymer network biomaterial composition characterized in that (i) the candidate polymeric biomaterial, upon administration, extends the release of the polynucleotide agent at the target site by at least 24 hours relative to administration of the same polynucleotide agent in a solution without the polymeric biomaterial, and / or (ii) no more than 50% of the polynucleotide agent is released from the candidate polymeric biomaterial in vivo 8 hours after the administration. The method of claim 10.

12. (a) whether the polynucleotide agent released from the candidate polymeric biomaterial is taken up by immune cells; and / or (b) the immune cells that take up the polynucleotide agent exhibit the following biological activity: (i) expressing said immunomodulatory polypeptide encoded by said polynucleotide agent; (ii) exhibiting increased expression of type 1 interferon in response to innate immune stimulation induced by the polynucleotide agent; and / or The method of claim 10 or claim 11, further comprising determining whether the antibody or antibody fragment exhibits at least one of the following: (i) exhibiting a change in the level and / or activity of the immunomodulatory polypeptide;

13. (c) whether cellular uptake of the polynucleotide agent released from the candidate polymeric biomaterial is delayed compared to contacting the cells with the polynucleotide agent in the absence of the candidate polymeric biomaterial; and / or The method of claim 12, further comprising (d) determining whether at least one of the biological activities induced by the polynucleotide agent is delayed compared to when the cells are contacted with the polynucleotide agent in the absence of the candidate polymeric biomaterial.

14. (a) the immune cells comprise myeloid cells and / or plasmacytoid dendritic cells, and / or (b) the method comprises: (e) whether the polynucleotide agent released from the candidate polymeric biomaterial is taken up by non-immune cells; and / or (f) determining whether the polynucleotide agent adversely affects the non-immune cells, e.g., the non-immune cells include fibroblasts and / or endothelial cells. The method of claim 12. (i) a polynucleotide agent carrier; and (ii) a polynucleotide agent that encodes or regulates an immunomodulatory polypeptide, for use in a method comprising administering the composition to a target site in a subject undergoing tumor resection, wherein the polynucleotide agent carrier optionally comprises a cationic agent and / or a lipid.

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