miR-155 harnesses Phf19 to potentiate cancer immunotherapy through epigenetic reprogramming of CD8+ T cell fate.

T cell senescence and exhaustion are major barriers to successful cancer immunotherapy. Here we show that miR-155 increases CD8+ T cell antitumor function by restraining T cell senescence and functional exhaustion through epigenetic silencing of drivers of terminal differentiation. miR-155 enhances Polycomb repressor complex 2 (PRC2) activity indirectly by promoting the expression of the PRC2-associated factor Phf19 through downregulation of the Akt inhibitor, Ship1. Phf19 orchestrates a transcriptional program extensively shared with miR-155 to restrain T cell senescence and sustain CD8+ T cell antitumor responses. These effects rely on Phf19 histone-binding capacity, which is critical for the recruitment of PRC2 to the target chromatin. These findings establish the miR-155-Phf19-PRC2 as a pivotal axis regulating CD8+ T cell differentiation, thereby paving new ways for potentiating cancer immunotherapy through epigenetic reprogramming of CD8+ T cell fate.

Differential Cytokine Utilization and Tissue Tropism Results in Distinct Repopulation Kinetics of Naïve vs. Memory T Cells in Mice.

Naïve and memory T cells co-exist in the peripheral T cell pool, but the cellular mechanisms that maintain the balance and homeostasis of these two populations remain mostly unclear. To address this question, here, we assessed homeostatic proliferation and repopulation kinetics of adoptively transferred naïve and memory T cells in lymphopenic host mice. We identified distinct kinetics of proliferation and tissue-distribution between naïve and memory donor T cells, which resulted in the occupancy of the peripheral T cell pool by mostly naïve-origin T cells in short term (<1 week), but, in a dramatic reversal, by mostly memory-origin T cells in long term (>4 weeks). To explain this finding, we assessed utilization of the homeostatic cytokines IL-7 and IL-15 by naïve and memory T cells. We found different efficiencies of IL-7 signaling between naïve and memory T cells, where memory T cells expressed larger amounts of IL-7Rα but were significantly less potent in activation of STAT5 that is downstream of IL-7 signaling. Nonetheless, memory T cells were superior in long-term repopulation of the peripheral T cell pool, presumably, because they preferentially migrated into non-lymphoid tissues upon adoptive transfer and additionally utilized tissue IL-15 for rapid expansion. Consequently, co-utilization of IL-7 and IL-15 provides memory T cells a long-term survival advantage. We consider this mechanism important, as it permits the memory T cell population to be maintained in face of constant influx of naïve T cells to the peripheral T cell pool and under competing conditions for survival cytokines.

Distinct IL-7 signaling in recent thymic emigrants versus mature naïve T cells controls T-cell homeostasis.

IL-7 is essential for T-cell survival but its availability is limited in vivo. Consequently, all peripheral T cells, including recent thymic emigrants (RTEs) are constantly competing for IL-7 to survive. RTEs are required to replenish TCR diversity and rejuvenate the peripheral T-cell pool. However, it remains unknown how RTEs successfully compete with resident mature T cells for IL-7. Moreover, RTEs express low levels of IL-7 receptors, presumably rendering them even less competitive. Here, we show that, surprisingly, RTEs are more responsive to IL-7 than mature naïve T cells as demonstrated by markedly increased STAT5 phosphorylation upon IL-7 stimulation. Nonetheless, adoptive transfer of RTE cells into lymphopenic host mice resulted in slower IL-7-induced homeostatic proliferation and diminished expansion compared to naïve donor T cells. Mechanistically, we found that IL-7 signaling in RTEs preferentially upregulated expression of Bcl-2, which is anti-apoptotic but also anti-proliferative. In contrast, naïve T cells showed diminished Bcl-2 induction but greater proliferative response to IL-7. Collectively, these data indicate that IL-7 responsiveness in RTE is designed to maximize survival at the expense of reduced proliferation, consistent with RTE serving as a subpopulation of T cells rich in diversity but not in frequency.

miR-155 augments CD8+ T-cell antitumor activity in lymphoreplete hosts by enhancing responsiveness to homeostatic γc cytokines.

Lymphodepleting regimens are used before adoptive immunotherapy to augment the antitumor efficacy of transferred T cells by removing endogenous homeostatic "cytokine sinks." These conditioning modalities, however, are often associated with severe toxicities. We found that microRNA-155 (miR-155) enabled tumor-specific CD8(+) T cells to mediate profound antitumor responses in lymphoreplete hosts that were not potentiated by immune-ablation. miR-155 enhanced T-cell responsiveness to limited amounts of homeostatic γc cytokines, resulting in delayed cellular contraction and sustained cytokine production. miR-155 restrained the expression of the inositol 5-phosphatase Ship1, an inhibitor of the serine-threonine protein kinase Akt, and multiple negative regulators of signal transducer and activator of transcription 5 (Stat5), including suppressor of cytokine signaling 1 (Socs1) and the protein tyrosine phosphatase Ptpn2. Expression of constitutively active Stat5a recapitulated the survival advantages conferred by miR-155, whereas constitutive Akt activation promoted sustained effector functions. Our results indicate that overexpression of miR-155 in tumor-specific T cells can be used to increase the effectiveness of adoptive immunotherapies in a cell-intrinsic manner without the need for life-threatening, lymphodepleting maneuvers.

CD200-expressing human basal cell carcinoma cells initiate tumor growth.

Smoothened antagonists directly target the genetic basis of human basal cell carcinoma (BCC), the most common of all cancers. These drugs inhibit BCC growth, but they are not curative. Although BCC cells are monomorphic, immunofluorescence microscopy reveals a complex hierarchical pattern of growth with inward differentiation along hair follicle lineages. Most BCC cells express the transcription factor KLF4 and are committed to terminal differentiation. A small CD200(+) CD45(-) BCC subpopulation that represents 1.63 ± 1.11% of all BCC cells resides in small clusters at the tumor periphery. By using reproducible in vivo xenograft growth assays, we determined that tumor initiating cell frequencies approximate one per 1.5 million unsorted BCC cells. The CD200(+) CD45(-) BCC subpopulation recreated BCC tumor growth in vivo with typical histological architecture and expression of sonic hedgehog-regulated genes. Reproducible in vivo BCC growth was achieved with as few as 10,000 CD200(+) CD45(-) cells, representing ~1,500-fold enrichment. CD200(-) CD45(-) BCC cells were unable to form tumors. These findings establish a platform to study the effects of Smoothened antagonists on BCC tumor initiating cell and also suggest that currently available anti-CD200 therapy be considered, either as monotherapy or an adjunct to Smoothened antagonists, in the treatment of inoperable BCC.