Maintenance therapy for treating cancer

Abstract

The present disclosure provides a method for treating cancer in a patient in need thereof comprising (i) an induction phase comprising administering one or more anticancer therapies to the patient in need thereof; and (ii) a maintenance phase comprising administering to the patient a nucleic acid construct encoding TUSC2.

Description

MAINTENANCE THERAPY FOR TREATING CANCERCROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of U.S. Provisional Application No. 63 / 571,848, filed March 29, 2024, which is incorporated herein by reference in its entirety.SEQUENCE LISTING

[0002] This application contains a Sequence Listing, which has been submitted electronically in xml format and is hereby incorporated by reference in its entirety. Said xml copy, created on March 28, 2025, is named SeqList-198628-46376.xml and is 2,984 bytes in size.FIELD

[0003] The field of the present disclosure relates to methods, vectors, and compositions for maintenance therapy in the treatment of cancers by expressing TUSC2 in the cancer cells. Also contemplated are compositions including nonviral or viral vectors for increasing TUSC2 expression in cancer cells.BACKGROUND

[0004] The development of cancer involves the deregulation of multiple cellular pathways that control normal cell growth. Healthy cells express a number of tumor suppressor genes, which act as molecular gatekeepers and prevent uncontrolled cell division. An important step in the development of cancer, therefore, is disruption of tumor suppressor signaling pathways. Therefore, one promising avenue for cancer therapy involves expression of tumor suppressor genes in cancer cells to restore cellular growth controls.

[0005] The tumor suppressor candidate 2 (TUSC2) gene encodes a small (110 amino acid), mitochondrial protein. TUSC2 is involved in a wide array of cellular processes, including induction of apoptosis, inhibition of many kinases, mitochondrial calcium homeostasis, inhibition of reactive oxygen species (ROS) response, and increase of immune stimulation.

[0006] TUSC2 has been shown to act as a tumor suppressor gene in a variety of cancers including lung cancer, breast cancer, and bone cancer. Expression of TUSC2 is reduced orabsent in more than 80 percent of all lung cancers. Expression of TUSC2 is absent in 41% of patients with small cell lung cancer (SCLC) and decreased in all. Patients with Extensive Stage SCLC (ES-SCLC) starting maintenance therapy with atezolizumab (atezo) have a median Progression Free Survival (PFS) of only 2.6 months. In addition, non-small cell lung cancer (NSCLC) patients with loss of TUSC2 expression have been reported to have a significantly worse overall survival (OS) than did those with normal TUSC2 expression.

[0007] For many types of cancer, the goal of treatment is to cure the cancer. Likewise, for many types of advanced cancer, the initial treatments can shrink the cancer and prevent it from spreading. Following the initial treatments, even when the patient is in remission or complete remission, cancer cells may remain in the patient’s body, and the cancer can recur. Accordingly, there is a need for maintenance therapies that delay or prevent recurrence and are more tolerable to patients than the initial therapies.SUMMARY

[0008] In one aspect, the disclosure provides a method for treating cancer in a patient in need thereof comprising (i) an induction phase comprising administering one or more anticancer therapies to the patient in need thereof; and (ii) a maintenance phase comprising administering to the patient a nucleic acid construct encoding TUSC2. The cancer may be, for example, colon cancer, pancreatic cancer, breast cancer, melanoma, osteosarcoma, rectal cancer, lung cancer, leukemia, and neuroblastoma. In embodiments, the cancer is non-small cell lung cancer (NSCLC). In embodiments, the cancer is small cell lung cancer (SCLC). In embodiments, the cancer is Extensive Stage SCLC (ES-SCLC)

[0009] In embodiments, the one or more anticancer therapies administered to the patient in the induction phase comprises one or more checkpoint inhibitors. In embodiments, the one or more checkpoint inhibitors is selected from an anti-PD-Ll antibody (e.g., atezolizumab, avelumab, and / or durvalumab) and / or an anti-PD-1 antibody (e.g., pembrolizumab, nivolumab, and / or cemiplimab). In embodiments, the one or more anticancer therapies administered to the patient in the induction phase comprises one or more tyrosine kinase inhibitors (TKIs).

[0010] In embodiments, the induction phase comprises administering one or more chemotherapies, such as one or more platinum based chemotherapeutic agents and / or one or more topoisomerase inhibitors. In embodiments, the one or more platinum-based chemotherapeutic agents are administered to the patient during the induction phase in addition to administration of one or more anticancer therapies disclosed herein (e.g., acheckpoint inhibitor, an EGFR tyrosine kinase inhibitor, and / or a TUSC2 expression construct). In embodiments, the one or more platinum-based chemotherapeutic agent is selected from cisplatin, carboplatin, oxaliplatin, nedaplatin, triplatin tetranitrate, phenanthriplatin, picoplatin, satraplatin, or combinations thereof. In embodiments, the platinum-based chemotherapeutic agent is cisplatin. In embodiments, the induction phase comprises administering one or more topoisomerase inhibitors (e.g., etoposide, irinotecan, teniposide, and topotecan).

[0011] In embodiments, the induction phase comprises administering a platinum based chemotherapeutic agent, a topoisomerase inhibitor, and a checkpoint inhibitor. In embodiments, the platinum based chemotherapeutic agent is selected from cisplatin, carboplatin, oxaliplatin, nedaplatin, triplatin tetranitrate, phenanthriplatin, picoplatin, satraplatin, and combinations thereof; the topoisomerase inhibitor is selected from etoposide, irinotecan, teniposide, topotecanand, and combinations thereof; and the checkpoint inhibitor is selected from an anti-PD-Ll antibody (e.g., atezolizumab, avelumab, and / or durvalumab) and / or an anti-PD-1 antibody (e.g., pembrolizumab, nivolumab, and / or cemiplimab). In embodiments, the induction phase comprises administering a platinum based chemotherapeutic agent, etoposide, and atezolizumab.

[0012] In embodiments, the induction phase comprises administering a nucleic acid construct encoding TUSC2 to the patient in need thereof in addition to administration of one or more anticancer therapies disclosed herein (e.g., a checkpoint inhibitor, a tyrosine kinase inhibitor, and / or a platinum-based chemotherapeutic agent).

[0013] In embodiments, the induction phase comprises administering atezolizumab and a platinum-based chemotherapeutic agent (e.g., cisplatin). In embodiments, the induction phase comprises administering pembrolizumab and a platinum-based chemotherapeutic agent (e.g., cisplatin). In embodiments, the induction phase comprises administering atezolizumab, a platinum-based chemotherapeutic agent (e.g., cisplatin) and a nucleic acid construct encoding TUSC2. In embodiments, the induction phase comprises administering pembrolizumab, a platinum-based chemotherapeutic agent (e.g., cisplatin), and a nucleic acid construct encoding TUSC2.

[0014] In embodiments, the maintenance phase comprises, administration of the nucleic acid encoding the TUSC2 protein as a monotherapy. In other embodiments, the maintenance phase comprises, in addition to the administration of the nucleic acid encoding the TUSC2 protein, also administering one or more checkpoint inhibitors. In embodiments, the one or more checkpoint inhibitors is selected from an anti-PD-Ll antibody (e.g., atezolizumab,avelumab, and / or durvalumab) and / or an anti-PD-1 antibody (e.g., pembrolizumab, nivolumab, and / or cemiplimab). In embodiments, the maintenance phase comprises, in addition to the administration of the nucleic acid encoding the TUSC2 protein, administration of atezolizumab. In embodiments, the maintenance phase comprises, in addition to the administration of the nucleic acid encoding the TUSC2 protein, also administering one or more tyrosine kinase inhibitors.

[0015] The nucleic acid construct encoding TUSC2 may be administered in a viral or nonviral vector. In embodiments, the nonviral vector comprises a DOTAP:cholesterol liposome. In embodiments, the DOTAP:cholesterol ratio is between about 3: 1 and about 1 :3.

[0016] In other embodiments, the polynucleotide construct is administered in a viral vector, for example an Adeno- Associated Virus (AAV) viral vector.

[0017] In embodiments, the nonviral vector comprising a nucleic acid construct encoding TUSC2 is quaratusugene ozeplasmid (REQORSA®).DETAILED DESCRIPTION

[0018] Provided herein are methods for treating cancer by expressing TUSC2 in the cancer cells of a patient as a maintenance phase therapy, or as part of a maintenance phase therapy, following induction phase therapy. TUSC2 can be expressed from a nucleic acid expression construct encoding a TUSC2 protein. Also contemplated are nonviral vectors (e.g., comprising DOTAP:cholesterol liposomes) having the nucleic acid constructs disclosed herein. Additionally contemplated are viral vectors (e.g., recombinant AAV vectors) having the nucleic acid constructs disclosed herein. Further contemplated herein are pharmaceutical compositions having the nonviral vectors or viral vectors disclosed herein.

[0019] I. Method of Treatment

[0020] Methods of treating cancer in a human subject are described herein. In one embodiment of the method of treating cancer, a composition comprising a nucleic acid construct comprising a TUSC2 encoding sequence as described herein is administered to a human subject as a maintenance phase therapy following induction phase therapy with one or more anticancer therapies. In another embodiment of the method of treating cancer, a composition comprising a vector as described herein is administered to a human subject as a maintenance phase therapy following induction phase therapy with one or more anticancer therapies. In a further embodiment of the method of treating cancer, a composition comprising a nonviral vector (comprising a nucleic acid construct comprising TUSC2encoding sequence) described herein is administered to a human subject as a maintenance phase therapy following induction phase therapy with one or more anticancer therapies.

[0021] References herein to “treating” or “treatment” include prophylaxis as well as the alleviation of established symptoms of a condition. “Treating” or “treatment” of the condition therefore includes (a) preventing the appearance of one or more clinical symptoms of the condition or (b) delaying the appearance of one or more clinical symptoms of the condition in a patient that may be afflicted with or predisposed to the condition, but does not currently experience or display clinical or subclinical symptoms related to the condition. “Treating” or “treatment” of the condition also includes inhibiting, arresting, reducing, or delaying the development of the disease and / or a relapse thereof or at least one clinical symptom thereof.

[0022] As used herein, the term “treating cancer” refers to administration of one or more therapeutic agents (e.g., nonviral vectors as described herein) to a patient having cancer with the purpose to cure, heal, alleviate, relieve, alter, remedy, ameliorate, or improve the disease, or one or more symptoms of the disease. The treatment methods described herein inhibit, delay, decrease, and / or reduce one or more adverse symptoms, disorders, illnesses, diseases and / or complications caused by or associated with cancer. The treatment methods described herein utilize a TUSC2 expression vector described herein to, for example, increase or restore TUSC2 levels to TUSC2 -deficient cancer cells, induce apoptosis of cancer cells, decrease tumor size or eliminate a tumor in a subject, and / or reduce or prevente metastasis. Methods of treating cancer generally include increasing or restoring TUSC2 signaling / expression to cancer cells that have reduced TUSC2 levels or inhibition of TUSC2 function.

[0023] The methods described herein can be used to treat human subjects suffering from various types of cancer, such as colon cancer, pancreatic cancer, breast cancer, melanoma, osteosarcoma, rectal cancer, lung cancer (e.g., small cell or Extensive Stage SCLC or nonsmall cell lung cancer), leukemia, and neuroblastoma. In embodiments, the methods disclosed herein can be used to treat skin cancer (including melanoma, squamous cell carcinoma and basal cell carcinoma), renal cell carcinoma, colorectal cancer, hepatocellular carcinoma, non-small cell lung cancer, small cell lung cancer, Extensive Stage SCLC, mesothelioma (e.g., malignant pleural mesothelioma), lymphoma (such as Hodgkin lymphoma), head and neck cancer, urothelial carcinoma, colorectal cancer, esophageal carcinoma, gastric cancer, cervical cancer, endometrial cancer, bladder cancer, breast cancer, or Merkel cell carcinoma. In preferred embodiments, the cancer is lung cancer (e.g., smallcell or Extensive Stage SCLC or non-small cell lung cancer), in particular non-small cell lung cancer.

[0024] Any suitable methods of administering nucleic acid constructs, nonviral vectors, viral vectors, and compositions to a subject in need thereof may be used. In these methods, the nucleic acid constructs, nonviral vectors, viral vectors and compositions can be administered to the human subject by any suitable route. In some embodiments, for example, they are administered intravenously (IV). If administered via IV injection, the nucleic acid constructs, nonviral vectors, and compositions may be administered in a single bolus, multiple injections, or by continuous infusion (e.g., intravenous pump infusion). In other embodiments, for example, they are administered intranasally. The nucleic acid constructs, nonviral vectors, and compositions can be administered to the human subject once (at one time point), or more than one time (e.g., two times, three times, four times, five times, six times, seven times, eight times, nine times, 10 times, etc.), i.e., at multiple time points. When the compositions are administered multiple times, the administrations may be separated by one day, three days, five days, one week, two weeks, three weeks, one month, two months, or six months.

[0025] i. Induction Phase

[0026] The present disclosure provides a method of treating cancer in a patient by providing induction phase therapy followed by a maintenance phase therapy comprising a nucleic acid construct encoding a TUSC2 protein. As used herein, “induction phase” refers to the initial anticancer therapy or therapies to decrease the signs and / or symptoms of cancer or make them disappear. Induction phase therapy is often part of a standard set of treatments, such as surgery followed by radiation and / or chemotherapy and / or additional anti -cancer therapies such as targeted therapy and / or immunotherapy.

[0027] In embodiments of the methods described herein, the induction phase therapy can include surgery, for example, for removal of diseased tissue, and / or radiation treatment. For example, in some embodiments of the methods described herein, the induction phase includes surgical resection of a tumor. As another example, in some embodiments of the methods described herein, the induction phase includes radiation treatment. As another example, in some embodiments of the methods described herein, the induction phase includes chemotherapy. In some embodiments of the methods described herein, the induction phase comprises surgery (e.g., resection of a tumor) and / or radiation treatment and / or chemotherapy.

[0028] In embodiments of the method of treating cancer disclosed herein, the induction phase comprises administering one or more anticancer therapies to a patient in need thereof, wherein the one or more anticancer therapies is selected from (i) a checkpoint inhibitor, (ii) a tyrosine kinase inhibitor, (iii) a nucleic acid construct, a nonviral vector, a viral vector, or a composition as described herein comprising a TUSC2 encoding sequence, and (iv) combinations thereof.

[0029] When multiple anti-cancer therapies are administered to the patient as part of the induction phase, the different anti-cancer agents may be administered multiple times over a time period in the induction phase. The time period over which the different anti-cancer agents are administered may be, for example, sequential (i.e., non-overlapping), overlapping, or coextensive.

[0030] In embodiments, the induction phase therapy comprises multiple cycles (e.g., 1 to 7 cycles, 2 to 6 cycles, or 4 to 6 cycles) of chemotherapy that may include administration of one or more of platinum-based chemotherapeutic agents in addition to one or more immunotherapies, or targeted therapies. In embodiments, the induction phase therapy comprises multiple cycles (e.g., 1 to 7 cycles, 2 to 6 cycles, or 4 to 6 cycles) of chemotherapy that may include administration of one or more of platinum-based chemotherapeutic agents, in addition to administration of one or more checkpoint inhibitors (e.g., anti-PD-1 or anti-PD- L1 antibodies) and / or one or more EGFR tyrosine kinase inhibitors.

[0031] In embodiments, the induction phase comprises administration of one or more checkpoint inhibitors (e.g., one or more checkpoint inhibitors disclosed herein). In embodiments, the one or more checkpoint inhibitors is or includes an anti-PD-Ll antibody (e.g., atezolizumab, avelumab, and / or durvalumab) and / or an anti-PD-1 antibody (e.g., pembrolizumab, nivolumab, and / or cemiplimab). In embodiments, the one or more checkpoint inhibitors is or includes an inhibitor of cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) (e.g., an anti-CTLA-4 antibody such as ipilimumab or tremelimumab). In embodiments, the one or more check-point inhibitors is or includes an inhibitor of lymphocyte activation gene-3 (LAG-3) (e.g., an anti-LAG-3 antibody such as relatlimab). In embodiments, the induction phase does not comprise administering an anti PD-1 antibody (e.g., pembrolizumab).

[0032] In embodiments, the induction phase comprises administration of one or more tyrosine kinase inhibitors such as inhibitors of EGFR tyrosine kinase (e.g., cetuximab, osimertinib, erlotinib, gefitinib, panitumumab, necitumumab, or mobocertinib). Inembodiments, the induction phase does not comprise administering an EGFR tyrosine kinase inhibitor (e.g., osimertinib).

[0033] In embodiments, the induction phase comprises administering a platinum based chemotherapeutic agent, a topoisomerase inhibitor, and a checkpoint inhibitor. In embodiments, the platinum based chemotherapeutic agent is selected from cisplatin, carboplatin, oxaliplatin, nedaplatin, triplatin tetranitrate, phenanthriplatin, picoplatin, satraplatin, and combinations thereof; the topoisomerase inhibitor is selected from etoposide, irinotecan, teniposide, topotecanand, and combinations thereof; and the checkpoint inhibitor is selected from an anti-PD-Ll antibody (e.g., atezolizumab, avelumab, and / or durvalumab) and / or an anti-PD-1 antibody (e.g., pembrolizumab, nivolumab, and / or cemiplimab). In embodiments, the induction phase comprises administering a platinum based chemotherapeutic agent, etoposide, and atezolizumab.

[0034] In embodiments of the method of treatment described herein, the induction phase includes administering to the human subject (i) a nucleic acid construct, a nonviral vector, a viral vector, or a composition as described herein comprising a TUSC2 encoding sequence, and (ii) administering one or more additional anti-cancer therapies (e.g., one or more check point inhibitors and / or one or more EGFR tyrosine kinase inhibitors). In embodiments, the induction phase comprises administering (i) a nucleic acid encoding TUSC2 and (ii) an anti- PD-Ll antibody (such as atezolizumab, avelumab, or durvalumab). In embodiments, the induction phase comprises administering (i) a nucleic acid encoding TUSC2 and (ii) an anti- PD-1 antibody (such as pembrolizumab, nivolumab, or cemiplimab). In embodiments, the induction phase comprises administering (i) a nucleic acid encoding TUSC2 and (ii) an EGFR tyrosine kinase inhibitor (e.g., cetuximab, osimertinib, erlotinib, gefitinib, panitumumab, necitumumab, or mobocertinib).

[0035] In embodiments, the induction phase comprises administration of (i) a TUSC2 expression construct packaged in a DOTAP:cholesterol liposome as disclosed herein and (ii) atezolizumab, and optionally (iii) a platinum-based chemotherapy (e.g., cisplatin).

[0036] In embodiments, the induction phase comprises administration of (i) a TUSC2 expression construct packaged in a DOTAP:cholesterol liposome as disclosed herein and (ii) pembrolizumab, and optionally (iii) a platinum-based chemotherapy (e.g., cisplatin).

[0037] In embodiments, the induction phase comprises administration of (i) a TUSC2 expression construct packaged in a DOTAP:cholesterol liposome as disclosed herein and (ii) an EGFR tyrosine kinase inhibitor (e.g., osimertinib, or erlotinib), and optionally (iii) a platinum-based chemotherapy (e.g., cisplatin).

[0038] In embodiments, during the induction phase, the nucleic acid encoding TUSC2 (e.g., a TUSC2 expression construct packaged in a DOTAP:cholesterol liposome) is administered intravenously once every 3 days, once every 5 days, once every 10 days, once a week, once every two weeks, once every three weeks, or once a month.

[0039] ii. Maintenance Phase

[0040] The present disclosure provides a method of treating cancer in a patient by providing to the patient (i) an induction phase therapy as disclosed herein, followed by (ii) a maintenance phase therapy comprising a nucleic acid construct, a nonviral vector, a viral vector, or a composition as described herein comprising a TUSC2 encoding sequence.

[0041] As used herein, the term “maintenance phase” refers to the stage in treatment of a patient, following the induction phase, in which the patient having responded to the induction phase therapy receives “maintenance phase therapy” or “maintenance treatment” to, for example, prolong remission and / or prevent or delay relapse of the disease. For example, a maintenance phase therapy may prevent or minimize growth of a cancer after it has been substantially reduced or eliminated following the induction phase therapy or therapies.

[0042] In embodiments, the goal of the maintenance phase therapy is to maintain a remission or prevent or delay the cancer's return if the cancer is in remission after the induction phase treatment. After the achievement of a response from the induction phase therapy disclosed herein, the maintenance therapy disclosed herein comprising administering a nucleic acid encoding a TUSC2 protein provides clinical benefit to the patient by delaying disease progression, delaying the need for additional rounds of toxic therapy (e.g., chemotherapy) and / or prolonging overall survival. In embodiments, the maintenance phase therapy is provided to patients whose cancer has responded to the induction phase treatment(s). In preferred embodiments, the maintenance phase therapy is provided to patients whose cancer is in partial or complete remission following the induction phase treatment(s).

[0043] The maintenance phase therapies disclosed herein can be administered periodically for extended time periods, up to the lifespan of the patient. Dosages used for maintenance therapy can vary and can include diminished dosages as compared to dosages used for induction phase therapy.

[0044] Maintenance phase therapy may be a continuous treatment where multiple doses are administered at spaced intervals such as every day, every other day, every week, every 2 weeks, every 3 weeks, every 4 weeks, every 6 weeks, or every 8 weeks. In someembodiments a maintenance therapy may continue for a predetermined length of time or may continue until unacceptable toxicity occurs and / or disease progression occurs. During maintenance phase therapy, treatment may be interrupted upon the occurrence of toxicity as indicated by an adverse event. If toxicity is appropriately resolved, the patient may restart maintenance treatment, which may include a dose level reduction.

[0045] Options for maintenance phase therapy include (i) continuing administration of one or more of the induction phase anticancer therapies disclosed herein (i.e., continuation maintenance) or introducing a new agent (i.e., switch maintenance therapy). In embodiments, at least one, but not all, of the induction phase therapies are continued in the maintenance phase and a new anti-cancer agent (i.e., an agent not used in the induction phase) is added to the maintenance phase.

[0046] In embodiments, the maintenance phase therapy comprises administration of a nucleic acid expressing TUSC2 without administration or coadministration of an additional anti-cancer therapy. In other words, in such embodiments, the maintenance phase comprises administration of a TUSC2 construct, vector, or composition disclosed herein as a monotherapy. In embodiments, the maintenance phase comprises administration of a TUSC2 expression construct packaged in a DOTAP:cholesterol liposome as disclosed herein as a monotherapy.

[0047] In embodiments of the method of treating cancer described herein, the maintenance phase comprises a combination therapy that includes (i) administering to the patient a nucleic acid construct, a nonviral vector, a viral vector, or a composition as described herein comprising a TUSC2 encoding sequence, and (ii) administering to the patient one or more additional anti-cancer therapies.

[0048] When multiple anti-cancer therapies are administered to the patient as part of the maintenance phase, the different anti -cancer agents may be administered multiple times over a time period in the maintenance phase. The time period over which the different anti-cancer agents are administered may be, for example, sequential (i.e., non-overlapping), overlapping, or coextensive.

[0049] In embodiments, the maintenance phase comprises administration of one or more checkpoint inhibitors (e.g., one or more checkpoint inhibitors disclosed herein). In embodiments, the one or more checkpoint inhibitors is or includes an anti-PD-Ll antibody (e.g., atezolizumab, avelumab, and / or durvalumab) and / or an anti-PD-1 antibody (e.g., pembrolizumab, nivolumab, and / or cemiplimab). In embodiments, the one or more checkpoint inhibitors is or includes an inhibitor of CTLA-4 (e.g., an anti-CTLA-4 antibody such asipilimumab or tremelimumab). In embodiments, the one or more check-point inhibitors is or includes an inhibitor of LAG-3 (e.g., an anti -LAG-3 antibody such as relatlimab).

[0050] In embodiments, the maintenance phase comprises administration of one or more tyrosine kinase inhibitors such as inhibitors of EGFR tyrosine kinase (e.g., cetuximab, osimertinib, erlotinib, gefitinib, panitumumab, necitumumab, or mobocertinib).

[0051] In embodiments, the maintenance phase comprises administration of (i) a TUSC2 expression construct packaged in a DOTAP:cholesterol liposome as disclosed herein and (ii) an anti-PD-Ll antibody (e.g., atezolizumab, avelumab, or durvalumab). In embodiments the anti-PD-Ll antibody is atezolizumab. In embodiments, the maintenance phase therapy is administered once about every two weeks, three weeks, four weeks; five weeks; six weeks; or eight weeks. In embodiments, the maintenance phase therapy is administered once about every three weeks; four weeks; or five weeks.

[0052] In embodiments, the maintenance phase comprises administration of (i) a TUSC2 expression construct packaged in a DOTAP:cholesterol liposome as disclosed herein and (ii) an anti-PD-1 antibody (e.g., pembrolizumab, nivolumab, or cemiplimab). In embodiments the anti-PD-1 antibody is pembrolizumab.

[0053] In embodiments, the maintenance phase comprises administration of (i) a TUSC2 expression construct packaged in a DOTAP:cholesterol liposome as disclosed herein and (ii) an EGFR tyrosine kinase inhibitor (e.g., cetuximab, osimertinib, erlotinib, gefitinib, panitumumab, necitumumab, or mobocertinib). In embodiments, the EGFR tyrosine kinas inhibitor is osimertinib or erlotinib.

[0054] In embodiments, when a TUSC2 expression construct packaged in a DOTAP:cholesterol liposome (e.g., quaratusugene ozeplasmid) is administered to a patient as a maintenance phase therapy (e.g., alone or in combination with a checkpoint inhibitor), the TUSC2 construct DOTAP:cholesterol liposome is administered to the patient at about 0.06 mg / kg; about 0.07 mg / kg; about 0.08 mg / kg; about 0.09 mg / kg; about 0.10 mg / kg; about 0.11 mg / kg; about 0.12 mg / kg; or about 0.15 mg / kg. In embodiments, the TUSC2 construct DOTAP:cholesterol liposome is administered at a dose between about 0.06 mg / kg to about 0.15 mg / kg; about 0.06 mg / kg to about 0.12 mg / kg; or about 0.09 mg / kg to about 0.12 mg / kg. The dose can be raised or lowered (e.g., in the current cycle or in later cycles) depending upon the patient’s response to the therapy.

[0055] In embodiments, the nucleic acid construct encoding TUSC2 is administered in the induction phase (e.g., in combination with one or more of: a platinum-based chemotherapy, a checkpoint inhibitor, and / or a tyrosine kinase inhibitor) and in the maintenance phase (e.g., asmonotherapy, or in combination with a checkpoint inhibitor or a tyrosine kinase inhibitor). In such embodiments, the nucleic acid construct encoding TUSC2 may be administered at a lower dose and / or at a reduced frequency in the maintenance phase than in the induction phase therapy. Alternatively, or in addition, in embodiments where the same checkpoint inhibitor or the same kinase inhibitor is administered to the patient during the induction phase and during the maintenance phase, the checkpoint inhibitor or kinase inhibitor may be administered at a lower dose and / or at a reduced frequency in the maintenance phase than in the induction phase therapy.

[0056] II. TUSC2 Therapy

[0057] The disclosure provides a method for treating cancer in a patient comprising an induction phase and a maintenance phase where a nucleic acid construct, a nonviral vector, a viral vector, or a composition as described herein comprising a TUSC2 encoding sequence is administered to the patient as part of the maintenance phase therapy or as part of the induction phase and maintenance phase therapies. In embodiment, dexamethasone and / or diphenhydramine are administered prior to administration of the TUSC2 therapy.

[0058] i. Nucleic Acid Constructs

[0059] Nucleic acid constructs described herein include a polynucleotide sequence encoding a TUSC2 protein (for example, human TUSC2 protein). The TUSC2 coding sequence is flanked by a 5' untranslated region (UTR) and a 3' UTR, and is operably linked to a promoter. In embodiments, the nucleic acid constructs further include a selectable marker. In a typical embodiment, the nucleic acid construct is used for recombinant production of human TUSC2 in a cancer cell (e.g., in a subject’s cancer cells). Nucleic acid constructs include expression constructs. The term “expression construct” refers to a nucleic acid construct that includes (i) a polynucleotide sequence coding for a TUSC2 protein; and (ii) one or more control / regulatory elements operably linked to the TUSC2 encoding sequence. Methods for constructing expression constructs and plasmids through standard recombinant techniques are known in the art. Methods for designing expression constructs / plasmids for gene therapy applications (e.g., DNA vaccines, immunotherapy), including antibiotic-free vector production, are also known. Various sequences and elements have been reported to increase and sustain therapeutic protein production (e.g., introns, Kozak consensus). Such sequences and elements are disclosed below under Control / Regulatory Sequences.

[0060] Expression constructs / plasmids for inclusion in the vectors described herein can be produced in suitable host producer cells (e.g., E. coif) using suitable methods, e.g., fed-batch fermentation, batch fermentation, etc. For example, the HyperGRO™ inducible fed-batch fermentation process is used commercially to manufacture plasmid DNA at Nature Technology Corporation (Lincoln, NE). The HyperGRO™ process yields plasmid productivity of up to 2,600 mg / L with low levels of nicking or multimerization. High yield of plasmid per gram of bacteria improves final product purity since plasmid is enriched relative to host cell impurities. Boehringer Ingelheim (Vienna, Austria) has developed an alternative high yield fermentation process which is commercially available for cGMP production of plasmid DNA vectors. Plasmid DNA can be extracted from producer cells using alkaline lysis. Commercial plasmid manufacture can utilize purification processes, such as anion exchange chromatography followed by hydrophobic interaction chromatography, that purify plasmid DNA away from impurities (e.g., endotoxin, bacterial RNA, genomic DNA).

[0061] In embodiments, the expression construct comprising the polynucleotide sequence encoding a TUSC2 protein is a covalently closed linear DNA (“doggybone DNA” or dbDNA”). dbDNA may be created starting with a circular double-stranded DNA molecule (e.g., a plasmid) containing a TUSC2 encoding sequence flanked on each side by 56 bp palindromic protelomerase recognition sequences. The DNA starting material is then denatured and Phi29 DNA polymerase is primed. Phi29 initiates rolling circle amplification of the template, creating double-stranded concatameric repeats of the original construct. Protelomerase is added, which binds to the recognition sites flanking the TUSC2 encoding sequence and performs a cleavage-joining reaction that results in monomeric doublestranded, linear, covalently closed DNA constructs. One of a panel of common restriction enzymes is added to cut undesired backbone DNA sequences, exposing open ended DNA that can be removed through digestion with exonuclease. dbDNA is purified from small fragments and reaction components with size separation to leave only the dbDNA construct comprising the TUSC2 encoding sequence. The resulting dbDNA construct can be used as a starting material for further amplification reactions. dbDNA constructs and methods of making them are disclosed in W02010086626 (PCT / GB2010 / 000165), incorporated herein by reference.

[0062] i.a. TUSC2 Polynucleotide and Amino Acid Sequences

[0063] The nucleic acid constructs described herein include a polynucleotide sequence encoding a TUSC2 protein. In embodiments, the TUSC2 protein is human TUSC2. The human TUSC2 amino acid sequence is provided as SEQ ID NO: 1, below.SEQ ID NO: 1 MGASGSKARGLWPFASAAGGGGSEAAGAEQALVRPRGRAVPPFVFTRRGSMFYDE DGDLAHEFYEETIVTKNGQKRAKLRRVHKNLIPQGIVKLDHPRIHVDFPVILYEV

[0064] The TUSC2 protein encoded by the TUSC2 polynucleotide sequence and expressed from the expression constructs described herein may be a truncated TUSC2 or may have one or more substitutions that retain a tumor suppressor function of the expressed protein. In embodiments the TUSC2 protein comprises amino acids 2-110 of SEQ ID NO: 1. In embodiments, the TUSC2 protein has greater than about 85%, greater than about 90%, greater than about 95%, or greater than about 99% sequence identity with the sequence of SEQ ID NO: 1.

[0065] In embodiments, the polynucleotide sequence encoding human TUSC2 is SEQ ID NO: 2, or is a polynucleotide sequence having greater than about 85%, greater than about 90%, or greater than about 95% sequence identity compared to the sequence of SEQ ID NO: 2. The human TUSC2 DNA coding sequence is provided as SEQ ID NO: 2, below.SEQ ID NO: 2 ATGGGCGCCAGCGGGTCCAAAGCTCGGGGCCTGTGGCCCTTCGCCTCGGCGGCC GGAGGCGGCGGCTCAGAGGCAGCAGGAGCTGAGCAAGCTTTGGTGCGGCCTCGG GGCCGAGCTGTGCCCCCCTTCGTATTCACGCGCCGCGGCTCTATGTTCTATGATG AGGATGGGGATCTGGCTCACGAGTTCTATGAGGAGACAATCGTCACCAAGAACG GGCAGAAGCGGGCCAAGCTGAGGCGAGTGCATAAGAATCTGATTCCTCAGGGCA TCGTGAAGCTGGATCACCCCCGCATCCACGTGGATTTCCCTGTGATCCTCTATGA GGTGTGA

[0066] In embodiments, the polynucleotide sequence encoding the TUSC2 protein has been codon optimized. In embodiments, the TUSC2 protein encoded by the polynucleotide sequence is human TUSC2 and includes the amino acid sequence of SEQ ID NO: 1, or an amino acid sequence having greater than about 90%, greater than about 95%, or greater than about 98%, or greater than about 99% sequence identity compared to the sequence of SEQ IDNO: 1. As used herein, the term “sequence identity” refers to the degree to which two sequences (e.g., peptide, polypeptide, nucleic acid, etc.) have the same sequential composition of monomer subunits.

[0067] i.b. Control / Regulatory Sequences

[0068] The nucleic acid constructs disclosed herein include control and regulatory sequences that are operably linked to the polynucleotide sequence encoding a TUSC2 protein. The nucleic acid constructs disclosed herein can include appropriate control sequences for expression of the human TUSC2 in human cancer cells. “Control sequences” include nucleic acid sequences necessary for replication of a vector in a producer cell (e.g., E. coli cell), as well as nucleic acid sequences necessary for, or involved in, transcription and / or translation of an operably linked polynucleotide coding sequence in a target cell (e.g., a human cancer cell). As used herein, the term “operably linked” refers to a physical or functional juxtaposition of the components so described as to permit them to function in their intended manner. In the example of an expression control element in operable linkage with a polynucleotide sequence encoding a TUSC2 protein, the relationship is such that the control element modulates expression of the TUSC2 protein encoding sequence. Examples of control / regulatory sequences include promoters, enhancers, translation initiation signals, termination signals, polyadenylation sequences (e.g., polyA signals derived from bovine growth hormone, SV40, rabbit P-globin), Kozak sequences (e.g., GCCACCATG; SEQ ID NO: 3), posttranslational regulatory elements, introns, splicing enhancers, nuclear targeting sequences, etc.

[0069] To facilitate expression of TUSC2 in the cancer cells, the nucleic acid constructs described herein include, for example, a suitable promoter operably linked to the TUSC2 transgene. In embodiments, the CMV promoter or a modified CMV promoter is used. Certain CMV promoters serve dual roles as a promoter and an enhancer. In embodiments, a mini-CMV promoter is used. In other embodiments, a chimeric promoter that is a fusion of two different promoter sequences or a fusion of a promoter sequence and an inducible element can be used. For example, a chicken P-actin / CMV enhancer combination can be used. Promoters, in addition to the CMV promoter, that can be used to promote transcription of the operably linked TUSC2 coding sequence include simian virus 40 (SV40) early promoter, elongation factor- la, (EFla), phosphoglycerate kinase (PGK), and human P-actin promoter (ACTB). In some embodiments, a tissue-specific promoter can be used. In some embodiments, a nucleic acid construct as described herein includes one or more (e.g., 1, 2, 3,4, 5, etc.) introns. For example, in a nucleic acid construct as disclosed herein, the 5' UTR, 3' UTR, and / or the TUSC2 coding sequence can include an intron (e.g., intron 2 of the human P globin gene). As another example, a chimeric intron (e.g., from the P-globulin and immunoglobulin heavy chain genes) upstream of the transgene can be used. Additionally or alternatively, the 5' UTR can include a HTLV-I R element for enhancement of mRNA translation efficiency and increasing transgene expression. Nuclear targeting sequences, which promote shuttling of the nucleic acid construct into the nucleus, can be included in the nucleic acid construct as described herein. MicroRNA target sites that mediate transgene expression in specific tissues or cell lineages and S / MAR regions that promote replication and long-term episomal transgene expression can also be included in some embodiments of a nucleic acid construct as described herein.

[0070] i.c. Selectable Markers

[0071] In embodiments, the nucleic acid constructs as disclosed herein include a selectable marker. A “selectable marker” as used herein is a nucleic acid sequence that confers a trait suitable for selection for a cell containing the nucleic acid construct.Selectable markers can include RNA selectable markers such as RNA-OUT (Luke et al., Vaccine 2009 vol. 27(46):6454-6459; Luke et al. Methods Mol Biol. 2014 vol. 1143:91-111), RNAI (US Patent No. 9297014), and suppressor tRNAs (Soubrier et al., Gene Therapy 1999 vol. 6: 1482-1488). RNA selectable markers are useful in applications where use of antibiotic-resistance markers is undesirable, including in production of nonviral vectors. For example, some regulatory agencies recommend avoiding inclusion of antibiotic resistance markers in DNA therapies administered to humans due to risk of unintended immune response and transmission of the antibiotic-resistant genes to the patient’s enteric bacteria. Thus, in some embodiments of a nucleic acid construct, the selectable marker is not an antibiotic resistance gene. In other embodiments, selectable markers can include an antibiotic resistance gene, for example, genes encoding resistance to ampicillin, chloramphenicol, tetracycline, or kanamycin. In some embodiments, the expression construct does not comprise a selectable marker (e.g., in certain embodiments in which a dbDNA construct is utilized).

[0072] ii. Nonviral Vectors

[0073] The term “vector” as used herein refers to a vehicle for delivering genetic material (e.g., RNA or DNA) to a cell, including for example, viral vectors (such as AAV andlentiviral vectors) and nonviral vectors. The term “nonviral vector” is used herein to refer to a nonviral vehicle for delivering genetic material to a cell. In embodiments, the nonviral vector comprises one or more carrier molecules (e.g., DOTAP: cholesterol liposome) complexed with a nucleic acid construct (e.g., a plasmid) as disclosed herein. The liposome formulations described herein deliver the nucleic acid construct into the target cell; entering target cells via endocytosis pathways to avoid lysosomal degradation. Once a liposome formulation binds to a negatively-charged cancer cell, the nucleic acid construct is transfected into the cell (endocytosis) and TUSC2 is expressed. The nonviral vectors described herein result in a high level of transfection efficiency with a low level of toxicity. The nonviral vectors display a high degree of specificity and protect against degradation of the nucleic acid construct by the target cell during transfection. The lipid formulations are designed for stability, increased half-life of the formulation and the prevention of aggregation of the lipid particles. In the liposomal nonviral vectors, the nucleic acid constructs can be added to liposomes in a range of concentrations. The ratio of the nucleic acid construct to lipids (liposomes) can be optimized for transfection efficiency. In embodiments, nucleic acid constructs are added to the liposomes at a concentration of 20, 25, 50, 75, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 225, 275, 300, 350, 375, 400, 450, 500, 550, 600, 650, 700, 750, 800, 850, 900, 950, or 1000 pg per 50, 100, 150, 200, 300, 400, 500, 600, 700, 800, 900, 1000, 1100, 1200, 1300, 1400, 1500, 2000, 2500, 3000, 3500, 4000, 4500, 5000, 5500, 6000, 7000, 8000, 9000, or 10,000 pl, as well as 15, 20, 25, 50 ml final volume. These concentrations can vary depending upon the ratio of the liposome components (e.g., DOTAP to cholesterol, cholesterol derivative or cholesterol mixture) in the particular liposome preparation. In some embodiments, equal volumes of nucleic acid construct and lipids (e.g., DOTAP:cholesterol liposome), at a concentration to obtain about 25 pg, 50 pg, 75 pg, 100 pg, 110 pg, 120 pg, 125 pg, 130 pg, 140 pg, 150 pg, 160 pg, 170 pg, 180 pg, 190 pg, 200 pg,210 pg, 220 pg, 225 pg, 230 pg, 240 pg, 250 pg, 260 pg, 270 pg, 275 pg, 280 pg, 290 pg,300 pg, 310 pg, 320 pg, 325 pg, 330 pg, 340 pg, 350 pg, 360 pg, 370 pg, 375 pg, 400 pg,425 pg, 450 pg, 500 pg, 550 pg, 600 pg, 650 pg, 700 pg, 750 pg, 800 pg, 850 pg, 900 pg,950 pg, or 1000 pg of nucleic acid per 1 mM, 2 mM, 3 mM, 4 mM, 5 mM, 6 mM, 7 mM, 8 mM, 9 mM, 10 mM, 11 mM, 12 mM, 13 mM, 14 mM, 15 mM, 16 mM, 17 mM, 18 mM, 19 mM, 20 mM, 22 mM, 24 mM, 26 mM, 28 mM, 30 mM, 32 mM, 34 mM, 36 mM, 38 mM, or 40 mM lipids per 50, 100, 150, 200, 300, 400, 500, 600, 700, 800, 900, 1000, 1100, 1200, 1300, 1400, 1500, 2000, 2500, 3000, 3500, 4000, 4500, 5000, 5500, 6000, 7000, 8000, 9000,or 10,000 pl, as well as 15, 20, 25, or 50 ml, are mixed by adding the nucleic acid construct rapidly to the surface of the lipid (e.g., DOTAP: cholesterol) solution followed by mixing.

[0074] The nonviral vectors disclosed herein are typically of an average particle size of between about 40 nm and about 250 nm (e.g., 39 nm, 40 nm, 50 nm, 60 nm, 70 nm, 80 nm, 90 nm, 100 nm, 110 nm, 120 nm, 130 nm, 140 nm, 150 nm, 160 nm, 170 nm, 180 nm, 190 nm, 200 nm, 250 nm, 251 nm). In some embodiments, the average mean particle size of the nonviral vector constructs is between 250 nm and 325 nm.

[0075] ii.a. DOTAP:Cholesterol Liposomes

[0076] DOTAP:cholesterol liposomes are nanoparticle liposomal formulations composed of l,2-bis(oleoyloxy)-3-(trimethyl ammonio) propane (DOTAP) and cholesterol (Templeton et al., Nat. Biotechnol., 1997 15:647-652, incorporated by reference herein). DOTAP:cholesterol liposomes form a stable structure and are efficient carriers of biologically active agents such as nucleic acid constructs. In embodiments, the liposomal formulation includes DOTAP in a concentration ranging from 1 to 8 millimolar (mM) (e.g., 1 mM, 2 to 7 mM, 3 to 6 mM, 4 to 5 mM, 8 mM). In embodiments, the liposomal formulation includes cholesterol or cholesterol derivative or cholesterol mixture in a concentration ranging from 1 to 8 mM (e.g., 1 mM, 2 to 7 mM, 3 to 6 mM, 4 to 5 mM, or 8 mM). In some embodiments of a nonviral vector, the DOTAP:cholesterol molar ratio is between about 3: 1 and about 1 :3 (e.g., about 3.1 : 1, about 3: 1, about 2.5: 1, about 2: 1, about 1.5: 1, about 1 : 1, about 1 : 1.5, about 1 :2, about 1 :2.5, about 1 :3, or about 1 :3.1). Methods of making DOTAP:cholesterol liposomes are known in the art. For example, extrusion, microfluidization, reverse phase evaporation, sonication, solvent (e.g., ethanol) injection, detergent dialysis, ether injection, and dehydration / rehydration may be utilized.

[0077] ii.b. Extrusion Techniques

[0078] The DOTAP:cholesterol liposomes described herein may be prepared, for example, by an extrusion method including the steps of heating, sonicating, and sequential extrusion of the lipids through filters of decreasing pore size, thereby resulting in the formation of small, stable liposome structures. In such methods, the production of liposomes often is accomplished by sonication or serial extrusion of liposomal mixtures after (i) reverse phase evaporation (ii) dehydration-rehydration (iii) detergent dialysis and (iv) thin film hydration. Methods of producing liposomes via extrusion are described in Templeton et al. (Nat. Biotechnol., 1997 15(7):647-52) and US Patent No. 10,293,056. In these methods,DNA:lipid complexes are prepared by diluting a given nucleic acid and lipids in 5% dextrose in water to obtain an appropriate concentration of nucleic acid and lipids in an isotonic solution. For example, DOTAP (cationic lipid) is mixed with cholesterol (neutral lipid) at about equimolar concentrations. This mixture of powdered lipids is then dissolved with a solvent such as chloroform. The lipid solution is dried to a thin film at 30°C for 30 minutes (using, e.g., a rotary evaporator). The thin film is further freeze dried under vacuum for 15 minutes. The film is hydrated with water containing 5% dextrose (w / v) to give a final concentration of about 20 mM DOTAP and about 20 mM cholesterol. The hydrated lipid film is rotated in a 50°C water bath for 45 minutes and then at 375°C for an additional 10 minutes. The mixture is left standing at room temperature overnight. The following day the mixture is sonicated for 5-8 minutes at 50°C. The sonicated mixture is transferred to a new vessel and is heated for 10 minutes at 50° C. This mixture is sequentially extruded through filters (e.g., syringe filters) of decreasing pore size (e.g., 1 pm, 0.45 pm, 0.2 pm, and 0.1 pm). The 0.2 pm and 0.1 pm filters can be, e.g., Whatman Anotop filters (Cat. #: 6809-2122 or equivalent). The filtrate can be stored at, e.g., 4°C under argon gas.

[0079] ii.c. Microfluidization Techniques

[0080] The DOTAP:cholesterol liposomes described herein may be produced using a microfluidization method. Microfluidization can be used, e.g., when consistently small (50 to 200 nm) and relatively uniform aggregates are desired. Large scale production of DOTAP:cholesterol liposomes by microfluidization are known in the art. Methods of manufacturing liposomes using microfluidization are described, for example, in US Patent Application No. 16 / 098619. In certain microfluidization methods, the liposomal suspension is pumped at high velocity through an inlet that is divided into two streams and progressively bifurcates. These streams eventually collide within an interaction chamber leading to the formation of smaller particles due to turbulence and pressure. Generally, in microfluidization methods, DOTAP:cholesterol liposomes are formed by a quick increase in polarity of the environment induced by rapid mixing of the two miscible phases. This rapid mixing induces supersaturation of lipid molecules which leads to the self-assembly of DOTAP: cholesterol liposomes. Microfluidic mixing methods include, for example: microfluidic mixing using a staggered herringbone mixer (SHM), in-line T-junction mixing, and microfluidic hydrodynamic mixing (MHF). MHF is a continuous-flow technique where, in the case of liposome production, lipids dissolved in an organic solvent are hydrodynamically focused using an aqueous phase. In T-junction mixing, rapid mixing occurs when the two inputstreams in the T-junction collide, resulting in a turbulent output flow. SHM is microfluidic mixing by chaotic advection. Similar to other microfluidic techniques, the main characteristic is controlled millisecond mixing of two miscible phases, for example, ethanol and an aqueous buffer. The structure of the SHM allows efficient wrapping of the two fluids around each other resulting in an exponential enlargement of the interface between the fluids ensuring rapid mixing.

[0081] ii.d. Methods of Making Nonviral Vectors

[0082] Once manufactured, DOTAP: cholesterol liposomes can be used to encapsulate nucleic acids (e.g., a polynucleotide construct as described herein) resulting in nonviral vectors as described herein. In some embodiments, a nonviral vector is prepared by diluting nucleic acid constructs and lipids (DOTAP:cholesterol) in 5% dextrose in water to obtain an appropriate concentration of nucleic acid constructs and lipids (DOTAP:cholesterol). The nucleic acid constructs can be added to the DOTAP: cholesterol liposomes in a range of concentrations as indicated above. For example, equal volumes of nucleic acid construct and DOTAP:cholesterol, at a concentration to obtain 100 pg of nucleic acid construct / 5 mM lipids / 100 pl, can be mixed by adding the nucleic acid construct rapidly to the DOTAP:cholesterol solution followed by rapid mixing.

[0083] In other methods, nonviral vectors can be produced using the heating, sonicating, and sequential extrusion methods described above. In some embodiments, nonviral vectors are produced using the microfluidization methods described above.

[0084] Once nonviral vectors are produced, they can be characterized using any suitable method. For example, mean particle size can be determined by dynamic light scattering using a particle size analyzer (e.g., a Malvern Zetasizer or Coulter N4 particle size analyzer).

[0085] iii. Viral Vectors

[0086] The term “viral vector” is used herein to refer to a recombinant viral vector for delivering genetic material (e.g., a polynucleotide sequence encoding a TUSC2 protein such as human TUSC2) into a cell. A recombinant viral vector comprises capsid or envelope proteins and a recombinant viral genome, which is a nucleic acid construct comprising components derived from a viral genome (e.g., AAV) and heterologous polynucleotide sequences (e.g., a polynucleotide sequence encoding TUSC2 protein). Examples of viral vectors include, but are not limited to, AAV vectors, retroviral vectors, lentiviral vectors, adenoviral vectors, herpesvirus vectors, alphavirus vectors, and the like.

[0087] A “recombinant AAV vector” or “rAAV vector” comprises a rAAV genome derived from the wild-type genome of AAV. Typically, for AAV, one or both inverted terminal repeat (ITR) sequences of the wild type AAV genome are retained in the rAAV vector. A recombinant viral genome can be packaged into a virus (also referred to herein as a “particle” or “virion”) for subsequent infection (transformation) of a cell, ex vivo, in vitro, or in vivo. Where a rAAV genome is encapsidated or packaged into an AAV particle, the particle can be referred to as a “rAAV.” Such particles or virions include proteins that encapsidate or package the viral genome. Particular examples include viral envelope proteins, and in the case of AAV, capsid proteins (VP1, VP2, VP3). As used herein, the term “serotype” refers to an AAV having a capsid that is serologically distinct from other AAV serotypes. Serologic distinctiveness is determined based on the lack of cross-reactivity between antibodies to one AAV as compared to another AAV. Such cross-reactivity differences are usually due to differences in capsid protein sequences / antigenic determinants (e.g., due to VP1, VP2, and / or VP3 sequence differences of AAV serotypes). Recombinant AAV vectors include AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAV11, RhlO, Rh74 or AAV-2i8, and variants thereof. Examples of rAAV can include capsid sequence of any of AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAV11, RhlO, Rh74 or AAV-2i8, or a capsid variant of AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAV11, RhlO, Rh74 or AAV-2i8, or a capsid variant of AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAV11, RhlO, Rh74 or AAV-2i8. Capsid variants include a capsid sequence with an amino acid substitution, deletion, or insertion / addition.

[0088] A rAAV vector can comprise a genome derived from an AAV serotype distinct from the AAV serotype of one or more of the capsid proteins that package the viral genome. rAAV particles (vectors) can include one or more capsid proteins from a different serotype, a mixture of serotypes, or hybrids or chimeras of different serotypes, such as a VP1, VP2 or VP3 capsid protein of AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAV11, RhlO, Rh74 or AAV-2i8 serotype. In some embodiments, an AAV serotype having a specific tissue tropism is used. rAAV can be produced using any suitable methods. Methods for large-scale production of rAAV are known and are described in Urabe M. J. (2006) Virol. 80: 1874-1885; Kotin R.M. (2011) Hum. Mol. Genet. 20:R2-6; Kohlbrenner E. et al. (2005) Mol. Ther. 12: 1217-1225; Mietzsch M. (2014) Hum. Gene Ther. 25:212-222; and U.S. Patent Nos. 6,436,392, 7,241,447, and 8,236,557.

[0089] III. Chemotherapy

[0090] In embodiments of the methods of treating cancer disclosed herein, the induction phase comprises administering one or more chemotherapeutic agents to the patient in need thereof. In embodiments, chemotherapeutic agents, include, but not limited to, an alkylating agent (e.g., cyclophosphamide, melphalan, temozolomide, or a platinum-based chemotherapeutic agent such as carboplatin, cisplatin, or oxaliplatin,), an antimetabolite (e.g., 5 -fluorouracil (5-FU), 6-mercaptopurine, cytarabine, gemcitabine, and methotrexate), an antitumor antibiotic (e.g., actinomycin-D, bleomycin, daunorubicin, and doxorubicin), and topoisomerase inhibitors (e.g., etoposide, irinotecan, teniposide, and topotecan).

[0091] In embodiments, the induction phase comprises administering to the patient a platinum-based chemotherapy selected from cisplatin, carboplatin, oxaliplatin, nedaplatin, triplatin tetranitrate, phenanthriplatin, picoplatin, satraplatin, or combinations thereof.

[0092] IV. Checkpoint Inhibitors

[0093] In embodiments, the induction phase and / or the maintenance phase of the methods of treating cancer disclosed herein include administration to the patient in need thereof one or more checkpoint inhibitors (also called immune checkpoint inhibitors). Examples of checkpoint inhibitors include PD-L1 inhibitors, PD-L2 inhibitors and PD-1 inhibitors. Other examples of checkpoint inhibitors include CTLA-4 inhibitors and LAG-3 inhibitors.

[0094] In embodiments, the one or more check point inhibitors is a PD-L1 inhibitor including for example an antibody that binds to PD-L1. In embodiments, the anti-PD-Ll antibody is atezolizumab (Tecentriq®), avelumab (Bavencio®), or durvalumab (Imfinzi®).

[0095] In embodiments, the one or more check point inhibitors is a PD-1 inhibitor including, for example, an antibody that binds PD-1. In embodiments, the anti -PD-1 antibody is pembrolizumab (Keytruda®), nivolumab (Opdivo®), cemiplimab (Libtayo®) or dostarlimab-gxly (Jemperli).

[0096] In embodiments, the one or more check point inhibitors is a CTLA-4 inhibitor including, for example, an antibody that binds to CTLA-4 such as ipilimumab (Yervoy®) and tremelimumab (Imjudo®). In embodiments, the CTLA-4 inhibitor (such as tremelimumab) is administered in combination with a PD-L1 inhibitor (such as durvalumab).

[0097] In embodiments, the one or more checkpoint inhibitors is a LAG-3 inhibitor including, for example, an antibody that binds to LAG-3 such as relatlimab. In embodiments, the anti -LAG-3 antibody is administered in combination with a PD-1 inhibitor, for exampleRelatlimab may be administered in combination with nivolumab (in a combination known as Opdualag™).

[0098] V. Tyrosine Kinase Inhibitors

[0099] In embodiments of the methods of treating cancer disclosed herein, the induction phase and / or the maintenance phase comprises administering one or more tyrosine kinase inhibitors to the patient in need thereof. In embodiments, the one or more tyrosine kinase inhibitors is an EGFR tyrosine kinase inhibitor such as cetuximab (Erbitux®), osimertinib (Tagrisso®), erlotinib (Tarceva®), gefitinib (Iressa®), panitumumab (Vectibix®), necitumumab (Portrazza™), mobocertinib (Exkivity™) and combinations thereof. In embodiments, the maintenance phase comprises, in addition to the administration of the nucleic acid encoding the TUSC2 protein, also administering one or more EGFR tyrosine kinase inhibitors. In embodiments, the one or more EGFR tyrosine kinase inhibitors is selected from cetuximab (Erbitux®), osimertinib (Tagrisso®), erlotinib (Tarceva®), gefitinib (Iressa®), panitumumab (Vectibix®), necitumumab (Portrazza™), and mobocertinib (Exkivity™).

[0100] In embodiments, the one or more tyrosine kinase inhibitors is a dual EGFR / HER2 tyrosine kinase inhibitor such as neratinib (Nerlynx®), dacomitinib (Vizimpro®), and lapatinib (Tykerb®).

[0101] VI. Compositions / Pharmaceutical Formulations

[0102] Compositions including the nucleic acid constructs, nonviral vectors, and viral vectors are described herein. In some embodiments, the composition includes a nonviral vector as described herein and dextrose, e.g., about 5% dextrose in water or saline. In other embodiments, the composition includes a nonviral vector as described herein and about 0.9% (e.g., 0.8%, 0.9%, 1.0%, etc.) sodium chloride. In additional embodiments, the composition includes a nonviral vector comprising a nucleic acid construct described herein and a combination of about 5% dextrose and about 0.9% sodium chloride.

[0103] The compositions, nucleic acid constructs, nonviral vectors and viral vectors described herein may be administered to mammals (e.g., rodents, humans, nonhuman primates, canines, felines, ovines, bovines) in a suitable formulation according to conventional pharmaceutical practice (see, e.g., Remington: The Science and Practice of Pharmacy (20th ed.), ed. A. R. Gennaro, Lippincott Williams & Wilkins, (2000) and Encyclopedia of Pharmaceutical Technology, eds. J. Swarbrick and J. C. Boylan, MarcelDekker, New York (1988-1999)). The nucleic acid constructs, nonviral vectors and viral vectors described herein are typically formulated in a composition comprising a pharmaceutically acceptable carrier. A description of exemplary pharmaceutically acceptable carriers and diluents, as well as pharmaceutical formulations, can be found in Remington. Other substances may be added to the compositions to stabilize and / or preserve the compositions. As used herein the terms “pharmaceutically acceptable” and “physiologically acceptable” mean a biologically acceptable formulation, gaseous, liquid, solid, or mixture thereof, which is suitable for one or more routes of administration, in vivo delivery or contact. A pharmaceutically acceptable or physiologically acceptable excipient is a material that is not biologically or otherwise undesirable, e.g., the material may be administered to a subject without causing substantial undesirable biological effects.

[0104] The compositions described herein may be in a form suitable for sterile injection. To prepare such a composition, the active therapeutic(s) (e.g., nonviral or viral vector) are dissolved or suspended in a parenterally acceptable liquid vehicle. Among acceptable vehicles, diluents and solvents that may be employed are water; water adjusted to a suitable pH by addition of an appropriate amount of a pH modifier (e.g., acid or base) or a suitable buffer; Ringer’s solution; isotonic sodium chloride solution; and dextrose solution. For example, in one embodiment, the vectors may be administered over 0.5 to several hours by infusion with a pharmaceutically acceptable diluent such as 5% dextrose in water, Ringer’s, and / or 0.5% NaCl. The aqueous formulation may also contain one or more preservatives (e.g., methyl, ethyl or n-propyl p-hydroxybenzoate). In cases where one of the therapeutics is only sparingly or slightly soluble in water, a dissolution enhancing or solubilizing agent can be added, or the solvent may include 10-60% w / w of propylene glycol or the like.

[0105] In other embodiments, the compositions described herein may be in a form suitable for intranasal administration. In one embodiment, the intranasal formulation is an aqueous formulation including a nucleic acid construct, nonviral vector or composition as described herein, a pH modifying agent, and a thickening agent. In the intranasal formulation, the pH modifying agent may provide or adjust the pH of the formulation to a suitable pH, e.g., a pH that assists in solubilizing an active agent in solution. In some embodiments, the intranasal formulation is administered as a stable intranasal spray that provides sufficient residence time on the nasal mucosa to allow trans-nasal absorption of the active agent(s). The thickening agent of the intranasal formulations described herein may modify the viscosity of the formulation to provide improved adherence of the formulation to the nasal mucosa without adversely affecting the ease of administration as an intranasal spray. The thickening agentmay additionally increase the residence time of the formulation on the nasal mucosa, reduce loss of the formulation via mucociliary clearance of the nasal passages and / or improve the trans-nasal absorption. Such intranasal formulations may provide a sustained release of a nonviral vector as described herein.

[0106] The nucleic acid constructs, nonviral vectors, viral vectors and compositions described herein are preferably administered to a mammal (e.g., human) in a therapeutically effective amount. By the phrases “therapeutically effective amount”, “effective amount” and “effective dosage” is meant an amount sufficient to produce a therapeutically (e.g., clinically) desirable result; for example, the result can include increasing or restoring TUSC2 express! on / signaling to TUSC2-deficient cancer cells, decreasing tumor size, eliminating a tumor, preventing or reducing metastasis, or maintaining remission in a subject. Dosage for a subject may depend on multiple factors, including the subject’s size, body surface area, age, the particular composition to be administered, time and route of administration, general health, and other drugs being administered concurrently. A delivery dose of a nucleic acid construct, nonviral vector, viral vector or composition as described herein is determined based on preclinical efficacy and safety. In some embodiments, a therapeutically effective amount of nonviral vector as described herein or a composition containing a therapeutically effective amount of the nonviral vector is injected intravenously. In other embodiments, a therapeutically effective amount of nonviral vector as described herein or a composition containing a therapeutically effective amount of the nonviral vector is administered intranasally. The nonviral vectors, viral vectors and compositions can be administered, for example, as a “unit dose.” A unit dose as used herein is defined as containing a predetermined quantity of the therapeutic agent calculated to produce the desired responses in association with its administration, i.e., the appropriate route and treatment regimen. A unit dose as described herein may be described in terms of nucleic acid mass (pg) of the nucleic acid construct in the lipid complex. Unit doses range from about 1, about 25, about 50, about 75, about 100, about 125, about 150, about 175, about 200, about 225, about 250, about 300, about 400, about 500, about 600, about 700, about 800, about 900, about 1000 pg, about 1500 pg, about 2000 pg, about 2500 pg, about 3000 pg and higher. In embodiments, unit doses range from about 500 pg to about 9000 pg; about 1000 pg to about 8000 pg; or about 2000 pg to about 5000 pg.

[0107] In embodiments, the TUSC2 nucleic acid construct is administered at about 5 to about 150, about 10 to about 120, about 20 to about 100, about 50 to about 90, about 10 to about 90, about 50 to about 100, or about 40 to about 80 pg / kg of body weight of the patient.

[0108] VII. Subject / Patient

[0109] The terms “patient,” “subject,” and “individual” are used interchangeably herein, and mean a mammalian (e.g., human) subject in need of treatment for cancer. Human subjects suffering from cancer include individuals suffering from various types of cancers, such as colon cancer, pancreatic cancer, breast cancer, melanoma, osteosarcoma, rectal cancer, lung cancer (e.g., small cell or Extensive Stage SCLC or non-small cell lung cancer), leukemia, and neuroblastoma. In the methods described herein, the subject can be undergoing surgery for any reason, such as for removal of diseased tissue, and / or radiation treatment. For example, in some embodiments of the methods described herein, the subject is undergoing, or has undergone, surgical resection of a tumor. As another example, in some embodiments of the methods described herein, the subject is undergoing, or has undergone, radiation treatment. As another example, in some embodiments of the methods described herein, the subject is undergoing, or has undergone, chemotherapy. In some embodiments of the methods described herein, the subject is undergoing, or has undergone, surgery (e.g., resection of a tumor) and / or radiation treatment and / or chemotherapy.

[0110] It is to be understood that this invention is not limited to the particular molecules, compositions, methodologies, or protocols described, as these may vary. Any methods and materials similar or equivalent to those described herein can be used in the practice or testing of embodiments of the present invention. It is further to be understood that the disclosure of the invention in this specification includes all possible combinations of such particular features. For example, where a particular feature is disclosed in the context of a particular aspect or embodiment of the invention, or a particular claim, that feature can also be used, to the extent possible, in combination with and / or in the context of other particular aspects and embodiments of the invention, and in the invention generally.[OHl] All other referenced patents and applications are incorporated herein by reference in their entirety. Furthermore, where a definition or use of a term in a reference, which is incorporated by reference herein is inconsistent or contrary to the definition of that term provided herein, the definition of that term provided herein applies and the definition of that term in the reference does not apply.

[0112] To facilitate a better understanding of the present invention, the following examples of specific embodiments are given. The following examples should not be read to limit or define the entire scope of the invention.EXAMPLES

[0113] Example 1: Phase 1 Clinical Trial of Liposome-Complexed TUSC2 Expression Construct and Atezolizumab Maintenance Therapy in Patients with Extensive Stage Small Cell Lung Cancer (ES-SCLC)

[0114] A Phase 1 clinical trial is conducted to evaluate the combination of a liposome- complexed TUSC2 expression construct (termed “L-TUSC2”) and atezolizumab as maintenance therapy for patients with ES-SCLC who did not develop tumor progression after receiving at least 3 cycles, and no more than 4 cycles, of induction therapy with carboplatin plus etoposide and atezolizumab. L-TUSC2 is a nonviral complex of DOTAP-C1: Cholesterol liposome solution (DC) and a DNA plasmid expressing TUSC2. The plasmid (pLJ 143 / KGB2 / TUSC2) is 3,895 base pairs (bp) and contains a kanamycin resistance gene, an origin of replication and the human wild-type TUSC2 gene driven by a cytomegalovirus promoter. The objectives and endpoints of the Phase 1 study are shown in Table 1.

[0115] Table 1. Study objectives and endpoints.

[0116] Study Design:

[0117] Up to 12 patients are enrolled. A 3+3 dose-finding design is used to determine the MTD / RP2D and evaluate the safety profile of L-TUSC2 in combination with a fixed dose of atezolizumab used as maintenance therapy. Patients receive L-TUSC2 up to 0.12 mg / kg (IV administration once every 21 days) and atezolizumab as maintenance therapy (1200 mg IV administration once every 21 days) until disease progression or unacceptable toxicity is experienced. The starting dose of L-TUSC2 is 0.09 mg / kg with potential escalation to 0.12 mg / kg. If L-TUSC2 0.09 mg / kg is not tolerated, de-escalation to 0.06 mg / kg is administered. All patients in each dose level (3-6 patients) are observed through the completion of the first 21 -day cycle before enrollment in the next dose level may begin. Atezolizumab is administered after L-TUSC2. In Cycle 1, the first dose of atezolizumab is administered on Day 2 following whole blood sample collection for PK; for all subsequent cycles, atezolizumab is administered on Day 1 following L-TUSC2 infusion and the 60-minute vital sign assessment.

[0118] Patients receive the following pre-treatment regimen no less than 30 minutes prior to each L-TUSC2 infusion:• Diphenhydramine 25-50 mg PO (oral) or IV (intravenous);• Acetaminophen 650-1000 mg PO or up to 10 mg / kg IV (or ibuprofen 200 mg tablets PO [max 400-600 mg]);• Dexamethasone 20 mg IV.

[0119] Study treatment is administered until disease progression, discontinuation due to adverse events, or discontinuation due to withdrawal of consent. Patients are followed for survival until death.

[0120] Inclusion Criteria:1. Male or female aged >18 years.2. Documented history of histologically or cytologically confirmed ES-SCLC, prior to starting treatment with the combination of atezolizumab, carboplatin, and etoposide.Note: ES-SCLC defined according to the Veterans Administration Lung Study Group (VLAG) staging system. Patients are not required to have measurable disease at the time of enrollment to be eligible. However, patients with measurable disease are evaluated using RECIST 1.1.3. Complete Response (CR), Partial Response (PR), or Stable Disease (SD) after receiving at least 3 cycles, and no more than 4 cycles, of atezolizumab, carboplatin, and etoposide.Note: No evidence of tumor progression after 3-4 cycles of atezolizumab, carboplatin, and etoposide.4. Eastern Cooperative Oncology Group performance status (ECOG PS) score from 0 to 1.5. Must be >28 days beyond major surgical procedures such as thoracotomy, laparotomy, or joint replacement, and must be >10 days beyond minor surgical procedures such as biopsy of subcutaneous tumors, pleuroscopy, etc., and must not have evidence of wound dehiscence, active wound infection, or comparable major residual complications of the surgery per Investigator assessment.Note: Placement of a pleural catheter despite being a minor surgical procedure, may be performed <10 days prior to enrollment.6. Asymptomatic brain metastases must meet ALL criteria of the following (a-d): a. No history of seizures in the preceding 6 months. b. Definitive treatment must be completed >21 days prior to enrollment. c. Must be off steroids administered because of brain metastases or related symptoms for >7 days. d. If had previous brain irradiation, post-treatment imaging must demonstrate stability or regression of the brain metastases.7. Absolute neutrophil count (ANC) >1500 / mm3, platelet count >100,000 / mm3within <21 days.8. Adequate renal function documented by serum creatinine of <1.5 mg / dL or calculated creatinine clearance >50 ml / min within <21 days.9. Adequate hepatic function as documented by serum bilirubin <1.5 mg / dL and aspartate aminotransferase (AST) and alanine aminotransferase (ALT) < 2.5 X upper limit of normal (ULN) within <21 days.10. Stable cardiac condition with a left ventricular ejection fraction >40% within <21 days.11. If female of childbearing potential (FOCBP), must have negative serum pregnancy test (serum beta-human chorionic gonadotropin [P-hCG]) within <7 days of first dose. Note: Non-childbearing is defined as greater than 1 year post-menopausal or surgically sterilized12. FOCBP and men who are sexually active with FOCBP must agree to use 2 forms of contraception including 1 highly effective and 1 effective methods during the study period and for 4 months following the last dose of study treatment.13. If male, must agree to no sperm donation during study treatment and for an additional 4 months following the last dose of study treatment.14. Must have voluntarily signed an informed consent in accordance with institutional policies.

[0121] Exclusion Criteria:1. Unable to tolerate atezolizumab treatment, leading to early treatment discontinuation or prolonged / frequent dosage modifications in previous atezolizumab treatment as determined by the Investigator.2. Received prior gene therapy.3. Received prophylactic cranial irradiation or consolidation thoracic radiation. Note: Prior cranial irradiation for brain metastases is allowed.4. Active systemic viral, bacterial, or fungal infection(s) requiring treatment.5. Serious concurrent illness or psychological, familial, sociological, geographical, or other concomitant conditions that, in the opinion of the Investigator, would not permit adequate follow-up and compliance with the study protocol.6. History of autoimmune disease requiring immunosuppression.7. History of myocardial infarction or unstable angina within <6 months.8. Known human immunodeficiency virus (HIV) infection or has active hepatitis infection.9. Female who is pregnant or breastfeeding.

[0122] The response criteria are shown in Table 2.

[0123] Table 2. Response Criteria.

[0124] It is expected that at least 1 evaluated dose for L-TUSC2 will have an estimated DLT (dose limiting toxicity) rate that is below 33%, when administered in combination with atezolizumab.

[0125] Example 2: Phase 2 Clinical Trial of Liposome-Complexed TUSC2 Expression Construct and Atezolizumab Maintenance Therapy in Patients with Extensive Stage Small Cell Lung Cancer (ES-SCLC)

[0126] A Phase 2 clinical trial is conducted to evaluate the efficacy of L-TUSC2 at the RP2D in combination with atezolizumab used as maintenance therapy. A single cohort of approximately 50 patients are enrolled and treated at the RP2D established in Phase 1, with the possibility of enrolling up to 10 additional patients with tumor biopsies after L-TUSC2administration (post-treatment biopsy). The objectives and endpoints of the Phase 2 study are shown in Table 3.

[0127] Table 3. Study objectives and endpoints.

[0128] Study Design:

[0129] Phase 2 patients receive L-TUSC2 at the RP2D (IV administration once every 21 days) in combination with atezolizumab 1200 mg (IV administration once every 21 days) until disease progression or unacceptable toxicity is experienced. Atezolizumab is administered after L-TUSC2. In Cycle 1, the first dose of atezolizumab is administered on Day 2 following whole blood sample collection for PK; for all subsequent cycles, atezolizumab is administered on Day 1 following L-TUSC2 infusion and the 60-minute vital sign assessment. Patients receive the following pre-treatment regimen no less than 30 minutes prior to each L-TUSC2 infusion:• Diphenhydramine 25-50 mg PO (oral) or IV (intravenous);• Acetaminophen 650-1000 mg PO or up to 10 mg / kg IV (or ibuprofen 200 mg tablets PO [max 400-600 mg]);• Dexamethasone 20 mg IV.

[0130] Ten patients in Phase 2 who consent to submit an archival tumor biopsy undergo a post-treatment biopsy on Cycle 1 Day 2, Day 3, or Day 4 after their first dose of L-TUSC2 (on C1D1). These archival and post-treatment biopsies are submitted to the central laboratory for IHC analysis to evaluate TUSC2 protein expression and NK cells. These 10 patients may also have an additional PK (pharmacokinetics) sample collected on the day of their posttreatment biopsy if the biopsy is completed on Cycle 1 Day 3 or Day 4. As necessary, enrollment in Phase 2 may continue (exceeding 50 patients) until a minimum of 10 posttreatment biopsies are submitted for IHC analysis.

[0131] The same inclusion and exclusion criteria described above in Example 1 is used for the phase 2 study. The response criteria is described above in Table 2.

[0132] It is expected that the addition of the MTD / RP2D of L-TUSC2 to atezolizumab will show improvement in the 18-week PFS rate compared to the historical rate for atezolizumab alone.

[0133] Example 3: Phase 1 Clinical Trial of Liposome-Complexed TUSC2 Expression Construct and Atezolizumab Maintenance Therapy in Patients with Extensive Stage Small Cell Lung Cancer (ES-SCLC).

[0134] Quaratusugene ozeplasmid (“quar oze”) gene therapy includes a lipoplex which delivers the TUSC2 gene to cancer cells, restoring TUSC2 expression. In this study quar oze was added to atezolizumab (“atezo”) maintenance therapy with the aim of improving PFS.

[0135] Methods:

[0136] Eligible patients had ES-SCLC and had completed 3-4 cycles of induction therapy with etoposide, a platin, and atezo. Maintenance therapy with quar oze was administered IV every 21 days in escalating doses and atezo 1200 mg was also administered IV every 21 days. Dexamethasone, acetaminophen, and diphenhydramine were given prior to each treatment to prevent post-infusion symptoms. Efficacy was evaluated after every even cycle of treatment using RECIST 1.1 criteria. Safety was evaluated using CTCAE v5, with dose limiting toxicities (DLTs) generally defined as > Gr 3 adverse events (AEs). Two dose levels (0.09, and 0.12 mg / kg) of quar oze were planned. A standard dose escalation with 3-6 pts / dose level was used. A Safety Review Committee (SRC) reviewed safety data and made recommendations on dose levels.

[0137] Results:

[0138] Six patients were enrolled (2M / 4F), median age 66, with 3 at 0.09 mg / kg, and 3 at 0.12 mg / kg. Adverse Events (AEs) that occurred during the first two cycles in more than 1 patient were chills, fever, hypotension, and sweating in 4, nausea in 3, and diarrhea, fatigue, and headache in 2 patients. There were no DLTs, and all Grade >3 AEs were considered to be unrelated. There was a delayed infusion-related reaction that included symptoms of chills, fever, hypotension, sweating, and nausea that started 3-4 hours after quar oze infusion and resolved over several hours with acetaminophen and diphenhydramine therapy. One patient at the 0.09 mg / kg dose level had a PR (30% decrease in target lesions) after 2 cycles of treatment. The decrease was maintained after 4 cycles of treatment, although progression of non-target lesions occurred after C4. A second patient had a 23% decrease in target lesions after 2 cycles of treatment, that was maintained after 4 cycles of treatment, and had a small change to 19% decrease after 6 cycles of treatment.

[0139] Table 4 - Efficacy in the Phase 1 Portion of the Acclaim-3 (ONC-005) Clinical Trial

[0140] Conclusions:

[0141] Quar oze was generally well tolerated with no DLTs. Quar oze administration was associated with a delayed infusion-related reaction including chills, fever, hypotension and sweating managed with prophylactic steroids, acetaminophen and diphenhydramine. The SRC met and determined that 0.12 mg / kg of quar oze combined with 1200 mg / kg of atezo was the RP2D. Among the 6 patients who received maintenance therapy with escalating doses of quar oze and standard doses of atezo, there were several patients with decreases in target lesions from the start of maintenance therapy which suggests activity of quar oze in this setting.

Claims

PCT ApplicationCLAIMSWe Claim:

1. A method for treating cancer in a patient comprising:(i) an induction phase comprising administering one or more anticancer therapies to the patient in need thereof; and(ii) a maintenance phase comprising administering to the patient a nucleic acid construct encoding TUSC2.

2. The method of claim 1, wherein the cancer is selected from colon cancer, pancreatic cancer, breast cancer, melanoma, osteosarcoma, rectal cancer, lung cancer, leukemia, and neuroblastoma.

3. The method of claim 2, wherein the cancer is non-small cell lung cancer (NSCLC).

4. The method of claim 2, wherein the cancer is small cell lung cancer (SCLC).

5. The method of claim 4, wherein the SCLC is Extensive Stage SCLC.

6. The method according to any one of claims 1 to 5, wherein the induction phase comprises administering one or more checkpoint inhibitors.

7. The method of claim 6, wherein the one or more checkpoint inhibitors is selected from an anti-PD-Ll antibody and / or an anti-PD-1 antibody.

8. The method of claim 7, wherein the one or more checkpoint inhibitors is an anti-PD- Ll antibody selected from atezolizumab, avelumab, and durvalumab.

9. The method of claim 7, wherein the one or more checkpoint inhibitors is an anti-PD-1 antibody selected from pembrolizumab, nivolumab, and cemiplimab.

10. The method according to any one of claims 1 to 9, wherein the induction phase comprises administering one or more platinum based chemotherapeutic agents selected from cisplatin, carboplatin, oxaliplatin, nedaplatin, triplatin tetranitrate, phenanthriplatin, picoplatin, and satraplatin.

11. The method according to any one of claims 1 to 10, wherein the induction phase comprises administering atezolizumab and a platinum-based chemotherapeutic agent (e.g., cisplatin).

12. The method of claim 1 or claim 2, wherein the induction phase comprises administering pembrolizumab and platinum-based chemotherapeutic agent (e.g., cisplatin).

13. The method according to any one of claims 1 to 12, wherein the induction phase comprises administering a nucleic acid construct encoding TUSC2 to the patient in need thereof.

14. The method according to any one of claims 1 to 13, wherein the maintenance phase comprises administering a checkpoint inhibitor.

15. The method of claim 14, wherein the one or more checkpoint inhibitors is selected from an anti-PD-Ll antibody and / or an anti-PD-1 antibody.

16. The method of claim 15, wherein the one or more checkpoint inhibitors is an anti-PD- Ll antibody selected from atezolizumab, avelumab, and durvalumab.

17. The method of claim 15, wherein the one or more checkpoint inhibitors is an anti-PD- 1 antibody selected from pembrolizumab, nivolumab, and cemiplimab.

18. The method according to any one of the preceding claims, wherein the nucleic acid construct encoding TUSC2 is administered in a viral or nonviral vector.

19. The method of claim 18, wherein the nonviral vector comprises a DOTAP: cholesterol liposome.

20. The method of claim 19, wherein the DOTAP:cholesterol ratio is between about 3 : 1 and about 1 :3.

21. The method of claim 18, wherein the polynucleotide construct is administered in a viral vector.

22. The method of claim 21, wherein the viral vector is an Adeno-Associated Virus (AAV) viral vector.

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