403 Tumor-infiltrating lymphocytes (TIL) with inducible and membrane-bound IL-12 exhibit superior antitumor activityin vitro

Background

TIL cell therapy has shown clinical benefit for patients with solid tumors.^{1 2} However, an immunosuppressive tumor microenvironment (TME) may abrogate the full potential of TIL cell therapy.³ The proinflammatory cytokine IL-12, known for its capability to increase IFN-γ production and promote type 1 immune responses, reshapes the TME and has a potential to augment antitumor activity. Here, we report genetic engineering of TIL with an inducible and membrane-bound IL-12, incorporated into a 22-day manufacturing process, which showed superior cytotoxic function in vitro.

Methods

Tumor tissue from several solid tumor types, including non-small cell lung, breast, head and neck, and ovarian cancers, was fragmented and cultured, followed by transduction with lentivirus containing a gene encoding membrane-bound IL-12 via an NFAT-promoter (TeIL-12), and a rapid expansion protocol (REP). Expression, biological function, and shedding of IL-12 molecule were assessed in vitro. Immune phenotype and in vitro cytotoxic activity of TeIL-12 gene-engineered TIL were examined in various assays.

Results

Lentiviral gene transfer resulted in TIL expressing IL-12 on their surface via a membrane anchor. This modification potentiated IL-12 receptor downstream signaling activation in a contact-dependent manner in assays using HEK-IL-12-Blue reporter cells. TeIL-12-TIL exhibited increased IFN-γ production and superior cytotoxicity in a KILR assay. An xCELLigence-based cytotoxicity assay further confirmed increased killing with TIL that were unstimulated or stimulated. Moreover, phenotype profiling revealed TeIL-12-TIL were less differentiated, with reduced expression of immune-inhibitory receptors and increased production of cytotoxic molecules associated with antitumor activity. IL-12 shedding in the co-culture supernatant was minimal.

Conclusions

The enhanced in vitro killing activity, combined with a less-differentiated phenotype of TIL with inducible and membrane-bound IL-12 suggests a potential for increased clinical efficacy. Further, minimal IL-12 shedding supports reduced potential for IL-12-associated toxicity.⁴ Together, these data support investigation of TeIL-12 in vivo and subsequent clinical development.

Acknowledgements

Editorial support was provided by Amanda Kelly (Iovance Biotherapeutics).

References

Chesney J, Lewis KD, Kluger H, et al. Efficacy and safety of lifileucel, a one-time autologous tumor-infiltrating lymphocyte (TIL) cell therapy, in patients with advanced melanoma after progression on immune checkpoint inhibitors and targeted therapies: pooled analysis of consecutive cohorts of the C- 144–01 study. Journal for ImmunoTherapy of Cancer 2022;10:e005755. Schoenfeld AJ, Lee S, Paz-Ares L, et al. First phase 2 results of autologous tumor-infiltrating lymphocyte (TIL; LN-145) monotherapy in patients with advanced, immune checkpoint inhibitor-treated, non-small cell lung cancer (NSCLC). SITC Annual Meeting 2021. Granhøj JS, Jensen AWP, Presti M, et al. Tumor-infiltrating lymphocytes for adoptive cell therapy: recent advances, challenges, and future directions. Expert Opin Biol Th. 2022;1–15. Zhang L, Morgan RA, Beane JD, et al. Tumor-Infiltrating Lymphocytes Genetically Engineered with an Inducible Gene Encoding Interleukin-12 for the Immunotherapy of Metastatic Melanoma. Clin Cancer Res. 2015;21:2278–88.

Ethics Approval

The study was approved by the institutional review board at each site and was conducted in accordance with the Declaration of Helsinki and Good Clinical Practice guidelines of the International Conference on Harmonization. All patients provided written informed consent.