Targeting the Peripheral Nervous System for Hepatocellular Carcinoma Treatment: Insights from a Preclinical Study

Hepatocellular carcinoma (HCC) is highly resistant to anti-PD-I immunotherapies due to its immunosuppressive tumor microenvironment. A principal feature of HCC is it expresses high inter- and intra-tumoral sympathetic nerve infiltration. Modulating sympathetic signaling to the liver may represent a new therapeutic approach for HCC. This study aimed to investigate in a preclinical model the effects of sympathetic neurolysis combined with systemic checkpoint inhibition in HCC tumors. 

Materials and Methods:

An HCC model was established using orthotopic subcapsular implantation of 1 x 10^7 HCA-1 hepatoma cells into the livers of C3H mice through mini-laparotomy. These mice were then randomly divided into control, neurolysis, and neurolysis with anti-PD I group. A neurotoxin, 6-hydroxydopamine (6-OHDA), and anti-mouse PD-I were administered intraperitoneally at a dose of 200 mg/kg and 250 µL/mL, respectively, for five consecutive days. On day 7, the animals were euthanized, and samples of blood, liver, and spleen tissues were collected for analysis. Liver tumor sections were processed for conventional H&E staining and spatial phenotyping using Lunaphore/Opal staining. Flow cytometry and ScRNA analysis were employed to analyze various immune cells and chemokine in the liver tumor.

Results:

We found numerous nerve fiber clusters and a thick lining of sympathetic nerve fibers in the hilum, central veins, and portal triads in HCC bearing mice. Neurolysis of the sympathetic nerve signaling to the liver resulted in a 10-fold increase in the M1/M2 polarization ratio of intrahepatic myeloid cells, attenuating the recruitment of myeloid-derived suppressor cells. The synergistic treatment of 6-OHDA and Anti-PD-1 effectively reactivated the immune environment within the HCC tumor microenvironment by facilitating dendritic cells (DCs) activation and increasing cytotoxic CD8 T and B cells presence, which are crucial for effective anti-tumor responses.

Conclusion:

Neurolysis effectively modulates the local tumor immune microenvironment by shifting the M2 macrophage population, thereby enhancing the efficacy of immune checkpoint inhibition in HCC tumors.