Treatment of Hepatocellular Carcinoma by Multimodal In Situ Vaccination Using Cryoablation and a Plant Virus Immunostimulant

Purpose: To test the hypothesis that cryoablation combined with intratumoral immunomodulating nanoparticles from cowpea mosaic virus (CPMV) as an in situ vaccination approach induces systemic antitumoral immunity in a murine model of hepatocellular carcinoma (HCC).Materials and Methods: Mice with bilateral, subcutaneous RIL-175 cell-derived HCCs were randomized to 4 groups: (a) phosphate-buffered saline (control), (b) cryoablation only (Cryo), (c) CPMV-treated only (CPMV), and (d) cryoablation plus CPMV-treated (Cryo + CPMV) (N = 11-14 per group). Intratumoral CPMV was administered every 3 days for 4 doses, with cryoablation performed on the third day. Contralateral tumors were monitored. Tumor growth and systemic chemokine/ cytokine levels were measured. A subset of tumors and spleens were harvested for immunohistochemistry (IHC) and flow cytometry. One-or 2-way analysis of variance was performed for statistical comparisons. A P value of <.05 was used as the threshold for statistical significance.Results: At 2 weeks after treatment, the Cryo and CPMV groups, alone or combined, outperformed the control group in the treated tumor; however, the Cryo + CPMV group showed the strongest reduction and lowest variance (1.6-fold & PLUSMN; 0.9 vs 6.3 fold & PLUSMN; 0.5, P < .0001). For the untreated tumor, only Cryo + CPMV significantly reduced tumor growth compared with control (9.2-fold & PLUSMN; 0.9 vs 17.8-fold & PLUSMN; 2.1, P = .01). The Cryo + CPMV group exhibited a transient increase in interleukin-10 and persistently decreased CXCL1. Flow cytometry revealed natural killer cell enrichment in the untreated tumor and increased PD-1 expression in the spleen. Tumor-infiltrating lymphocytes increased in Cryo + CPMV-treated tumors by IHC.Conclusions: Cryoablation and intratumoral CPMV, alone or combined, demonstrated potent efficacy against treated HCC tumors; however, only cryoablation combined with CPMV slowed the growth of untreated tumors, consistent with an abscopal effect.

Automated summary

The study aimed to test the effectiveness of cryoablation combined with intratumoral immunomodulating nanoparticles (CPMV) in inducing systemic antitumoral immunity in a murine model of hepatocellular carcinoma (HCC). The results showed that cryoablation combined with intratumoral CPMV was the most effective treatment for the treated tumors and also slowed the growth of untreated tumors.