Randomized dose-finding clinical trial of oncolytic immunotherapeutic vaccinia JX-594 in liver cancer

Abstract

Oncolytic viruses and active immunotherapeutics have complementary mechanisms of action (MOA) that are both self amplifying in tumors, yet the impact of dose on subject outcome is unclear. JX-594 (Pexa-Vec) is an oncolytic and immunotherapeutic vaccinia virus. To determine the optimal JX-594 dose in subjects with advanced hepatocellular carcinoma (HCC), we conducted a randomized phase 2 dose-finding trial (n=30). Radiologists infused low- or high-dose JX-594 into liver tumors (days 1, 15 and 29); infusions resulted in acute detectable intravascular JX-594 genomes. Objective intrahepatic Modified Response Evaluation Criteria in Solid Tumors (mRECIST) (15%) and Choi (62%) response rates and intrahepatic disease control (50%) were equivalent in injected and distant noninjected tumors at both doses. JX-594 replication and granulocyte-macrophage colony-stimulating factor (GM-CSF) expression preceded the induction of anticancer immunity. In contrast to tumor response rate and immune endpoints, subject survival duration was significantly related to dose (median survival of 14.1 months compared to 6.7 months on the high and low dose, respectively; hazard ratio 0.39; P=0.020). JX-594 demonstrated oncolytic and immunotherapy MOA, tumor responses and dose-related survival in individuals with HCC.

Trial registration: ClinicalTrials.gov NCT00554372.

Figure 1

Radiographic antitumor activity after JX-594 therapy determined by dynamic MRI with a central radiographic reader blinded to treatment group. (a) Greatest decrease in the sum of the longest diameter (LD) of target tumors from baseline (mRECIST criteria) in livers of individual patients after JX-594 treatment. n = 22 patients who had measurable and evaluable tumors at baseline and at least one follow-up time point. A tumor with an increase >100% is indicated with a + above the bar. (b) Greatest decrease in tumor contrast enhancement or longest diameter from baseline (Choi criteria) in target tumors in livers of individual patients after JX-594 treatment. n = 24 patients who had measurable and evaluable tumors at baseline and at least one follow-up time point. A tumor with an increase >100% is indicated with a + above the bar. (c) Greatest decrease in the sum of the longest tumor diameter from baseline (RECIST criteria) in noninjected liver tumors of individual patients after JX-594 treatment. n = 8 evaluable. (d) Greatest decrease in tumor contrast enhancement or longest diameter from baseline (Choi criteria) in noninjected liver tumors of individual patients after JX-594 treatment. n = 8 evaluable. (e) Example of the effects of JX-594 on contrast enhancement and perfusion in an injected tumor (JX7-1401; high dose). (f) Example of the effects of JX-594 on the longest diameter of an injected tumor (complete mRECIST response) (JX7-1715; low dose). (g) Example of the effects of JX-594 on contrast enhancement and perfusion in a noninjected (distant) tumor (JX7-1403; high dose). The red circles (e–g) indicate the same (responding) tumors over time.

Figure 2

Laboratory evidence for JX-594 replication, transgene expression and GM-CSF protein function. (a) The mean (± s.e.m.) peak concentration of JX-594 (genomes measured by quantitative PCR (qPCR)) in blood after each treatment cycle (using blood obtained 15 min after the completion of treatment) by dose group (t test). IT, intratumoral. (b) The percentage of patients with evidence of β-gal transgene expression (+95% confidence interval (CI)) after JX-594 treatment (generation of antibodies (Ab) to the β-gal transgene product within 29 d of treatment is indicative of JX-594 replication, as β-gal protein expression is associated with virus replication) (Fisher’s exact test). (c) The percentage of patients with evidence of hGM-CSF transgene expression (+95% CI) on day 5 after JX-594 treatment (Fisher’s exact test). (d) Maximum induction of neutrophil concentration in blood after treatment cycle 1 by dose group (using blood obtained on days 5 and 15 after treatment). Black bars, low-dose JX-594; white bars, high-dose JX-594.

Figure 3

Laboratory, radiographic and biopsy evidence for JX-594– associated induction of anticancer immunity. (a) Antibody-mediated complement-dependent cytotoxicity induction after JX-594 therapy in HCC (n = 4), normal (n = 2; HUVEC and MRC-5) and non-HCC (RCC, n = 3; melanoma, n = 4) cell lines. Each graph shows the mean percentage cell viability (+s.d.) after incubation with each individual patient’s serum (diluted to 5%) collected on day 43 after the initiation of treatment compared to baseline. JX7-1704 and JX7-1702, two high-dose patients. (b) Antibody-mediated CDC induction against HCC cell lines in individual patients over time after JX-594 therapy (serum diluted to 5%; mean ± s.d.). The data shown are from serum of patients that induced ≤50% cell viability (>50% cell killing) on at least one follow-up time point. (c) Radiographic evidence of progressive necrosis and peripheral enhancement over time in noninjected tumors (JX7-0307; low dose). (d) H&E staining of a biopsy sample from a tumor collected from patient JX7-0301 (low dose) 1.5 years after the initiation of JX-594 treatment. Scale bars, 100 αm (low magnification); 50 αm (high-magnification inset). (e) Radiographic evidence of progressive necrosis and peripheral enhancement over time in a noninjected tumor (JX7-0301; low dose). Red circles (c,e) indicate the same (responding) tumors over time. (f) ELISPOT analysis detecting T cells producing interferon-γ in response to stimulation with β-gal peptides at baseline and after JX-594 treatment; data are expressed as the mean number of spot-forming cells (SFC) per 10⁵ cells (+s.d.) (JX7-0310, low dose; JX7-0311, high dose). (−), negative control peptide. The P values in a and f were calculated by t test.

Figure 4

Kaplan-Meier analysis of overall survival. (a) Overall survival in the entire evaluable study population (dashed line) and by dose group (n = 29 total). (b) Overall survival in subjects with multiple tumors at baseline (ten high-dose and nine low-dose subjects). (c) Overall survival in the entire evaluable study population by the presence or absence of baseline neutralizing antibody status. (d) Overall survival in high-dose subjects having previously failed systemic therapy (n = 6).