IL-12 conditioning improves retrovirally mediated transduction efficiency of CD8+ T cells

Abstract

The ability to genetically modify T cells is a critical component to many immunotherapeutic strategies and research studies. However, the success of these approaches is often limited by transduction efficiency. Since retroviral vectors require cell division for integration, transduction efficiency is dependent on the appropriate activation and culture conditions for T cells. Naïve CD8+ T cells which are quiescent must be first activated to induce cell division to allow genetic modification. To optimize this process, we activated mouse T cells with a panel of different cytokines, including IL-2, IL-4, IL-6, IL-7, IL-12, IL-15 and IL-23, known to act on T cells. After activation, cytokines were removed, and activated T cells were retrovirally transduced. We found that IL-12 pre-conditioning of mouse T cells greatly enhanced transduction efficiency while preserving function and expansion potential. We also observed a similar transduction enhancing effect of IL-12 pre-conditioning on human T cells. These findings provide a simple method to improve the transduction efficiencies of CD8+ T cells.

Single-cell imaging of CAR T cell activity in vivo reveals extensive functional and anatomical heterogeneity

“…CD8 + T cells were isolated from lymph nodes of Ubi-GFP OT-I TCR RAG1 −/− or Ubi-GFP mice using a CD8-negative selection kit (Miltenyi). T cells were activated in plates coated with 2.5 µg anti-CD3 mAb (clone 17.A2; BioLegend) in the presence of 2.5 µg/ml soluble anti-CD28 mAb (clone 37.51; BioLegend) and 10 ng/ml murine IL-12 (Andrijauskaite et al, 2015; SRP3204; Sigma-Aldrich). Two rounds of spin-infection were performed at 24 and 48 h after T cell activation using retroviral particles supplemented with 8 µg/ml polybrene (Merck).…”

A cross-talk between CAR T cell subsets and the tumor microenvironment is essential for sustained cytotoxic activity

“…Polyclonal CD4 + and CD8 + T cells were purified from lymph nodes and spleens of male WT, UBC-GFP, Ifng −/− mice using CD4-negative and CD8-negative selection kits (Miltenyi Biotec). T cells were activated in plates coated with anti-CD3 monoclonal antibody (mAb) (2.5 g ml −1 ; clone 17.A2; BioLegend) in the presence of soluble anti-CD28 mAb (2.5 g ml −1 ; clone 37.51; BioLegend) and murine IL-12 (10 ng ml −1 ; I8523; Sigma-Aldrich) (60). Two rounds of spin infection were performed at 24 and 48 hours after T cell activation using retroviral particles supplemented with polybrene (8 g ml −1 ; Merck).…”

New Developments in T Cell Immunometabolism and Implications for Cancer Immunotherapy

“…IL-12, one of the critical cytokines known to promote the Th1 subset of T cells, has also improved T cell-mediated tumor control in pre-clinical models [ 176 ]. It enhances the retroviral transduction efficiency of TCRs while preserving the effector function and expansion potential of the transduced T cells [ 177 ]. CD8 + T cells activated with exogenous IL-12 have elevated IL-7 receptor expression and rely on IL-7 for persistence and antitumor immunity [ 178 ].…”