Biology subjects

Lesinski, G. B.

Publications and source records attributed to Lesinski, G. B..

11 recordsLinked to original sources

bioRxiv2026

Immune determinants of leuprolide-associated cardiovascular risk in prostate cancer

Leuprolide, a widely used androgen-deprivation therapy (ADT) for prostate cancer (PC), has been associated with increased cardiovascular events (CVE), although the underlying mechanisms remain unclear. Through integrated spatial transcriptomic analyses of murine heart and immune-stromal profiling in patient blood, we identified immune programs linked to leuprolide-associated CVE risk. Spatial analyses revealed the distinct organization of lymphoid (T+B cells) aggregates and inflammatory macrophages in the plaque following leuprolide exposure that accompanied by ventricular hypertrophy, relative to normal vasculature. Parallel profiling of circulatory soluble and cellular components in patients demonstrated CD8+T cell enriched pattern in ~44% of patients, including memory and type 1 cytotoxic subsets that linked to treatment-induced CVE, such as ventricular dysfunctions, but no contribution of adipocytokines. These translational findings highlight the stratification of leuprolide-associated CVE risk in PC patients based on circulating T cell profiling and could guide the risk mitigation with alternative ADT agents.

immunology↗

bioRxiv2026

MetScore and classical-basal subtype intersect to define chemotherapy response and tumor microenvironment architecture in pancreatic ductal adenocarcinoma

Background Transcriptomic heterogeneity in pancreatic ductal adenocarcinoma (PDAC) is conventionally modeled along a single classical-basal axis, but this framework incompletely explains clinical behavior. We hypothesized that MetScore, a single-sample measure of metastatic colonization potential, defines a complementary axis. Objective To determine how MetScore and classical-basal identity jointly shape PDAC biology and clinical outcomes. Design We integrated RNA sequencing, DNA sequencing, clinical, and digital pathology data from 512 tumor biopsies from 510 patients in the Pancreatic Cancer Action Network Know Your Tumor real-world cohort. We assessed associations with biopsy site, survival, chemotherapy response, and tumor microenvironment composition; validated metastatic discrimination in an independent cohort; and examined cancer cell-intrinsic drug sensitivity in vitro. Results MetScore distinguished metastatic from primary biopsies, a finding validated externally. High MetScore and basal identity were independently associated with inferior survival. High MetScore and classical identity were each associated with preferential response to 5-fluorouracil (5-FU)-based rather than gemcitabine-based therapy, with the strongest enrichment for 5-FU responses among classical/MetScore-high patients. In vitro drug-sensitivity experiments did not recapitulate the clinical treatment-response pattern, motivating investigation of cancer cell-extrinsic factors. Transcriptomic deconvolution and histopathologic machine learning-based cell annotation demonstrated that high-MetScore and basal states converged on macrophage/monocyte enrichment but diverged in their associations with cancer-associated fibroblasts. Conclusion Integrating MetScore with classical-basal identity establishes a two-axis framework that better explains metastatic behavior, prognosis, treatment response, and microenvironmental composition than the conventional one-axis model and provides a clinically testable strategy for transcriptome-guided patient stratification.

cancer biology↗

bioRxiv2025

Neoantigens drive adoptively transferred CD8 T cells to long-lived effectors mediated by lymph node trafficking

Adoptive transfer of neoantigen-reactive T lymphocytes mediates potent responses against solid tumors in patients while often limiting toxicity to normal tissues; however, the mechanisms governing the trafficking, activation, and superior antitumor function of infused T cells remain unknown. Using a clinically relevant TCR-transgenic T cell therapy model, we examined CD8 T cell responses to melanoma expressing either wild-type or mutated antigen. Neoantigen expression conferred robust tumor regression, durable cures, and long-term protective immunity. Mechanistically, T cells reacting to neoantigen exhibited enhanced cytokine and chemokine production, heightened effector function, and sustained persistence within the blood, tumor, and lymph nodes. Notably, trafficking through secondary lymphoid organs was necessary for T cell persistence and antitumor efficacy. These findings highlight the critical role of T cell trafficking to the lymph nodes in shaping neoantigen-specific antitumor responses and offers insight for improving adoptive cellular therapies.

cancer biology↗

bioRxiv2025

High-Dimensional Protein Analysis Uncovers Distinct Immunological and Stromal Signatures Between Primary and Metastatic Pancreatic Ductal Adenocarcinoma

Our understanding of the pancreatic ductal adenocarcinoma (PDAC) tumor microenvironment (TME) primarily stems from murine models or primary patient tumors. While metastatic tumors have generally less immune infiltration compared to primary tumors, the specific cellular features of metastatic PDAC remain understudied. This knowledge gap is impactful as most patients present with metastatic disease and are most often enrolled in clinical trials. We hypothesized PDAC tumors harbor distinct immunologic and stromal features depending on their anatomical site. Using multiplex immunohistochemistry (mIHC), spatial analysis, and single-cell mass cytometry (CyTOF), we uncover dominant immune and stromal cell populations in tumors derived from 27 primary and 26 liver metastases. Metastatic liver tumors from PDAC patients contained fewer T cells and alpha-smooth muscle actin (-SMA+) activated fibroblasts than primary lesions, while CD68+ cells were more abundant. Spatial analyses revealed distinct immune cell communities in primary and metastatic PDAC, whereby CK19+ cells clustered differentially with -SMA+, CD3+, and CD68+ cells, depending on tumor site. When comparing tumor-associated regions, the proportion of peritumoral CK19- cells remained consistent, but their composition varied by disease site. CD8+ T cells were significantly less frequent in metastatic tumors, while both CD4+ and CD8+ T cells present in primary tumors expressed more transcription factors (TFs) associated with suppressive properties, including FoxP3 and ROR{gamma}t. CyTOF revealed that T cells co-expressed multiple inhibitory checkpoint receptors, with LAG-3 and PD-1 predominating. This report reveals that primary and metastatic tumors from PDAC patients harbor vastly distinct immunologic and stromal features at the protein level. Statement of SignificanceProtein level analysis reveals distinct immunological and stromal features between primary and metastatic PDAC tumors, offering a rationale for immunotherapies that target myeloid cells and increase T cell abundance in metastatic disease.

cancer biology↗

bioRxiv2024

VPAC2 receptor signaling promotes pancreatic cancer cell growth and decreases the immunogenicity of the tumor microenvironment.

Identifying mechanisms underlying tumor growth and immune resistance is needed to treat pancreatic ductal adenocarcinoma (PDAC) effectively. The complexity of the tumor microenvironment (TME) suggests that the crosstalk between cells in the TME could drive drug resistance and relapse in PDAC. We have previously determined that vasoactive intestinal peptide (VIP) is overexpressed in PDAC and that VIP receptors expressed on T cells are a targetable pathway that sensitizes PDAC to anti-PD1 therapy. In this study, we show that pancreatic cancer cells engage in autocrine signaling of VIP through VIP-receptor 2 (VPAC2), and that high co-expression of VIP with VPAC2 leads to reduced relapse-free survival in PDAC patients. Mechanistically, we identified piwi-like RNA-mediated gene silencing2 (Piwil2) as a tumor-cell intrinsic protein downstream of VPAC2 that regulates cancer cell growth. In addition, we discovered TGF{beta}-1 as a potential tumor-extrinsic inhibitor of T cell function induced by VPAC2 signaling. In vivo, knock out and knockdown of VPAC2 on PDAC cells led to reduced tumor growth rate and increased sensitivity to anti-PD-1 therapy in various mouse models of PDAC that were T-cell dependent. Overall, these findings emphasize the implications of VIP/VPAC2 signaling in the PDAC tumor microenvironment and further support the rationale for developing VPAC2-specific antagonists. SignificanceThe autocrine VIP signaling via VPAC2 promotes cancer cell growth and dampens T cell function in pancreatic ductal adenocarcinoma and thus represents a potential therapeutic target in PDAC.

cancer biology↗

bioRxiv2023

Distinct host preconditioning regimens differentially impact the antitumor potency of adoptively transferred Th17 cells.

BackgroundMechanisms by which distinct methods of host preconditioning impact the efficacy of adoptively transferred antitumor T helper cells is unknown. MethodsCD4+ T cells with a transgenic TCR that recognize TRP-1 melanoma antigen were polarized to the T helper 17 (Th17) phenotype and then transferred into melanoma-bearing mice preconditioned with either total body irradiation or chemotherapy. ResultsWe found that preconditioning mice with a non-myeloablative dose of total body irradiation (TBI of 5 Gy) was more effective than using an equivalently dosed non-myeloablative chemotherapy (CTX at 200 mg/kg) at augmenting therapeutic activity of anti-tumor TRP-1 Th17 cells. Anti-tumor Th17 cells engrafted better following preconditioning with TBI and regressed large established melanoma in all animals. Conversely, only half of mice survived long-term when preconditioned with CTX and infused with anti-melanoma Th17 cells. IL-17 and IFN-g produced by the infused Th17 cells, were detected in animals given either TBI or CTX preconditioning. Interestingly, inflammatory cytokines (G-CSF, IL-6, MCP-1, IL-5, and KC) were significantly elevated in the serum of mice preconditioned with TBI versus CTX after Th17 therapy. ConclusionsOur results indicate, for the first time, that the antitumor response, persistence, and cytokine profiles resulting from Th17 therapy are impacted by the specific regimen of host preconditioning. This work is important for understanding mechanisms that promote long-lived responses by ACT, particularly as CD4+ based T cell therapies are now emerging in the clinic.

cancer biology↗

bioRxiv2023

Clinically relevant orthotopic pancreatic cancer models for adoptive T cell therapy

Pancreatic ductal adenocarcinoma (PDAC) is an aggressive tumor. Prognosis is poor and survival is low in patients diagnosed with this disease; [~]12% at 5 years. Immunotherapy, including adoptive T cell transfer therapy, has not impacted outcomes in PDAC patients, due in part to the hostile tumor microenvironment (TME) which limits T cell trafficking and persistence. We posit that murine models serve as useful tools to study the fate of T cell therapy. Currently, genetically engineered mouse models (GEMM) for PDAC are considered a "gold-standard" as they recapitulate many aspects of human disease. However, these models have limitations, including marked tumor variability across individual mice and cost of colony maintenance. We characterized the immunologic features and trafficking patterns of adoptively transferred T cells in orthotopic PDAC models using two mouse cell lines, KPC-Luc and MT-5, isolated from KPC-GEMM mouse models (KrasLSL-G12D/+p53-/- and KrasLSL-G12D/+p53LSL-R172H/+, respectively). The MT-5 orthotopic model best recapitulates the cellular and stromal features of the TME in the PDAC GEMM. In contrast, far more host immune cells infiltrate KPC-Luc tumors, which have less stroma. Albeit CD4+ T cells were similarly detected in MT-5 tumors compared to KPC-GEMM in mice. Interestingly, we found that CAR T cells redirected to recognize mesothelin on these tumors that signal via CD3{sigma} and 41BB (Meso-41BB{sigma}-CAR T cells) post antigen recognition infiltrated the tumors of mice bearing stroma-devoid KPC-Luc orthotopic tumors, but not MT-5 tumors. Our data establish for the first time a reproducible and realistic clinical system useful for modeling stroma-rich and stroma-devoid PDAC tumors. These models shall serve in-depth study of how to overcome barriers that limit anti-tumor activity of adoptively transferred T cells.

cancer biology↗

bioRxiv2023

The degree of T cell stemness differentially impacts the potency of adoptive cancer immunotherapy in a Lef-1 and Tcf-1 dependent manner.

Generating stem memory T cells (TSCM) is a key goal for improving cancer immunotherapy. Yet, the optimal way to modulate signaling pathways that enrich TSCM properties remains elusive. Here, we discovered that the degree to which the PI3K{delta} pathway is blocked pharmaceutically can generate T cells with differential levels of stemness properties. This observation was based on the progressive enrichment of transcriptional factors of stemness (Tcf-1 and Lef-1). Additional investigation revealed that T cells with high stemness features had enhanced metabolic plasticity, marked by heightened mitochondrial function and glucose uptake. Conversely, T cells with low or medium features of stemness expressed more inhibitory checkpoint receptors (Tim-3, CD39) and were vulnerable to antigen-induced cell death. Only TCR-antigen specific T cells with high stemness persisted following adoptive transfer in vivo and mounted protective immunity to melanoma tumors. Likewise, the strongest level of PI3K{delta} blockade in vitro generated human tumor infiltrating lymphocytes (TILs) and CAR T cells with heightened stemness properties, in turn bolstering their capacity to regress human mesothelioma tumors. We find that the level of stemness T cells possess in vitro differentially impacts their potency upon transfer in three tumor models. Mechanistically, both Lef-1 and Tcf-1 sustain anti-tumor protection by high TSCM, as deletion of either one compromised cellular therapy. Collectively, these findings highlight the therapeutic potential of carefully modulating PI3K{delta} signaling in T cells to confer high stemness and mediate protective responses to solid tumors.

immunology↗

bioRxiv2022

Augmenting TCR signal strength and ICOS costimulation results in metabolically fit and therapeutically potent human CAR Th17 cell therapy

Adoptive cell transfer (ACT) therapy with IL-17 producing human T cells elicits potent antitumor activity in preclinical models. However, further refinement of this novel approach is needed to position it for clinical application. While activation signal strength differentially regulates IL-17 production by human CD4+ T cells, the degree to which TCR and co-stimulation signal strength impacts antitumor Th17 cell immunity remains poorly understood. We discovered that decreasing TCR/co-stimulation signal strength by incremental reduction of CD3/co-stimulation beads in a Th17 culture progressively diminished their effector memory phenotype but enhanced their polyfunctionality. Additional investigation revealed that Th17 cells stimulated with CD3/ICOS beads produced more IL-17A, IFN{gamma}, IL-2 and IL-22 than those stimulated with CD3/CD28 beads, regardless of signal strength. Th17 cells propagated with 30-fold fewer CD3/ICOS beads (weak signal strength, 1 bead per 10 T cells) were less reliant on glucose for growth compared to those stimulated with the standard, strong signal strength (3 beads per T cell). Further metabolomic analysis revealed Th17 cells weakly simulated with CD3/ICOS beads favored the central carbon pathway through increased gluconeogenesis for bioenergetics, marked by abundant intracellular phosphoenoylpyruvate (PEP). Importantly, Th17 cells weakly stimulated with CD3/ICOS beads and redirected with a chimeric antigen receptor (CAR) that recognizes mesothelin were more effective at clearing large human mesothelioma tumors when infused into mice than those manufactured using the standard FDA-approved protocols. Taken together, these data indicate Th17 ACT therapy can be improved by using fewer activation beads during T cell manufacturing, a finding that is both cost effective and directly translatable to patients.

immunology↗

bioRxiv2020

Optical Control of Cytokine Signaling via Bioinspired, Polymer-Induced Latency

Cytokine signaling is challenging to study and therapeutically exploit as the effects of these protein are often pleiotropic. A subset of cytokines can, however, exert signal specificity via association with latency-inducing proteins which cage the cytokine until disrupted by discreet biological stimuli. Inspired by this precision, here we describe a strategy for synthetic induction of cytokine latency via modification with photo-labile polymers that mimic latency while attached, then restore protein activity in response to light, thus controlling the magnitude, duration, and location of cytokine signals. We characterize the high dynamic range of latent cytokine activity modulation and find that polymer-induced latency, alone, can prolong in vivo circulation and bias receptor subunit binding. We further show that protein de-repression can be achieved with near single-cell resolution and demonstrate the feasibility of transcutaneous photoactivation. Future extensions of this approach could enable multicolor, optical reprogramming of cytokine signaling networks and more precise immunotherapies.

bioengineering↗

bioRxiv2020

Dual blockade of IL-6 and CTLA-4 regresses pancreatic tumors in a CD4+ T cell-dependent manner.

Pancreatic ductal adenocarcinoma (PDAC) is exceptionally resistant to immune checkpoint inhibition (ICI). We previously reported that elevated systemic interleukin-6 (IL-6) and increased numbers of T cells positive for circulating cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) correlate with worse overall survival in patients with PDAC. We postulated that combined blockade of IL-6 and CTLA-4 would significantly enhance anti-tumor immune responses to PDAC. Dual blockade of IL-6 and CTLA-4 in immune competent mice bearing subcutaneously injected pancreatic tumors significantly inhibited tumor growth, accompanied by overwhelming T cell infiltration. Therapeutic efficacy was confirmed in an orthotopic murine model of pancreatic cancer and T cell depletion studies unveiled a unique dependence on CD4+ T cells for anti-tumor activity of dual IL-6 and CTLA-4 blockade. In vitro studies utilizing T cells from a TRP-1 transgenic mouse as an antigen-specific model system demonstrate this combination therapy elicits increased IFN-{gamma} production by activated CD4+ T cells. Additionally, IFN-{gamma} stimulation of pancreatic tumor cells in vitro profoundly increased tumor cell production of CXCR3 specific chemokines (CXCL10 and CXCL9). Further studies blocking CXCR3 in the presence of combined IL-6 and CTLA-4 blockade prevented orthotopic tumor regression, demonstrating a dependence on the CXCR3 axis for anti-tumor efficacy. We also found combination therapy increased intratumoral CD4+ T cells and elicited systemic changes in T-helper subsets. These data represent the first report of IL-6 and CTLA-4 blockade as a means to regress pancreatic tumors with defined operative mechanisms of efficacy. Given these results, this therapeutic combination has potential for immediate clinical translation. One Sentence SummaryBlockade of interleukin-6 in pancreatic cancer enhances CTLA-4 immune checkpoint inhibition to regress tumors in a CD4+ T cell and CXCR3-dependent manner.

cancer biology↗