Biology subjects

Kane, L. P.

Publications and source records attributed to Kane, L. P..

10 recordsLinked to original sources

bioRxiv2026

ATR kinase inhibitors induce mitochondrial fission in CD8+ T cells and impair immune memory in vivo

The DNA damage response kinase ATR restrains CDK1 activity during S and G2 phases of the cell cycle, confining CDK1-driven processes to mitosis. ATR kinase inhibitors were originally developed to potentiate chemotherapy-induced DNA damage at stalled replication forks and to disrupt DNA damage-induced cell cycle checkpoints. Recent evidence, however, reveals that these inhibitors also disrupt cell cycle organization in cells that have not sustained any DNA damage. We show that ATR kinase inhibitors potently trigger unscheduled mitochondrial fission, causing loss of mitochondrial mass in actively dividing CD8+ T cells that persists in memory CD8+ T cells. Moreover, ATR inhibition during the peak of CD8+ T cell expansion in a mouse model of LCMV Armstrong infection impairs the formation of immune memory. These findings carry significant clinical implications. ATR kinase inhibitors are currently being evaluated in clinical trials in combination with chemotherapy, radiation, and immune checkpoint inhibitors in patients where anti-tumor immune responses are recognized as a determinant of durable response. Our results identify an unexpected consequence of ATR inhibition that disrupts cellular metabolism with broad implications for both preclinical research and clinical application.

cancer biology↗

bioRxiv2026

T cell-specific loss of IRF1 results in defective CD8 T cell activation and antitumor immunity

Interferon regulatory factor 1 (IRF1) has long been recognized as a tumor suppressor; however, recent studies have revealed context-specific and sometimes opposing roles in cancer progression. Here, we describe a T cell-specific mechanism underlying the antitumor activity of IRF1. Unlike germline Irf1-deficient mice, T cell-specific loss of IRF1 does not lead to a deficiency in cytotoxic CD8 T cells. Nevertheless, tumor burden remains elevated in these mice, associated with reduced CD8 T cell infiltration driven by impaired activation and proliferation in the absence of IRF1. Transcriptomic analysis of activated Irf1-deficient T cells identified NFATc1 as a key gene significantly downregulated upon IRF1 loss. Analysis of human melanoma datasets further corroborated this finding, highlighting a previously unappreciated role for IRF1 in regulating T cell activation and antitumor immunity.

immunology↗

bioRxiv2025

Pik3ip1/TrIP Regulation of PI3K Restricts CD8 T Cell Anti-Tumor Immunity

BackgroundThe protein PI3K-interacting protein (PIK3IP1), or transmembrane inhibitor of PI3K (TrIP), is highly expressed by T cells and can modulate PI3K activity in these cells. Several studies have also revealed that TrIP is rapidly downregulated following T cell activation and can play important roles in T cell differentiation. MethodsWe generated mice with CD8-specific TrIP deficiency. We then implanted these mice, and Cre-only control animals, with B16 melanoma or MC38 colon carcinoma tumors. Tumor growth and anti-tumor immunity were then followed. We also assessed the effects of TrIP deficiency on transcriptional programs in CD8 T cells stimulated in vivo or derived from tumor- bearing mice. ResultsWe found that activated TrIP KO CD8 T cells display an increased inflammatory transcriptional profile in the absence of TrIP. Consistent with these effects, we also found that knockout of TrIP specifically in CD8 T cells resulted in reduced growth of syngeneic tumors. When characterizing the tumor-infiltrating cells, TrIP KO led to an increase in the number of tumor-infiltrating T cells, as well as a delay in the acquisition of an exhausted phenotype, based on phenotypic and transcriptomic analyses. Finally, our data suggest that TrIP regulates the diversity of T cell clonal responses to tumors, since we observed an increase in the number of distinct T cell clonotypes responding to a tumor neoantigen. ConclusionsTaken together, our findings demonstrate that TrIP intrinsically restricts the CD8 T cell response to tumors, and that targeting TrIP may augment the anti-tumor response in a way that is distinct from established checkpoint therapies.

immunology↗

bioRxiv2025

Tim-3 Promotes Early Differentiation of Tbet+ Effector T Cells During Acute Viral Infection

The transmembrane protein Tim-3 has received significant attention in recent years as a possible immunotherapy target. This is due to its robust expression on dysfunctional, exhausted, T cells found in the settings of cancer and chronic infection and biochemical evidence suggesting an inhibitory function of Tim-3. However, numerous clinical trials of putative Tim-3 blocking antibodies have yielded modest benefits, at best, in clinical trials for various cancers. Thus, there is a need to more fully understand the function of Tim-3 in vivo. Here we have studied the function of Tim-3 early during a T cell response to LCMV Armstrong, which causes an acute viral infection in mice. We show that Tim-3 is rapidly expressed after infection and that the expression of Tim-3 is associated with acquisition of a type I effector phenotype, including expression of T-bet and downregulation of Tcf-1, by both CD4+ and CD8+ T cells. In addition, we demonstrate that knockout or cytoplasmic truncation of Tim-3 attenuates the acquisition of the effector program by T cells after LCMV Armstrong infection. Together, these data help to clarify the role of Tim-3 during acute infection.

immunology↗

bioRxiv2024

Downregulation of PIK3IP1/TrIP on T cells is controlled by TCR signal strength, PKC, and metalloprotease-mediated cleavage

The protein known as PI3K-interacting protein (PIK3IP1), or transmembrane inhibitor of PI3K (TrIP), is highly expressed by T cells and can modulate PI3K activity in these cells. Several studies have also revealed that TrIP is rapidly downregulated following T cell activation. However, it is unclear as to how this downregulation is controlled. Using a novel monoclonal antibody that robustly stains cell-surface TrIP, we demonstrate that TrIP is lost from the surface of activated T cells in a manner dependent on the strength of signaling through the T cell receptor (TCR) and specific downstream signaling pathways, in particular classical PKC isoforms. TrIP expression returns by 24 hours after stimulation, suggesting that it may play a role in resetting TCR signaling at later time points. We also provide evidence that ADAM family proteases are required for both constitutive and stimulation-induced downregulation of TrIP in T cells. Finally, by expressing truncated forms of TrIP in cells, we identify the region in the extracellular stalk domain of TrIP that is targeted for proteolytic cleavage.

cell biology↗

bioRxiv2020

Expression of Tim-3 drives naive Treg to an effector-like state with enhanced suppressive activity

Regulatory T cells (Treg) are critical mediators of self-tolerance but can also limit effective anti-tumor immunity. We and others previously reported that 40-60% percent of Treg-infiltrating head and neck cancer (HNC) and other tumors highly express Tim-3, compared with about 5% in lymphoid organs. Tumor-infiltrating Tim-3+ Treg also have enhanced suppressive function and display a more effector-like phenotype. Using a novel mouse model with cell type-specific Tim-3 expression, we show here that expression of Tim-3 by Treg is sufficient to drive Treg to a more effector-like phenotype, resulting in enhanced suppressive activity and increased tumor growth. These findings may help to reconcile previous reports that some Tim-3 antibodies enhance T cell responses in vivo, while expression of Tim-3 has a cell-intrinsic ability to enhance TCR signaling and T cell activation. Thus, we propose that Tim-3 regulates anti-tumor immunity at least in part through enhancement of Treg function. To our knowledge, this is the first example in which expression of a single co-stimulatory molecule is sufficient to drive differentiation of Treg in this manner.

immunology↗

bioRxiv2019

The co-stimulatory activity of Tim-3 requires Akt and MAPK signaling and immune synapse recruitment

Expression of the transmembrane protein Tim-3 is increased on dysregulated T cells undergoing chronic T cell activation, including in chronic infection and solid tumors. We and others previously reported that Tim-3 exerts apparently paradoxical co-stimulatory activity in T cells (and other cells), including enhancement of ribosomal S6 protein phosphorylation (pS6). Here we examined the upstream signaling pathways that control Tim3-mediated increases in pS6 in T cells. We have also defined the localization of Tim-3 relative to the T cell immune synapse and impacts on downstream signaling. Recruitment of Tim-3 to the immune synapse was mediated exclusively by the transmembrane domain, replacement of which impaired Tim-3 co-stimulation of pS6. Strikingly, enforced localization of the Tim-3 cytoplasmic domain to the immune synapse in the context of a chimeric antigen receptor still allowed for robust T cell activation. Our findings are consistent with a model whereby Tim-3 enhances TCR-proximal signaling under acute conditions. One Sentence SummaryHere we define elements of signaling and localization associated with Tim-3 co-stimulatory function in T cells.

immunology↗

bioRxiv2019

Polyubiquitin-dependent recruitment of NEMO/IKKγ into T cell receptor signaling microclusters

The NF-{kappa}B essential modulator protein (NEMO) is required for activation of canonical NF-{kappa}B by the T cell antigen receptor (TCR). However, the subcellular localization of NEMO during this process is not well understood. By dynamically imaging fluorescent NEMO chimeras in live human T cells, we demonstrate that NEMO is rapidly recruited into TCR microclusters via domains previously implicated in the recognition of linear and K63-linked polyubiquitin. The recruitment of NEMO into TCR microclusters requires the activities of the tyrosine kinases Lck and ZAP-70, but not the adaptor proteins LAT or SLP-76. Thus, our findings reveal that the pathways leading from TCR to NF-{kappa}B bifurcate downstream of ZAP-70 to independently control the recruitment and activation of NEMO.

immunology↗

bioRxiv2017

PIK3IP1/TrIP restricts activation of T cells through inhibition of PI3K/Akt

Phosphatidylinositol-3 kinases (PI3Ks) modulate numerous cellular functions, including growth, proliferation and survival. Dysregulation of the PI3K pathway can lead to autoimmune disease and cancer. PIK3IP1 (or Transmembrane Inhibitor of PI3K - TrIP) is a novel transmembrane regulator of PI3K. TrIP contains an extracellular kringle domain and an intracellular \"p85-like\" domain with homology to the inter-SH2 domain of the regulatory subunit of PI3K. Although TrIP has been shown to bind to the p110 catalytic subunit of PI3K in fibroblasts, the mechanism by which TrIP functions is poorly understood. We show that both the kringle and \"p85-like\" domains are necessary for TrIP inhibition of PI3K. We also demonstrate that TrIP protein is down-modulated from the surface of T cells to allow T cell activation. In addition, we present evidence that the kringle domain may modulate TrIP function by binding an as-yet-unidentified ligand. Using an inducible knockout mouse model that we developed, we show that TrIP-deficient T cells exhibit more robust T cell activation, show a preference for Th1 polarization and can mediate clearance of Listeria monocytogenes infection faster than WT mice. Thus, TrIP is an important negative regulator of T cell activation and may represent a novel target for immune modulation therapies.

immunology↗

bioRxiv2017

Tim-3 co-stimulation promotes short-term effector T cells, restricts memory precursors and is dispensable for T cell exhaustion

Tim-3 is highly expressed on a subset of T cells during T cell exhaustion, in settings of chronic viral infection and tumors (1, 2). Using LCMV Clone-13, a model for chronic infection, we have found that Tim-3 is neither necessary nor sufficient for the development of T cell exhaustion. Nonetheless, expression of Tim-3 was sufficient to drive resistance to PD-L1 blockade therapy during chronic infection. Strikingly, expression of Tim-3 promoted development of short-term effector T cells, at the expense of memory precursor development, after acute LCMV infection. These effects were accompanied by increased Akt/mTOR signaling in T cells expressing endogenous or ectopic Tim-3. Conversely, Akt/mTOR signaling was reduced in effector T cells from Tim-3 deficient mice. Thus, Tim-3 is essential for optimal effector T cell responses, but may also contribute to exhaustion, by restricting development of long-lived memory T cells, including PD-1int \"stem-like\" exhausted T cells that expand during PD-1 pathway blockade. Taken together, our results suggest that Tim-3 is actually more similar to co-stimulatory receptors that are upregulated after T cell activation, rather than a dominant inhibitory protein like PD-1. These findings have significant implications for the development of anti-Tim-3 antibodies as therapeutic agents.\n\nSignificanceDuring a chronic viral infection, prolonged exposure to viral antigens leads to dysfunction or \"exhaustion\" of T cells specific to the virus, a condition also observed in T cells that infiltrate tumors. The exhausted state is associated with expression of specific cell-surface proteins, some of which may inhibit T cell activation. Expression of Tim-3 is associated with acquisition of T cell exhaustion, although it is also expressed transiently during acute infection. Here we provide evidence that a major function of Tim-3 is to enhance T cell activation, during either acute or chronic viral infection. However, Tim-3 is not required for development of exhaustion. Thus, we propose that Tim-3 would be better described as a stimulatory, rather than inhibitory, protein.

immunology↗