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Immune cells have intensely physical lifestyles characterized by structural plasticity and force exertion. To investigate whether specific immune functions require stereotyped mechanical outputs, we used super-resolution traction force microscopy to compare the immune synapses formed by cytotoxic T cells with contacts formed by other T cell subsets and by macrophages. T cell synapses were globally compressive, which was fundamentally different from the pulling and pinching associated with macrophage phagocytosis. Spectral decomposition of force exertion patterns from each cell type linked cytotoxicity to compressive strength, local protrusiveness, and the induction of complex, asymmetric topography. These features were validated as cytotoxic drivers by genetic disruption of cytoskeletal regulators, live imaging of synaptic secretion, and in silico analysis of interfacial distortion. Synapse architecture and force exertion were sensitive to target stiffness and size, suggesting that the mechanical potentiation of killing is biophysically adaptive. We conclude that cellular cytotoxicity and, by implication, other effector responses are supported by specialized patterns of efferent force. Editor's summary: Immune cells apply mechanical forces to their surrounding environment, but whether their biomechanical output is tuned to support distinct immunological functions remains unclear. Using super-resolution traction force microscopy, de Jesus et al. compared the patterns of interfacial forces exerted by cytotoxic T lymphocytes (CTLs) with other immune cells. Within the immune synapse, CTLs formed a complex compressive crater, a biomechanical signature that was distinct from those of macrophages initiating phagocytosis and T cells engaging in nonlytic interactions. These findings demonstrate that immune cells exert specialized patterns of force on their targets to carry out distinct immunological functions. —Claire Olingy [ABSTRACT FROM AUTHOR]

Tumors evade attacks from the immune system through various mechanisms. Here, we identify a component of tumor immune evasion mediated by YTH domain–containing family protein 2 (YTHDF2), a reader protein that usually destabilizes m6A-modified mRNA. Loss of tumoral YTHDF2 inhibits tumor growth and prolongs survival in immunocompetent tumor models. Mechanistically, tumoral YTHDF2 deficiency promotes the recruitment of macrophages via CX3CL1 and enhances mitochondrial respiration of CD8+ T cells by impairing tumor glycolysis metabolism. Tumoral YTHDF2 deficiency promotes inflammatory macrophage polarization and antigen presentation in the presence of IFN-γ. In addition, IFN-γ induces autophagic degradation of tumoral YTHDF2, thereby sensitizing tumor cells to CD8+ T cell–mediated cytotoxicity. Last, we identified a small molecule compound that preferentially induces YTHDF2 degradation, which shows a potent antitumor effect alone but a better effect when combined with anti–PD-L1 or anti–PD-1 antibodies. Collectively, YTHDF2 appears to be a tumor-intrinsic regulator that orchestrates immune evasion, representing a promising target for enhancing cancer immunotherapy. Editor's Summary: Immune evasion mechanisms facilitate tumor growth and limit immunotherapy efficacy. Xiao et al. identified a role for YTH domain–containing family protein 2 (YTHDF2), a N6-methyladenosine reader protein, in regulating immune evasion in solid tumors. YTHDF2 can target CX3CL1 mRNA to limit expression, which prevents inflammatory macrophage recruitment to tumors and cytotoxic T cell responses. CD8 T cell–derived IFN-γ can mediate autophagic degradation of YTHDF2, thus sensitizing tumor cells to T cell–mediated destruction. They also identified a small molecule compound that degrades YTHDF2 in tumor cells, which exhibits antitumor effects that are further enhanced with PD-1/PD-L1 blockade. —Christiana Fogg [ABSTRACT FROM AUTHOR]

Immune checkpoint blockade is a promising approach to activate antitumor immunity and improve the survival of patients with cancer. V-domain immunoglobulin suppressor of T cell activation (VISTA) is an immune checkpoint target; however, the downstream signaling mechanisms are elusive. Here, we identify leucine-rich repeats and immunoglobulin-like domains 1 (LRIG1) as a VISTA binding partner, which acts as an inhibitory receptor by engaging VISTA and suppressing T cell receptor signaling pathways. Mice with T cell–specific LRIG1 deletion developed superior antitumor responses because of expansion of tumor-specific cytotoxic T lymphocytes (CTLs) with increased effector function and survival. Sustained tumor control was associated with a reduction of quiescent CTLs (TCF1+ CD62Lhi PD-1low) and a reciprocal increase in progenitor and memory-like CTLs (TCF1+ PD-1+). In patients with melanoma, elevated LRIG1 expression on tumor-infiltrating CD8+ CTLs correlated with resistance to immunotherapies. These results delineate the role of LRIG1 as an inhibitory immune checkpoint receptor and propose a rationale for targeting the VISTA/LRIG1 axis for cancer immunotherapy. Editor's summary: The negative checkpoint regulator VISTA has emerged as a promising target for cancer immunotherapy. VISTA plays an important role in maintaining T cell quiescence; however, the molecular mechanisms through which VISTA regulates antitumor immunity are not well defined. In this study, Ta et al. identified LRIG1 as a VISTA-binding partner and co-inhibitory receptor. VISTA engagement by LRIG1 impaired murine antitumor immunity by promoting quiescence in tumor-specific CD8 T cells and limited the abundance of progenitor and memory-like cells with the ability to differentiate into effector T cells. In patients with melanoma, expression of LRIG1 on tumor-infiltrating lymphocytes correlated with resistance to immunotherapy. Together, these findings suggest that selectively inhibiting VISTA/LRIG1 may be an effective cancer immunotherapy. —Hannah Isles [ABSTRACT FROM AUTHOR]

Programmed cell death-1 (PD-1) is a potent immune checkpoint receptor on T lymphocytes. Upon engagement by its ligands, PD-L1 or PD-L2, PD-1 inhibits T cell activation and can promote immune tolerance. Antagonism of PD-1 signaling has proven effective in cancer immunotherapy, and conversely, agonists of the receptor may have a role in treating autoimmune disease. Some immune receptors function as dimers, but PD-1 has been considered monomeric. Here, we show that PD-1 and its ligands form dimers as a consequence of transmembrane domain interactions and that propensity for dimerization correlates with the ability of PD-1 to inhibit immune responses, antitumor immunity, cytotoxic T cell function, and autoimmune tissue destruction. These observations contribute to our understanding of the PD-1 axis and how it can potentially be manipulated for improved treatment of cancer and autoimmune diseases. Editor's Summary: Programmed cell death-1 (PD-1) is well known as an immune checkpoint receptor on T cells that can inhibit activation upon engagement with its ligands PD-L1 or PD-L2. PD-1 has largely been considered to function as a monomer, but Philips et al. now show that PD-1 can form dimers in cis via transmembrane domain interactions. They observed that limiting PD-1 dimerization weakens T cell inhibition upon ligand engagement, which enhances antitumor and cytotoxic T cell function and promotes autoimmunity. These findings may shape ongoing efforts to more effectively block PD-1 function or, conversely, to develop PD-1 agonists. —Christiana Fogg [ABSTRACT FROM AUTHOR]

Familial hemophagocytic lymphohistiocytosis (FHL) is an inherited, often fatal immune deficiency characterized by severe systemic hyperinflammation. Although allogeneic bone marrow transplantation can be curative, more effective therapies are urgently needed. FHL is caused by inactivating mutations in proteins that regulate cellular immunity. Here, we used an adeno-associated virus–based CRISPR-Cas9 system with an inhibitor of nonhomologous end joining to repair such mutations in potentially long-lived T cells ex vivo. Repaired CD8 memory T cells efficiently cured lethal hyperinflammation in a mouse model of Epstein-Barr virus–triggered FHL2, a subtype caused by perforin-1 (Prf1) deficiency. Furthermore, repair of PRF1 and Munc13-4 (UNC13D)—whose deficiency causes the FHL subtype FHL3—in mutant memory T cells from two critically ill patients with FHL restored T cell cytotoxicity. These results provide a starting point for the treatment of genetic T cell immune dysregulation syndromes with repaired autologous T cells. Editor's summary: Familial hemophagocytic lymphohistiocytosis (FHL) is a spectrum of inherited, potentially fatal inflammatory diseases caused by loss-of-function mutations in the NK cell and T cell cytotoxic machinery. Although FHL can be treated with hematopoietic stem cell transplantation (HSCT), mortality is still high and more effective therapeutic approaches are needed. Li et al. developed an adeno-associated virus (AAV)–based CRISPR-Cas9 system combined with nonhomologous end joining (NHEJ) inhibition to repair T cells from a perforin (Prf1)–deficient Epstein-Barr virus (EBV)–triggered mouse model of FHL (FHL2). Repaired T cells could prevent or cure FHL2-like disease caused by EBV-driven B cell hyperproliferation. Moreover, this CRISPR-Cas9 system could be used to mend T cells from pediatric patients with FHL2 (PRF1 deficiency) and FHL3 (Munc13-4/UNC13D deficiency). This work demonstrates that gene repair of autologous T cells is a potentially promising treatment for human FHL. —Seth Thomas Scanlon [ABSTRACT FROM AUTHOR]

CD8+ T cells are key antiviral effectors against hepatitis B virus (HBV), yet their number and function can be compromised in chronic infections. Preclinical HBV models displaying CD8+ T cell dysfunction showed that interleukin-2 (IL-2)–based treatment, unlike programmed cell death ligand 1 (PD-L1) checkpoint blockade, could reverse this defect, suggesting its therapeutic potential against HBV. However, IL-2's effectiveness is hindered by its pleiotropic nature, because its receptor is found on various immune cells, including regulatory T (Treg) cells and natural killer (NK) cells, which can counteract antiviral responses or contribute to toxicity, respectively. To address this, we developed a cis-targeted CD8-IL2 fusion protein, aiming to selectively stimulate dysfunctional CD8+ T cells in chronic HBV. In a mouse model, CD8-IL2 boosted the number of HBV-reactive CD8+ T cells in the liver without substantially altering Treg or NK cell counts. These expanded CD8+ T cells exhibited increased interferon-γ and granzyme B production, demonstrating enhanced functionality. CD8-IL2 treatment resulted in substantial antiviral effects, evidenced by marked reductions in viremia and antigenemia and HBV core antigen–positive hepatocytes. In contrast, an untargeted CTRL-IL2 led to predominant NK cell expansion, minimal CD8+ T cell expansion, negligible changes in effector molecules, and minimal antiviral activity. Human CD8-IL2 trials in cynomolgus monkeys mirrored these results, achieving a roughly 20-fold increase in peripheral blood CD8+ T cells without affecting NK or Treg cell numbers. These data support the development of CD8-IL2 as a therapy for chronic HBV infection. Editor's summary: Promoting CD8+ T cell responses to hepatitis B virus (HBV) with IL-2 may offer an approach to clear the virus; however, traditional IL-2–based therapies are plagued by toxicities and unintended activation of other immune cell populations. Here, Andreata et al. developed and tested a CD8 cis-targeted IL-2 (CD8-IL2) that selectively activates CD8+ T cells without affecting regulatory T cells or natural killer cells. The authors found that CD8-IL2 treatment exhibited antiviral activity in a preclinical model of chronic HBV, reducing viremia; further data in human cells and cynomolgus monkeys supported the specificity and safety of this molecule. Together, these data support further development of CD8-IL2 for chronic HBV. —Courtney Malo [ABSTRACT FROM AUTHOR]

Whereas CD4+ T cells conventionally mediate antitumor immunity by providing help to CD8+ T cells, recent clinical studies have implied an important role for cytotoxic CD4+ T cells in cancer immunity. Using an orthotopic melanoma model, we provide a detailed account of antitumoral CD4+ T cell responses and their regulation by major histocompatibility complex class II (MHC II) in the skin. Intravital imaging revealed prominent interactions of CD4+ T cells with tumor debris-laden MHC II+ host antigen-presenting cells that accumulated around tumor cell nests, although direct recognition of MHC II+ melanoma cells alone could also promote CD4+ T cell control. CD4+ T cells stably suppressed or eradicated tumors even in the absence of other lymphocytes by using tumor necrosis factor–α and Fas ligand (FasL) but not perforin-mediated cytotoxicity. Interferon-γ was critical for protection, acting both directly on melanoma cells and via induction of nitric oxide synthase in myeloid cells. Our results illustrate multifaceted and context-specific aspects of MHC II–dependent CD4+ T cell immunity against cutaneous melanoma, emphasizing modulation of this axis as a potential avenue for immunotherapies. Editor's summary: CD4+ T cells have been primarily appreciated for their helper functions in antitumor immune responses, but the extent to which they directly contribute to tumor killing remains unclear. Using a mouse model of cutaneous melanoma, Bawden et al. characterized the effector functions of CD4+ T cells in generating protective antitumor immunity. Melanoma-specific CD4+ T cells infiltrated tumors, adopted diverse effector states, and could provide tumor protection independent of other lymphocytes. Whereas MHC II expression by melanoma cells was sufficient to activate protective CD4+ T cell–mediated immunity, host antigen-presenting cells could also drive protection. CD4+ T cells provided protection through multiple partially redundant cytotoxicity pathways, including TNF and Fas ligand, but also by activating IFN-γ–dependent nitric oxide production in myeloid cells. Together, these results demonstrate that tumor-infiltrating CD4+ T cells are equipped to contribute to tumor control through multiple helper and effector modes. —Claire Olingy [ABSTRACT FROM AUTHOR]

Engagement of the receptor programmed cell death molecule 1 (PD-1) by its ligands PD-L1 and PD-L2 inhibits T cell–mediated immune responses. Blocking such signaling provides the clinical effects of PD-1–targeted immunotherapy. Here, we investigated the mechanisms underlying PD-1–mediated inhibition. Because dynamic actin remodeling is crucial for T cell functions, we characterized the effects of PD-1 engagement on actin remodeling at the immunological synapse, the interface between a T cell and an antigen-presenting cell (APC) or target cell. We used microscopy to analyze the formation of immunological synapses between PD-1+ Jurkat cells or primary human CD8+ cytotoxic T cells and APCs that presented T cell–activating antibodies and were either positive or negative for PD-L1. PD-1 binding to PD-L1 inhibited T cell spreading induced by antibody-mediated activation, which was characterized by the absence of the F-actin–dense distal lamellipodial network at the immunological synapse and the Arp2/3 complex, which mediates branched actin formation. PD-1–induced inhibition of actin remodeling also prevented the characteristic deformation of T cells that contact APCs and the release of cytotoxic granules. We showed that the effects of PD-1 on actin remodeling did not require its tyrosine-based signaling motifs, which are thought to mediate the co-inhibitory effects of PD-1. Our study highlights a previously unappreciated mechanism of PD-1–mediated suppression of T cell activity, which depends on the regulation of actin cytoskeleton dynamics in a signaling motif–independent manner. Editor's summary: When engaged by its ligands, the receptor PD-1 inhibits the activation of T cells stimulated through the T cell receptor. Alleviating this effect is the basis of checkpoint inhibitors that promote antitumor immunity (see the Focus by Acharya and Kumari). Paillon et al. used imaging techniques to show that PD-1 signaling inhibited the formation of an immunological synapse between T cells and their target cells by preventing remodeling of the actin cytoskeleton in a manner independent of its intracellular, tyrosine-based signaling motifs. These findings suggest an alternative mechanism by which PD-1 regulates immune responses. —John F. Foley [ABSTRACT FROM AUTHOR]

T cell immunoglobulin and mucin-containing molecule 3 (Tim-3), expressed in dysfunctional and exhausted T cells, has been widely acknowledged as a promising immune checkpoint target for tumor immunotherapy. Here, using a strategy combining virtual and functional screening, we identified a compound named ML-T7 that targets the FG-CC′ cleft of Tim-3, a highly conserved binding site of phosphatidylserine (PtdSer) and carcinoembryonic antigen–related cell adhesion molecule 1 (CEACAM1). ML-T7 enhanced the survival and antitumor activity of primary CD8+ cytotoxic T lymphocytes (CTLs) and human chimeric antigen receptor (CAR) T cells and reduced their exhaustion in vitro and in vivo. In addition, ML-T7 promoted NK cells' killing activity and DC antigen-presenting capacity, consistent with the reported activity of Tim-3. ML-T7 strengthened DCs' functions through both Tim-3 and Tim-4, which is consistent with the fact that Tim-4 contains a similar FG-CC′ loop. Intraperitoneal dosing of ML-T7 showed comparable tumor inhibitory effects to the Tim-3 blocking antibody. ML-T7 reduced syngeneic tumor progression in both wild-type and Tim-3 humanized mice and alleviated the immunosuppressive microenvironment. Furthermore, combined ML-T7 and anti–PD-1 therapy had greater therapeutic efficacy than monotherapy in mice, supporting further development of ML-T7 for tumor immunotherapy. Our study demonstrates a potential small molecule for selectively blocking Tim-3 and warrants further study. Editor's summary: T cell immunoglobulin and mucin-containing molecule 3 (Tim-3) has been suggested as a promising immune checkpoint target for immunotherapy, but clinical translation of Tim-3 antibodies has not performed well. Here, Ma et al. identified a small-molecule Tim-3 inhibitor, ML-T7, through a virtual and functional screening strategy. They saw effective antitumor activity alone or in combination with anti–PD-1 therapy in both syngeneic and humanized mouse models, warranting further study of this small molecule. —Dorothy Hallberg [ABSTRACT FROM AUTHOR]

Autoimmune vasculitis of the medium and large elastic arteries can cause blindness, stroke, aortic arch syndrome, and aortic aneurysm. The disease is often refractory to immunosuppressive therapy and progresses over decades as smoldering aortitis. How the granulomatous infiltrates in the vessel wall are maintained and how tissue-infiltrating T cells and macrophages are replenished are unknown. Single-cell and whole-tissue transcriptomic studies of immune cell populations in vasculitic arteries identified a CD4+ T cell population with stem cell–like features. CD4+ T cells supplying the tissue-infiltrating and tissue-damaging effector T cells survived in tertiary lymphoid structures around adventitial vasa vasora, expressed the transcription factor T cell factor 1 (TCF1), had high proliferative potential, and gave rise to two effector populations, Eomesodermin (EOMES)+ cytotoxic T cells and B cell lymphoma 6 (BCL6)+ T follicular helper-like cells. TCF1hiCD4+ T cells expressing the interleukin 7 receptor (IL-7R) sustained vasculitis in serial transplantation experiments. Thus, TCF1hiCD4+ T cells function as disease stem cells and promote chronicity and autonomy of autoimmune tissue inflammation. Remission-inducing therapies will require targeting stem-like CD4+ T cells instead of only effector T cells. Editor's summary: Giant cell arteritis (GCA) is an autoimmune disease of the medium and large arteries that affects older adults and is often refractory to standard immunosuppression. Although T cell infiltrates occupy the vessel walls in patients with GCA, the source of these cells, and their importance in driving disease, is unclear. Here, Sato et al. found, using both human aortic tissues and serial transplantation experiments in mice, that a population of stem-like CD4+ T cells residing in tertiary lymphoid structures replenishes pathogenic effector cells independent of secondary lymphoid organs. Targeting such stem-like CD4+ T cells may represent a therapeutic opportunity to halt the chronic progression of autoimmune vasculitis. —Courtney Malo [ABSTRACT FROM AUTHOR]

This first-in-human study evaluated RO7122290, a bispecific fusion protein carrying a split trimeric 4-1BB (CD137) ligand and a fibroblast activation protein α (FAP) binding site that costimulates T cells for improved tumor cell killing in FAP-expressing tumors. Patients with advanced or metastatic solid tumors received escalating weekly intravenous doses of RO7122290 as a single agent (n = 65) or in combination with a 1200-milligram fixed dose of the anti–programmed death-ligand 1 (anti–PD-L1) antibody atezolizumab given every 3 weeks (n = 50), across a tested RO7122290 dose range of 5 to 2000 milligrams and 45 to 2000 milligrams, respectively. Three dose-limiting toxicities were reported, two at different RO7122290 single-agent doses (grade 3 febrile neutropenia and grade 3 cytokine release syndrome) and one for the combination (grade 3 pneumonitis). No maximum tolerated dose was identified. The pharmacokinetic profile of RO7122290 suggested nonlinearity in elimination. The observed changes in peripheral and tissue pharmacodynamic (PD) biomarkers were consistent with the postulated mechanism of action. Treatment-induced PD changes included an increase in proliferating and activated T cells in peripheral blood both in the single-agent and combination arms. Increased infiltration of intratumoral CD8+ and Ki67+CD8+ T cells was observed for both treatment regimens, accompanied by the up-regulation of T cell activation genes and gene signatures. Eleven patients experienced a complete or partial response, six of whom were confirmed to be immune checkpoint inhibitor naive. These results support further evaluation of RO7122290 in combination with atezolizumab or other immune-oncology agents for the treatment of solid tumors. Costimulating T cells to treat tumors: Most patients with cancer do not respond to immune checkpoint inhibitors (ICIs). Co-stimulatory agents targeted to tumors could lead to enhanced immune system response and increased antitumor immunity in these patients. Here, Melero and colleagues studied the safety, mechanism of action, and activity of RO7122290, a fibroblast activation protein-α-targeted 4-1BB agonist designed to co-stimulate activated CD8+ T cells and natural killer cells, either as a single agent or when given with the ICI atezolizumab in patients with advanced solid tumors. Treatment resulted in increased proliferating and activated T cells in the peripheral blood and increased infiltration of intratumoral CD8+ cells. Eleven patients experienced complete or partial response. These results support further clinical evaluation of RO7122290 in combination with ICIs. —MN [ABSTRACT FROM AUTHOR]

Interleukin-33 (IL-33) functions both as a secreted cytokine and as a nuclear factor, with pleiotropic roles in cancer and immunity. Here, we explored its role in hepatocellular carcinoma (HCC) and identified that a posttranslational modification altered its nuclear activity and promoted immune escape for HCC. IL-33 abundance was overall decreased but more frequently localized to the nucleus in patient HCC tissues than in normal liver tissues. In human and mouse HCC cells in culture and in vivo, IL-33 overexpression inhibited proliferation and repressed the abundance of programmed death ligand 1 (PD-L1) at the transcriptional level by promoting the ubiquitin-dependent degradation of interferon regulatory factor 1 (IRF1). However, this interaction was disrupted by SUMOylation of IL-33 at Lys54 mediated by the E3 ligase RanBP2. IL-33 SUMOylation correlated with its nuclear localization in HCC cells and tumors. An increase in SUMOylated IL-33 in HCC cells in cocultures and in vivo stabilized IRF1 and increased PD-L1 abundance and chemokine IL-8 secretion, which prevented the activation of cytotoxic T cells and promoted the M2 polarization of macrophages, respectively. Mutating the SUMOylation site in IL-33 reversed these effects and suppressed tumor growth. These findings indicate that SUMOylation of nuclear IL-33 in HCC cells impairs antitumor immunity. SUMO switches off immunosurveillance in liver cancer: Interleukin-33 (IL-33) is a dual function protein. As an extracellular cytokine, it is associated with the progression of liver cancer. However, Wang et al. found that as an intracellular factor, IL-33 is tumor suppressive, a function that is blocked by posttranslational modification in liver cancer cells. In cell lines and tissues from patients with hepatocellular carcinoma (HCC), intracellular IL-33 was less abundant but more frequently nuclear and SUMOylated. SUMOylation prevented IL-33 from facilitating the degradation of transcription factor IRF1, thus enabling IRF1-dependent production of the chemokine IL-8 and the immune checkpoint protein PD-L1, which suppressed antitumor activity in T cells and macrophages in vivo. The findings suggest that immunosurveillance of HCC is impaired in part through the loss of the nuclear factor function of IL-33. —LKF [ABSTRACT FROM AUTHOR]

Anti–PD-1/PD-L1 therapy, either by anti–PD-1 antibody or anti–PD-L1 antibody, has efficacy by reinvigorating tumor-infiltrating CD8+ T cells in a subset of patients with cancer, but it has unequal effects on heterogeneous CD8+ T cell populations. Hence, the subset crucial to efficacious PD-1 blockade therapy remains elusive. Here, we found an increase in tumor-infiltrating CD200+ cytotoxic T lymphocytes (CTLs) upon PD-1/PD-L1 blockade, with higher proportions of CD200+ T cells positively related to a favorable clinical outcome to anti–PD-1/PD-L1 therapy in three independent cohorts of patients with cancer. Using multiple mouse tumor models, we demonstrated that CD200+ CTLs are essential for efficacious anti–PD-L1 therapy. Mechanistically, we observed a unique chromatin landscape in CD200+ CTLs and found that these cells are enriched for tumor antigen–specific CTLs and have antitumor effector functions. Coinoculation of CD200+ CTLs with tumor cells led to robust tumor regression in two transplanted mouse models. Clinically, we found that infiltration of CD200+ CTLs into tumors could predict immunotherapy efficacy in six patient cohorts. Together, our findings reveal that CD200+ CTLs in the tumor microenvironment are crucial for efficacious anti–PD-1/PD-L1 therapy and could serve as a predictor of successful immunotherapy in the clinic. Identifying responders: A challenge for the field of cancer immunotherapy is identifying which patients will respond to immune checkpoint blockade treatments such as anti-PD-1 or anti-PD-L1. Here, Wang et al. showed using murine tumor models that CD200+ CD8+ T cells were crucial for the efficacy of anti-PD-1/PD-L1 therapy. These CD200+ cytotoxic T lymphocytes had a unique chromatin landscape and possessed superior anti-tumor effector functions as compared to CD200- CD8+ T cells. Further, the authors showed, using data from multiple patient cohorts, that CD200+ T cells could predict immunotherapy efficacy, suggesting that this cell population could serve as a potential marker in the clinic. —CM [ABSTRACT FROM AUTHOR]

Despite the success of cancer immunotherapies such as programmed cell death–1 (PD-1) and PD-1 ligand 1 (PD-L1) inhibitors, patients often develop resistance. New combination therapies with PD-1/PD-L1 inhibitors are needed to overcome this issue. Bezafibrate, a ligand of peroxisome proliferator–activated receptor–γ coactivator 1α/peroxisome proliferator–activated receptor complexes, has shown a synergistic antitumor effect with PD-1 blockade in mice that is mediated by activation of mitochondria in T cells. We have therefore now performed a phase 1 trial (UMIN000017854) of bezafibrate with nivolumab in previously treated patients with advanced non–small cell lung cancer. The primary end point was the percentage of patients who experience dose-limiting toxicity, and this combination regimen was found to be well tolerated. Preplanned comprehensive analysis of plasma metabolites and gene expression in peripheral cytotoxic T cells indicated that bezafibrate promoted T cell function through up-regulation of mitochondrial metabolism including fatty acid oxidation and may thereby have prolonged the duration of response. This combination strategy targeting T cell metabolism thus has the potential to maintain antitumor activity of immune checkpoint inhibitors and warrants further validation. Targeting T cell metabolism: Targeting T cell metabolismImmune checkpoint inhibition (ICI) has improved survival for patients with non-small cell lung cancer (NSCLC), however, 30-50% of patients are unresponsive or acquire resistance. To improve survival of patients, Tanaka et al. have taken guidance from previous preclinical studies where Bezafibrate has shown synergy with ICIs and have implemented a phase I trial (UMIN000017854) of bezafibrate with nivolumab in previously treated patients with advanced NSCLC. The authors found that the regimen was well tolerated, saw partial response in 12 out of 18 patients, and promoted T cell function. This combination targeting T cell metabolism shows promise at potentiating the effects of ICIs and warrants further study.– DH [ABSTRACT FROM AUTHOR]

Chimeric antigen receptors (CARs) repurpose natural signaling components to retarget T cells to refractory cancers but have shown limited efficacy in persistent, recurrent malignancies. Here, we introduce "CAR Pooling," a multiplexed approach to rapidly identify CAR designs with clinical potential. Forty CARs with signaling domains derived from a range of immune cell lineages were evaluated in pooled assays for their ability to stimulate critical T cell effector functions during repetitive stimulation that mimics long-term tumor antigen exposure. Several domains were identified from the tumor necrosis factor (TNF) receptor family that have been primarily associated with B cells. CD40 enhanced proliferation, whereas B cell–activating factor receptor (BAFF-R) and transmembrane activator and CAML interactor (TACI) promoted cytotoxicity. These functions were enhanced relative to clinical benchmarks after prolonged antigen stimulation, and CAR T cell signaling through these domains fell into distinct states of memory, cytotoxicity, and metabolism. BAFF-R CAR T cells were enriched for a highly cytotoxic transcriptional signature previously associated with positive clinical outcomes. We also observed that replacing the 4-1BB intracellular signaling domain with the BAFF-R signaling domain in a clinically validated B cell maturation antigen (BCMA)–specific CAR resulted in enhanced activity in a xenotransplant model of multiple myeloma. Together, these results show that CAR Pooling is a general approach for rapid exploration of CAR architecture and activity to improve the efficacy of CAR T cell therapies. Test driving CAR T cells: CAR T cells have already been through multiple generations, with each iteration improving signaling through the receptor. However, a strategy to broadly test multiple candidate intracellular signaling domains would allow for even better optimization. Here, Goodman et al. have developed CAR Pooling, a multiplexed approach to rapidly identify the top CAR signaling domains. The authors found using CAR Pooling that BAFF-R and 4-1BB intracellular signaling domains improved CAR T cell activity; these results translated into control of tumor burden in vivo. Thus, CAR Pooling represents an effective approach to rapidly identify strategies to improve CAR T cells. [ABSTRACT FROM AUTHOR]

Although many patients with diffuse large B cell lymphoma (DLBCL) may achieve a complete response to frontline chemoimmunotherapy, patients with relapsed/refractory disease typically have poor outcomes. Odronextamab, a CD20xCD3 bispecific antibody that provides "signal 1" through the activation of the T cell receptor/CD3 complex, has exhibited early, promising activity for patients with highly refractory DLBCL in phase 1 trials. However, not all patients achieve complete responses, and many relapse, thus representing a high unmet medical need. Here, we investigated whether adding a costimulatory "signal 2" by engaging CD28 receptors on T cells could augment odronextamab activity. We demonstrate that REGN5837, a bispecific antibody that cross-links CD22-expressing tumor cells with CD28-expressing T cells, enhances odronextamab by potentiating T cell activation and cytolytic function. In preclinical DLBCL studies using human immune system–reconstituted animals, REGN5837 promotes the antitumor activity of odronextamab and induces intratumoral expansion of reprogrammable T cells while skewing away from a dysfunctional state. Although REGN5837 monotherapy shows limited activity and no toxicity in primate studies, it augments T cell activation when dosed in combination with odronextamab. In addition, analysis of non-Hodgkin lymphoma clinical samples reveals an increase in CD28+CD8+ T cells after odronextamab treatment, demonstrating the presence of a population that could potentially be targeted by REGN5837. Collectively, our data demonstrate that REGN5837 can markedly enhance the antitumor activity of odronextamab in preclinical NHL models, and the combination of these two bispecific antibodies may provide a chemotherapy-free approach for the treatment of DLBCL. Dynamic bispecific duo: Bispecific antibodies are being used for cancer immunotherapy because they target two different antigens thus facilitating T cell targeting to tumors. Odronextamab is a CD20xCD3 bispecific antibody that shows promising activity in patients with diffuse large B cell lymphoma (DLBCL), but not all patients achieve complete responses, and there remains a high unmet need in the setting of relapsed/refractory disease. Wei et al. (p. 0000) demonstrate that REGN5837, a CD22xCD28 bispecific antibody, enhanced the antitumor activity of odronextamab in preclinical models of DLBCL and that the combination exhibited no toxicity in primates while augmenting T cell activation. CD28+CD8+ T cells were expanded in non-Hodgkin lymphoma samples from a phase 1 odronextamab trial, suggesting that addition of REGN5837 could enhance antitumor activity. These findings highlight the potential clinical usefulness of a bispecific antibody combination for treating DLBCL. [ABSTRACT FROM AUTHOR]

Immune checkpoint inhibitors (ICIs) have been increasingly used in combination for cancer treatment but are associated with myocarditis. Here, we report that tumor-bearing mice exhibited response to treatment with combinatorial anti–programmed cell death 1 and anti–cytotoxic T lymphocyte antigen–4 antibodies but also presented with cardiovascular toxicities observed clinically with ICI therapy, including myocarditis and arrhythmia. Female mice were preferentially affected with myocarditis compared to male mice, consistent with a previously described genetic model of ICI myocarditis and emerging clinical data. Mechanistically, myocardial tissue from ICI-treated mice, the genetic mouse model, and human heart tissue from affected patients with ICI myocarditis all exhibited down-regulation of MANF (mesencephalic astrocyte–derived neurotrophic factor) and HSPA5 (heat shock 70-kDa protein 5) in the heart; this down-regulation was particularly notable in female mice. ICI myocarditis was amplified by heart-specific genetic deletion of mouse Manf and was attenuated by administration of recombinant MANF protein, suggesting a causal role. Ironically, both MANF and HSPA5 were transcriptionally induced by liganded estrogen receptor β and inhibited by androgen receptor. However, ICI treatment reduced serum estradiol concentration to a greater extent in female compared to male mice. Treatment with an estrogen receptor β–specific agonist and androgen depletion therapy attenuated ICI-associated cardiac effects. Together, our data suggest that ICI treatment inhibits estradiol-dependent expression of MANF/HSPA5 in the heart, curtailing the cardiomyocyte response to immune injury. This endocrine-cardiac-immune pathway offers new insights into the mechanisms of sex differences in cardiac disease and may offer treatment strategies for ICI myocarditis. Of myocarditis and women: Inflammation of the heart muscle, or myocarditis, is an off-target effect of anticancer treatments such as immune checkpoint inhibitor (ICI) therapy. Here, Zhang et al. studied sex differences in ICI myocarditis. They observed greater T cell infiltration and cardiac dysfunction in female mice with ICI myocarditis and identified decreased MANF and HSPA5 expression in the heart. Cardiac depletion of Manf worsened ICI myocarditis, whereas recombinant MANF improved heart function. Treating cardiomyocytes derived from human induced pluripotent stem cells with estrogen or estrogen receptor β agonist induced MANF and HSPA5 expression. Treating female tumor-bearing mice with estrogen receptor β agonist during ICI treatment reduced T cell infiltration and preserved heart function. Results suggest that hormone therapy could limit ICI myocarditis by promoting the protective effects of MANF. [ABSTRACT FROM AUTHOR]

Upon chronic antigen exposure, CD8+ T cells become exhausted, acquiring a dysfunctional state correlated with the inability to control infection or tumor progression. In contrast, stem-like CD8+ T progenitors maintain the ability to promote and sustain effective immunity. Adenovirus (Ad)–vectored vaccines encoding tumor neoantigens have been shown to eradicate large tumors when combined with anti–programmed cell death protein 1 (αPD-1) in murine models; however, the mechanisms and translational potential have not yet been elucidated. Here, we show that gorilla Ad vaccine targeting tumor neoepitopes enhances responses to αPD-1 therapy by improving immunogenicity and antitumor efficacy. Single-cell RNA sequencing demonstrated that the combination of Ad vaccine and αPD-1 increased the number of murine polyfunctional neoantigen-specific CD8+ T cells over αPD-1 monotherapy, with an accumulation of Tcf1+ stem-like progenitors in draining lymph nodes and effector CD8+ T cells in tumors. Combined T cell receptor (TCR) sequencing analysis highlighted a broader spectrum of neoantigen-specific CD8+ T cells upon vaccination compared to αPD-1 monotherapy. The translational relevance of these data is supported by results obtained in the first 12 patients with metastatic deficient mismatch repair (dMMR) tumors vaccinated with an Ad vaccine encoding shared neoantigens. Expansion and diversification of TCRs were observed in post-treatment biopsies of patients with clinical response, as well as an increase in tumor-infiltrating T cells with an effector memory signature. These findings indicate a promising mechanism to overcome resistance to PD-1 blockade by promoting immunogenicity and broadening the spectrum and magnitude of neoantigen-specific T cells infiltrating tumors. Immunotherapy that is not monkeying around: Adenoviral vaccines encoding for tumor neoantigens have shown promise treating solid tumors when combined with anti–programmed cell death protein 1 (αPD-1) preclinically; however, the mechanism is not well understood. To elucidate this, Chen et al. generated Great Ape adenovirus (GAd) vaccines and treated tumor-bearing mice in combination with αPD-1 to elicit an accumulation of Tcf1+ stem-like CD8+ T cell progenitors, improving immunogenicity and antitumor efficacy. In addition, they performed a first-in-human trial on patients with metastatic mismatch repair–deficient tumors and saw a clinical response, suggesting this as a promising therapy to overcome resistance to αPD-1 treatment. [ABSTRACT FROM AUTHOR]

Adoptive immunotherapy with T cells engineered with tumor-specific T cell receptors (TCRs) holds promise for cancer treatment. However, suppressive cues generated in the tumor microenvironment (TME) can hinder the efficacy of these therapies, prompting the search for strategies to overcome these detrimental conditions and improve cellular therapeutic approaches. CD1d-restricted invariant natural killer T (iNKT) cells actively participate in tumor immunosurveillance by restricting suppressive myeloid populations in the TME. Here, we showed that harnessing iNKT cells with a second TCR specific for a tumor-associated peptide generated bispecific effectors for CD1d- and major histocompatibility complex (MHC)–restricted antigens in vitro. Upon in vivo transfer, TCR-engineered iNKT (TCR-iNKT) cells showed the highest efficacy in restraining the progression of multiple tumors that expressed the cognate antigen compared with nontransduced iNKT cells or CD8+ T cells engineered with the same TCR. TCR-iNKT cells achieved robust cancer control by simultaneously modulating intratumoral suppressive myeloid populations and killing malignant cells. This dual antitumor function was further enhanced when the iNKT cell agonist α-galactosyl ceramide (α-GalCer) was administered as a therapeutic booster through a platform that ensured controlled delivery at the tumor site, named multistage vector (MSV). These preclinical results support the combination of tumor-redirected TCR-iNKT cells and local α-GalCer boosting as a potential therapy for patients with cancer. TCR-engineered iNKT cells: Adoptive cell therapy has had success in treating various blood cancers. However, in solid tumors, most of these therapies show little efficacy due to exhaustion and poor migration to tumors. Here, Delfanti et al. leveraged natural killer T cells as an adoptive cell therapy platform, specifically producing them to express a TCR specific for a tumor-associated antigen. In mouse models, they found that these TCR-engineered iNKT robustly delayed tumor growth, leading to rejection and prolonged antitumor effects in some mice. These cells migrated well into tumors, killed tumor cells directly, and helped to shift intratumorally myeloid cells to an antitumor phenotype. Boosting TCR-engineered iNKTs with α-GalCer improved antitumor immune responses. Thus, TCR-engineered iNKT cells might be a promising therapy for patients with cancer. [ABSTRACT FROM AUTHOR]

Sex bias exists in the development and progression of nonreproductive organ cancers, but the underlying mechanisms are enigmatic. Studies so far have focused largely on sexual dimorphisms in cancer biology and socioeconomic factors. Here, we establish a role for CD8+ T cell–dependent antitumor immunity in mediating sex differences in tumor aggressiveness, which is driven by the gonadal androgen but not sex chromosomes. A male bias exists in the frequency of intratumoral antigen-experienced Tcf7/TCF1+ progenitor exhausted CD8+ T cells that are devoid of effector activity as a consequence of intrinsic androgen receptor (AR) function. Mechanistically, we identify a novel sex-specific regulon in progenitor exhausted CD8+ T cells and a pertinent contribution from AR as a direct transcriptional transactivator of Tcf7/TCF1. The T cell–intrinsic function of AR in promoting CD8+ T cell exhaustion in vivo was established using multiple approaches including loss-of-function studies with CD8-specific Ar knockout mice. Moreover, ablation of the androgen-AR axis rewires the tumor microenvironment to favor effector T cell differentiation and potentiates the efficacy of anti–PD-1 immune checkpoint blockade. Collectively, our findings highlight androgen-mediated promotion of CD8+ T cell dysfunction in cancer and imply broader opportunities for therapeutic development from understanding sex disparities in health and disease. Antitumor CD8+ T cells curtailed by androgen: Cancer epidemiology statistics reveal a male sex bias toward increased incidence and worse clinical outcomes. Kwon et al. used mouse models of bladder cancer showing male sex bias to investigate whether androgen acting on CD8+ T cells is part of the biological explanation. Single-cell RNA sequencing analysis of intratumoral CD8+ T cells revealed male-biased expression of a progenitor exhausted (PE) signature and higher levels of effector genes in female tumors. The skew toward a PE state in male T cells was dependent on both androgen availability and androgen receptor expression by T cells. These advances in understanding the immunological basis of sex bias in cancer create opportunities to design precision medicine therapies that factor in functional differences between male and female cytotoxic T cells. [ABSTRACT FROM AUTHOR]

Gammaherpesviruses (γHVs) have coevolved with their host, leading to a remarkably high infection prevalence and establishment of latency. The lifelong persistence of γHVs in hosts appears to broadly shape host immunity, and we show here that pulmonary infection with Murid herpesvirus 4 (MuHV-4), a mouse γHV, drives the recruitment of Ly6Chi monocytes (MOs) into the airway, thereby modulating the host immune response. The absence of Ly6Chi MOs is associated with severe virus-induced immunopathology and the systemic release of inflammatory mediators. Mechanistically, MuHV-4–imprinted MOs recruit CD4 T cells to the airways and trigger immunosuppressive signaling pathways through the PD-L1/PD-1 axis, thereby dampening the deleterious activation of cytotoxic CD4 T cells. These results uncover a role for Ly6Chi MOs in modulating CD4 T cell functions and reveal pathways that could be targeted therapeutically to reduce detrimental immunopathological responses associated with respiratory viral infections. Monocyte mediator: Gammaherpesviruses (γHVs) coevolved with their hosts in a manner that allows for the lifelong persistence of these viruses in immunocompetent individuals. The presence of latent γHVs in host organisms has been shown to shape adaptive immunity, and here, Maquet et al. observed that pulmonary infection of mice with a murine γHV (MuHV-4) leads to recruitment of Ly6Chi monocytes (MOs) to airways. These γHV-imprinted Ly6Chi MOs recruit CD4 T cells to airways followed by engagement of the immunosuppressive PD-1/PD-L1 signaling pathway, which limits pathological cytotoxic CD4 T cell responses. These findings highlight a role for γHV-imprinted Ly6Chi MOs in shaping CD4 T cell functions and suggest a potential pathway for the development of therapeutics that target respiratory viral infections. [ABSTRACT FROM AUTHOR]

The rising global HIV-1 burden urgently requires vaccines capable of providing heterologous protection. Here, we developed a clade C HIV-1 vaccine consisting of priming with modified vaccinia Ankara (MVA) and boosting with cyclically permuted trimeric gp120 (CycP-gp120) protein, delivered either orally using a needle-free injector or through parenteral injection. We tested protective efficacy of the vaccine against intrarectal challenges with a pathogenic heterologous clade C SHIV infection in rhesus macaques. Both routes of vaccination induced a strong envelope-specific IgG in serum and rectal secretions directed against V1V2 scaffolds from a global panel of viruses with polyfunctional activities. Envelope-specific IgG showed lower fucosylation compared with total IgG at baseline, and most of the vaccine-induced proliferating blood CD4+ T cells did not express CCR5 and α4β7, markers associated with HIV target cells. After SHIV challenge, both routes of vaccination conferred significant and equivalent protection, with 40% of animals remaining uninfected at the end of six weekly repeated challenges with an estimated efficacy of 68% per exposure. Induction of envelope-specific IgG correlated positively with G1FB glycosylation, and G2S2F glycosylation correlated negatively with protection. Vaccine-induced TNF-α+ IFN-γ+ CD8+ T cells and TNF-α+ CD4+ T cells expressing low levels of CCR5 in the rectum at prechallenge were associated with decreased risk of SHIV acquisition. These results demonstrate that the clade C MVA/CycP-gp120 vaccine provides heterologous protection against a tier2 SHIV rectal challenge by inducing a polyfunctional antibody response with distinct Fc glycosylation profile, as well as cytotoxic CD8 T cell response and CCR5-negative T helper response in the rectum. A clade C HIV-1 vaccine: Clade C HIV-1 infections are responsible for a large proportion of global HIV-1 burden, and a vaccine is urgently required. Sahoo et al. created a clade C HIV-1 vaccination regimen that they tested in rhesus macaques, consisting of priming with modified vaccinia Ankara (MVA) and boosting with cyclically permuted trimeric gp120 (CycP-gp120). To assess delivery methods, vaccines were administered by parenteral injection or oral administration using a needle-free injector. Both routes of MVA/CycP-gp120 administration provided equivalent protection against mucosal SHIV infection. Reduced risk of infection correlated with distinct env-specific IgG Fc glycosylation profiles, as well as with vaccine-induced TNF-α+ IFN-γ+ CD8+ T cells and TNF-α+ CCR5lo CD4+ T cells. These findings demonstrate that the MVA/CycP-gp120 immunization regimen provides significant protection against heterologous SHIV infection. [ABSTRACT FROM AUTHOR]

Immunomodulators that remodel the tumor immunosuppressive microenvironment have been combined with anti–programmed death 1 (α-PD1) or anti–programmed death ligand 1 (α-PDL1) immunotherapy but have shown limited success in clinical trials. However, therapeutic strategies to modulate the immunosuppressive microenvironment of lymph nodes have been largely overlooked. Here, we designed an albumin nanoparticle, Nano-PI, containing the immunomodulators PI3Kγ inhibitor (IPI-549) and paclitaxel (PTX). We treated two breast cancer mouse models with Nano-PI in combination with α-PD1, which remodeled the tumor microenvironment in both lymph nodes and tumors. This combination achieved long-term tumor remission in mouse models and eliminated lung metastases. PTX combined with IPI-549 enabled the formation of a stable nanoparticle and enhanced the repolarization of M2 to M1 macrophages. Nano-PI not only enhanced the delivery of both immunomodulators to lymph nodes and tumors but also improved the drug accumulation in the macrophages of these two tissues. Immune cell profiling revealed that the combination of Nano-PI with α-PD1 remodeled the immune microenvironment by polarizing M2 to M1 macrophages, increasing CD4+ and CD8+ T cells, B cells, and dendritic cells, decreasing regulatory T cells, and preventing T cell exhaustion. Our data suggest that Nano-PI in combination with α-PD1 modulates the immune microenvironment in both lymph nodes and tumors to achieve long-term remission in mice with metastatic breast cancer, and represents a promising candidate for future clinical trials. Honey I shrunk the immunotherapy: Triple-negative breast cancer is known to have an immunosuppressive tumor microenvironment, which results in poor response to combined immunotherapy and chemotherapy. To overcome this, Song et al. focused on improving drug delivery using albumin-encapsulated nanoparticles containing a PI3Kγ inhibitor and paclitaxel, called Nano-PI, targeting tumors and adjacent lymph nodes. They combined Nano-PI with anti–programmed death 1 (α-PD1) to treat mouse models of breast cancer and induced long-term tumor remission. Their combination treatment remodeled the tumor microenvironment by repolarizing immunosuppressive M2 macrophages to immune-promoting M1 macrophages. This study represents a promising step forward in immunotherapy treatment for breast cancer and warrants further clinical investigation. [ABSTRACT FROM AUTHOR]

Neoadjuvant immunotherapy with anti-cytotoxic T lymphocyte–associated protein 4 (CTLA4) + anti–programmed cell death protein 1 (PD1) monoclonal antibodies has demonstrated remarkable pathological responses and relapse-free survival in ~80% of patients with clinically detectable stage III melanoma. However, about 20% of the treated patients do not respond. In pretreatment biopsies of patients with melanoma, we found that resistance to neoadjuvant CTLA4 + PD1 blockade was associated with a low CD4/interleukin-2 (IL-2) gene signature. Ex vivo, addition of IL-2 to CTLA4 + PD1 blockade induced T cell activation and deep immunological responses in anti-CTLA4 + anti-PD1–resistant human tumor specimens. In the 4T1.2 breast cancer mouse model of neoadjuvant immunotherapy, triple combination of anti-CTLA4 + anti-PD1 + IL-2 cured almost twice as many mice as compared with dual checkpoint inhibitor therapy. This improved efficacy was due to the expansion of tumor-specific CD8+ T cells and improved proinflammatory cytokine polyfunctionality of both CD4+ and CD8+ T effector cells and regulatory T cells. Depletion studies suggested that CD4+ T cells were critical for priming of CD8+ T cell immunity against 4T1.2 and helped in the expansion of tumor-specific CD8+ T cells early after neoadjuvant triple immunotherapy. Our results suggest that the addition of IL-2 can overcome resistance to neoadjuvant anti-CTLA4 + anti-PD1, providing the rationale for testing this combination as a neoadjuvant therapy in patients with early-stage cancer. Immunotherapy triple threat: Some patients with melanoma do not respond to dual immunotherapy of anti-cytotoxic T lymphocyte–associated protein 4 (CTLA4) and anti–programmed cell death protein 1 (PD1). Kaptein et al. identified a low interleukin 2 (IL-2) gene signature associated with resistance in patients with melanoma. The addition of IL-2 to dual immunotherapy treatment of resistant ex vivo patient samples and mouse models induced immunological responses and extended survival. Therapy expanded tumor-specific CD8+ T cells and improved proinflammatory cytokine polyfunctionality of T cells in vivo. Triple-combination immunotherapy can overcome resistance to dual neoadjuvant immunotherapy and warrants further testing in patients with early-stage melanoma. [ABSTRACT FROM AUTHOR]

Melanoma is an immunogenic cancer with a high response rate to immune checkpoint inhibitors (ICIs). It harbors a high mutation burden compared with other cancers and, as a result, has abundant tumor-infiltrating lymphocytes (TILs) within its microenvironment. However, understanding the complex interplay between the stroma, tumor cells, and distinct TIL subsets remains a substantial challenge in immune oncology. To properly study this interplay, quantifying spatial relationships of multiple cell types within the tumor microenvironment is crucial. To address this, we used cytometry time-of-flight (CyTOF) imaging mass cytometry (IMC) to simultaneously quantify the expression of 35 protein markers, characterizing the microenvironment of 5 benign nevi and 67 melanomas. We profiled more than 220,000 individual cells to identify melanoma, lymphocyte subsets, macrophage/monocyte, and stromal cell populations, allowing for in-depth spatial quantification of the melanoma microenvironment. We found that within pretreatment melanomas, the abundance of proliferating antigen-experienced cytotoxic T cells (CD8+CD45RO+Ki67+) and the proximity of antigen-experienced cytotoxic T cells to melanoma cells were associated with positive response to ICIs. Our study highlights the potential of multiplexed single-cell technology to quantify spatial cell-cell interactions within the tumor microenvironment to understand immune therapy responses. Spacing of immune cells matters: A better understanding of the tumor immune microenvironment can determine biomarkers of immunotherapy effectiveness. Moldoveanu et al. used cytometry time-of-flight (CyTOF) imaging mass cytometry (IMC) to characterize the immune microenvironment of melanoma patient samples. CyTOF IMC identified the spatial arrangement and proximity of immune cells to each other. The researchers observed that a closer proximity of antigen-experienced cytotoxic T cells to melanoma cells intratumorally correlated to a patient's responsiveness to immune checkpoint blockade. Thus, spatial characterization of the tumor immune microenvironment using CyTOF IMC allows for the identification of prognostic biomarkers for immunotherapy. [ABSTRACT FROM AUTHOR]

Type 1 conventional dendritic cells (cDC1s) are rare immune cells critical for the induction of antigen-specific cytotoxic CD8+ T cells, although the genetic program driving human cDC1 specification remains largely unexplored. We previously identified PU.1, IRF8, and BATF3 transcription factors as sufficient to induce cDC1 fate in mouse fibroblasts, but reprogramming of human somatic cells was limited by low efficiency. Here, we investigated single-cell transcriptional dynamics during human cDC1 reprogramming. Human induced cDC1s (hiDC1s) generated from embryonic fibroblasts gradually acquired a global cDC1 transcriptional profile and expressed antigen presentation signatures, whereas other DC subsets were not induced at the single-cell level during the reprogramming process. We extracted gene modules associated with successful reprogramming and identified inflammatory signaling and the cDC1-inducing transcription factor network as key drivers of the process. Combining IFN-γ, IFN-β, and TNF-α with constitutive expression of cDC1-inducing transcription factors led to improvement of reprogramming efficiency by 190-fold. hiDC1s engulfed dead cells, secreted inflammatory cytokines, and performed antigen cross-presentation, key cDC1 functions. This approach allowed efficient hiDC1 generation from adult fibroblasts and mesenchymal stromal cells. Mechanistically, PU.1 showed dominant and independent chromatin targeting at early phases of reprogramming, recruiting IRF8 and BATF3 to shared binding sites. The cooperative binding at open enhancers and promoters led to silencing of fibroblast genes and activation of a cDC1 program. These findings provide mechanistic insights into human cDC1 specification and reprogramming and represent a platform for generating patient-tailored cDC1s, a long-sought DC subset for vaccination strategies in cancer immunotherapy. Encoding human dendritic cell fate: Recent advances in human dendritic cell (DC) biology have been accelerated by single-cell technologies, but DCs remain challenging to study because of their rarity and localization within tissues. Rosa et al. used single-cell transcriptomics to guide reprogramming of human fibroblasts into induced DCs and benchmarked their function against blood-derived DCs. Enforced expression of the transcription factors PU.1, IRF8, and BATF3 induced a small population resembling type 1 conventional DCs (cDC1s), which could be improved to more than 75% efficiency with a constitutive promoter and the inflammatory cytokines IFN-γ, IFN-β, and TNF-α. PU.1 chromatin binding early during reprogramming recruited IRF8 and BATF3 to facilitate cDC1-associated gene expression. Systems to generate functional patient-specific DCs represent a promising tool for studying human DCs and developing novel immunotherapies. [ABSTRACT FROM AUTHOR]

The use of coronavirus disease 2019 (COVID-19) vaccines will play the major role in helping to end the pandemic that has killed millions worldwide. COVID-19 vaccines have resulted in robust humoral responses and protective efficacy in human trials, but efficacy trials excluded individuals with a prior diagnosis of COVID-19. As a result, little is known about how immune responses induced by mRNA vaccines differ in individuals who recovered from COVID-19. Here, we evaluated longitudinal immune responses to two-dose BNT162b2 mRNA vaccination in 15 adults who had experienced COVID-19, compared to 21 adults who did not have prior COVID-19. Consistent with prior studies of mRNA vaccines, we observed robust cytotoxic CD8+ T cell responses in both cohorts after the second dose. Furthermore, SARS-CoV-2–naive individuals had progressive increases in humoral and antigen-specific antibody-secreting cell (ASC) responses after each dose of vaccine, whereas SARS-CoV-2–experienced individuals demonstrated strong humoral and antigen-specific ASC responses to the first dose but these responses were not further enhanced after the second dose of the vaccine at the time points studied. Together, these data highlight the relevance of immunological history for understanding vaccine immune responses and may have implications for personalizing mRNA vaccination regimens used to prevent COVID-19, including for the deployment of booster shots. The impact of prior immunity: Prior infection with SARS-CoV-2 elicits antibodies and T cell responses that influence subsequent responses to vaccines. Here, Samanovic et al. evaluated adaptive immune responses in individuals who received two doses of the BNT162b2 COVID-19 vaccine with or without prior confirmed SARS-CoV-2 infection. The authors found that humoral and cellular immune responses were elicited in both cohorts. Individuals who were vaccinated after prior infection exhibited increased humoral immunity after the first dose of vaccine that was stable after the second dose. Together, these findings show that immunological history may be important for vaccine and booster regimens. [ABSTRACT FROM AUTHOR]

T cell receptor (TCR)–based therapy has the potential to induce durable clinical responses in patients with cancer by targeting intracellular tumor antigens with high sensitivity and by promoting T cell survival. However, the need for TCRs specific for shared oncogenic antigens and the need for manufacturing protocols able to redirect T cell specificity while preserving T cell fitness remain limiting factors. By longitudinal monitoring of T cell functionality and dynamics in 15 healthy donors, we isolated 19 TCRs specific for Wilms' tumor antigen 1 (WT1), which is overexpressed by several tumor types. TCRs recognized several peptides restricted by common human leukocyte antigen (HLA) alleles and displayed a wide range of functional avidities. We selected five high-avidity HLA-A*02:01–restricted TCRs, three that were specific to the less explored immunodominant WT137–45 and two that were specific to the noncanonical WT1−78–64 epitopes, both naturally processed by primary acute myeloid leukemia (AML) blasts. With CRISPR-Cas9 genome editing tools, we combined TCR-targeted integration into the TCR α constant (TRAC) locus with TCR β constant (TRBC) knockout, thus avoiding TCRαβ mispairing and maximizing TCR expression and function. The engineered lymphocytes were enriched in memory stem T cells. A unique WT137–45-specific TCR showed antigen-specific responses and efficiently killed AML blasts, acute lymphoblastic leukemia blasts, and glioblastoma cells in vitro and in vivo in the absence of off-tumor toxicity. T cells engineered to express this receptor are being advanced into clinical development for AML immunotherapy and represent a candidate therapy for other WT1-expressing tumors. Alternative WT1 targets: Adoptive cell therapy using T cell receptor (TCR)–modified T cells has shown clinical success. However, it remains a challenge to find TCRs that respond to shared tumor antigens presented in common human leukocyte antigen molecules. To that end, Ruggiero et al. and Lahman et al. identified and tested TCR constructs targeting alternative epitopes in the Wilms' tumor antigen 1 (WT1) protein. Ruggiero and colleagues identified WT137–45 as a candidate peptide for TCR-based therapies and used CRISPR-Cas9 editing to target WT137–45-specific TCR sequences to the TRAC locus. Lahman and colleagues showed that, whereas a TCR-targeted WT1 epitope was lost because of immunoproteasome down-regulation, WT137–45 retains its expression and could be targeted by T cells expressing TCRs specific to that epitope, even in cancer cells that lose immunoproteasome activity. In both cases, T cells expressing WT137–45-specific TCRs controlled tumor burden in murine models. Thus, WT137–45 represents a candidate target for TCR-based immunotherapies that is resistant to immunoproteasome down-regulation. [ABSTRACT FROM AUTHOR]

Immunotherapy with chimeric antigen receptor (CAR)–engineered T cells showed exceptional successes in patients with refractory B cell malignancies. However, first-in-human studies in solid tumors revealed unique hurdles contributing to poor demonstration of efficacy. Understanding the determinants of tumor recognition by CAR T cells should translate into the design of strategies that can overcome resistance. Here, we show that multiple carcinomas express extracellular N-glycans, whose abundance negatively correlates with CAR T cell killing. By knocking out mannoside acetyl-glucosaminyltransferase 5 (MGAT5) in pancreatic adenocarcinoma (PAC), we showed that N-glycans protect tumors from CAR T cell killing by interfering with proper immunological synapse formation and reducing transcriptional activation, cytokine production, and cytotoxicity. To overcome this barrier, we exploited the high metabolic demand of tumors to safely inhibit N-glycans synthesis with the glucose/mannose analog 2-deoxy-d-glucose (2DG). Treatment with 2DG disrupts the N-glycan cover on tumor cells and results in enhanced CAR T cell activity in different xenograft mouse models of PAC. Moreover, 2DG treatment interferes with the PD-1–PD-L1 axis and results in a reduced exhaustion profile of tumor-infiltrating CAR T cells in vivo. The combined 2DG and CAR T cell therapy was successful against multiple carcinomas besides PAC, including those arising from the lung, ovary, and bladder, and with different clinically relevant CAR specificities, such as CD44v6 and CEA. Overall, our results indicate that tumor N-glycosylation regulates the quality and magnitude of CAR T cell responses, paving the way for the rational design of improved therapies against solid malignancies. Going after glycosylation: Chimeric antigen receptor (CAR) T cells have not performed as well against solid tumors as they have against hematological malignancies. One mechanism of resistance that tumor cells used to prevent CAR T cell killing is by glycosylating surface proteins. To reverse this resistance, Greco et al. genetically ablated out mannoside acetyl-glucosaminyltransferase 5 (MGAT5) in tumor cells or treated mice with the glucose/mannose analog 2-deoxy-d-glucose (2DG), both of which disrupted N-glycan expression on the surface of pancreatic adenocarcinoma cells. Disrupting N-glycan synthesis and expression enhanced CAR T cell activity and led to improved killing of not only pancreatic tumors but also tumors of the lung, ovary, and bladder. Together, these findings highlight tumor N-glycosylation as a therapeutic target to improve immunotherapies. [ABSTRACT FROM AUTHOR]

Interleukin-12 (IL-12) plays a critical role in modulating the tumor immune environment, including activating and expanding tumor-infiltrating lymphocytes (TILs). (C) Tumor growth curves of C57BL/6N mice (n = 8 per group) bearing MC38 tumors that were depleted of T cells and then treated with PBS or 3.33 pmol of pro-IL-12 by intraperitoneal injection on days 13, 16, and 19. (D) Tumor growth curves of MC38 tumor-bearing mice (n = 11 per group) given FTY720 and then treated with 3.33 pmol of pro-IL-12 by intraperitoneal injection on days 11, 14, and 17. A next-generation tumor-targeting IL-2 preferentially promotes tumor-infiltrating CD8+ T-cell response and effective tumor control. [Extracted from the article]

γδ T cells and malaria immunity: Repeated infections with Plasmodium falciparum can induce protective immunity against malaria, but mechanisms mediating protection are not completely understood. von Borstel et al. observed that repeated P. falciparum infection altered the ratios of human γδ T cells in children and adults with naturally occurring infections in Mali and found enrichment and maintenance of cytotoxic Vδ1effector T cells compared with naïve controls who had innate-like Vδ2+ γδ T cells as a dominant subset. U.S. adults subjected to sequential controlled human malaria infections (CHMI) initially expanded Vδ2+ γδ T cells, but repeated infection promoted clonal selection, differentiation, and expansion of cytotoxic Vδ1effector T cells. These Vδ1+ γδ T cells from U.S. and Malian individuals reacted to P. falciparum parasites in vitro, suggesting a role for this subset in malaria immunity. Repeated Plasmodium falciparum infections drive the development of clinical immunity to malaria in humans; however, the immunological mechanisms that underpin this response are only partially understood. We investigated the impact of repeated P. falciparum infections on human γδ T cells in the context of natural infection in Malian children and adults, as well as serial controlled human malaria infection (CHMI) of U.S. adults, some of whom became clinically immune to malaria. In contrast to the predominant Vδ2+ T cell population in malaria-naïve Australian individuals, clonally expanded cytotoxic Vδ1effector T cells were enriched in the γδ T cell compartment of Malian subjects. Malaria-naïve U.S. adults exposed to four sequential CHMIs defined the precise impact of P. falciparum on the γδ T cell repertoire. Specifically, innate-like Vδ2+ T cells exhibited an initial robust polyclonal response to P. falciparum infection that was not sustained with repeated infections, whereas Vδ1+ T cells increased in frequency with repeated infections. Moreover, repeated P. falciparum infection drove waves of clonal selection in the Vδ1+ T cell receptor repertoire that coincided with the differentiation of Vδ1naïve T cells into cytotoxic Vδ1effector T cells. Vδ1+ T cells of malaria-exposed Malian and U.S. individuals were licensed for reactivity to P. falciparum parasites in vitro. Together, our study indicates that repeated P. falciparum infection drives the clonal expansion of an adaptive γδ T cell repertoire and establishes a role for Vδ1+ T cells in the human immune response to malaria. [ABSTRACT FROM AUTHOR]

Inducing immunosuppression: Type 1 regulatory T cells (Tr1 cells) are a population of inducible regulatory T cells that suppress graft-versus-host disease (GvHD) after allogeneic stem cell transplantation. To determine mechanisms of suppression by Tr1 cells, Chen et al. compared Tr1 cells from a human Tr1–enriched cell product, T-allo10, with non-Tr1 cells from the same product. The authors found that Tr1 cells had a distinct T cell receptor repertoire and transcriptional program. In addition, Tr1 cells suppressed both through IL-10 production and through expression of CTLA-4 and PD-1. Last, the authors showed that, in patients receiving T-allo10, Tr1 cells persist up to 1 year after transfer. These findings further characterize the function of Tr1 cells and demonstrate their clinical relevance for GvHD. Type 1 regulatory T (Tr1) cells are inducible, interleukin (IL)-10+FOXP3− regulatory T cells that can suppress graft-versus-host disease (GvHD) after allogeneic hematopoietic stem cell transplantation (allo-HSCT). We have optimized an in vitro protocol to generate a Tr1-enriched cell product called T-allo10, which is undergoing clinical evaluation in patients with hematological malignancies receiving a human leukocyte antigen (HLA)–mismatched allo-HSCT. Donor-derived T-allo10 cells are specific for host alloantigens, are anergic, and mediate alloantigen-specific suppression. In this study, we determined the mechanism of action of T-allo10 cells and evaluated survival of adoptively transferred Tr1 cells in patients. We showed that Tr1 cells, in contrast to the non-Tr1 population, displayed a restricted T cell receptor (TCR) repertoire, indicating alloantigen-induced clonal expansion. Tr1 cells also had a distinct transcriptome, including high expression of cytotoxic T lymphocyte–associated protein 4 (CTLA-4) and programmed cell death protein 1 (PD-1). Blockade of CTLA-4 or PD-1/PD-L1 abrogated T-allo10–mediated suppression, confirming that these proteins, in addition to IL-10, play key roles in Tr1-suppressive function and that Tr1 cells represent the active component of the T-allo10 product. Furthermore, T-allo10–derived Tr1 cells were detectable in the peripheral blood of HSCT patients up to 1 year after T-allo10 transfer. Collectively, we revealed a distinct molecular phenotype, mechanisms of action, and in vivo persistence of alloantigen-specific Tr1 cells. These results further characterize Tr1 cell biology and provide essential knowledge for the design and tracking of Tr1-based cell therapies. [ABSTRACT FROM AUTHOR]

keyimage.jpg Tailoring cancer vaccines to patients: Immunotherapies boost the body's immune system to detect and kill cancer cells. The immune response can be enhanced by molecules released from dying tumor cells, leading to proposals to use dead tumor cells as adjuvants in immunotherapeutics and cancer vaccines. However, Sriram et al. found that, in some cases, it is the remaining live tumor cells that are immunogenic. Treating cultured tumor cells with select DNA-damaging chemotherapies (and specific doses thereof) stimulated cytotoxic T cell activation when the live, not dead, cell fraction was injected back into the tumor in mice. Combining these cells with clinically used immune checkpoint inhibitors resulted in tumor regression and antitumor immune memory. The findings potentially identify a patient-specific approach to making immunotherapies more durably and broadly effective. Although immune checkpoint blockade (ICB) has strong clinical benefit for treating some tumor types, it fails in others, indicating a need for additional modalities to enhance the ICB effect. Here, we identified one such modality by using DNA damage to create a live, injured tumor cell adjuvant. Using an optimized ex vivo coculture system, we found that treating tumor cells with specific concentrations of etoposide, mitoxantrone, or doxorubicin markedly enhanced dendritic cell–mediated T cell activation. These immune-enhancing effects of DNA damage did not correlate with immunogenic cell death markers or with the extent of apoptosis or necroptosis; instead, these effects were mediated by live injured cells with activation of the DNA-PK, ATR, NF-κB, p38 MAPK, and RIPK1 signaling pathways. In mice, intratumoral injection of ex vivo etoposide–treated tumor cells in combination with systemic ICB (by anti-PD-1 and anti-CTLA4 antibodies) increased the number of intratumoral CD103+ dendritic cells and circulating tumor-antigen–specific CD8+ T cells, decreased tumor growth, and improved survival. These effects were absent in Batf3−/− mice and in mice in which the DNA-damaging drug was injected directly into the tumor, due to DNA damage in the immune cells. The combination treatment induced complete tumor regression in a subset of mice that were then able to reject tumor rechallenge, indicating that the injured cell adjuvant treatment induced durable antitumor immunological memory. These results provide a strategy for enhancing the efficacy of immune checkpoint inhibition in tumor types that do not respond to this treatment modality by itself. [ABSTRACT FROM AUTHOR]

keyimage.jpg Activating γδ T cells: Gamma delta T (γδ T) cells exhibit potent cytotoxic properties that can be activated against both solid and liquid tumors. Here, De Gassart et al. have developed a humanized monoclonal antibody, ICT01, which targets BTN3A, a plasma membrane receptor responsible for Vγ9Vδ2 T cell activation. Treatment with ICT01 prolonged survival of xenograft mouse models adoptively transferred with human Vγ9Vδ2 T cells and was well tolerated in cynomolgus macaques. In addition, treatment in a first-in-human, phase 1/2a clinical study of ICT01 showed tolerance and pharmacodynamic results in patients. ICT01 represents a promising clinical strategy to activate circulating Vγ9Vδ2 T cells for efficient killing of tumors. Gamma delta T (γδ T) cells are among the most potent cytotoxic lymphocytes. Activating anti–butyrophilin 3A (BTN3A) antibodies prime diverse tumor cell types to be killed by Vγ9Vδ2 T cells, the predominant γδ T cell subset in peripheral circulation, by mechanisms independent of tumor antigen–major histocompatibility complex (MHC) complexes. In this report, we describe the development of a humanized monoclonal antibody, ICT01, with subnanomolar affinity for the three isoforms of BTN3A. We demonstrate that ICT01-activated Vγ9Vδ2 T cells kill multiple tumor cell lines and primary tumor cells, but not normal healthy cells, in an efficient process requiring approximately 20% target occupancy. We show that ICT01 activity is dependent on BTN3A and BTN2A but independent of the phosphoantigen (pAg)–binding B30.2 domain. ICT01 delays the growth of hematologic and solid tumor xenografts and prolongs survival of NOD/SCID/IL2rγnull (NSG) mice adoptively transferred with human Vγ9Vδ2 T cells. In single- and multiple-dose safety studies in cynomolgus macaques that received up to 100 mg/kg once weekly, ICT01 was well tolerated. With respect to pharmacodynamic endpoints, ICT01 selectively activated Vγ9Vδ2 T cells without affecting other BTN3A-expressing lymphocytes such as αβ T or B cells. A first-in-human, phase 1/2a, open-label, clinical study of ICT01 was thus initiated in patients with advanced-stage solid tumors (EVICTION: NCT04243499; EudraCT: 2019-003847-31). Preliminary results show that ICT01 was well tolerated and pharmacodynamically active in the first patients. Digital pathology analysis of tumor biopsies of a patient with melanoma suggests that ICT01 may promote immune cell infiltration within the tumor microenvironment. [ABSTRACT FROM AUTHOR]

PGE2 signaling networks: The lipid mediator PGE2 suppresses antitumor immunity by activating its four related GPCRs on T cells. Lone et al. used quantitative phosphoproteomics and phosphoflow cytometry to analyze downstream signaling elicited by the stimulation of all receptors simultaneously or individually in different T cell subsets. The analysis revealed G protein–dependent and G protein–independent pathways that were activated by each receptor in all T cells, as well as pathways that were activated by only a subset of receptors, in only a subset of cells, or with receptor-specific kinetics. Network modeling predicted mechanisms of cross-talk and signal integration downstream of the receptors. These data are a comprehensive resource for future explorations of the functional consequences of PGE2 receptor–specific signaling in immune homeostasis, inflammation, and tumor-associated immunosuppression. Prostaglandin E2 (PGE2) promotes an immunosuppressive microenvironment in cancer, partly by signaling through four receptors (EP1, EP2, EP3, and EP4) on T cells. Here, we comprehensively characterized PGE2 signaling networks in helper, cytotoxic, and regulatory T cells using a phosphoproteomics and phosphoflow cytometry approach. We identified ~1500 PGE2-regulated phosphosites and several important EP1–4 signaling nodes, including PKC, CK2, PKA, PI3K, and Src. T cell subtypes exhibited distinct signaling pathways, with the strongest signaling in EP2-stimulated CD8+ cells. EP2 and EP4, both of which signal through Gαs, induced similar signaling outputs, but with distinct kinetics and intensity. Functional predictions from the observed phosphosite changes revealed PGE2 regulation of key cellular and immunological processes. Last, network modeling suggested signal integration between the receptors and a substantial contribution from G protein–independent signaling. This study offers a comprehensive view of the different PGE2-regulated phosphoproteomes in T cell subsets, providing a valuable resource for further research on this physiologically and pathophysiologically important signaling system. [ABSTRACT FROM AUTHOR]

keyimage.jpg Baseline killers aid immune checkpoint blockade: CD8+ T cells are crucial for the efficacy of immune checkpoint blockades (ICB), but it is unclear how specific T cell subsets or clones affect efficacy. Here, Watson et al. used transcriptomic approaches to phenotype the subsets and TCR clones of peripheral CD8+ T cells in eight patients with metastatic melanoma before and during ICB. They found that many of the expanded T cell clones during ICB were from a pool of cytotoxic T cell clones present pretreatment. In a larger independent clinical dataset of patients with melanoma, the authors found that the clone size and cytotoxicity of CD8+ T cells before treatment positively correlated to ICB efficacy. Together, these data suggest that targeting preexisting cytotoxic CD8+ T cell clones might improve the antitumor responses of ICB. The antitumor action of immune checkpoint blockade (ICB) is primarily mediated by CD8+ T cells. How sensitivity to ICB varies across CD8+ T cell subsets and clonotypes and the relationship of these with clinical outcome is unclear. To explore this, we used single-cell V(D)J and RNA-sequencing to track gene expression changes elicited by ICB across individual peripheral CD8+ T cell clones, identify baseline markers of CD8+ T cell clonal sensitivity, and chart how CD8+ T cell transcriptional changes vary according to phenotypic subset and clonal size. We identified seven subsets of CD8+ T cells with divergent reactivity to ICB and found that the cytotoxic effector subset showed the greatest number of differentially expressed genes while remaining stable in clonal size after ICB. At the level of CD8+ T cell clonotypes, we found a relationship between transcriptional changes and clone size, with large clones showing a greater number of differentially regulated genes enriched for pathways including T cell receptor (TCR) signaling. Cytotoxic CD8+ effector clones were more likely to persist following ICB and were more likely to correspond with public tumor-infiltrating lymphocyte clonotypes. Last, we demonstrated that individuals whose CD8+ T cell pretreatment showed low cytotoxicity and had fewer expanded clones typically had worse outcomes after ICB treatment. This work further advances understanding of the molecular determinants of ICB response, assisting in the search for peripheral prognostic biomarkers and highlighting the importance of the baseline CD8+ immune landscape in determining ICB response in metastatic melanoma. [ABSTRACT FROM AUTHOR]

Taking a BiTE out of tumors: Bispecific T cell engagers (BiTE) and other forms of T cell engager therapies represent a promising off-the-shelf treatment for many types of cancer. However, it is unclear why some types of tumors are particularly resistant to BiTE treatment. To address this, Belmontes et al. evaluated pharmacokinetics, pharmacodynamics, and immune correlates of responsiveness to BiTE treatment. The authors found that, in multiple mouse tumor models, pretreatment T cell density in tumors associated with BiTE efficacy. Further, in T cell–cold tumors, which lack abundant T cells, coadministration of immune checkpoint blockade antibodies or a 4-1BB agonist overcame resistance to BiTE treatment. Together, these findings identify combination therapeutic strategies for BiTE and other T cell engagers. Therapeutic approaches are needed to promote T cell–mediated destruction of poorly immunogenic, "cold" tumors typically associated with minimal response to immune checkpoint blockade (ICB) therapy. Bispecific T cell engager (BiTE) molecules induce redirected lysis of cancer cells by polyclonal T cells and have demonstrated promising clinical activity against solid tumors in some patients. However, little is understood about the key factors that govern clinical responses to these therapies. Using an immunocompetent mouse model expressing a humanized CD3ε chain (huCD3e mice) and BiTE molecules directed against mouse CD19, mouse CLDN18.2, or human EPCAM antigens, we investigated the pharmacokinetic and pharmacodynamic parameters and immune correlates associated with BiTE efficacy across multiple syngeneic solid-tumor models. These studies demonstrated that pretreatment tumor-associated T cell density is a critical determinant of response to BiTE therapy, identified CD8+ T cells as important targets and mediators of BiTE activity, and revealed an antagonistic role for CD4+ T cells in BiTE efficacy. We also identified therapeutic combinations, including ICB and 4-1BB agonism, that synergized with BiTE treatment in poorly T cell–infiltrated, immunotherapy-refractory tumors. In these models, BiTE efficacy was dependent on local expansion of tumor-associated CD8+ T cells, rather than their recruitment from circulation. Our findings highlight the relative contributions of baseline T cell infiltration, local T cell proliferation, and peripheral T cell trafficking for BiTE molecule–mediated efficacy, identify combination strategies capable of overcoming resistance to BiTE therapy, and have clinical relevance for the development of BiTE and other T cell engager therapies. [ABSTRACT FROM AUTHOR]

But in cell culture studies, Li and colleagues found, human CD8 T cells carrying KIR proteins killed the gliadin-detecting helper T cells. CD8 T cells are best known for killing infected or cancerous cells, but in mice some of them also kill T cells that orchestrate autoimmune reactions. The price for a vigilant immune system that can pounce on tumor cells or pathogens is occasional friendly fire - an autoimmune attack. [Extracted from the article]