Jacob S. Bowers

Resident memory T cells in the skin mediate durable immunity to melanoma

Resident memory CD8 T cells maintained in vitiligo-affected skin mediate long-lived protection against melanoma. Resident memory to cancer Melanoma patients with vitiligo are more likely to have a positive outcome, but the mechanism behind this association has remained unclear. Now, Malik et al. report that skin-resident memory T (TRM) cells specific to melanoma antigens are maintained in vitiligo-affected skin. These cells persist and function independently of the lymphoid compartment, suggesting that the vitiligo lesions provide a niche for the TRM cells. The TRM cells provide durable memory to the tumor, even in pigmented skin. These data suggest that skin TRM cells are critical for maintaining antitumor immunity. Tissue-resident memory T (TRM) cells have been widely characterized in infectious disease settings; however, their role in mediating immunity to cancer remains unknown. We report that skin-resident memory T cell responses to melanoma are generated naturally as a result of autoimmune vitiligo. Melanoma antigen–specific TRM cells resided predominantly in melanocyte-depleted hair follicles and were maintained without recirculation or replenishment from the lymphoid compartment. These cells expressed CD103, CD69, and CLA (cutaneous lymphocyte antigen), but lacked PD-1 (programmed cell death protein–1) or LAG-3 (lymphocyte activation gene–3), and were capable of making IFN-γ (interferon-γ). CD103 expression on CD8 T cells was required for the establishment of TRM cells in the skin but was dispensable for vitiligo development. CD103+ CD8 TRM cells were critical for protection against melanoma rechallenge. This work establishes that CD103-dependent TRM cells play a key role in perpetuating antitumor immunity.

The Inducible Costimulator Augments Tc17 Cell Responses to Self and Tumor Tissue

The inducible costimulator (ICOS) plays a key role in the development of Th17 cells, but its role in the development and antitumor activity of IL-17–producing CD8+ T cells (Tc17) remains unknown. We found that ICOS costimulation was important for the functional maintenance, but not differentiation, of Tc17 cells in vitro. Blocking the ICOS pathway using an antagonist mAb or by using recipient mice genetically deficient in the ICOS ligand reduced the antitumor activity of adoptively transferred Tc17 cells. Conversely, activating Tc17 cells with an ICOS agonist in vitro enhanced their capacity to eradicate melanoma and induce autoimmune vitiligo when infused into mice. However, ICOS stimulation did not augment the antitumor activity of IL-2 expanded T cells. Additional investigation revealed that ICOS stimulation not only increased IL-2Rα, CXCR3, and IL-23R expression on Tc17 cells, but also dampened their expression of suppressive molecule CD39. Although Tc17 cells activated with an ICOS agonist cosecreted heightened IL-17A, IL-9, and IFN-γ, their therapeutic effectiveness was critically dependent on IFN-γ production. Depletion of IL-17A and IL-9 had little impact on antitumor Tc17 cells activated with an ICOS agonist. Collectively, our work reveals that the ICOS pathway potentiates the antitumor activity of adoptively transferred Tc17 cells. This work has major implications for the design of vaccine, Ab and cell-based therapies for autoimmunity, infectious disease, and cancer.

Dendritic Cells in Irradiated Mice Trigger the Functional Plasticity and Antitumor Activity of Adoptively Transferred Tc17 Cells via IL12 Signaling

Purpose: The adoptive cell transfer (ACT) of CD8+ T cells is a promising treatment for advanced malignancies. Lymphodepletion before ACT enhances IFNγ+CD8+ T cell (Tc0)–mediated tumor regression. Yet, how lymphodepletion regulates the function and antitumor activity of IL17A+CD8+ T cells (Tc17) is unknown. Experimental Design: To address this question, pmel-1 CD8+ T cells were polarized to secrete either IL17A or IFNγ. These subsets were then infused into mice with B16F10 melanoma that were lymphoreplete [no total body irradiation (TBI)], or lymphodepleted with nonmyeloablative (5 Gy) or myeloablative (9 Gy with hematopoietic stem cell transplantation) TBI. The activation of innate immune cells and function of donor T-cell subsets were monitored in recipient mice. Results: Tc17 cells regress melanoma in myeloablated mice to a greater extent than in lymphoreplete or nonmyeloablated mice. TBI induced functional plasticity in Tc17 cells, causing conversion from IL17A to IFNγ producers. Additional investigation revealed that Tc17 plasticity and antitumor activity were mediated by IL12 secreted by irradiated host dendritic cells (DC). Neutralization of endogenous IL12 reduced the antitumor activity of Tc17 cells in myeloablated mice, whereas ex vivo priming with IL12 enhanced their capacity to regress melanoma in nonmyeloablated animals. This, coupled with exogenous administration of low-dose IL12, obviated the need for host preconditioning, creating curative responses in nonirradiated mice. Conclusions: Our findings indicate that TBI-induced IL12 augments Tc17 cell–mediated tumor immunity and underline the substantial implications of in vitro preparation of antitumor Tc17 cells with IL12 in the design of T-cell immunotherapies. Clin Cancer Res; 21(11); 2546–57. ©2015 AACR.

Human CD26 high T cells elicit tumor immunity against multiple malignancies via enhanced migration and persistence

CD8+ T lymphocytes mediate potent immune responses against tumor, but the role of human CD4+ T cell subsets in cancer immunotherapy remains ill-defined. Herein, we exhibit that CD26 identifies three T helper subsets with distinct immunological properties in both healthy individuals and cancer patients. Although CD26neg T cells possess a regulatory phenotype, CD26int T cells are mainly naive and CD26high T cells appear terminally differentiated and exhausted. Paradoxically, CD26high T cells persist in and regress multiple solid tumors following adoptive cell transfer. Further analysis revealed that CD26high cells have a rich chemokine receptor profile (including CCR2 and CCR5), profound cytotoxicity (Granzyme B and CD107A), resistance to apoptosis (c-KIT and Bcl2), and enhanced stemness (β-catenin and Lef1). These properties license CD26high T cells with a natural capacity to traffic to, regress and survive in solid tumors. Collectively, these findings identify CD4+ T cell subsets with properties critical for improving cancer immunotherapy.The role of human CD4+ T cell subsets in cancer immunotherapy is still unclear. Here, the authors show that CD26 identifies three CD4+ T cell subsets with distinct immunological properties in both healthy individuals and cancer patients.

Th17 cells are refractory to senescence and retain robust antitumor activity after long-term ex vivo expansion

Adoptive immunotherapy for solid tumors relies on infusing large numbers of T cells to mediate successful antitumor responses in patients. While long-term rapid-expansion protocols (REPs) produce sufficient numbers of CD8+ T cells for treatment, they also cause decline in the cells therapeutic fitness. In contrast, we discovered that IL-17-producing CD4+ T cells (Th17 cells) do not require REPs to expand 5,000-fold over 3 weeks. Also, unlike Th1 cells, Th17 cells do not exhibit hallmarks of senescence or apoptosis, retaining robust antitumor efficacy in vivo. Three-week-expanded Th17 cells eliminated melanoma as effectively as Th17 cells expanded for 1 week when infused in equal numbers into mice. However, treating mice with large recalcitrant tumors required the infusion of all cells generated after 2 or 3 weeks of expansion, while the cell yield obtained after 1-week expansion was insufficient. Long-term-expanded Th17 cells also protected mice from tumor rechallenge including lung metastasis. Importantly, 2-week-expanded human chimeric antigen receptor-positive (CAR+) Th17 cells also retained their ability to regress human mesothelioma, while CAR+ Th1 cells did not. Our results indicate that tumor-reactive Th17 cells are an effective cell therapy for cancer, remaining uncompromised when expanded for a long duration owing to their resistance to senescence.

Exploiting IL-17-producing CD4+ and CD8+ T cells to improve cancer immunotherapy in the clinic.

Cancer immunotherapy is one the most effective approaches for treating patients with tumors, as it bolsters the generation and persistence of memory T cells. In preclinical work, it has been reported that adoptively transferred CD4+ and CD8+ lymphocytes that secrete IL-17A (i.e., Th17 and Tc17 cells) regress tumors to a greater extent than IFN-γ+Th1 or Tc1 cells in vivo. Herein, we review the mechanisms underlying how infused Th17 and Tc17 cells regress established malignancies in clinically relevant mouse models of cancer. We also discuss how unique signaling cues—such as co-stimulatory molecules (ICOS and 41BB), cytokines (IL-12 and IL-23) or pharmaceutical reagents (Akt inhibitors, etc.)—can be exploited to bolster the therapeutic potential of IL-17+ lymphocytes with an emphasis on using this knowledge to improve next-generation clinical trials for patients with cancer.

β -catenin and PI3K δ inhibition expands precursor Th17 cells with heightened stemness and antitumor activity

ICOS costimulation generates Th17 cells with durable memory responses to tumor. Herein, we found that ICOS induces PI3K/p110δ/Akt and Wnt/β-catenin pathways in Th17 cells. Coinhibiting PI3Kδ and β-catenin altered the biological fate of Th17 cells. Th17 cells inhibited of both pathways expressed less RORγt, which, in turn, reduced their ability to secrete IL-17. Unexpectedly, these cells were more effective (than uninhibited cells) at regressing tumor when infused into mice, leading to long-term curative responses. PI3Kδ inhibition expanded precursor Th17 cells with a central memory phenotype that expressed nominal regulatory properties (low FoxP3), while β-catenin inhibition enhanced Th17 multifunctionality in vivo. Remarkably, upon TCR restimulation, RORγt and IL-17 rebounded in Th17 cells treated with PI3Kδ and β-catenin inhibitors. Moreover, these cells regained β-catenin, Tcf7, and Akt expression, licensing them to secrete heightened IL-2, persist, and eradicate solid tumors without help from endogenous NK and CD8 T cells. This finding shines a light on ways to repurpose FDA-approved drugs to augment T cell-based cancer immunotherapies.

Adoptive Transfer of Ceramide Synthase 6 Deficient Splenocytes Reduces the Development of Colitis

Sphingolipids regulate critical cellular processes including inflammation. Ceramide, which serves a central role in sphingolipid metabolism, is generated by six ceramide synthases (CerS) that differ in substrate specificity. CerS6 preferentially generates C16-ceramide and its mRNA is highly expressed in immune tissues. In this study we analyzed how deficiency of CerS6 impacts on the development of colitis using an adoptive transfer model. Adoptive transfer of CerS6-deficient splenocytes, which have significantly decreased levels of C16-ceramide, showed that CerS6-deficiency protected against the development of colitis. However, adoptively transferred cells isolated from the lamina propria of the large intestine from wild type or CerS6-deficient groups showed no differences in the percentages of immune-suppressive regulatory T cells, pro-inflammatory Th17 cells, or their ability to express IL-17. In vitro polarization of wild type or CerS6-deficient splenocytes also revealed no defects in the development of T cell subsets. Our data suggest that protection from colitis following adoptive transfer of CerS6-deficient splenocytes maybe related to their ability to migrate and proliferate in vivo rather than subset development or cytokine expression.

PI3Kδ Inhibition Enhances the Antitumor Fitness of Adoptively Transferred CD8+ T Cells

Phosphatidylinositol-3-kinase p110δ (PI3Kδ) inhibition by Idelalisib (CAL-101) in hematological malignancies directly induces apoptosis in cancer cells and disrupts immunological tolerance by depleting regulatory T cells. Yet, little is known about the direct impact of PI3Kδ blockade on effector T cells from CAL-101 therapy. Herein, we demonstrate a direct effect of p110δ inactivation via CAL-101 on murine and human CD8+ T cells that promotes a strong undifferentiated phenotype (elevated CD62L/CCR7, CD127, and Tcf7). These CAL-101 T cells also persisted longer after transfer into tumor bearing mice in both the murine syngeneic and human xenograft mouse models. The less differentiated phenotype and improved engraftment of CAL-101 T cells resulted in stronger antitumor immunity compared to traditionally expanded CD8+ T cells in both tumor models. Thus, this report describes a novel direct enhancement of CD8+ T cells by a p110δ inhibitor that leads to markedly improved tumor regression. This finding has significant implications to improve outcomes from next generation cancer immunotherapies.

PI3Kδ inhibition supports memory T cells with enhanced antitumor fitness

Phosphatidylinositol-3-kinase p110δ (PI3Kδ) inhibition by Idelalisib (CAL-101) in hematological malignancies directly induces apoptosis in cancer cells and disrupts immunological tolerance by depleting regulatory T cells (Tregs). Yet, little is known about the direct impact of PI3Kδ blockade on effector T cells from CAL-101 therapy. Herein, we demonstrate a direct effect of p110δ inactivation via CAL-101 on murine and human CD8+ T cells that promotes a strong undifferentiated memory phenotype (elevated CD62L/CCR7, CD127 and Tcf7). These CAL-101 T cells also persisted longer after transfer and exerted stronger antitumor immunity compared to traditionally expanded CD8+ T cells in two solid tumor models. Thus, this report describes a novel direct enhancement of CD8+ T cell memory by a p110δ inhibitor that leads to markedly improved tumor regression. This finding has significant implications to improve outcomes from next generation cancer immunotherapies. Highlights In vitro blockade of PI3K p110δ with CAL-101 endows antitumor T cells with a stronger memory phenotype than those treated with AKTi The strong memory phenotype of CAL-101 treated cells translates into improved survival of mice bearing aggressive tumors after adoptive transfer of these T cells Human CAR engineered T cells treated with CAL-101 possess an enhanced memory phenotype and robust antitumor efficacy The antitumor efficacy of CAL-101 primed T cells is not mediated by high CD62L or CD127 expression, but is likely driven by their stem memory phenotype eTOC Blurb Bowers et al report a novel function of PI3K blockade using the p110δ subunit inhibitor CAL-101 to induce memory and antitumor potency in CD8+ T cells. Ex vivo treatment of T cells with CAL-101 leads to improved antitumor control and subject survival in both murine transgenic T cell and human CAR T cell models.