Abstract 1793: Regional administration of class C CpG Oligodeoxynucleotides results in superior intrahepatic TLR9 activation and immunomodulation compared to systemic infusion
Authors: Chandra Ghosh, Kara Heatherton, Kyle O'Connell, Jason Laporte, Prajna Guha, David Jaroch, Bryan Cox, Steven C. Katz
Journal: Cancer Research (2021)

Abstract CpG oligodeoxynucleotides (ODNs) are TLR9 agonists (TLR9As) that stimulate pDC and B cells. Yet, multiple other cell types in the liver express TLR9. The effect of TLR9A in reshaping the tumor microenvironment (TME), particularly in the myeloid-derived suppressor cells (MDSC) compartment, is yet to be determined. Clinical success of TLR9As has been limited by toxicity when given systemically and delivery limitations when administered by direct needle injection. Here, we hypothesized that regional intra-vascular infusion of TRL9As would reprogram the liver MDSC compartment to enable immune control of liver metastases (LM). In this study, we evaluated the effect of class C ODN-2395 as TLR9A in inhibiting LM progression and its impact on the liver MDSC population. C57/BL6 mice were challenged with 2.5e6 MC38-CEA-Luc cells via intra-splenic route. After a week, mice were treated with 1, 3, 10 or 30 µg ODN2395 via PV or IV (30 µg). Bioluminescence was measured at 24 and 48 hrs post-ODN administration. CD45+ cells were isolated, and FACS analysis was performed to quantify MDSCs, monocytic MDSCs (M-MDSC), and M1-macrophage subsets, along with downstream signaling. We found that 30 µg ODN2395 delivered via PV compared to IV reduced tumor burden at 24 hrs and persisted up to 48 hrs. Also, 30 µg ODN2395 infused via PV was optimal in controlling MDSC, M-MDSC cells and polarized CD11b+F4/80+ monocytic cells towards pro-inflammatory/anti-tumorigenic M1 macrophage. Using an NFκB-dependent soluble alkaline phosphatase assay (SEAP), we determined that ODN2395 dose-dependently enhanced NFκB transcription factor activity (p&amp;lt;0.001). Western Blot data of tumor lysates show that ODN delivery by PV increased NFκB (pP65) activity and production of IL-6, significantly relative to IV. Overall, our data suggest that the regional delivery of a TLR9A has the potential to provide superior modulation of the TME within the liver. Table:Readout30µg IV30µg PVP valuenBioluminescence-24 h (photon/s)4e71e7&amp;lt;0.0113Bioluminescence-48h (photon/s)7e72e7&amp;lt;0.0513FACSCD11b+Gr1+ cells/CD45+ NPC (MDSC)29.70 %20.75%&amp;lt;0.0113CD11b+LY6C+ (M-MDSC)60.03%38.98%&amp;lt;0.0113CD11b+F4/80+CD38+EGR2-(M1 macrophage)34.82%58.20%&amp;lt;0.0113Western BlottingpNFκB/NFκB (pP65s276/pP65): Fold change0.862.028&amp;lt;0.016IL6/GAPDH Fold change12.387&amp;lt;0.056 Citation Format: Chandra Ghosh, Kara Heatherton, Kyle O'Connell, Jason Laporte, Prajna Guha, David Jaroch, Bryan Cox, Steven C. Katz. Regional administration of class C CpG Oligodeoxynucleotides results in superior intrahepatic TLR9 activation and immunomodulation compared to systemic infusion [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1793.