Compositions and methods for topical delivery of therapy to brain tumors

Abstract

Compositions for topical delivery of a drug to an intracranial region, such as brain tissue or brain tumor. Methods for local delivery of drugs or therapies to an intracranial region. The composition may include a hydrogel in which a chemotherapeutic drug or an immunotherapeutic drug is dispersed. Kits comprising a composition or solutions that can be combined to form a composition.

Description

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0002] This application claims priority to U.S. Provisional Patent Application No. 63 / 334,643, filed on April 25, 2022, which is incorporated herein by reference. Background Art

[0003] Current treatments for brain cancer patients, such as glioblastoma (GBM) patients, are limited for a variety of reasons that often result in a poor prognosis for the patients. From 1971 to 2011, the survival rate for brain cancer increased from about 8% to about 13%, while the average survival rate for all cancers increased from about 25% to 50% during the same time period.

[0004] Glioblastoma is the most common and aggressive primary central nervous system tumor, accounting for approximately 48% of all malignant brain tumors, with a median age of 65 years at diagnosis. Standard treatment for GBM includes surgical resection followed by concomitant chemotherapy and radiation. Surgery provides clinical relief, allows collection of diagnostic tissue samples, and improves survival rates. However, it is not possible to remove all cells because GBM has a diffuse infiltration pattern and resection must be balanced with the preservation of healthy tissue. Therefore, radiation therapy and chemotherapy are used to eliminate residual tumor cells after surgery, however, these techniques are limited by their inability to reach all tumor cells, their lack of specificity for tumor cells, and the ability of tumor cells to develop resistance to these therapies.

[0005] Approximately 95% of patients are diagnosed after the age of 40 (median age = 65 years). Primary GBM tumors arise de novo and account for 90% of cases, while secondary tumors arise from low-grade gliomas and account for 10% of all tumors. Patients with both primary and secondary tumors often present with symptoms of increased intracranial pressure, such as headaches, neurological deficits, seizures, etc. The diagnosis of GBM is based on the presence of several histological features, including dysplasia, mitotic activity, microvascular proliferation, and necrosis.

[0006] Current GBM therapy typically includes surgical resection, radiation therapy, and oral temozolomide chemotherapy, and this treatment regimen results in a median overall survival time of approximately 15 months.

[0007] The ability to achieve more effective therapies is limited by many factors, such as drug toxicity, drug delivery, and tumor biology. Toxicity from certain drugs may result in hypophysitis, uveitis and orbital inflammation, pneumonitis, adrenal insufficiency, enterocolitis, arthralgia, pancreatitis and autoimmune diabetes, rash and vitiligo, hepatitis, hypothyroidism, dry mouth, or a combination thereof.

[0008] It is believed that Wafers are the only FDA-approved products that directly address the challenges associated with therapeutic delivery to the brain, but they suffer from several drawbacks, including wafer migration, mechanical mismatch with soft tissue, rapid drug release, slow material degradation, and / or use of less effective chemotherapies that may be susceptible to resistance. Figure 1 The polyanhydride wafers (i.e., doxorubicin-loaded) were used to treat rats bearing 9L glioma. Delivery technology) treatments.

[0009] There remains a need for improved compositions and methods for treating brain cancers such as GBM, including compositions and methods for local delivery of various therapies to brain tumors that overcome one or more of the aforementioned disadvantages. Summary of the invention

[0010] Provided herein are methods and compositions that can effectively deliver therapeutic agents to brain tumors or brain tissues, such as hydrogels (e.g., adhesive hydrogels). In some embodiments, the composition includes a hydrogel loaded with spray-dispersed drugs. The compositions described herein can be injected during biopsy or through intraventricular shunts. The compositions and methods described herein can increase drug diffusion from the resection cavity to spread tumor lesions, provide an alternative therapy for temozolomide, have a safe product profile after treatment with radiotherapy, promote wound healing, and / or reduce or eliminate scar formation.

[0011] Embodiments of the compositions provided herein include hydrogels that can be soft and / or adhesive and can provide advantages over The compositions provided herein may provide a competitive advantage over other products such as wafers (Azity Pharmtics, USA). For example, the compositions provided herein may provide a higher hydrogel: tissue interface, improved stability at the excision site due to adhesion, mechanical stiffness equivalent to soft tissue, slower and / or more controlled drug delivery than wafers, and / or the potential for combined delivery. The compositions provided herein may also deliver doxorubicin, which may be more effective and less prone to resistance than current therapies, and / or have multiple mechanisms of action, including the ability to induce immunogenic cell death.

[0012] In one aspect, a drug delivery composition is provided. In some embodiments, the drug delivery composition includes a hydrogel and a drug, and the drug is dispersed in the hydrogel. The drug may include a chemotherapeutic drug, an immunotherapy drug, or a combination thereof. In some embodiments, the hydrogel includes a polymer component, wherein the polymer component includes a polymer having three or more aldehyde groups; and a dendritic polymer component, wherein the dendritic polymer component includes a dendritic polymer having at least 2 branches and one or more surface groups. The one or more surface groups can react reversibly or irreversibly with the three or more aldehyde groups.

[0013] In another aspect, a method for treating a patient is provided. In certain embodiments, the method comprises locally delivering the drug delivery composition described herein to the intracranial region of the patient. The local delivery of the drug delivery composition may include injecting or spraying the drug delivery composition in the intracranial region, which may include brain tissue, brain tumor, previously removed tumor site or a combination thereof.

[0014] In yet another aspect, a kit is provided. In some embodiments, the kit comprises a first part, the first part comprising a first solution, the first solution comprising a polymer component, wherein the polymer component comprises a polymer; and a second part, the second part comprising a second solution, the second solution comprising a dendritic polymer component, wherein the dendritic polymer component comprises a dendritic polymer having at least 2 branches and one or more surface groups, wherein at least one drug is disposed in the first solution, the second solution, or both the first solution and the second solution, and wherein the drug comprises a chemotherapeutic drug, an immunotherapy drug, or a combination thereof.

[0015] Additional aspects will be set forth in part in the following description and will become apparent in part from the description, or may be understood by practicing the various aspects described herein. The advantages described herein may be realized and obtained by the elements and combinations particularly noted in the appended claims. It should be understood that the foregoing general description and the following detailed description are merely exemplary and explanatory, and not restrictive. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 Percent survival for various treatments is depicted.

[0017] Figure 2 An embodiment of a multi-compartment syringe is depicted.

[0018] Figure 3 Plots of cell viability versus concentration for various treatments are depicted.

[0019] Figure 4 Degradation rates of examples of hydrogels are depicted.

[0020] Figure 5A , Figure 5B , Fig. 6A , Figure 6B , Fig. 7A , Figure 7B , Fig. 8A and Figure 8B Release profiles of examples of compositions are depicted.

[0021] Fig. 9 Cumulative release of doxorubicin from examples of the compositions is depicted.

[0022] Fig.10 Cumulative release of capped doxorubicin from examples of the compositions is depicted.

[0023] Fig.11 Cumulative release of doxorubicin and examples of encapsulated doxorubicin from examples of the compositions are depicted.

[0024] Fig. 12A and Fig. 12B Depicted are the drug release profiles in Examples from compositions with different drug loadings.

[0025] Fig.13 A graph depicting the average radiant efficiency of embodiments of the compositions.

[0026] Fig.14 A graph depicting the probability of survival of embodiments of the compositions.

[0027] Fig.15 Depicted are graphs of the probability of survival for examples of the composition and examples of empty hydrogels.

[0028] Fig.16 A graph depicting the probability of survival of embodiments of the compositions.

[0029] Fig.17 A graph depicting total flux of examples of compositions.

[0030] Fig.18 A graph depicting total flux of examples of compositions.

[0031] Fig.19 A graph depicting the percent survival of examples of compositions.

[0032] Fig. 20A and Fig. 20B Graphs depicting cell viability of examples of compositions.

[0033] Fig.21Depicted is a graph of the probability of survival following administration of examples of the compositions.

[0034] Fig. 22 Depicted is a graph of the probability of survival following administration of examples of the compositions.

[0035] Fig.23 Depicted is a graph of the probability of survival following administration of examples of the compositions.

[0036] Fig.24 Depicted is a graph of the probability of survival following administration of examples of the compositions.

[0037] Fig.25A Depicted is a graph of the probability of survival following administration of an Example of a composition containing 7.5 micrograms of a drug.

[0038] Fig.25B Depicted is a graph of the probability of survival following administration of an Example of a composition containing 15 micrograms of a drug.

[0039] Fig.25C Depicted is a graph of the probability of survival following administration of an Example of a composition containing 7.5 micrograms of a drug.

[0040] Fig.26 Depicted is a graph of percent survival following administration of examples of the compositions.

[0041] Fig. 27 Depicted is a graph of the probability of survival following administration of examples of the compositions.

[0042] Fig.28 Depicted is a graph of the probability of survival following administration of examples of the compositions.

[0043] Fig.29 Graph depicting total flux following injection of an example of a hydrogel.

[0044] Fig.30 Depicted are data collected from tests used to determine the effects of embodiments of compositions on T cells in lymph nodes.

[0045] Fig.31 Depicted are data collected from tests used to determine the effects of embodiments of compositions on central memory CD8 T cells in the spleen.

[0046] Fig.32 Depicted are data collected from tests to determine whether macrophages lead to an increase in CD45+ cells in tumors when treated with embodiments of the compositions.

[0047] Fig.33Depicted are the results of testing for M1 polarization markers in CD80+ / Macs 206 tumors treated with examples of the compositions.

[0048] Fig.34 Depicted are the results of a test configured to determine the translocation of a population of dendritic cells.

[0049] Fig.35 is a schematic diagram of an example of anti-cancer immune activity.

[0050] Fig.36 Schematic diagram of the possible behavior of nanoparticles in the cGAS-STING pathway.

[0051] Fig.37 is a schematic diagram of the possible mechanism of action.

[0052] Fig.38 Examples of nanoparticle-based STING agonist formulations are depicted.

[0053] Fig.39 Depicted are data demonstrating stimulation of dendritic cells and macrophages by embodiments of cyclic dinucleotide nanoparticles.

[0054] Fig.40A and Fig.40B The total release profile and the cumulative release profile of the examples of the compositions are depicted separately.

[0055] Fig.41 The results of a pilot study of local immunotherapy are depicted.

[0056] Fig.42 is a schematic diagram of treatment mechanisms and outcomes.

[0057] Fig.43A and Fig.43B Results regarding cell viability and CRT expression upon treatment with examples of the compositions are depicted, respectively.

[0058] Fig.44 is a schematic diagram of the research described in this article.

[0059] Fig.45 Depicted are the results of a study of BMDM CD86 activation marker expression.

[0060] Fig.46 Results are depicted, indicating that combination therapy increased the ratio of pro-inflammatory to anti-inflammatory BMDMs.

[0061] Fig.47 is a schematic diagram of an embodiment of a treatment plan. DETAILED DESCRIPTION

[0062] The drug delivery composition described herein may include a hydrogel and a drug. The drug may be selected from a chemotherapeutic drug, an immunotherapy drug, or a combination thereof. In certain embodiments, the drug includes a nucleic acid, a nanoparticle, an antibody, a small molecule, an immunomodulator, or a combination thereof. The hydrogel may include a hydrogel known in the art. Non-limiting examples of hydrogels are described in, for example, U.S. Patent No. 8,802,072 and U.S. Patent No. 10,736,914, which are incorporated herein by reference.

[0063] The drug delivery composition described herein may include a hydrogel and a drug, wherein the drug is dispersed in the hydrogel. The drug may be substantially uniformly dispersed in the hydrogel. In some embodiments, the drug is dispersed in the hydrogel in a manner that produces a concentration gradient of the drug.

[0064]

[0013] A hydrogel as described herein can include a polymer component, wherein the polymer component includes a polymer having three or more aldehyde groups; and a dendrimer component, wherein the dendrimer component includes a dendrimer having at least 2 branches and one or more surface groups.

[0065] drug

[0066] The drug of the drug delivery composition described herein may include a chemotherapeutic drug, an immunotherapy drug, or a combination thereof.

[0067] Chemotherapeutic drugs may include any known chemotherapeutic drugs, including but not limited to bleomycin, busulfan, carboplatin, carmustine, cisplatin, cladbrbine, dactinomycin, daunorubicin, doxorubicin, estramustine, interferon, irinotecan, levamisole, methotrexate, mitomycin, paclitaxel, pentostatin, plicamycin, tamoxifen, temozolomide, vinblastine, vindesine, etc. Additionally or alternatively, the chemotherapy drugs may include one or more radiosensitizers including 5-halo-uracil, anti-angiogenic compounds including thalidomide and tranilast, natural or synthetic peptide hormones including octreotide, and apoptosis-inducing compounds including butyrate. In some embodiments, the chemotherapy drugs include doxorubicin.

[0068] Immunotherapy drugs can include any known immunotherapy drugs. As used herein, the phrase immunotherapy drug refers to any drug that can induce, enhance, inhibit or otherwise change an immune response. In some embodiments, the immunotherapy drug is selected from drugs targeting (a) CTLA-4, such as ipilimumab or tremelimumab; drugs targeting (b) PD-1, such as nivolumab, pidilizumab or pembrolizumab, AMP-224; drugs targeting (c) PD-L1, such as MPDL-3280A, MSB0010718C or MEDI4736; or drugs targeting (d) GITR, such as TRX518 or MK4166. U.S. Patent No. 11,186,640 discloses non-limiting examples of immunotherapy drugs.

[0069] In some embodiments, immunotherapy drugs include cyclic dinucleotides. "Cyclic dinucleotides" or "CDNs" may include a class of molecules comprising 2'-5' and / or 3'-5' phosphodiester bonds between two purine nucleotides. This bond includes 2'-5'-2', 5', 2'-5'-3'5' and 3', 5'-3', 5' bonds. CDNs can activate cytoplasmic surveillance pathways through direct binding of two cytoplasmic pattern recognition receptors (PRRs) (i.e., DEAD (aspartate-glutamate-alanine-aspartate) box helicase 41 (DDX41) and interferon gene stimulator (STING)). The response of type I interferons to infections caused by intracellular bacteria may be caused by cyclic diadenosine monophosphate (cdAMP) or its related cyclic dinucleotides (CDNs), cyclic diguanine monophosphate (cdGMP). CDN can bind to DDX41 with high affinity and complex with the STING adapter protein, thereby causing activation of the TBK1 / IRF3 signaling pathway and induction of IFN-β and other IRF-3-dependent gene products that strongly activate innate immunity. CDN can include second messengers expressed by most bacteria and regulate various processes, including movement and biofilm formation. Endogenous CDN can also be produced by the host enzyme cyclic guanosine monophosphate-adenylate monophosphate synthase (cGAS) in response to cytoplasmic DNA in tumors or during infection. In some embodiments, CDN is a typical bacterial CDN, i.e., cyclic diguanine monophosphate (cdGMP). In some embodiments, CDN is an endogenous product of cGAS. In some embodiments, CDN is an agonist of STING.

[0070] Cyclic dinucleotides that can be used include those known in the art, such as U.S. Pat. Nos. 7,709,458 and 7,592,326; WO 2007 / 054279; U.S. Patent Application Publication No. 2014 / 0205653; and Yan et al., Bioorg. Med. Chem Lett., 18:5631 (2008). Non-limiting examples of cyclic dinucleotides include cdAMP, cdGMP, cdIMP, c-AMP-GMP, c-AMP-IMP, and c-GMP-IMP, and analogs thereof, including but not limited to phosphorothioate analogs.

[0071] In some embodiments, the cyclic dinucleotide is an agonist of STING (stimulator of interferon genes). As described herein, the STING signaling pathway in immune cells can be a central mediator of the innate immune response and, when stimulated, can induce the expression of amplification and enhanced immune activity of various interferons, cytokines, and T cell recruitment factors. Recent studies have shown that STING agonists can be effective adjuvants and efficiently induce immune responses, such as Dubensky, T. et al., Therapeutic Advances in Vaccines, Vol. 1 (4), 131-143 (2013); and Hanson, M. et al., The Journal of Clinical Investigation, Vol. 125 (6), 2532-2546 (2015). In some embodiments, the STING agonist is chemically synthesized. In some embodiments, the STING agonist is an analog of a naturally occurring cyclic dinucleotide. STING agonists (including analogs of cyclic dinucleotides) suitable for use in the present disclosure are provided in U.S. Patent Nos. 7,709,458 and 7,592,326 and U.S. Patent Application Publication No. 2014 / 0205653.

[0072] Drugs such as chemotherapeutic drugs and / or immunotherapy drugs (e.g., cyclic dinucleotides) can be encapsulated in a liquid. Encapsulation can occur before the drug is dispersed in the hydrogel. For example, a chemotherapeutic drug can be encapsulated in a liquid, an immunotherapy drug (e.g., cyclic dinucleotides) can be encapsulated in a liquid, or a chemotherapeutic drug and an immunotherapy drug (e.g., cyclic dinucleotides) can be encapsulated together in the same liquid or separately in different liquids. The encapsulated drug can be in the form of liquid nanoparticles (e.g., nanospheres), such as any liquid nanoparticles known in the art. U.S. Patent No. 11,207,418 discloses non-limiting examples of encapsulated liquid nanoparticles, which are incorporated by reference. In some embodiments, the chemotherapeutic drug is doxorubicin, and doxorubicin can be encapsulated in liquid particles (e.g., liquid spheres). Encapsulated doxorubicin can be Anthracycline topoisomerase inhibitors (Baxter, USA). The nanoparticles may have any suitable average diameter, such as about 1 nm to about 100 nm, about 10 nm to about 90 nm, about 20 nm to about 80 nm, about 30 nm to about 70 nm, or about 40 nm to about 60 nm.

[0073] The drugs described herein may be present in any amount in the drug delivery composition. In some embodiments, the drug is present in a total amount of about 1 μg to about 5000 μg, about 1 μg to about 1000 μg, about 1 μg to about 500 μg, about 50 μg to about 500 μg, or about 50 μg to about 500 μg in the drug delivery composition. For example, a composition including a "total amount" of 500 μg of a drug may include 500 μg of a chemotherapy drug, 500 μg of an immunotherapy drug, 400 μg of a chemotherapy drug, and 100 μg of an immunotherapy drug; etc. The drugs described herein may be present in the drug delivery composition at any concentration. In some embodiments, the drug is present in the drug delivery composition at a total concentration of about 0.1 μg / μL to about 100 μg / μL, about 1 μg / μL to about 80 μg / μL, about 1 μg / μL to about 60 μg / μL, about 1 μg / μL to about 40 μg / μL, about 1 μg / μL to about 30 μg / μL, about 1 μg / μL to about 20 μg / μL, about 1 μg / μL to about 10 μg / μL, about 2 μg / μL to about 8 μg / μL, or about 2 μg / μL to about 5 μg / μL.

[0074] Treatment

[0075] Provided herein are methods for treating patients, including patients with brain tumors. The patient can be any mammal, such as a human. In some embodiments, the method for treating a patient comprises locally delivering a drug delivery composition as described herein to an intracranial region of the patient.

[0076] Local delivery of the drug delivery composition can be achieved in any manner. In some embodiments, local delivery of the drug delivery composition comprises injection or spraying of the drug delivery composition. Injection of the drug delivery composition can occur during a biopsy, through an intraventricular shunt, or a combination thereof.

[0077] The intracranial region may include brain tissue, a brain tumor, the site of a previously resected tumor, or a combination thereof.

[0078] In some embodiments, the method also includes administering a second drug or therapy to the patient before, during and / or after the local delivery of the drug delivery composition. The administration of the second drug or therapy can be achieved in any way, such as oral administration of the second drug or therapy. The second drug can include a second chemotherapy drug or therapy, such as temozolomide or PD1 checkpoint blocking therapy.

[0079] Hydrogels as described herein can provide controlled drug release, such as sustained drug release. The components of the hydrogel can be customized to achieve a desired release profile. The drug can be released continuously or intermittently. In certain embodiments, the drug is continuously released from the drug delivery composition for at least 12 hours, at least 24 hours, at least 36 hours, at least 48 hours, at least 72 hours, at least 100 hours, at least 200 hours, or at least 300 hours.

[0080] In some embodiments, the cumulative percentage of drug released from the drug delivery composition is at least 80%, 85%, or 90% by weight of the drug within 10 days or less, 12 days or less, 14 days or less, 16 days or less, 18 days or less, 20 days or less, 22 days or less, 24 days or less, 26 days or less, 28 days or less, 30 days or less, 32 days or less, 34 days or less, 36 days or less, 38 days or less, 40 days or less, 50 days or less, 60 days or less, 80 days or less, or 100 days or less after local delivery of the drug delivery composition.

[0081] Dendrimer components

[0082] In some embodiments, the dendritic polymer component comprises a dendritic polymer having amines on at least a portion of its surface groups, which are often referred to as "terminal groups" or "end groups". The dendritic polymer may have amines on 20% to 100% of its surface groups. In some embodiments, the dendritic polymer has amines on 100% of its surface groups. In some embodiments, the dendritic polymer component comprises a dendritic polymer having amines on less than 75% of its surface groups. As used herein, the term "dendritic polymer" refers to any compound having a multivalent core covalently bonded to two or more dendritic branches. In some embodiments, the multivalent core is covalently bonded to three or more dendritic branches. In some embodiments, the amine is a primary amine. In some embodiments, the amine is a secondary amine. In some embodiments, one or more surface groups have at least one primary amine and at least one secondary amine.

[0083] In some embodiments, the dendrimer extends to at least 2 passages. In some embodiments, the dendrimer extends to at least 3 passages. In some embodiments, the dendrimer extends to at least 4 passages. In some embodiments, the dendrimer extends to at least 5 passages. In some embodiments, the dendrimer extends to at least 6 passages. In some embodiments, the dendrimer extends to at least 7 passages.

[0084] In some embodiments, the molecular weight of the dendritic polymer is from about 1,000 Daltons to about 1,000,000 Daltons. In some embodiments, the molecular weight of the dendritic polymer is from about 3,000 Daltons to about 120,000 Daltons. In some embodiments, the molecular weight of the dendritic polymer is from about 10,000 Daltons to about 100,000 Daltons. In some embodiments, the molecular weight of the dendritic polymer is from about 20,000 Daltons to about 40,000 Daltons. Unless otherwise indicated, the "molecular weight" of a dendritic polymer refers to the weight average molecular weight.

[0085] In general, dendritic polymers can be prepared using any known method. In some embodiments, the dendritic polymer is prepared by oxidizing a starting dendritic polymer having surface groups comprising at least one hydroxyl group such that at least a portion of the surface groups comprise at least one amine. In some embodiments, the dendritic polymer is prepared by oxidizing a starting 5th generation (G5) dendritic polymer having surface groups comprising at least one hydroxyl group such that at least a portion of the surface groups comprise at least one amine. In some embodiments, the dendritic polymer is prepared by oxidizing a starting G5 dendritic polymer having surface groups comprising at least one hydroxyl group such that about 25% of the surface groups comprise at least one amine. In some embodiments, the dendritic polymer is a G5 dendritic polymer having primary amines on about 25% of the surface groups of the dendritic polymer.

[0086] In some embodiments, the dendrimer is a poly(aminoamine) derived (PAMAM) dendrimer. In some embodiments, the dendrimer is a G5 PAMAM derived dendrimer. In some embodiments, the dendrimer is a G5 PAMAM derived dendrimer having primary amines on about 25% of the surface groups of the dendrimer.

[0087] In some embodiments, the dendrimer is a poly(propyleneimine) derived dendrimer. In some embodiments, the dendrimer component is combined with a liquid to form a dendrimer component solution. In some embodiments, the dendrimer component solution is an aqueous solution. In some embodiments, the solution comprises water, phosphate buffered saline (PBS), Dulbecco's Modified Eagle's Medium (DMEM), or any combination thereof. In some embodiments, the dendrimer component concentration in the dendrimer component solution is about 5 wt % to about 25 wt %. In some embodiments, the dendrimer component concentration in the dendrimer component solution is about 10 wt % to about 20 wt %. In some embodiments, the dendrimer component concentration in the dendrimer component solution is about 11 wt % to about 15 wt %.

[0088] In some embodiments, the dendritic polymer component or dendritic polymer component solution further includes one or more additives. Generally, the amount of the additive can vary depending on the application, tissue type, concentration of the dendritic polymer component solution, type of dendritic polymer component, concentration of the polymer component solution, and / or type of polymer component. Examples of suitable additives include, but are not limited to, pH adjusters, thickeners, antimicrobial agents, colorants, surfactants, and radiopaque compounds. Specific examples of these types of additives are described herein. In one embodiment, the dendritic polymer component solution includes a foaming additive.

[0089] Polymer components

[0090] Typically, the polymer component comprises a polymer and / or oligomer having one or more functional groups that are capable of reacting with one or more functional groups on biological tissue and / or one or more functional groups on the dendrimer component.

[0091] In some embodiments, the polymer is at least one polysaccharide. In some embodiments, at least one polysaccharide can be linear, branched, or have linear segments and branched segments in its structure. Typically, at least one polysaccharide can be natural, synthetic or modified, such as by cross-linking, changing the substituents of the polysaccharide, or both. In one embodiment, at least one polysaccharide is plant-based. In some embodiments, at least one polysaccharide is animal-based. In some embodiments, at least one polysaccharide is a combination of plant-based polysaccharides and animal-based polysaccharides. Non-limiting examples of polysaccharides include, but are not limited to, dextran, chitin, starch, agar, cellulose, hyaluronic acid, or a combination thereof.

[0092] In some embodiments, at least one polymer has a molecular weight of about 1,000 Daltons to about 1,000,000 Daltons. In some embodiments, at least one polymer has a molecular weight of about 5,000 Daltons to about 15,000 Daltons. Unless otherwise indicated, "molecular weight" of a polymer refers to weight average molecular weight.

[0093] In some embodiments, the polymer is functionalized such that its structure includes one or more functional groups that will react with one or more functional groups on biological tissue and / or one or more functional groups on a dendrimer component. In some embodiments, the polymer is functionalized such that its structure includes three or more functional groups that will react with one or more functional groups on biological tissue and / or one or more functional groups on a dendrimer component. In some embodiments, the functional group incorporated into the structure of the polymer is an aldehyde.

[0094] In some embodiments, the functionalization degree of the polymer is adjustable. "Functionalization degree" generally refers to the number or percentage of groups on the polymer that are replaced or converted into the desired one or more functional groups. In a specific embodiment, one or more functional groups include aldehydes, substituents capable of photoreversible dimerization, or a combination thereof. In some embodiments, the functionalization degree is regulated based on the type of tissue to which the adhesive is applied, the concentration of the components, and / or the type of polymer or dendrimer used in the adhesive. In some embodiments, the functionalization degree is about 10% to about 75%. In some embodiments, the functionalization degree is about 15% to about 50%. In some embodiments, the functionalization degree is about 20% to about 30%.

[0095] In some embodiments, the polymer is a dextran having a molecular weight of about 10 kDa. In some embodiments, the polymer is a dextran having about 50% of its hydroxyl groups converted to aldehydes. In some embodiments, the polymer is a dextran having a molecular weight of about 10 kDa and about 50% of its hydroxyl groups converted to aldehydes.

[0096] In certain embodiments, polysaccharide is oxidized to include one or more aldehyde functional groups of desired percentage. For example, polysaccharide can be oxidized to convert about 10% to about 100%, about 10% to about 90%, about 20% to about 80%, about 30% to about 70%, about 40% to about 60% or about 50% (mol %) of its hydroxyl group into aldehyde. Usually, this oxidation can be carried out using any known means. For example, suitable oxidizing agents include but are not limited to periodate, hypochlorite, ozone, peroxide, hydroperoxide, persulfate and percarbonate. In one embodiment, oxidation is carried out using sodium periodate. Usually, different amounts of oxidizing agents can be used to change the degree of functionalization.

[0097] In some embodiments, the polymer component is combined with a liquid to form a polymer component solution. In some embodiments, the polymer component solution is an aqueous solution. In some embodiments, the solution comprises water, PBS, DMEM, or any combination thereof.

[0098] In general, the polymer component solution can have any suitable concentration of polymer components. In some embodiments, the polymer component concentration in the polymer component solution is from about 5% to about 40% by weight. In some embodiments, the polymer component concentration in the polymer component solution is from about 5% to about 30% by weight. In some embodiments, the polymer component concentration in the polymer component solution is from about 5% to about 25% by weight. In general, the concentration can be customized and / or adjusted based on the specific application, tissue type, and / or the type and concentration of the dendritic polymer component used.

[0099] The polymer component or polymer component solution may also include one or more additives. In some embodiments, the additive is compatible with the polymer component. In some embodiments, the additive does not contain primary or secondary amines. Typically, the amount of the additive varies depending on the application, tissue type, concentration of the polymer component solution, type of polymer component and / or dendrimer component. Examples of suitable additives include, but are not limited to, pH regulators, thickeners, antimicrobial agents, colorants, surfactants, radiopaque compounds, and other additives described herein. In other embodiments, the polymer component solution includes a foaming agent.

[0100] In some embodiments, the pH adjusting agent is an acidic compound. Examples of acidic pH adjusting agents include, but are not limited to, carboxylic acids, inorganic acids, and sulfonic acids. In some embodiments, the pH adjusting agent is a basic compound. Examples of basic pH adjusting agents include, but are not limited to, hydroxides, alkoxides, nitrogen-containing compounds other than primary and secondary amines, basic carbonates, and basic phosphates.

[0101] Generally, the thickener may be selected from any known viscosity adjusting compound including, but not limited to, polysaccharides and their derivatives, such as starch or hydroxyethylcellulose.

[0102] Typically, the surfactant can be any compound that reduces the surface tension of water. In one embodiment, the surfactant is an ionic surfactant, for example, sodium lauryl sulfate. In another embodiment, the surfactant is a neutral surfactant. Examples of neutral surfactants include, but are not limited to, polyoxyethylene ether, polyoxyethylene esters, and polyoxyethylene sorbitan.

[0103] In some embodiments, the radiopaque compound is barium sulfate, gold particles, or a combination thereof.

[0104] Hydrogel

[0105] In general, the hydrogels described herein can be formed by combining a polymer component or a polymer component solution with a dendritic polymer component or a dendritic polymer component solution in any manner. In some embodiments, the polymer component or polymer component solution is combined with the dendritic polymer component or a dendritic polymer component solution prior to local delivery of the drug delivery composition. In some embodiments, the polymer component or polymer component solution is combined with the dendritic polymer component or a dendritic polymer component solution in any order in the intracranial region. In some embodiments, local delivery of the composition comprises local delivery of the polymer component or polymer component solution to the intracranial region, and local delivery of the dendritic polymer component or a dendritic polymer component solution to the intracranial region, wherein the polymer component and the dendritic polymer component are in contact with each other in the intracranial region.

[0106] The hydrogel may be delivered topically using any suitable means and methods.Dual barrel syringes with rigid or flexible discharge tips and optional extension tubes, needles, stents, catheters and other devices known in the art are contemplated.

[0107] As used herein, a hydrogel is "therapeutic" when it prevents, reverses, or reduces the rate of tumor growth, improves the probability of survival, prolongs the life of a patient, or a combination thereof.

[0108] After contacting one or more biological tissues in the intracranial region, the hydrogel can be allowed sufficient time to solidify or gel. When the hydrogel "solidifies" or "gels", as the term is used herein, the hydrogel means that the reactive groups on the polymer component, the dendrimer component, and the one or more biological tissues have undergone one or more reactions. Without wishing to be bound by any particular theory, it is believed that the hydrogels described herein are effective because the polymer component reacts with both the surface of the dendrimer component and the biological tissue. In some embodiments, the aldehyde functional group of the polymer component reacts with one or more amines in (i) the drug (such as doxorubicin), (ii) the dendrimer component and / or (iii) one or more biological tissues to form an imine bond. The formation of the imine bond can be reversible, and the formation and hydrolysis of the imine bond can control or promote the release kinetics of the drug from the hydrogel; therefore, the number / ratio of functional groups involved in the imine bond formation can be configured to achieve the desired release kinetics. In some embodiments, it is believed that the amines on the dendrimer component react with a high percentage of aldehydes on the polymer component, thereby reducing toxicity and improving the biocompatibility of the hydrogel. In general, the time required for the hydrogel to solidify or gel will vary based on many factors, including but not limited to the properties of the polymer component and / or the dendritic polymer component, the concentration of the polymer component solution and / or the dendritic polymer component solution, and the properties of one or more biological tissues. In some embodiments, the hydrogel will be sufficiently solidified to provide the desired bonding or sealing shortly after the components are combined. The gelation or solidification time should be such that the mixture of components can be delivered to the target area in a fluid form before becoming too viscous or solidifying, and then quickly set after being applied to the target area. In one embodiment, the gelation or solidification time is less than 120 seconds. In some embodiments, the gelation or solidification time is between 3 seconds and 60 seconds. In some embodiments, the gelation or solidification time is between 5 seconds and 30 seconds. Before or after the hydrogel solidifies, the substituents capable of reversible dimerization can be activated or deactivated as needed.

[0109] In certain embodiments, one or more blowing agents are added to the polymer component solution and / or the dendrimer component solution prior to combining the solutions. In one embodiment, the blowing agent comprises a two-part liquid system including part 1 and part 2, wherein part 1 comprises a bicarbonate salt and part 2 comprises an aqueous solution of a dialdehyde or polyaldehyde and a titrant. There is a wide range of dialdehydes or polyaldehydes, and their usefulness is largely limited by availability and solubility in water. For example, aqueous glyoxal / ethanedial is useful, as is aqueous glutaraldehyde / pentadial. Water-soluble mixtures of dialdehydes and polyaldehydes prepared by oxidative cleavage of appropriate carbohydrates with periodates, ozone, etc. are also useful.

[0110] The titrant is most preferably used for the liquid solution of Part 2. More specifically, the titrant is an organic or inorganic acid, buffer, salt or salt solution that is capable of reacting with the bicarbonate component of Part 1 to produce carbon dioxide and water as reaction byproducts. The generated carbon dioxide gas produces a foamy structure of the hydrogel and also expands the volume of the hydrogel.

[0111] Most preferably, the titrant is an inorganic or organic acid present in an amount to impart an acidic pH to the resulting mixture of the components of Part 1 and Part 2. Preferred acids that may be used in the practice of the present invention include phosphoric acid, sulfuric acid, hydrochloric acid, acetic acid, and citric acid.

[0112] Hydrogel Kit

[0113] In another aspect, a kit is provided, comprising a first part comprising a polymer component or a polymer component solution and a second part comprising a dendrimer component or a dendrimer component solution. The kit may further comprise an applicator or other device means, such as a multi-compartment syringe for storing, combining the two parts and / or the resulting hydrogel, and delivering the two parts and / or the resulting hydrogel to the intracranial region.

[0114] As described herein, at least one drug may be disposed in the polymer component solution, the dendrimer component solution, or both the polymer component solution and the dendrimer component solution.As described herein, the drug may include a chemotherapeutic drug, an immunotherapy drug, or a combination thereof.

[0115] In some embodiments, the kit comprises separate reservoirs for the polymer component solution and the dendrimer component solution. In some embodiments, the kit comprises reservoirs for polymer component solutions of varying concentrations. In some embodiments, the kit comprises reservoirs for dendrimer component solutions of varying concentrations.

[0116] In some embodiments, the kit comprises instructions for selecting an appropriate concentration or amount of at least one of a polymer component, a polymer component solution, a dendrimer component, a dendrimer component solution, or a drug to compensate for or address at least one property of one or more biological tissues, treatment plans, and the like.

[0117] In some embodiments, the kit comprises at least one syringe. In some embodiments, the syringe comprises separate reservoirs for the polymer component solution and the dendrimer component solution. The syringe may also comprise a mixing tip that combines the two solutions when the plunger is pressed. The mixing tip may be releasably fixed to the syringe (to enable exchange of the mixing tip), and the mixing tip may comprise a static mixer. In some embodiments, the reservoirs in the syringe may have different sizes or may hold solutions of different volumes. In some embodiments, the reservoirs in the syringe may have the same size or may hold solutions of the same volume. In some embodiments, one reservoir may comprise part 1 of the foaming composition described above, and a second reservoir may comprise part 2 of the foaming composition.

[0118] Figure 2 One embodiment of a syringe 100 is depicted. The syringe 100 includes a body 110 having two reservoirs (130, 140). A dendrimer component solution is disposed in a first reservoir 130, and a polymer component solution is disposed in a second reservoir 140. Both reservoirs (130, 140) are emptied by depressing a plunger 120, which pushes the contents of both reservoirs (130, 140) into a mixing tip 150 and out of the syringe 100.

[0119] In a further embodiment, one or more of the reservoirs of the syringe may be removable. In this embodiment, the removable reservoir may be replaced with a reservoir containing a polymer component solution or a dendrimer component solution at a desired concentration.

[0120] In a preferred embodiment, the kit is sterile. For example, the components of the kit can be packaged together, such as in a tray, pouch, and / or box. The packaged kit can be sterilized using known techniques such as electron beam radiation, gamma radiation, ethylene oxide sterilization, etc., at a suitable wavelength (where applicable), or other suitable techniques.

[0121] All cited publications are incorporated herein by reference in their entirety. In addition, where a definition or use of a term in a reference incorporated herein by reference is inconsistent or contrary to the definition of the term provided herein, the definition of the term provided herein applies, and the definition of the term in the reference does not apply.

[0122] Although certain aspects of conventional technology have been discussed to facilitate the disclosure of various embodiments, applicants in no way disclaim these aspects and contemplate that the present disclosure may encompass one or more of the conventional technology aspects discussed herein.

[0123] The present disclosure can solve one or more of the problems and defects of known methods and processes. However, it is contemplated that various embodiments may prove useful in solving other problems and defects in many technical fields. Therefore, the present disclosure should not necessarily be interpreted as being limited to solving any of the specific problems or defects discussed herein.

[0124] Where documents, acts or items of knowledge are referred to or discussed in this specification, such reference or discussion does not constitute an admission that the documents, acts or items of knowledge, or any combination thereof, were, at the priority date, publicly available, known to the public, part of the common general knowledge, or otherwise constituted prior art under applicable legal provisions; or were known to be relevant to attempts to solve any problem addressed in this specification.

[0125] In the description provided herein, the terms "comprising," "is," "containing," "having," and "including" are used in an open-ended manner, and thus should be interpreted to mean "including, but not limited to." When a composition, kit, or method is claimed or described in terms of "comprising" various steps or components, the composition, kit, or method may also "consist essentially of" or "consist of" the various steps or components, unless otherwise specified.

[0126] The terms "a," "an," "the," and the like are intended to include plural alternatives, e.g., at least one. For example, unless otherwise indicated, disclosure of "a polymer," "a drug," and the like is intended to encompass a mixture or combination of one polymer, drug, and the like or more than one polymer, drug, and the like.

[0127] Each numerical range may be disclosed herein. Unless otherwise specified, when the applicant discloses or claims any type of range, the applicant's intention is to disclose or claim individually each possible number that such range can reasonably cover, including the endpoints of the range and any sub-ranges and sub-range combinations covered therein. In addition, all numerical endpoints of the range disclosed herein are approximate values. As a representative example, in some embodiments, the applicant discloses that the drug is present in the drug delivery composition at a total concentration of about 1 μg / μL to about 10 μg / μL. This range should be interpreted as covering about 1 μg / μL and about 10 μg / μL, and further covering "about" 2 μg / μL, 3 μg / μL, 4 μg / μL, 5 μg / μL, 6 μg / μL, 7 μg / μL, 8 μg / μL and 9 μg / μL, including any ranges and sub-ranges between these values.

[0128] As used herein, the term "about" means ±10% of the numerical value of the number being used.

[0129] List of Examples

[0130] The following is a non-limiting list of embodiments of the present disclosure:

[0131] Example 1: A drug delivery composition comprising a hydrogel such as an adhesive hydrogel and a drug dispersed in the hydrogel, wherein the drug comprises a chemotherapeutic drug, an immunotherapy drug, or a combination thereof.

[0132] Embodiment 2: The composition of embodiment 1, wherein the hydrogel comprises a polymer component, such as a polymer component comprising a polymer having three or more aldehyde groups.

[0133] Embodiment 3: The composition of any preceding embodiment, wherein the hydrogel comprises a dendritic polymer component, such as a dendritic polymer component comprising a dendritic polymer having at least 2 branches and one or more surface groups.

[0134] Embodiment 4: The composition of any preceding embodiment, wherein 100% of the one or more surface groups comprise at least one primary or secondary amine.

[0135] Embodiment 5: The composition of any preceding embodiment, wherein less than 95%, less than 90%, less than 85%, less than 80%, or less than 75% of the one or more surface groups comprise at least one primary or secondary amine.

[0136] Embodiment 6: The composition of any of the preceding embodiments, wherein the dendritic polymer is a Generation 5 polyamidoamine (G5 PAMAM) dendritic polymer.

[0137] Embodiment 7: The composition of any preceding embodiment, wherein the polymer having three or more aldehyde groups comprises a polysaccharide, such as an oxidized polysaccharide.

[0138] Embodiment 8: A composition according to any of the preceding embodiments, wherein about 10% to about 100%, about 10% to about 90%, about 20% to about 80%, about 30% to about 70%, about 40% to about 60%, or about 50% (molar %) of the hydroxyl groups of the polysaccharide are oxidized to aldehydes.

[0139] Embodiment 9: The composition according to any of the preceding embodiments, wherein the polymer comprises dextran.

[0140] Embodiment 10: The composition according to any one of the preceding embodiments, wherein the drug comprises or consists of the chemotherapeutic drug.

[0141] Embodiment 11: The composition according to any of the preceding embodiments, wherein the chemotherapy drug comprises bleomycin, busulfan, carboplatin, carmustine, cisplatin, cladribine, dactinomycin, daunorubicin, doxorubicin, estramustine, interferon, irinotecan, levamisole, methotrexate, mitomycin, paclitaxel, pentostatin, plicamycin, tamoxifen, temozolomide, vinblastine, vindesine or a combination thereof.

[0142] Embodiment 12: A composition according to any of the preceding embodiments, wherein the chemotherapy drug comprises one or more radiosensitizers (such as 5-halo-uracil), anti-angiogenic compounds (such as thalidomide, tranilast, natural or synthetic peptide hormones including octreotide and / or apoptosis-inducing compounds including butyrate).

[0143] Embodiment 13: The composition according to any of the preceding embodiments, wherein the chemotherapy drug comprises or consists of doxorubicin.

[0144] Embodiment 14: A composition according to any of the preceding embodiments, wherein the immunotherapy drug comprises one or more agents targeting (a) CTLA-4, such as ipilimumab or tremelimumab; agents targeting (b) PD-1, such as nivolumab, pidilizumab or pembrolizumab, AMP-224; agents targeting (c) PD-L1, such as MPDL-3280A, MSB0010718C or MEDI4736; agents targeting (d) GITR, such as TRX518 or MK4166; or drugs targeting (e) or a combination thereof.

[0145] Embodiment 15: The composition according to any of the preceding embodiments, wherein the immunotherapy drug comprises or consists of a cyclic dinucleotide.

[0146] Embodiment 16: A composition according to any one of the preceding embodiments, wherein the chemotherapy drug, the immunotherapy drug, or the chemotherapy drug and the immunotherapy drug are encapsulated in a liquid, wherein the encapsulation can form liquid particles (e.g., liquid nanoparticles), such as liquid spheres (e.g., liquid nanospheres).

[0147] Embodiment 17: The composition according to any one of the preceding embodiments, wherein the chemotherapeutic drug and the immunotherapy drug are encapsulated together in the same liquid particles, or the chemotherapeutic drug and the immunotherapy drug are separately encapsulated in different liquid particles.

[0148] Embodiment 18: The composition of any preceding embodiment, wherein the liquid particles such as liquid nanoparticles comprise poly-β-amino-ester.

[0149] Embodiment 19: The composition of embodiment 18, wherein the cyclic dinucleotide is conjugated to the poly-β-amino-ester via a cathepsin-sensitive bond, and wherein the poly-β-amino-ester is optionally modified with arginine.

[0150] Embodiment 20: A composition according to any of the preceding embodiments, wherein the liquid particles are liquid nanoparticles (e.g., liquid nanospheres) having an average diameter of about 1 nm to about 100 nm, about 10 nm to about 90 nm, about 20 nm to about 80 nm, about 30 nm to about 70 nm, or about 40 nm to about 60 nm.

[0151] Embodiment 21: The composition of any preceding embodiment, wherein the hydrogel comprises phosphate buffered saline (PBS).

[0152] Embodiment 22: A composition according to any of the preceding embodiments, wherein the solids content of the hydrogel is from about 5 wt % to about 40 wt %, from about 5 wt % to about 35 wt %, from about 5 wt % to about 30 wt %, from about 5 wt % to about 25 wt %, from about 8 wt % to about 25 wt %, from about 10 wt % to about 20 wt % or from about 12 wt % to about 18 wt %.

[0153] Embodiment 23: A composition according to any of the preceding embodiments, wherein the drug is present in the drug delivery composition in a total amount of about 1 μg to about 5000 μg, about 1 μg to about 1000 μg, about 1 μg to about 500 μg, about 50 μg to about 500 μg, or about 50 μg to about 500 μg.

[0154] Embodiment 24: A composition according to any one of the preceding embodiments, wherein

[0155] The drug is present in the drug delivery composition at a total concentration of about 0.1 μg / μL to about 100 μg / μL, about 1 μg / μL to about 80 μg / μL, about 1 μg / μL to about 60 μg / μL, about 1 μg / μL to about 40 μg / μL, about 1 μg / μL to about 30 μg / μL, about 1 μg / μL to about 20 μg / μL, about 1 μg / μL to about 10 μg / μL, about 2 μg / μL to about 8 μg / μL, or about 2 μg / μL to about 5 μg / μL.

[0156] Embodiment 25: A method of treating a patient, the method comprising: locally delivering the drug delivery composition according to any one of the preceding embodiments to the patient, such as to the intracranial region of the patient, wherein the patient is optionally a human.

[0157] Embodiment 26: A method according to any one of the preceding embodiments, wherein -

[0158] (A) the local delivery of the drug delivery composition comprises injecting or spraying the drug delivery composition; and / or

[0159] (B) contacting the polymer component / solution and the dendrimer component / solution prior to topically delivering the composition to the intracranial region; and / or

[0160] (C) said local delivery of said drug delivery composition comprises local delivery of said polymer component / solution to said intracranial region and local delivery of said dendrimer component / solution to said intracranial region in any order, sequentially, at least partially simultaneously or a combination thereof.

[0161] Embodiment 27: The method according to any one of the preceding embodiments, wherein said injection of said drug delivery composition occurs during a biopsy, through an intraventricular shunt, or a combination thereof.

[0162] Embodiment 28: The method according to any one of the preceding embodiments, wherein the intracranial region comprises brain tissue, a brain tumor, a site of a previously resected tumor, or a combination thereof.

[0163] Embodiment 29: The method according to any of the preceding embodiments, further comprising administering a second drug or therapy to the patient before, during and / or after the local delivery of the drug delivery composition.

[0164] Embodiment 30: The method according to any one of the preceding embodiments, wherein said administering of said second drug or therapy comprises orally administering said second drug or therapy.

[0165] Embodiment 31: A method according to any of the preceding embodiments, wherein the second drug or therapy comprises a second chemotherapy drug or therapy, which includes but is not limited to any chemotherapy drug or therapy disclosed herein, such as any chemotherapy drug or therapy of Embodiment 11.

[0166] Embodiment 32: The method according to any of the preceding embodiments, wherein the second chemotherapeutic drug or therapy is not present in the drug delivery composition.

[0167] Embodiment 33: The method according to any of the preceding embodiments, wherein the second chemotherapy drug or treatment comprises or consists of temozolomide.

[0168] Embodiment 34: The method according to any of the preceding embodiments, wherein the second drug or therapy comprises an immunotherapy drug, such as any immunotherapy drug disclosed herein, such as any immunotherapy drug of Embodiment 14 or 15.

[0169] Embodiment 35: The method according to any one of the preceding embodiments, wherein the second drug or therapy comprises aPD1 checkpoint blockade therapy.

[0170] Embodiment 36: A method according to any of the preceding embodiments, wherein the drug is continuously released from the drug delivery composition for at least 12 hours, at least 24 hours, at least 36 hours, at least 48 hours, at least 72 hours, at least 100 hours, at least 200 hours or at least 300 hours.

[0171] Embodiment 37: The method of any of the preceding embodiments, wherein the cumulative percentage of the drug released from the drug delivery composition is at least 80 wt % within 10 days or less, 12 days or less, 14 days or less, 16 days or less, 18 days or less, 20 days or less, 22 days or less, 24 days or less, 26 days or less, 28 days or less, 30 days or less, 32 days or less, 34 days or less, 36 days or less, 38 days or less, 40 days or less, 50 days or less, 60 days or less, 80 days or less, or 100 days or less after the local delivery of the drug delivery composition.

[0172] Embodiment 38: The method of any of the preceding embodiments, wherein the cumulative percentage of the drug released from the drug delivery composition is at least 90 weight percent within 10 days or less, 12 days or less, 14 days or less, 16 days or less, 18 days or less, 20 days or less, 22 days or less, 24 days or less, 26 days or less, 28 days or less, 30 days or less, 32 days or less, 34 days or less, 36 days or less, 38 days or less, 40 days or less, 50 days or less, 60 days or less, 80 days or less, or 100 days or less after the local delivery of the drug delivery composition.

[0173] Embodiment 39: A kit for preparing a drug delivery composition, the kit comprising: a first part, the first part comprising a first solution, the first solution comprising the polymer component of any one of the preceding embodiments; and a second part, the second part comprising a second solution, the second part comprising the dendritic polymer component of any one of the preceding embodiments.

[0174] Embodiment 40: The kit according to any one of the preceding embodiments, wherein at least one drug is disposed in the first solution, the second solution, or both the first solution and the second solution.

[0175] Embodiment 41: A kit according to Embodiment 40, wherein (i) the drug comprises or consists of a chemotherapeutic drug, and the chemotherapeutic drug is disposed in the first solution, the second solution, or both the first solution and the second solution, (ii) the drug comprises or consists of an immunotherapy drug, and the immunotherapy drug is disposed in the first solution, the second solution, or both the first solution and the second solution, or (iii) the drug comprises or consists of a chemotherapeutic drug and an immunotherapy drug, and the chemotherapeutic drug and the immunotherapy drug are both disposed in the first solution, both disposed in the second solution, both disposed in the first solution and the second solution, or the chemotherapeutic drug is disposed in one solution of the first solution and the second solution, and the immunotherapy drug is disposed in the other solution.

[0176] Embodiment 42: The kit according to any one of the preceding embodiments, wherein the kit comprises a syringe.

[0177] Embodiment 43: A kit according to any one of the preceding embodiments, wherein the syringe comprises a mixing tip such as Figure 2 Mixed tip depicted at the premises.

[0178] Embodiment 44: The kit according to any one of the preceding embodiments, wherein the syringe comprises a first reservoir and a second reservoir, wherein the first solution and the second solution are disposed in the first reservoir and the second reservoir, respectively.

[0179] Embodiment 45: A composition, kit or method according to any of the preceding embodiments, wherein the aldehyde group of the polymer component reacts (e.g., reversibly reacts) with (i) the drug (e.g., doxorubicin), (ii) the dendritic polymer component and / or and (iii) one or more amines of one or more biological tissues to form an imine bond.

[0180] Embodiment 46: The composition, kit or method of Embodiment 45, wherein said formation of said imine bond and hydrolysis of said imine bond controls or contributes to the release kinetics of said drug from said hydrogel.

[0181] Embodiment 47: The composition, kit or method of Embodiment 45, wherein said forming of said imine bond and hydrolysis of said imine bond achieves sustained delivery of a drug.

[0182] Examples

[0183] The present invention is further illustrated by the following examples, which should not be construed in any way as imposing limitations on its scope. On the contrary, it should be clearly understood that, without departing from the spirit of the present invention or the scope of the appended claims, after reading the description herein, various other aspects, embodiments, modifications and equivalents thereof that may be associated with those of ordinary skill in the art may be resorted to. Therefore, in view of the description and practice of the present invention disclosed herein, other aspects of the present invention will be apparent to those of ordinary skill in the art.

[0184] Example 1 - Testing of various therapies

[0185] Several therapies were measured for efficacy in GBM cell lines. Doxorubicin demonstrated increased efficacy in GBM cell lines compared to standard of care chemotherapy.

[0186] The efficacy of various drugs

[0187] drug <![CDATA[IC in GL261 50 (μM)]]> <![CDATA[IC in CT-2A 50 (μM)]]> Temozolomide >750 >750 Irinotecan 106.8 150.1 Carmustine (BCNU) 145.9 155.8 Acriflavine 2.7 7.3 Adriamycin 0.8 0.1

[0188] Figure 3 Plots of cell viability versus concentration are depicted for each of the treatments listed in the preceding tables.

[0189] Testing has shown that doxorubicin is more effective in eliminating GBM spheroids in vitro than other chemotherapies tested. However, other chemotherapies tested may be used in the methods and compositions described herein.

[0190] However, based on these tests, an initial screening of dextran-dendrimer hydrogels was performed.

[0191] Examples of compositions

[0192]

[0193]

[0194] like Figure 4 As depicted, testing of the compositions in the preceding table indicated that Sample 5 had a faster degradation time than Sample 1.

[0195] Based on the results of the initial screening, a number of hydrogel formulations were generated and tested, including those described in the following table:

[0196]

[0197] The doxorubicin release profiles of the aforementioned compositions were tested. Figure 5A and Figure 5B As depicted, increasing the w / v % of dextran did not substantially affect the doxorubicin release profile.

[0198] The tests in this example also show that changing the solid content of the tested composition does not substantially affect the doxorubicin release profile, such as Fig. 6A and Figure 6B Place depicted.

[0199] Further testing in this example also showed that loading doxorubicin with dextran or dendrimers did not substantially change the doxorubicin release profile, e.g. Fig. 7A and Figure 7B Place depicted.

[0200] However, varying the amount of doxorubicin loaded into the aforementioned compositions slightly altered the doxorubicin release profile, e.g. Fig. 8A and Figure 8B Place depicted.

[0201] However, each of the hydrogels tested in this example provided sustained release of doxorubicin, e.g. Fig. 9 After 19 days, the cumulative release percentage of the "12% dendrimer, 10% dextran" hydrogel was slightly less than 90%, the cumulative release percentage of the "15% dendrimer, 12.5% ​​dextran" hydrogel was greater than 90%, and the cumulative release percentage of the "18% dendrimer, 15% dextran" hydrogel was 100%.

[0202] The release of primary amine-capped doxorubicin was also tested. According to the test, the capped doxorubicin achieved a higher cumulative release percentage compared to doxorubicin, such as Fig.10 The cumulative release percentages of doxil and adriamycin were also compared. Fig.11 As depicted in Figure 3, a higher cumulative percentage of doxorubicin was released over 200 hours.

[0203] The in vitro release of acriflavine was also tested. Injectable hydrogel formulations (75% oxidized dextran, 12.5% ​​w / v; PAMAM dendrimer, 15% w / v, pH 9.2) with four different loading concentrations (0.625 mg, 1.25 mg, 2.5 mg, and 5 mg, each of which had been previously injected locally into the brain) were prepared. The cumulative release profiles of the different loading amounts were similar, as shown in Table 1. Fig. 12A and Fig. 12B As expected, acriflavine retained its activity after release.

[0204] After the hydrogel of this example was injected into the brain of mice, magnetic resonance imaging (MRI) was used to evaluate the persistence of the hydrogel. According to MRI scans collected at the time of injection and on days 2, 12, and 20 thereafter, the hydrogel lasted for at least 20 days.

[0205] In addition to its durability, the degradation of the hydrogels was also studied, especially in vivo hydrogel degradation of formulations with different solid contents by IVIS. The mean radiation efficiency is plotted against the number of days from hydrogel injection in Fig.13 Place description.

[0206] Therapeutic retention and tissue penetration studies were designed and performed. The purpose of this study was to evaluate the retention, cellular uptake, and tissue penetration of doxorubicin when released from a hydrogel and to compare these data with data from intratumoral injection of the therapy. The following table describes the study:

[0207]

[0208] The results of this test showed that hydrogel delivery of doxorubicin improved doxorubicin retention compared to intratumoral injection. An increase in doxorubicin tissue permeation was also observed over time as doxorubicin was released from the hydrogel.

[0209] Further testing showed that the solids content of the hydrogel formulation did not significantly affect the outcome of topical doxorubicin therapy, as Fig.14 Place depicted.

[0210] Another test was designed and performed to evaluate whether sustained local delivery of doxorubicin using the hydrogel of this example can prolong the survival of mice bearing orthotopic GL261 tumors. The following table describes the study, and these steps were applied to four groups (untreated control, empty hydrogel, intratumoral injection of doxorubicin, and hydrogel delivery of doxorubicin (N = 6-10 / group; C57BL / 6 mice)):

[0211] Days Activity 0 GL261 tumor implantation 7 IVIS imaging of tumor size 10-14 Administration of treatment 15-50 Monitoring mouse health, survival, and tumor burden

[0212] This testing showed that local hydrogel delivery of doxorubicin significantly improved the survival of mice bearing orthotopic tumors compared with intratumoral (IT) injection. Fig.15 The results also showed that the survival efficacy of hydrogel-delivered doxorubicin was dose-dependent, as shown in Fig.16 Place depicted. Fig.16 It was shown that the survival probability of the control and empty hydrogels was similar, and the hydrogel containing 15 μg of doxorubicin resulted in a higher survival probability than the hydrogel containing 7.5 μg of doxorubicin.

[0213] Another study was designed and tested to determine whether local delivery of doxorubicin could improve the efficacy of current oral temozolomide chemotherapy. The following table describes the study applied to six groups (untreated control, empty hydrogel, oral temozolomide (TMZ), hydrogel-delivered doxorubicin, hydrogel-delivered doxorubicin plus oral TMZ) (N=6-10 / group; C57BL / 6 mice):

[0214] Days Activity 0 GL261 tumor implantation 7 IVIS imaging of tumor size 10-14 Treatment administration (systemic TMZ delivered daily on days 10-14; hydrogel delivery of doxorubicin on day 10) 15-50 Monitoring mouse health, survival, and tumor progression

[0215] The results showed that local delivery of doxorubicin reduced tumor burden in mice bearing intracranial tumors, e.g. Fig.17 Tumor burden bioluminescence analysis was performed throughout the assay and the data are presented in Fig.18 Describe the place.

[0216] For the tests in this example, mice receiving a combination of oral TMZ and hydroadriamycin therapy had no clear benefit in terms of median OS. This conclusion is based, at least in part, on Fig.19 The figure is obtained.

[0217] Comparative testing showed that doxil was as effective as doxorubicin in eliminating glioblastoma cells in vitro. Fig. 20A and Fig. 20B As demonstrated, the graph depicts cell viability for the following treatments:

[0218] therapy GL261 CT-2A Adriamycin 0.83μM 0.17μM Doxil 1.40μM 0.60μM

[0219] Further testing showed that clinically available nanoparticle doxorubicin ( Chemotherapy) when delivered to tumor-bearing mice using the hydrogel of this example resulted in improved survival. This conclusion is based, at least in part, on Fig.21 At half the concentration (7.5 μg), clinically available nanoparticle doxorubicin ( chemotherapy) was as effective as doxorubicin without the hydrogel, e.g. Fig. 22 Place depicted.

[0220] A study was also designed to test whether topical doxorubicin delivered via hydrogel could eradicate patient-derived xenograft orthotopic tumors in nude mice. The table below describes the study for the six groups (untreated control, empty hydrogel, oral TMZ, hydrogel-delivered doxorubicin (7.5 μg), hydrogel-delivered doxorubicin (15 μg), and hydrogel-delivered Chemotherapy (7.5 μg) (N=6-10 / group, nude mice) (PDX characteristics: MGMT methylation, EGFR not amplified, TP53 not mutated).

[0221]

[0222] like Fig.23 Results from the studies described herein demonstrate that mice bearing orthotopic PDX tumors treated with hydrogel doxorubicin therapy had significantly prolonged survival, as seen in syngeneic models. Chemotherapy (7.5 μg) resulted in a survival probability greater than 50%, and hydrogel-delivered doxorubicin (15 μg) resulted in a survival probability slightly less than 50%.

[0223] In this example, NSG mice bearing orthotopic PDX tumors treated with various hydrogel doxorubicin therapies had significantly prolonged survival. Fig.24 Describe the place.

[0224] The test in this example also showed that there was no significant difference between the survival rate of PDX nude mice and the survival rate of GL261 C57BL6 mice (see Fig.25A ( Chemotherapy hydrogel) (7.5 μg), Fig.25B (Adriamycin hydrogel) (15 μg), Fig.25C (Adriamycin hydrogel) (7.5 μg). For all treatments, non-significant differences were obtained between the survival curves of nude mice and C57BL6 mice, but it should be noted that different cells were used. These results suggest that the adaptive immune response may not play a significant role in eliminating tumors at the time of primary challenge.

[0225] Another test was performed to evaluate whether topical doxorubicin therapy confers protection against tumor rechallenge in long-term surviving mice. This study followed the following parameters:

[0226] Days Activity 0 Primary excitation (left hemisphere of the brain) 10-13 treat 70 Tumor reactivation (right hemisphere of the brain)

[0227] All long-term syngeneic mice treated with topical hydrogel therapy rejected tumors upon rechallenge in the contralateral hemisphere. Fig.26 The data depicted are derived from

[0228] Furthermore, long-term surviving PDX nude mice did not reject the tumor when the contralateral hemisphere was rechallenged, e.g. Fig. 27 Place depicted.

[0229] Yet another study was conducted to determine whether topical hydrogel doxorubicin chemotherapy could synergize with systemic aPD1 checkpoint blockade therapy. The dose for this study was 200 μg aPD1 given intraperitoneally, and four groups were tested (untreated, systemic aPD1, intracranial doxorubicin hydrogel, intracranial doxorubicin hydrogel plus systemic aPD1) (C57BL / 6 mice, N=6-10 / group):

[0230] Days Activity 0 <![CDATA[Primary excitation (1.3x 10 5 GL261)]]> 10 Doxorubicin hydrogel injection / aPD1 injection 12 aPD1 injection 14 aPD1 injection 60 Tracking tumor growth / Tracking weight

[0231] The combination of local doxorubicin therapy and systemic aPD1 did not improve the survival of mice bearing glioblastoma GL261, as Fig.28 Without wishing to be bound by any particular theory, it is believed that one or more factors may drive the lack of synergy between local doxorubicin chemotherapy and aPD1. For example, with respect to the drug release and biodistribution profile of the therapy, doxorubicin may reduce infiltrating T cells, and / or systemic aPD1 may not be able to reach the target tissue with the necessary pharmacokinetic profile. With respect to the immunosuppressive tumor microenvironment (TME), the factors may include limited infiltration and penetration of T cells into the TME using chemotherapy, and / or the inability of the chemotherapy to eliminate or reverse immunosuppressive macrophages.

[0232] Primary Tumor Immunophenotyping - Five mice were tested per treatment group (untreated, doxorubicin hydrogel) and mice were sacrificed 14 days after tumor inoculation (4 days post-therapy). Tissues tested included tumor, spleen, bone marrow, blood, and tumor draining lymph nodes. The panel used for this test included memory T cells, macrophages, and dendritic cells / MDSCs.

[0233]

[0234] Most untreated tumors did not have significant numbers of immune cells for the T cell panel. In 3 / 5 untreated tumors, the CD45+ and CD3+ subsets of viable cells were <1000 events. Different enrichment steps were tested to increase the number of immune cells in the sample (magnetic beads, different Percoll gradients). A plot of total flux (photons / second) versus days after tumor implantation is shown in Fig.29 Describe the place.

[0235] Compared with untreated mice, the increase in naive T cells in the lymph nodes was Fig.30 No other significant differences in T cells were noted in the tumor-draining lymph nodes. Only a slight decrease in central memory CD8 T cells was observed in the spleen, as shown in Figure 2. Fig.31 The only other difference in splenocyte immune infiltration was a slight decrease in CD45+ cells in the treated groups.

[0236] Macrophages promoted the increase of CD45+ cells in tumors after treatment with doxorubicin hydrogels, such as Fig.32 A slight increase in the M1 polarization marker CD80+ was observed in doxorubicin-treated tumors, as shown in Fig.33 Place depicted.

[0237] Initial analysis showed a significant shift in the dendritic cell population, e.g. Fig.34 However, it should be noted that the low number of DCs (approximately 1000) in untreated tumors complicates the analysis of cDC / pDC populations.

[0238] Example 2 - Further testing and analysis

[0239] STING (cGAS / stimulator of interferon genes antagonist) is understood to drive anticancer immune activity through the production of type I interferons and other proinflammatory cytokines ( Fig.35 ). Nanoparticles can be used in the cGAS-STING pathway. Cyclic dinucleotides (CDNs) can be potential cancer immunotherapy drugs and can be delivered using nanoparticle-mediated delivery ( Fig.36 ).

[0240] It has been hypothesized that topical hydrogel delivery of combination chemoimmunotherapy could (1) modulate key components of the tumor microenvironment, (2) enhance antitumor immune sensitization, and / or (3) improve the efficacy of checkpoint blockade therapy ( Fig.37 ).

[0241] Nanoparticle-based STING agonist formulations can possess or promote high biodegradability, low toxicity, nucleic acid encapsulation (amine groups), high endosomal escape (good buffering capacity), ease of synthesis (high versatility) ( Fig.38 ).

[0242] CDN nanoparticles stimulate dendritic cells and macrophages, such as Fig.39 The hydrogel composition of the example mediates the release of CDN nanoparticles in vitro, such as Fig.40A and Fig.40B Place depicted.

[0243] The high hydrophilicity and negative charge of CDNs may hinder their delivery into cells and thus their clinical potential, limiting their delivery to tumors to intratumoral injections. Although CDNs are currently undergoing clinical trials, there are concerns that simple intratumoral CDN injections are a suboptimal way to stimulate the cytoplasmic STING signaling pathway. However, this example describes polymer-based CDN-conjugated nanoparticles for allowing systemic delivery of CDNs that are otherwise cleared from the circulation within minutes, which are programmed to be released in the cell cytoplasm, and whose safety, efficacy, and mechanism of action have been studied in multiple mouse tumor models.

[0244] These NPs are made of PBAE modified with arginine residues to enhance the biocompatibility and endosomal escape ability of the NPs and complexed with CDN-conjugated PBAE chains via cathepsin-sensitive bonds, enabling the CDN release in the cytoplasm.

[0245] A local immunotherapy MTD pilot study was designed and conducted. The study parameters appear in the table below. Mice bearing GL261-luc tumors were tested, and each group included 3 mice. Mice received a 10 μL hydrogel injection and IVIS imaging was used to evaluate luciferase-tumor burden, AF568-hydrogel degradation, and AF647-therapy release.

[0246] Days Activity 0 Tumor implantation 7 Hydrogel injection After Day 7 IVIS imaging - monitoring body weight, tumor growth, survival, hydrogel degradation and therapeutic release

[0247]

[0248] *CDN-NP = PAMAM dendrimer + NHS-PEG-SH linker + Aduro CDN-Mal (about 30CDN / dendrimer)

[0249] Results of a pilot study of local immunotherapy Fig.41 IVIS bioluminescence-based imaging of tumor burden (day 18) was collected for empty hydrogels, aPD-1 hydrogels (150 μg), aPD-L1 hydrogels (150 μg), and CDN dendrimer NP hydrogels (40 μg CDN).

[0250] We determined that doxorubicin treatment increased surface expression of calreticulin on GBM cells but did not affect HMGB1 excretion at relevant concentrations. Fig.42 Doxorubicin induces exposure of calreticulin (CRT) that directs immunogenic cell death, as depicted in FIG. 43 .

[0251] We also tested whether chemoimmunotherapy could synergize to enhance bone marrow-derived macrophage activation ( Fig.44 ).like Fig.45 As depicted in Figure 2, BMDM CD86 activation marker expression was enhanced by chemoimmunotherapy. Fig.46 As depicted, combination therapy resulted in an increase in the ratio of pro-inflammatory BMDM to anti-inflammatory BMDM.

[0252] It was determined that the hydrogels (including adhesive hydrogels) of the aforementioned examples effectively delivered local therapy to treat malignant brain tumors in mice. For example, hydrogel delivery of free doxorubicin or nanoparticle doxorubicin prolonged the survival of mice with orthotopic synergistic tumors of PDX tumors. In addition, 100% of the long-lived mice treated with local hydrogel doxorubicin therapy rejected the tumor upon re-stimulation. Hydrogel delivery of CDN nanoparticles is another strategy for treating GBM alone or in combination with doxorubicin chemotherapy.

[0253] The compositions and methods described herein can allow for the engineering of immunity using materials to enhance therapeutic efficacy ( Fig.47 ).

Claims

1. A drug delivery composition comprising: A hydrogel, wherein the hydrogel is an adhesive hydrogel; and a drug, wherein the drug is dispersed in the hydrogel, The drug comprises a chemotherapy drug, an immunotherapy drug, or a combination thereof.

2. The drug delivery composition according to claim 1, wherein - The drug comprises the chemotherapy drug, and the chemotherapy drug comprises doxorubicin; and The hydrogel comprises: (i) a polymer component, wherein the polymer component comprises a polymer having three or more aldehyde groups; and (ii) a dendrimer component, wherein the dendrimer component comprises a dendrimer having at least two branches and one or more surface groups.

3. The drug delivery composition of claim 1, wherein the hydrogel comprises: a polymer component, wherein the polymer component comprises a polymer having three or more aldehyde groups; and A dendritic polymer component, wherein the dendritic polymer component comprises a dendritic polymer having at least 2 branches and one or more surface groups.

4. The drug delivery composition of claim 3, wherein 100% of the one or more surface groups comprise at least one primary or secondary amine.

5. The drug delivery composition of claim 3, wherein less than 75% of the one or more surface groups comprise at least one primary or secondary amine.

6. The drug delivery composition of claim 1, wherein the dendritic polymer is a fifth generation polyamidoamine (G5 PAMAM) dendritic polymer.

7. The drug delivery composition of claim 1, wherein the polymer comprises dextran.

8. The drug delivery composition of claim 1, wherein the chemotherapy drug comprises doxorubicin, and the doxorubicin is encapsulated in liquid spheres.

9. The drug delivery composition according to claim 8, wherein the liquid sphere is a liquid nanosphere.

10. The drug delivery composition of claim 1, wherein the immunotherapy drug comprises a cyclic dinucleotide.

11. The drug delivery composition according to claim 10, wherein the cyclic dinucleotide is encapsulated in nanoparticles, wherein the nanoparticles- (i) comprising a poly-β-amino-ester to which the cyclic dinucleotide is conjugated via a cathepsin-sensitive bond, wherein the poly-β-amino-ester is optionally modified with arginine; (ii) has an average diameter of about 30 nm to about 70 nm; or (iii) combinations thereof.

12. The drug delivery composition of claim 1, wherein the solids content of the drug delivery composition is from about 8 wt% to about 25 wt%.

13. The drug delivery composition of claim 1, wherein the hydrogel comprises phosphate buffered saline (PBS).

14. The drug delivery composition of claim 1, wherein the drug is present in the drug delivery composition in (i) a total amount of about 50 μg to about 500 μg, (ii) a total concentration of about 1 μg / μL to about 10 μg / μL, or (iii) a combination thereof.

15. The drug delivery composition of claim 3, wherein the three or more aldehyde groups of the polymer reversibly react with (i) the drug, (ii) the dendrimer, and / or (iii) one or more amines in one or more biological tissues to form imine bonds, and wherein the formation of the imine bonds and the hydrolysis of the imine bonds control or contribute to the release kinetics of the drug from the hydrogel.

16. A method of treating a patient, the method comprising: The drug delivery composition according to any one of claims 1 to 15 is locally delivered to the intracranial region of the patient.

17. The method of claim 16, wherein the local delivery of the drug delivery composition comprises injecting or spraying the drug delivery composition.

18. The method of claim 17, wherein the injection of the drug delivery composition occurs during a biopsy, through an intraventricular shunt, or a combination thereof.

19. The method of claim 16, wherein the intracranial region comprises brain tissue, a brain tumor, a site of a previously resected tumor, or a combination thereof.

20. The method of claim 16, further comprising administering a second drug or therapy to the patient before, during, and / or after the local delivery of the drug delivery composition.

21. The method of claim 20, wherein the administering of the second drug or therapy comprises orally administering the second drug or therapy.

22. The method of claim 20, wherein the second drug or therapy comprises a second chemotherapy drug.

23. The method of claim 22, wherein the second chemotherapy drug or treatment comprises temozolomide.

24. The method of claim 20, wherein the second drug or therapy comprises aPD1 checkpoint blockade therapy.

25. The method of claim 16, wherein the drug is continuously released from the drug delivery composition for at least 24 hours, at least 48 hours, at least 36 hours, at least 72 hours, at least 100 hours, at least 200 hours, or at least 300 hours.

26. The method of claim 16, wherein the cumulative percentage of the drug released from the drug delivery composition is at least 80% by weight of the drug, preferably at least 85% by weight of the drug, within 10 days or less, 12 days or less, 14 days or less, 16 days or less, 18 days or less, 20 days or less, 22 days or less, 24 days or less, 26 days or less, 28 days or less, 30 days or less, 32 days or less, 34 days or less, 36 days or less, 38 days or less, 40 days or less, 50 days or less, 60 days or less, 80 days or less, or 100 days or less after the local delivery of the drug delivery composition.

27. A kit for preparing a drug delivery composition, the kit comprising: a first part, the first part comprising a first solution, the first solution comprising a polymer component, wherein the polymer component comprises a polymer; and a second part, the second part comprising a second solution, the second solution comprising a dendritic polymer component, wherein the dendritic polymer component comprises a dendritic polymer having at least 2 branches and one or more surface groups; wherein at least one drug is disposed in the first solution, the second solution, or both the first solution and the second solution, and The drug comprises a chemotherapy drug, an immunotherapy drug, or a combination thereof.

28. The kit of claim 27, wherein the chemotherapy drug comprises doxorubicin.

29. The kit of claim 28, wherein the doxorubicin is encapsulated in liquid spheres.

30. The kit of claim 29, wherein the liquid spheres are liquid nanospheres.

31. The kit of any one of claims 27 to 30, wherein the immunotherapy drug comprises a cyclic dinucleotide.

32. The kit of claim 31, wherein the cyclic dinucleotide is encapsulated in nanoparticles having an average diameter of about 30 nm to about 70 nm.

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