ENTPD1/CD39 as a predictive marker of treatment response to gemogenovatucel-T as maintenance therapy in newly diagnosed ovarian cancer.

Background: Broadened use of predictive molecular and phenotypic profiling amongst oncologists has facilitated optimal integration of targeted- and immuno-therapeutics into clinical care. However, the use of predictive immunomarkers in ovarian cancer (OC) has not consistently translated into clinical benefit. Vigil (gemogenovatucel-T) is a novel plasmid engineered autologous tumor cell immunotherapy designed to knock down the tumor suppressor cytokines, TGFß1 and TGFß2, augment local immune function via increased GMCSF expression and enhance presentation of clonal neoantigen epitopes. Methods: All patients enrolled in the VITAL trial (NCT02346747) of maintenance Vigil vs. placebo as front-line therapy with homologous recombination proficient (HRP) stage IIIB-IV newly diagnosed ovarian cancer underwent NanoString gene expression analysis. Tissue was obtained from surgically resected ovarian tumor tissue following surgical debulking. A statistical algorithm was used to analyze the NanoString gene expression data. Results: Using the NanoString Statistical Algorithm (NSA), we identify high expression of ENTPD1/CD39 (which functions as the rate-limiting step in the production of the immune suppressor adenosine from ATP to ADP) as a presumptive predictor of response to Vigil versus placebo regardless of HRP status on the basis of relapse free survival (median not achieved vs 8.1 months, p = 0.00007) and overall survival (median not achieved vs 41.4 months, p = 0.013) extension. Conclusion: NSA should be considered for application to investigational targeted therapies in order to identify populations most likely to benefit from treatment, in preparation for efficacy conclusive trials.

Treatment options are limited in patients with advanced stage ovarian cancer. Treatments that stimulate the immune system to target the cancer are sometimes effective, however determining which patients will have benefit has been difficult. It is therefore important to develop new markers to predict which patients will respond to therapy. In this study, we looked at the levels of a large number of genes in tumors from patients treated with Vigil (gemogenovatucel-T), a treatment that modifies patient's own tumor cells to activate the immune system. We demonstrate that high expression of a gene named ENTPD1/CD39 predicts a positive response to Vigil therapy. This finding could help clinicians to determine which patients might benefit from Vigil treatment and therefore might guide decisions on who should receive this treatment.

Corrigendum to "Long-term follow-up of Phase 2A trial results involving advanced ovarian cancer patients treated with Vigil® in frontline maintenance" [Gynecol. Oncol. Rep. 34 (2020) 100648].

[This corrects the article DOI: 10.1016/j.gore.2020.100648.].

Gemogenovatucel-T (Vigil) immunotherapy demonstrates clinical benefit in homologous recombination proficient (HRP) ovarian cancer.

OBJECTIVE: Recently, Vigil showed significant clinical benefit with improvement in relapse free (RFS) and overall survival (OS) in pre-planned subgroup analysis in stage III/IV newly diagnosed ovarian cancer patients with BRCA wild type (BRCA-wt) molecular profile. Here we analyze homologous recombination (HR) status of patients enrolled in the Phase 2b VITAL study and determine clinical benefit of Vigil in HR proficient (P) patients. METHODS: Patients were previously enrolled in a Phase 2b, double-blind, placebo-controlled trial. All were in complete response with Stage III/IV high grade serious, endometroid or clear cell ovarian cancer. HR status was determined using MyChoice®CDx score (<42 = HRP) (Myriad Genetics, Inc., UT). Post-hoc analyses were carried out using Kaplan Meier and restricted mean survival time (RMST) analysis to evaluate RFS and OS based on HR deficiency (D) status. RESULTS: RFS was improved with Vigil (n = 25) in HRP patients compared to placebo (n = 20) (HR = 0.386; 90% CI 0.199-0.750; p = 0.007), results were verified by RMST (p = 0.017). Similarly, OS benefit was observed in Vigil group compared to placebo (HR = 0.342; 90% CI 0.141-0.832; p = 0.019). Results with OS were also verified with RMST (p = 0.008). CONCLUSION: Vigil exhibited clinical benefit in HRP molecular profile patients.

Chimeric Antigen Receptor T-Cell Therapy: Reach to Solid Tumor Experience.

Chimeric antigen receptor (CAR) modified T-cell therapy, a unique platform technology highlighting precision medicine through utilization of molecular biology and cell-based therapeutics has shown unprecedented rates in patients with hematological malignancies such as acute lymphocyte leukemia, non-Hodgkin's lymphoma and multiple myeloma (MM). With the approval of CD19-targeted CAR T-cells by the Food and Drug Administration in acute lymphoblastic leukemia (ALL) and NHL, this technology is positioned for aggressive expansion to combination therapeutic opportunities and proof of principle towards utility in other malignant disorders. However, despite the impressive results seen with hematological malignancies, CAR T-cells have shown limited efficacy in solid tumors with several unsuccessful preclinical studies. Regardless, these attempts have provided us with a better understanding of the imminent challenges specific to solid tumors even if they have not so far led to expanded clinical treatment opportunities outside ALL/NHL/MM. This review summarizes our current understanding of CAR T-cell mechanism of action, while presenting the major limitations of CAR T-cell derived treatments in solid tumors. We further discuss recent findings and present new potential strategies to overcome the challenges facing solid tumor targeting by CAR T-cell platforms.

Bromodomain and extra-terminal motif inhibitors: a review of preclinical and clinical advances in cancer therapy.

Histone lysine acetylation is critical in regulating transcription. Dysregulation of this process results in aberrant gene expression in various diseases, including cancer. The bromodomain, present in several proteins, recognizes promotor lysine acetylation and recruits other transcription factors. The bromodomain extra-terminal (BET) family of proteins consists of four conserved mammalian members that regulate transcription of oncogenes such as MYC and the NUT fusion oncoprotein. Targeting the acetyl-lysine-binding property of BET proteins is a potential therapeutic approach of cancer. Consequently, following the demonstration that thienotriazolodiazepine small molecules effectively inhibit BET, clinical trials were initiated. We thus discuss the mechanisms of action of various BET inhibitors and the prospects for their clinical use as cancer therapeutics.

Evofosfamide for the treatment of human papillomavirus-negative head and neck squamous cell carcinoma.

Evofosfamide (TH-302) is a clinical-stage hypoxia-activated prodrug of a DNA-crosslinking nitrogen mustard that has potential utility for human papillomavirus (HPV) negative head and neck squamous cell carcinoma (HNSCC), in which tumor hypoxia limits treatment outcome. We report the preclinical efficacy, target engagement, preliminary predictive biomarkers and initial clinical activity of evofosfamide for HPV-negative HNSCC. Evofosfamide was assessed in 22 genomically characterized cell lines and 7 cell line-derived xenograft (CDX), patient-derived xenograft (PDX), orthotopic, and syngeneic tumor models. Biomarker analysis used RNA sequencing, whole-exome sequencing, and whole-genome CRISPR knockout screens. Five advanced/metastatic HNSCC patients received evofosfamide monotherapy (480 mg/m2 qw × 3 each month) in a phase 2 study. Evofosfamide was potent and highly selective for hypoxic HNSCC cells. Proliferative rate was a predominant evofosfamide sensitivity determinant and a proliferation metagene correlated with activity in CDX models. Evofosfamide showed efficacy as monotherapy and with radiotherapy in PDX models, augmented CTLA-4 blockade in syngeneic tumors, and reduced hypoxia in nodes disseminated from an orthotopic model. Of 5 advanced HNSCC patients treated with evofosfamide, 2 showed partial responses while 3 had stable disease. In conclusion, evofosfamide shows promising efficacy in aggressive HPV-negative HNSCC, with predictive biomarkers in development to support further clinical evaluation in this indication.

KRAS mutant allele-specific expression knockdown in pancreatic cancer model with systemically delivered bi-shRNA KRAS lipoplex.

The KRAS oncogene, present in over 90% of pancreatic ductal adenocarcinomas, is most frequently the result of one of three gain-of-function substitution mutations of codon 12 glycine. Thus far, RAS mutations have been clinically refractory to both direct and selective inhibition by systemic therapeutics. This report presents the results of pre-clinical assessment of a lipoplex comprising a plasmid-encoded, modular bi-functional shRNA (bi-shRNA), which executes selective and multi-mutant allelic KRASG12mut gene silencing, encased within a fusogenic liposome systemic delivery vehicle. Using both a dual luciferase reporter system and a Restriction Fragment Length Polymorphism (RFLP) assay, selective discrimination of KRASG12mut from KRASwt was confirmed in vitro in PANC1 cells. Subsequently, systemic administration of the bi-shRNAKRAS fusogenic lipoplex into female athymic Nu/Nu mice bearing PANC1 xenografts demonstrated intratumoral plasmid delivery, KRASG12mut knockdown, and inhibition of tumor growth, without adverse effect. Clinical trials with the bi-shRNA lipoplex have been implemented.

Case Report: Immune-mediated Complete Response in a Patient With Recurrent Advanced Ewing Sarcoma (EWS) After Vigil Immunotherapy.

Ewing sarcoma is a highly resistant disease with a <10% chance of survival at 5 years after failure of frontline chemotherapy. This is a case report of an Ewing sarcoma patient with metastatic disease recurrence <2 years after standard chemotherapy/radiation who achieved a durable and sustained complete response after 2 series of treatments with Vigil (GMCSF/bi-shRNA furin DNA autologous tumor immunotherapy) serially manufactured from first and second recurrences with ELISPOT assay correlation. Results support justification of further testing of Vigil with ELISPOT assay as a biomarker to assess level of immune response and correlation with disease control.

Preclinical Biodistribution and Safety Evaluation of a pbi-shRNA STMN1 Lipoplex after Subcutaneous Delivery.

Stathmin-1 (STMN1) is a microtubule-destabilizing protein which is overexpressed in cancer. Its overexpression is associated with poor prognosis and also serves as a predictive marker to taxane therapy. We have developed a proprietary bi-functional shRNA (bi-shRNA) platform to execute RNA interference (RNAi)-mediated gene silencing and a liposome-carrier complex to systemically deliver the pbi-shRNA plasmids. In vitro and in vivo testing demonstrated efficacy and specificity of pbi-shRNA plasmid in targeting STMN1 (Phadke, A. P., Jay, C. M., Wang, Z., Chen, S., Liu, S., Haddock, C., Kumar, P., Pappen, B. O., Rao, D. D., Templeton, N. S., et al. (2011). In vivo safety and antitumor efficacy of bifunctional small hairpin RNAs specific for the human Stathmin 1 oncoprotein. DNA Cell Biol. 30, 715-726.). Biodistribution and toxicology studies in bio-relevant Sprague Dawley rats with pbi-shRNA STMN1 lipoplex revealed that the plasmid DNA was delivered to a broad distribution of organs after a single subcutaneous injection. Specifically, plasmid was detected within the first week using QPCR (threshold 50 copies plasmid/1 µg genomic DNA) at the injection site, lung, spleen, blood, skin, ovary (limited), lymph nodes, and liver. It was not detected in the heart, testis or bone marrow. No plasmid was detected from any organ 30 days after injection. Treatment was well tolerated. Minimal inflammation/erythema was observed at the injection site. Circulating cytokine response was also examined by ELISA. The IL-6 levels were induced within 6 h then declined to the vehicle control level 72 h after the injection. TNFα induction was transiently observed 4 days after the DNA lipoplex treatment. In summary, the pbi-shRNA STMN1 lipoplex was well tolerated and displayed broad distribution after a single subcutaneous injection. The pre-clinical data has been filed to FDA and the pbi-shRNA STMN1 lipoplex is being investigated in a phase I clinical study.

Phase II study of Vigil® DNA engineered immunotherapy as maintenance in advanced stage ovarian cancer.

OBJECTIVES: The majority of women with Stage III/IV ovarian cancer who achieve clinical complete response with frontline standard of care will relapse within 2years. Vigil immunotherapy, a GMCSF/bi-shRNA furin DNA engineered autologous tumor cell (EATC) product, demonstrated safety and induction of circulating activated T-cells against autologous tumor in Phase I trial Senzer et al. (2012, 2013) . Our objectives for this study include evaluation of safety, immune response and recurrence free survival (RFS). METHODS: This is a Phase II crossover trial of Vigil (1.0×107 cells/intradermal injection/month for 4 to 12 doses) in Stage III/IV ovarian cancer patients achieving cCR (normal imaging, CA-125≤35units/ml, physical exam, and no symptoms suggestive of the presence of active disease) following primary surgical debulking and carboplatin/paclitaxel adjuvant or neoadjuvant chemotherapy. Patients received Vigil or standard of care during the maintenance period. RESULTS: Forty-two patients were entered into trial, 31 received Vigil and 11 received standard of care. No≥Grade 3 toxicity related to product was observed. A marked induction of circulating activated T-cell population was observed against individual, pre-processed autologous tumor in the Vigil arm as compared to pre-Vigil baseline using IFNγ ELISPOT response (30/31 negative ELISPOT pre Vigil to 31/31 positive ELISPOT post Vigil, median 134 spots). Moreover, in correlation with ELISPOT response, RFS from time of procurement was improved (mean 826days/median 604days in the Vigil arm from mean 481days/median 377days in the control arm, p=0.033). CONCLUSION: In conjunction with the demonstrated safety, the high rate of induction of T-cell activation and correlation with improvement in RFS justify further Phase II/III assessment of Vigil.

Preclinical Justification of pbi-shRNA EWS/FLI1 Lipoplex (LPX) Treatment for Ewing's Sarcoma.

The EWS/FLI1 fusion gene is well characterized as a driver of Ewing's sarcoma. Bi-shRNA EWS/FLI1 is a functional plasmid DNA construct that transcribes both siRNA and miRNA-like effectors each of which targets the identical type 1 translocation junction region of the EWS/FLI1 transcribed mRNA sequence. Previous preclinical and clinical studies confirm the safety of this RNA interference platform technology and consistently demonstrate designated mRNA and protein target knockdown at greater than 90% efficiency. We initiated development of pbi-shRNA EWS/FLI1 lipoplex (LPX) for the treatment of type 1 Ewing's sarcoma. Clinical-grade plasmid was manufactured and both sequence and activity verified. Target protein and RNA knockdown of 85-92% was demonstrated in vitro in type 1 human Ewing's sarcoma tumor cell lines with the optimal bi-shRNA EWS/FLI1 plasmid. This functional plasmid was placed in a clinically tested, liposomal (LP) delivery vehicle followed by in vivo verification of activity. Type 1 Ewing's sarcoma xenograft modeling confirmed dose related safety and tumor response to pbi-shRNA EWS/FLI1 LPX. Toxicology studies in mini-pigs with doses comparable to the demonstrated in vivo efficacy dose resulted in transient fever, occasional limited hypertension at low- and high-dose assessment and transient liver enzyme elevation at high dose. These results provide the justification to initiate clinical testing.

Three-year Follow up of GMCSF/bi-shRNA(furin) DNA-transfected Autologous Tumor Immunotherapy (Vigil) in Metastatic Advanced Ewing's Sarcoma.

Ewing's sarcoma is a devastating rare pediatric cancer of the bone. Intense chemotherapy temporarily controls disease in most patients at presentation but has limited effect in patients with progressive or recurrent disease. We previously described preliminary results of a novel immunotherapy, FANG (Vigil) vaccine, in which 12 advanced stage Ewing's patients were safely treated and went on to achieve a predicted immune response (IFNγ ELISPOT). We describe follow-up through year 3 of a prospective, nonrandomized study comparing an expanded group of Vigil-treated advanced disease Ewing's sarcoma patients (n = 16) with a contemporaneous group of Ewing's sarcoma patients (n = 14) not treated with Vigil. Long-term follow-up results show a survival benefit without evidence of significant toxicity (no ≥ grade 3) to Vigil when administered once monthly by intradermal injection (1 × 10e(6) cells/injection to 1 × 10e(7) cells/injection). Specifically, we report a 1-year actual survival of 73% for Vigil-treated patients compared to 23% in those not treated with Vigil. In addition, there was a 17.2-month difference in overall survival (OS; Kaplan-Meier) between the Vigil (median OS 731 days) and no Vigil patient groups (median OS 207 days). In conclusion, these results supply the rational for further testing of Vigil in advanced stage Ewing's sarcoma.

A Phase II Efficacy and Safety, Open-Label, Multicenter Study of Imprime PGG Injection in Combination With Cetuximab in Patients With Stage IV KRAS-Mutant Colorectal Cancer.

BACKGROUND: Imprime PGG (ß(1,6)-[poly-(1,3)-D-glucopyranosyl]-poly-ß(1,3)-D-glucopyranose) is an innate immune cell modulator that primes neutrophils and monocytes/macrophages to exert antitumor activity against complement opsonized tumor cells. In patients with KRAS-mutant colorectal cancer (CRC), cetuximab alone is ineffective; however, it can bind to tumor cells and induce opsonization for recognition by Imprime PGG-bound innate immune cells. The primary objective of this study was to determine the antitumor activity of Imprime PGG in combination with cetuximab in patients with KRAS-mutant metastatic CRC. PATIENTS AND METHODS: The study had a 2-stage Simon optimal design with 80% power to detect a target objective response rate (ORR) of ≥10% at a 10% significance level. Patients received weekly Imprime PGG (4 mg/kg) and cetuximab (loading dose, 400 mg/m(2), then 250 mg/m(2)) intravenously. The primary end point was ORR; secondary end points included duration of response (DOR), time to progression (TTP), overall survival (OS), disease control rate, progression-free survival, and safety. Stage 1 of the study was to enroll 17 evaluable patients. RESULTS: One partial response (5.6%) was observed among 18 patients enrolled into stage 1. Median DOR was 4.2 months, TTP 2.7 months, and OS 6.6 months. Overall, observed toxicity was as expected from cetuximab alone. The most common (≥20%) adverse events related to Imprime PGG were fatigue (7 patients; 38.9%), infusion reaction (4 patients; 22.2%), and headache (4 patients; 22.2%). There was no Grade 4 toxicity nor treatment-related deaths. CONCLUSION: Imprime PGG in combination with cetuximab treatment in patients with KRAS-mutant CRC showed compelling, albeit modest, clinical activity. This study provides proof of principle that Imprime PGG, in combination with complement-activating antibodies, is associated with clinical activity.

Systemic versus local responses in melanoma patients treated with talimogene laherparepvec from a multi-institutional phase II study.

BACKGROUND: We previously reported that talimogene laherparepvec, an oncolytic herpes virus encoding granulocyte-macrophage colony-stimulating factor (GM-CSF), resulted in an objective response rate of 26 % in patients with advanced melanoma in a phase II clinical trial. The response of individual lesions, however, was not reported. Since talimogene laherparepvec is thought to mediate anti-tumor activity through both direct tumor cytolysis and induction of systemic tumor-specific immunity, we sought to determine the independent response rate in virus-injected and non-injected lesions. METHODS: Fifty patients with stage IIIC or IV melanoma were treated with talimogene laherparepvec in a multi-institutional single-arm open-label phase II clinical trial. In this study patients were treated until a complete response was achieved, all accessible tumors disappeared, clinically significant disease progression, or unacceptable toxicity. This report is a post hoc analysis of the systemic effects of talimogene laherparepvec in injected lesions and two types of uninjected lesions-non-visceral lesions and visceral lesions. RESULTS: Eleven of 23 patients (47.8 %) had a ≥ 30 % reduction in the total burden of uninjected non-visceral lesions, and 2 of 12 patients (16.7 %) had a ≥ 30 % reduction in the total burden of visceral lesions. Among 128 evaluable lesions directly injected with talimogene laherparepvec, 86 (67.2 %) decreased in size by ≥ 30 % and 59 (46.1 %) completely resolved. Of 146 uninjected non-visceral lesions, 60 (41.1 %) decreased in size by ≥ 30 %, the majority of which (44 [30.1 %]) completely resolved. Of 32 visceral lesions, 4 (12.5 %) decreased in size by ≥ 30 %, and 3 (9.4 %) completely resolved. The median time to lesion response was shortest for lesions that were directly injected (18.4 weeks), followed by uninjected non-visceral lesions (23.1 weeks) and visceral lesions (51.3 weeks), consistent with initiation of a delayed regional and systemic anti-tumor immune response to talimogene laherparepvec. CONCLUSIONS: These results support a regional and systemic effect of talimogene laherparepvec immunotherapy in patients with advanced melanoma.

Talimogene Laherparepvec Improves Durable Response Rate in Patients With Advanced Melanoma.

PURPOSE: Talimogene laherparepvec (T-VEC) is a herpes simplex virus type 1-derived oncolytic immunotherapy designed to selectively replicate within tumors and produce granulocyte macrophage colony-stimulating factor (GM-CSF) to enhance systemic antitumor immune responses. T-VEC was compared with GM-CSF in patients with unresected stage IIIB to IV melanoma in a randomized open-label phase III trial. PATIENTS AND METHODS: Patients with injectable melanoma that was not surgically resectable were randomly assigned at a two-to-one ratio to intralesional T-VEC or subcutaneous GM-CSF. The primary end point was durable response rate (DRR; objective response lasting continuously ≥ 6 months) per independent assessment. Key secondary end points included overall survival (OS) and overall response rate. RESULTS: Among 436 patients randomly assigned, DRR was significantly higher with T-VEC (16.3%; 95% CI, 12.1% to 20.5%) than GM-CSF (2.1%; 95% CI, 0% to 4.5%]; odds ratio, 8.9; P < .001). Overall response rate was also higher in the T-VEC arm (26.4%; 95% CI, 21.4% to 31.5% v 5.7%; 95% CI, 1.9% to 9.5%). Median OS was 23.3 months (95% CI, 19.5 to 29.6 months) with T-VEC and 18.9 months (95% CI, 16.0 to 23.7 months) with GM-CSF (hazard ratio, 0.79; 95% CI, 0.62 to 1.00; P = .051). T-VEC efficacy was most pronounced in patients with stage IIIB, IIIC, or IVM1a disease and in patients with treatment-naive disease. The most common adverse events (AEs) with T-VEC were fatigue, chills, and pyrexia. The only grade 3 or 4 AE occurring in ≥ 2% of T-VEC-treated patients was cellulitis (2.1%). No fatal treatment-related AEs occurred. CONCLUSION: T-VEC is the first oncolytic immunotherapy to demonstrate therapeutic benefit against melanoma in a phase III clinical trial. T-VEC was well tolerated and resulted in a higher DRR (P < .001) and longer median OS (P = .051), particularly in untreated patients or those with stage IIIB, IIIC, or IVM1a disease. T-VEC represents a novel potential therapy for patients with metastatic melanoma.