A B S T R A C T PurposeDespite aggressive therapies, median survival for malignant gliomas is less than 15 months. Patients with unmethylated O 6 -methylguanine-DNA methyltransferase (MGMT) fare worse, presumably because of temozolomide resistance. AdV-tk, an adenoviral vector containing the herpes simplex virus thymidine kinase gene, plus prodrug synergizes with surgery and chemoradiotherapy, kills tumor cells, has not shown MGMT dependency, and elicits an antitumor vaccine effect. Patients and MethodsPatients with newly diagnosed malignant glioma received AdV-tk at 3 ϫ 10 10 , 1 ϫ 10 11 , or 3 ϫ 10 11 vector particles (vp) via tumor bed injection at time of surgery followed by 14 days of valacyclovir. Radiation was initiated within 9 days after AdV-tk injection to overlap with AdV-tk activity.Temozolomide was administered after completing valacyclovir treatment. ResultsAccrual began December 2005 and was completed in 13 months. Thirteen patients were enrolled and 12 completed therapy, three at dose levels 1 and 2 and six at dose level 3. There were no dose-limiting or significant added toxicities. One patient withdrew before completing prodrug because of an unrelated surgical complication. Survival at 2 years was 33% and at 3 years was 25%. Patient-reported quality of life assessed with the Functional Assessment of Cancer Therapy-Brain (FACT-Br) was stable or improved after treatment. A significant CD3 ϩ T-cell infiltrate was found in four of four tumors analyzed after treatment. Three patients with MGMT unmethylated glioblastoma multiforme survived 6.5, 8.7, and 46.4 months. ConclusionAdV-tk plus valacyclovir can be safely delivered with surgery and accelerated radiation in newly diagnosed malignant gliomas. Temozolomide did not prevent immune responses. Although not powered for efficacy, the survival and MGMT independence trends are encouraging. A phase II trial is ongoing.

ClinicalTrials.gov NCT00589875.

AdV-tk can be safely combined with pancreatic cancer SOC without added toxicity. Response and survival compare favorably to expected outcomes and immune activity increased. These results support further evaluation of GMCI with more modern chemoradiation and surgery as well as PD-1/PD-L1 inhibitors in pancreatic cancer.

Background. Gene-mediated cytotoxic immunotherapy (GMCI) is a tumor-specific immune stimulatory strategy implemented through local delivery of aglatimagene besadenovec (AdV-tk) followed by anti-herpetic prodrug. GMCI induces T-cell dependent tumor immunity and synergizes with radiotherapy. Clinical trials in adult malignant gliomas demonstrated safety and potential efficacy. This is the first trial of GMCI in pediatric brain tumors. Methods. This phase I dose escalation study was conducted to evaluate GMCI in patients 3 years of age or older with malignant glioma or recurrent ependymoma. AdV-tk at doses of 1 × 10 11 and 3 × 10 11 vector particles (vp) was injected into the tumor bed at the time of surgery followed by 14 days of valacyclovir. Radiation started within 8 days of surgery, and if indicated, chemotherapy began after completion of valacyclovir. Results. Eight patients (6 glioblastoma, 1 anaplastic astrocytoma, 1 recurrent ependymoma) were enrolled and completed therapy: 3 on dose level 1 and 5 on dose level 2. Median age was 12.5 years (range 7-17) and Lansky/Karnofsky performance scores were 60-100. Five patients had multifocal/extensive tumors that could not be resected completely and 3 had gross total resection. There were no dose-limiting toxicities. The most common possibly GMCI-related adverse events included Common Terminology Criteria for Adverse Events grade 1-2 fever, fatigue, and nausea/vomiting. Three patients, in dose level 2, lived more than 24 months, with 2 alive without progression 37.3 and 47.7 months after AdV-tk injection. Conclusions. GMCI can be safely combined with radiation therapy with or without temozolomide in pediatric patients with brain tumors and the present results strongly support further investigation. Clinical trial registry. ClinicalTrials.gov NCT00634231 Key Points1. Gene-mediated cytotoxic immunotherapy can be safely used in pediatric patients with brain tumors. 2. Durable progression-free responses are achievable in 2 of 5 patients at the highest dose level.

Gene-mediated cytotoxic immunotherapy (GMCI) is an immune strategy implemented through local delivery of an adenovirus-based vector expressing the thymidine kinase gene (aglatimagene besadenovec, AdV-tk) followed by anti-herpetic prodrug valacyclovir. A phase I dose escalation trial of GMCI followed by chemotherapy was conducted in patients with malignant pleural effusion (MPE). AdV-tk was administered intrapleurally (IP) in three cohorts at a dose of 1 × 10 to 10 vector particles. Primary endpoint was safety; secondary endpoints included response rate, progression-free survival, and overall survival. Nineteen patients were enrolled: median age 67 years; 14 with malignant mesothelioma, 4 non-small-cell lung cancer (NSCLC), and 1 breast cancer. There were no dose limiting toxicities. All 3 patients in cohort 2 experienced transient cytokine release syndrome (CRS). Addition of celecoxib in cohort 3 reduced the incidence and severity of CRS (none > grade 2). Three patients are alive (23-33 months after GMCI), and 3 of 4 NSCLC patients had prolonged disease stabilization; one is alive 29 months after GMCI, 3.6 years after initial diagnosis. GMCI was safe and well tolerated in combination with chemotherapy in patients with MPE and showed encouraging response. Further studies are warranted to determine efficacy.

A phase I-II study to evaluate gene mediated cytotoxic immunotherapy in newly diagnosed prostate cancer before radical prostatectomy was conducted in Monterrey, Mexico.MethodsTo investigate delivery of adenovirus to the prostate, fluorescently labeled vector was injected into fresh prostatectomy specimens and distribution visually analyzed. The optimal volume and site instillation was then used for transrectal ultrasound guided intraprostatic injection in 10 patients with adenocarcinoma scheduled for radical prostatectomy. Each received 2-apical and 2-basal 0.5 ml injections of AdV-tk for a total of 1×1011 vp followed by 14 days of prodrug. Nine patients continued to tumor resection: 6 high-risk, 1 intermediate and 2 low-risk. In-vivo vector distribution was analyzed from resected tissue of four patients. Patients were monitored for tumor progression and acute and long-term safety.ResultsTwo apical and two basal injections of 0.5ml led to optimal organ-wide distribution of an adenoviral vector ex-vivo and in-vivo. Cytotoxicity was evidenced by transient rise in PSA and tumor histology. There were no significant adverse events deemed related to the treatment and no late toxicities after median follow up of 11.3 years. All six high-risk patients had positive surgical margins and one had seminal vesicle involvement. Despite slow PSA rise post-surgery in 3 of these patients, none developed metastases. The intermediate and low-risk patients had complete resections and none have progressed.ConclusionIn-vivo transrectal ultrasound guided instillation of an adenoviral vector into four sites in the prostate was practical as an outpatient procedure, well tolerated and led to distribution throughout the intraprostatic tumor mass. AdV-tk demonstrated no significant acute or late toxicities. Trends in PSA and disease progression conveyed the possibility of a sustained immune response against residual disease.

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Gene-mediated cytotoxic immunotherapy (GMCI) is an immuno-oncology approach involving local delivery of a replication-deficient adenovirus expressing herpes simplex thymidine kinase (AdV-tk) followed by anti-herpetic prodrug activation that promotes immunogenic tumor cell death, antigen-presenting cell activation, and T cell stimulation. This phase I dose-escalation pilot trial assessed bronchoscopic delivery of AdV-tk in patients with suspected lung cancer who were candidates for surgery. A single intra-tumoral AdV-tk injection in three dose cohorts (maximum 10 12 viral particles) was performed during diagnostic staging, followed by a 14-day course of the prodrug valacyclovir, and subsequent surgery 1 week later. Twelve patients participated after appropriate informed consent. Vector-related adverse events were minimal. Immune biomarkers were evaluated in tumor and blood before and after GMCI. Significantly increased infiltration of CD8 + T cells was found in resected tumors. Expression of activation, inhibitory, and proliferation markers, such as human leukocyte antigen (HLA)-DR, CD38, Ki67, PD-1, CD39, and CTLA-4, were significantly increased in both the tumor and peripheral CD8 + T cells. Thus, intratumoral AdV-tk injection into non-small-cell lung cancer (NSCLC) proved safe and feasible, and it effectively induced CD8 + T cell activation. These data provide a foundation for additional clinical trials of GMCI for lung cancer patients with potential benefit if combined with other immune therapies.