Abstract Current immune checkpoint therapies targeting PD-1 and PD-L1 are ineffective in treating Acute Myeloid Leukemia (AML). Vasoactive Intestinal Peptide (VIP) is an immunosuppressive 28-amino acid neuropeptide overexpressed in nearly 30% AML patients, suggesting that some AML tumors may be using VIP expression as a mechanism to evade immune surveillance. We hypothesize that AML cells secrete VIP which signals through the VIP-R (receptor) on T cells to limit anti-cancer immunity. Therefore, targeting the VIP signaling with a VIP-R antagonist can reverse immunosuppression and potentiate anti-cancer immunity. Using in silico modeling, in vitro screening for potentiation of T cell activation, and proof-of-principle screening in mouse leukemia models, we identified a novel VIP-R antagonist, ANT308. ANT308 shows stronger binding affinity to VIP-R as compared to VIP and potentiates activation of human and mouse T cells with an EC50 in the range of 200-400 nM. Immunofluorescence assays show that ANT308 colocalizes with the VIP receptor, VPAC1. Co-IP and flow cytometry confirmed ANT308 binding to VPAC1. In addition, ANT308 treatment of leukemic mice induced strong T cell-mediated anti-leukemic activity. To improve the pharmacokinetics of ANT308, we have developed a novel fusion peptide combining ANT308 and Fc fragment of IgG4 named CAMV-01. In a Jurkat T-cell activation assay, CAMV-01 has an EC50 of 100-200 nM and reduces T cell exhaustion. Furthermore, CAMV-01 significantly improved survival in multiple murine leukemia models, including those that secrete VIP (C1498) and the VIP negative cell line P815. In vitro stability studies show that CAMV-01 remains stable in plasma for > 4 days, whereas the ANT308 peptide is stable for only 15 minutes. PK studies with intravenous (i.v.) or subcutaneous (s.c.) injection of 15mg/kg CAMV-01 show a half-life of 2 weeks in mice, compared to 15 minutes for the ANT308 peptide. Using Sprague-Dawley rats receiving a single i.v. or s.c injection of 5mg/kg CAMV-01, the Cmax and t1/2 were estimated to be 82 nM and 165 hours (i.v.) respectively and 0.7 nM and 235 hours (s.c.) respectively. In a P815 mouse model, s.c. administration of CAMV-01 induced a stronger tetramer-positive CD8+ T-cell response compared to ANT308. In conclusion, CAMV-01 is a novel, first-in-class immunotherapeutic targeting the VIP immune checkpoint, with a robust pharmacokinetic and pharmacodynamic profile and promising potential for the treatment of AML. Citation Format: Swapnaa Balaji, Antonio Ward, Yuou Wang, Jian-Ming Li, Srijon Sarkar, Sonia Mecorapaj, Tenzin Kalsang, Shuhua Wang, Cynthia Giver, Edmund Waller. A first in class VIP receptor antagonist as an immunotherapeutic for acute myeloid leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 2808.

BACKGROUND:Impaired endogenous vascular regenerative capacity, reflected by reduced circulating progenitor cell (CPC) counts, has been linked to age-related diseases, particularly adverse cardiovascular outcomes. Lower CPC counts have also been associated with accelerated age-related cognitive decline in otherwise healthy individuals, but their relationships with cognitive impairment and neuroimaging markers of vascular brain injury and neurodegeneration remain unclear. We investigated cross-sectional associations between CPC subsets, cognitive performance, and neuroimaging phenotypes, hypothesizing that lower CPC levels would be associated with worse cognition and adverse brain markers. METHODS:In 283 community-dwelling participants (mean age, 65 years; 59% female, 39% Black) enrolled in the Brain Stress, Hypertension, and Aging Research Program, cognitive assessments (including Montreal Cognitive Assessment), brain magnetic resonance imaging-derived white matter hyperintensity volumes, and whole-brain cortical thickness were measured. Flow cytometry was used to enumerate CPCs as CD45med mononuclear cells expressing CD34 with coexpression of either CD133, chemokine CXCR4 (CXC motif receptor 4), or VEGFR2+ (vascular endothelial growth factor receptor-2). Linear regression models were adjusted for demographic and vascular risk factors. RESULTS:In fully adjusted models, lower CD34+/CD133+ CPC levels were associated with worse global cognition (Montreal Cognitive Assessment: β=0.59; P=0.01), lower mean cortical thickness (β=0.01; P=0.01), and greater white matter hyperintensity burden (β=-0.15; P=0.01). Similarly, lower CD34+ and lower CD34+/CXCR4+ CPC levels were significantly associated with greater white matter hyperintensity volume (CD34+: β=-0.27, P<0.01; CD34+/CXCR4+: β=-0.14, P=0.03). CD34+/VEGFR2+ CPC levels were associated with Montreal Cognitive Assessment (β=0.37, P<0.01) and Boston Naming Test performance (β=0.01, P=0.03), but not with neuroimaging phenotypes. CONCLUSIONS:Reduced regenerative capacity was associated with worse global cognitive performance and markers of vascular brain injury, including greater white matter hyperintensity burden and cortical thinning. These findings should be validated in longitudinal studies to clarify temporality and potential causality.

ABSTRACT:Belumosudil was approved by the Food and Drug Administration in the United States for the treatment of relapsed/refractory chronic graft-versus-host disease (cGVHD) based on a randomized phase 2 trial comparing 2 belumosudil doses. The efficacy and safety of belumosudil vs the best available therapy (BAT) has not been studied. Applying rigorous statistical methodology to real-world data, this study estimated the efficacy of belumosudil vs BAT in cGVHD patients whose disease failed to respond to 2 to 5 prior lines of therapy (LOTs). Retrospective data between March 2015 and 2024 were collected across 8 US sites for 196 patients, contributing 113 belumosudil and 245 BAT LOTs. The primary outcome was 6-month overall response rate (ORR), defined as the proportion of complete or partial responses based on 2014 National Institutes of Health consensus criteria, physician assessment, or corticosteroid dose taper of ≥50% without cGVHD progression. Death, relapse, and beginning a new LOT were considered a lack of response. Targeted maximum likelihood estimation (TMLE) was used to estimate the 6-month ORR following belumosudil vs BAT (38.7% vs 26.8%, respectively) or 44.2% improvement with belumosudil (1-sided 95% confidence interval [CI], [4.4 to ∞]; P = .031). TMLE was also used to estimate 1-year failure-free survival when treated with belumosudil (61.2%) or BAT (47.8%), a 13.5% difference (95% CI, 1.5-100; P = .032). Descriptive assessment of safety showed adverse events recorded in 27% of belumosudil and 36% of BAT LOTs. Findings demonstrated that belumosudil improved clinical outcomes compared to BAT in cGVHD patients with 2 to 5 prior LOTs, and safety was consistent with belumosudil's established profile.

Orca-T is a cellular immunotherapy comprising purified donor hematopoietic progenitor stem cells, regulatory T cells (Tregs), and conventional T cells (Tcons). Both Orca-T and post-transplant cyclophosphamide (PTCy) have demonstrated superior graft-versus-host disease (GVHD) control compared to tacrolimus and methotrexate in randomized clinical trials. Orca-T achieves immune tolerance through high-purity regulatory T cells in contrast to broad pharmacological immunosuppression with PTCy regimens. The principal goal of this study was to evaluate the long-term overall survival (OS) of Orca-T compared to PTCy. In this retrospective analysis, long-term survival follow-up was collected from a multicenter Phase 1b study of Orca-T that was initiated in 2019. OS outcomes were evaluated against a cohort of patients receiving PTCy-based GVHD prophylaxis, using a registry dataset obtained from the Center for International Blood and Marrow Transplant Research/NMDP. To ensure comparability, inclusion criteria were aligned with Orca-T Ph3 eligibility, specifically age ≤65 yr, diagnosis of intermediate- or high-risk acute myeloid leukemia or acute lymphoblastic leukemia in complete remission or myelodysplastic syndrome, myeloablative conditioning, and an 8/8 HLA-matched donor. The analysis included 76 Orca-T patients and 360 PTCy patients. Orca-T was associated with significantly higher OS over a 3-yr follow-up period compared to PTCy (hazard ratio [HR] = 0.41; log-rank P = .003). OS at 1, 2, and 3 yr for Orca-T was 96% (95% CI: 88% to 99%), 86% (76% to 92%), and 83% (73% to 90%), respectively. For the PTCy cohort, OS at 1, 2, and 3 yr was 81% (77% to 85%), 72% (67% to 77%), and 66% (60% to 71%), respectively. A propensity score-matched analysis (n = 76/group) confirmed the primary findings (HR = 0.40; log-rank test P = .010), as did a multivariable Cox model adjusted for recipient age, sex, disease, disease risk index, HCT-CI, and donor type (adjusted HR = 0.38; 95% CI 0.21 to 0.71; P = .002). The advantage persisted in sensitivity analyses excluding bone marrow grafts from the comparator and restricting both arms to the overlapping 2019 to 2021 transplant era (HR range across all specifications, 0.36 to 0.48). Three-yr nonrelapse mortality was lower with Orca-T (3.1% versus 10%; Gray's P = .0497), while relapse did not differ significantly; within the PTCy cohort, outcomes did not differ by mycophenolate mofetil use. The OS advantage for Orca-T was consistently observed across age, disease type, and other clinical subgroups. Within the limitations of this retrospective analysis, these results suggest that survival following allogeneic hematopoietic stem cell transplantation may be improved with Orca-T relative to PTCy.

Introduction: The clinical efficacy of chimeric antigen receptor (CAR) T cell therapy remains limited in solid tumors, with immunosuppressive and hypoxic tumor microenvironments. Enhancing CAR T cell function and harnessing the endogenous immune response through novel signaling pathways may help overcome this limitation. We previously showed that targeting the PI3K signaling pathway improves CAR T cell efficacy in hematologic cancers, however, its therapeutic potential in solid tumors has yet to be fully explored. Vasoactive intestinal peptide (VIP) is a neuropeptide which exerts its function via interaction with two receptors, VPAC1 and VPAC2. VIP signaling in T cells is known to suppress T cell proliferation and effector function. Therefore, making this pathway an intriguing target for cancer immunotherapy, and pharmacological inhibition of PI3K and VPAC increased the expansion of TCMin vitro. Thus, dual targeting of PI3K and VIP signaling pathways may be a novel approach to enhance CAR T cell function against solid tumors. CAR T cells generated from VIP-knockout (VIP-KO), and VPAC-knockout (VPAC-KO) mice were expanded using PI3Kδ/γ inhibition,metabolism and in vivo anti-cancer activity were assessed. Methods: Splenocytes isolated from C57BL/6 wild-type (WT), VIP-KO, and VPAC-KO mice were activated ex vivo using concanavalin A and IL-2, followed by retroviral transduction via spinoculation with a CAR-expressing vector. During expansion, cells were cultured in the presence or absence of the PI3Kδ/γ inhibitor, Duvelisib. Transduction efficiency and T cell phenotypes—including central memory, stem-like, and exhausted subsets—were analyzed by spectral flow cytometry using surface markers. Metabolic function was assessed using a Seahorse extracellular flux analyzer. CAR T cell function was assessed using syngeneic pancreatic ductal adenocarcinoma (PDAC) tumor model. Result: VIP-KO and VPAC-KO CAR T cells have higher frequencies of CD4⁺ and CD8⁺ naïve and stem-like memory Tscm subsets compared to WT CAR T cells. Treatment with Duvelisib further enhanced these memory subsets in both VIP-KO and VPAC-KO CAR T cells. Duvelsisib also significantly reduced effector memory Tcells and reduced phenotypically-exhausted CAR T cell populations. Seahorse metabolic analysis revealed that VIP-KO and VIPKO-DUV CAR T cells had increased basal respiration and greater ATP production through mitochondrial oxidative phosphorylation compared to WT CAR T cells. This metabolic reprogramming promoted a more quiescent phenotype in VIP-KO CAR T cells, characterized by enhanced mitochondrial function and greater metabolism up-regulation upon antigen stimulation. Despite enhanced metabolic fitness and increased frequencies of memory T cell subsets in VIPKO-DUV and VPACKO-DUV CAR T cells, in vivo studies using a syngeneic PDAC mouse model demonstrated only modest tumor control and no significant improvement in overall survival of any group compared to WT CAR T cells. Co-culture impedance experiments revealed that the addition of macrophages enhanced the function of DUV-CAR T cells, suggesting that endogenous immune cells within the tumor microenvironment may be critical for full therapeutic efficacy. Conclusion: Dual targeting of PI3Kδ/γ and VIP signaling enhances CAR T cell memory phenotype and mitochondrial metabolic fitness, promoting a less exhausted and more quiescent T cell state. These in vivo results demonstrate that infusion of greater frequencies of memory CAR T cells with greater reliance on oxidative phosphorylation were insufficient to achieve durable tumor control in a solid tumor model, highlighting the critical role of the tumor microenvironment and endogenous immune cells in CAR T cell efficacy. The genetic absence of VIP/VPAC in the context of PI3K inhibitors may have a more profound effect on T cell survial than pharmacological inhibition of VPAC and PI3K. Future studies will include evaluation of CAR T cell infiltration and persistence in solid tumors, as well as profiling of immune cell populations in tumors from responders versus non-responders to better understand the cellular context that supports effective tumor control. These findings demonstrated the need for integrated strategies that consider both intrinsic T cell programming and extrinsic immune context to optimize CAR T cell therapy for solid tumors.

Cardiovascular disease is a leading cause of death in men with localized prostate cancer, and treatment with androgen deprivation therapy (ADT) is known to exacerbate cardiovascular risk; however, the underlying mechanisms driving ADT-associated cardiotoxicity remain unclear. center dot In this case series, we observed significant alterations in circulating adipocytokine levels following 6 months of therapy with the gonadotropin-releasing hormone agonist, leuprolide, in 10 men with localized prostate cancer. center dot The significantly elevated adipocytokine levels following treatment with leuprolide highlight the potential role of ADT-induced systemic inflammation and metabolic dysregulation in the leptin (adipose)-osteopontin (bone) axis in men with localized prostate cancer treated with ADT.

For myelofibrosis (MF) patient's undergoing hematopoietic stem cell transplant (HSCT) engraftment failure is a life-threatening challenge, largely due to primary graft failure (PGF) and/or prolonged poor graft function (PGFxn). Although medical management of MF patients has improved with the introduction of JAK2 inhibitors, HSCT remains the only potentially curative option. In this single center retrospective analysis, we studied a cohort of 50 consecutive patients with primary MF or secondary MF (post ET/PV) who underwent allogeneic HSCT at our institution between 2012 and 2023. PGF was defined according to the CIBMTR definition as failure to achieve a sustained ANC ≥ 500/uL by posttransplant day 30. We defined PGFxn as prolonged platelet transfusion dependence and/or growth factor dependence necessitating a CD-34 selected stem cell boost (SCB). Survival outcomes were censored at 2 yrs post-transplant to avoid time bias. The median age at transplant of 63.5 years (range 35-74 years). The cohort was predominantly male (n=28; 56%), Caucasian (n=41; 82%) and most had JAK2 mutated (n=36, 72%) primary MF (n=47; 94%). The median Dynamic International Prognostic Scoring System (DIPSS) score at transplant was 2 (Intermediate-1 risk). The majority of patients received a JAK2 inhibitor prior to transplant (80%, n=40). The graft source was peripheral blood stem cells for 45 patients (90%), and bone marrow for 5 patients (10%). Reduced intensity conditioning (RIC) regimens were used in 38 cases (76%), and myeloablative regimens in 12 cases (24%). Donors included matched unrelated donors in 28 cases (56%), matched related donors in 15 (30%), mismatched unrelated donors in 6 (12%), and a haploidentical related donor in 1 case (2%). GVHD prophylaxis consisted of MTX/tacrolimus-based regimens (n=35; 70%), post-transplant cyclophosphamide (PTCy)-based prophylaxis (n=9; 18%) or other regimens (n=6; 12%). Eight patients (16%) met criteria for PGF, and 5 additional patients (10%) had PGFxn for which they received a CD-34 selected stem cell boost. Primary Graft Failure Group (n=8): All patients had donor predominant chimerism in blood and/or marrow. Two PGF patients received early SCB (on days 93 and 151 post-transplant) with subsequent improvement in hematopoiesis, and both survived beyond 2 yrs post-transplant. One patient received a second transplant from a different donor and died shortly thereafter without engraftment. Among patients with PGF, 3 patients (37.5%) died before posttransplant day 100 with cause of death being infection. The 2-year OS of the PGF group was 40%. Poor Graft Function Group (n=5): All patients had donor predominant chimerism in blood and/or marrow. PGFxn patients all received a SCB at a median of post-transplant day 99 (day range 34-141), and all had improvement in hematopoietic function. 2 year OS for the PGFxn group was 75%. Predictive Factors for Impaired Graft Function: In univariate analysis, the only significant associations with either PGF or PGFxn were DIPSS score >3 (p=0.026), and PTCy based GVHD prophylaxis [p=0.019; OR = 7.07, 95% CI: 1.57-31.86]. Notably, graft source, HLA matching and conditioning intensity were not predictive of impaired graft function in our series. Conclusion: These findings confirm previous observations that impaired graft function is relatively common after allogeneic transplantation for MF. Our data suggests that PGF and PGFxn may be associated with advanced disease burden reflected by the higher DIPSS scores, as well as the use of PTCy-based GVHD prophylaxis. SCB was used as a rescue intervention in both PGF and PGFxn groups, with generally favorable outcomes. Unfortunately, the feasibility of arranging a prompt SCB can be challenging, particularly with registry donors. Further analyses of larger data sets are warranted to inform selection of the best GVHD prophylaxis strategies as well as optimization of selection criteria for earlier consideration of stem cell boost in this challenging patient population.

e18561 Background: The introduction of JAK2 inhibitors has improved symptom control for patients with myelofibrosis (MF), but allogeneic hematopoietic stem cell transplant (HSCT) remains the only curative option. A significant challenge for patients with MF undergoing HSCT is poor or delayed hematopoietic engraftment (PGF), presumably reflecting the challenge of establishing a stable hematopoietic niche in a fibrotic marrow space. Methods: We retrospectively studied a cohort of 41 consecutive patients with primary MF and post-ET/PV MF who received an HSCT from 2012 to 2023 at our institution. We examined patient and HSCT-specific variables influencing poor engraftment necessitating a CD 34+ stem cell boost (SCB) for poor graft function following transplantation. We used a logistic regression model to analyze various factors, including age, graft source, donor type, match grade, CMV seropositivity, acute and chronic graft vs host disease (GVHD), conditioning regimens, GVHD prophylaxis, DIPSS, HCT-CI score, and mutations in JAK2, MPL, and CALR. Results: Six of the 41 patients (15%) received a SCB because of poor engraftment. This rate is significantly higher compared to the 1.4% of acute myeloid leukemia/acute lymphoblastic leukemia (AML/ALL) patients and 3.8% of myelodysplastic syndrome (MDS) patients who required an SCB at our institution. The most common reason for SCB was for pancytopenia in 5 patients and bicytopenia (anemia and thrombocytopenia) in 1 patient. All six MF patients receiving SCB had originally received a peripheral blood stem cell graft, and all had received a JAK2 inhibitor prior to transplant. The median time from transplant to SCB was 112 days. However, we did not identify any statistically significant predictors of the need for an SCB. Two year overall survival was 83% in the SCB group vs 54% in the non-SCB group (p=NS). Infection was the primary cause of non-relapse mortality among patients receiving SCB. We also found that the SCB group had numerically lower rates of acute GVHD but had slightly higher rates of cGVHD (p=NS). Conclusions: We acknowledge that our analysis is limited by the small sample size. However, despite the challenges of poor graft function in MF patients undergoing HSCT, our findings suggest that favorable outcomes are possible if hematopoietic and immune reconstitution is successfully achieved following SCB.

The immune microenvironment is pivotal in regulating two complementary adverse outcomes of allogeneic hematopoietic stem cell transplantation (allo-HSCT): graft-versus-host disease (GVHD) and tumor progression. While GVHD manifests as immune hyperactivation, causing tissue injury, cancer subverts immune surveillance by immunosuppressive strategies. Although initial studies focused on distinct mechanisms involving T-B interactions in GVHD and T-tumor cell interactions in cancer immunotherapy, it is increasingly clear that dysregulated T cell-macrophage interactions drive immunopathology in GVHD, with comparative insights into cancer immunotherapy. Here, we discuss recent advances and elucidate three core regulatory paradigms governing these immunostimulatory vs immunosuppressive interactions: (1) cytokine networks, (2) immune checkpoint regulation, and (3) metabolic reprogramming. Based on functional pre-clinical studies and clinical evidence, we describe the dynamic, reciprocal immune crosstalk between T cells and macrophages that underlies GVHD after allo-HSCT and the immunosuppressive tumor microenvironment. Finally, we outline emerging therapeutic approaches that target T cell and macrophage interactions to prevent GVHD and overcome an immunologically “cold” microenvironment in the cancer.

Acute myeloid leukemia (AML) is a heterogeneous malignant disorder of myeloid precursor cells, with high relapse rates, particularly in patients who fail to achieve morphological remission after induction therapy. Allogeneic hematopoietic cell transplantation (allo-HCT) can induce durable remissions through the graft-versus-leukemia (GVL) effect, yet current approaches of allo-HSCT often fail, with relapse rates of ∼40% within 6 months post-transplant. Outcomes following allo-HCT are inversely proportional to leukemia burden at the time of transplant. Both morphological relapse (≥5% blasts in the marrow) or minimal residual disease (MRD) positivity predict significantly reduced overall survival rates. Emerging strategies to improve outcomes in patients with high leukemic burden include aggressive bridging therapies (encompassing intensive salvage chemotherapy, hypomethylating agents, targeted inhibitors, and sequential induction-conditioning approaches such as FLAMSA), tailored conditioning regimens, post-transplant maintenance therapy, and innovative graft engineering methods. Graft engineering strategies, such as ORCA-T, which engineers stem cell grafts with a defined ratio of T-regulatory cells to effector T cells, are particularly promising and under evaluation in phase III clinical trials. These approaches aim to improve upon the poor outcomes for patients with persistent/relapsed AML undergoing allo-HCT.

BACKGROUND Allogeneic hematopoietic stem cell transplantation (alloHSCT) remains the only potentially curative option for several high-risk blood cancers and benign blood disorders. Conventional stem cell sources include high doses of T cells that contribute to graft-versus-leukemia (GVL) and graft-versus-infection (GVI) effects, but they also mediate graft-versus-host disease (GVHD). Preventing GVHD following alloHSCT typically requires multi-agent immunosuppression, commonly involving methotrexate or post-transplant cyclophosphamide (PTCy) in combination with a calcineurin inhibitor and other agents. However, pharmacologic immunosuppression impairs immune reconstitution and heightens the risk of organ toxicity, infection, and disease relapse. Orca-Q is a proprietary, investigational precision engineered allogeneic T-cell immunotherapy designed to promote GVL and GVI and minimize GVHD. Here, we report data from Orca-Q patients with 8/8 HLA matched related donors. Patients recieved either single agent tacrolimus (Arm A) or no immune suppression (Arm C) in a multicenter Phase 1 clinical study (NCT03802695). METHODS Adult patients with high-risk hematologic malignancies eligible for myeloablative conditioning alloHSCT with 8/8 HLA-matched donor were first enrolled on the dose expansion Arm A (with single agent tacrolimus for GVHD prevention; n=18). Due to a low incidence of GVHD, Arm C (with no immunosuppression; n=26) was opened and Arm A closed to enrollment of patients with 8/8 HLA matched donors. Orca-Q was centrally manufactured at an Orca Bio GMP facility (Sacramento, CA) from G-CSF mobilized peripheral blood apheresis. Patients were followed for adverse events for one year post-transplant and for survival thereafter. GVHD-free, relapse-free survival (GRFS) was defined as the time from transplant to the first occurrence of any of these events: grade 3-4 acute GVHD (aGVHD), moderate to severe chronic GVHD (cGVHD), disease relapse, or death from any cause. RESULTS Orca-Q was successfully manufactured and administered to all recipients with a vein-to-vein time of less than 72 hours. As of July 14, 2025, a total of 44 patients with hematological malignancies had been treated, including acute myeloid leukemia, acute lymphocytic leukemia, chronic myeloid leukemia, myelofibrosis, myelodysplastic syndrome, and mixed phenotype leukemia. On Arm A, 14 donors were related and 4 unrelated, compared to Arm B where 12 were related and 14 unrelated. Other patient baseline characteristics were as follows: Arm A: Median age 41 years (range 27-55), 56% male and median follow-up 1531 days (range 66-1825), and Arm C: Median age 60 years (range 20-70), 50% male and median follow-up 234 days (range 5-1418). All patients received myeloablative conditioning. Orca-Q was well tolerated. Overall clinical outcomes were:All patients engrafted neutrophils by day +20 (median 12 days)Overall survival and 1 and 2 years: 90% at 1 year and 86% at 2 yearsRelapse-free survival at 1-year: 87%Moderate-to-severe cGVHD-free survival at 1-year: 85%GRFS at 1-year: 78%Non-relapse mortality at 1-year: 3.4%Relapse incidence at 1-year: 12.7%Grade 3-4 aGVHD at Day +180: 7.1% (6% for Arm A and 8% for Arm C)Moderate-to-severe chronic GVHD at 1 year: 5.9% (12% for Arm A and 0% for Arm C) Orca-Q patients treated without pharmacological prophylaxis (Arm C) demonstrated more rapid immune reconstitution and improved control of infections. Specifically, BMT CTN Grade 2+ infections at 1-year: 33% for Arm A and 17% for Arm C;BMT CTN Grade 3 infection at 1-year: 6% for Arm A and 0% for Arm CMedian absolute CD4 / CD8 cell counts per µL at Day +28: 97 / 94 for Arm A and 102 / 245 for Arm C CONCLUSIONS Orca-Q demonstrates promising efficacy and safety as a precision-engineered T-cell immunotherapy for alloHSCT in patients with high-risk hematologic malignancies. The therapy achieved rapid neutrophil recovery, while maintaining low rates of severe acute GVHD, relapse, and non-relapse mortality across both treatment arms. The low infection rates observed suggest that the cellular composition of Orca-Q can control GVHD and accelerate immune reconstitution even without pharmacological immunosuppression. The Phase 1 study continues to enroll patients to further confirm these results.

Cancer is one of the leading causes of mortality globally. Current treatments, such as radiotherapy and chemotherapy, have numerous side effects. To create cancer therapies, various studies have investigated the anticancer effects of plants, but present literature fails to mention the potential anticancer effects of Uvularia perfoliata. Despite being used by Native American tribes in salves, no scientific investigation has studied the potential medicinal properties of U. perfoliata. The objective of this study was to identify whether U. perfoliata could be a viable option for future drug discovery research. We hypothesized that if we applied an ethanol extract of U. perfoliata to uveal melanoma cells, 70% or more of the cells would die. Although our findings did not match our a priori value for the magnitude of cancer cell killing, the 0.1% concentration U. perfoliata did significantly decrease melanoma cell viability (p < 0.05). Future testing must be conducted with non-malignant cell lines and higher concentration extracts to understand if U. perfoliata possesses anticancer properties. These additional experiments must be conducted because our extracts were significantly more dilute than similar studies. In addition, the cytotoxic properties of U. perfoliata may also apply to non-malignant cultures, so future experiments should include non-malignant cell lines as a control. However, this is the first study to highlight the potential anticancer effects of U. perfoliata.

ABSTRACT:Tisagenlecleucel is a CD19-directed autologous chimeric antigen receptor (CAR) T-cell therapy. Quantitative polymerase chain reaction assays are highly sensitive in defining in vivo kinetics by measuring CAR transgene in peripheral blood. This study aimed to identify clinically meaningful CAR T-cell blood levels that correlated with response/relapse. In pediatric/young adult patients with relapsed/refractory (R/R) B-cell acute lymphoblastic leukemia (B-ALL), maximum CAR T-cell blood levels were higher in patients with ongoing complete remission and in patients with CD19- relapse relative to those with CD19+ relapse. In adult patients with R/R diffuse large B-cell lymphoma (DLBCL), no apparent association between in vivo kinetics and response was noted, with a wide range of transgene levels at relapse. In B-ALL, patients with B-cell aplasia sustained >6 months had higher CAR T-cell expansion relative to those with early B-cell recovery (BCR) (<6 months after infusion); however, a definitive cutoff for BCR-associated expansion level could not be identified. In most patients with B-ALL, BCR >6 months maintained favorable responses. However, early BCR could not be confirmed as a potential indicator of relapse due to high censoring from transplant, following presumed risk of relapse. However, allografting in these patients may potentially mitigate the poor prognosis related to early BCR. In DLBCL, BCR was not associated with relapse. These findings suggest that blood CAR transgene levels may be associated with long-term responses; however, they lack robust predictive potential for relapses. As reported earlier, next-generation sequencing for minimal residual disease appears to be a reliable biomarker predictive of relapse for B-ALL.

Achieving an adequate dose of genetically modified hematopoietic stem cells (HSCs) for gene therapy in patients with sickle cell disease (SCD) remains a challenge due to limitations related to stem cell mobilization using plerixafor alone, reduced apheresis collection efficiency, and losses during ex vivo cell manipulation. To date, 39 SCD patients were enrolled in NHLBI-funded Phase I (NCT03282656; PMC7962145) and NHLBI/CIRM-funded multi-site Phase II (NCT05353647) gene therapy trials to receive autologous CD34+ HSCs transduced with a lentiviral vector encoding a short hairpin RNA embedded in a microRNA (shmiR) targeting BCL11A. To collect HSCs as the starting material for drug product manufacture (minimum 4×10⁶ cells/kg), apheresis (minimum 4 blood volumes or up to 8 hours) was performed within 3 hours of daily plerixafor for 2 consecutive days. A third day of collection was used to generate a back-up product as needed. Mean mobilized, pre-apheresis peripheral blood (PB) CD34+ count was 40 cells/μL (range 7–126). Among 35 evaluable patients (4 excluded: 1 pending release, 3 withdrew), 74% (26/35) reached target CD34+ collection in a single mobilization cycle (mean 2.6 procedures, range 2–6). Manufacturing was achieved with a mean of 11.92×10⁶ CD34+ cells/kg collected resulting in a drug product of 7.04×10⁶ CD34+ cells/kg and a net cell recovery from apheresis to drug product of 62% (range 38–91%). Drug products were successfully generated for all patients with a mean product vector copy number of 4.04 copies/cell (range 1-7). The time interval from first collection cycle to completion of product testing was a median of 39 days (mean 56) for all patients, and a median of 37 days (mean 38) for those collected in 1 cycle. Preparative transfusions were given before collections to bridge patients after stopping hydroxyurea or to mitigate stress erythropoiesis. We examined the impact of preparative transfusion on mobilization and apheresis efficiency in 16 evaluable patients at a single site. All underwent ≥ 3 months of preparative transfusion with a HbS target of ≤30% before mobilization using simple transfusion or automated red cell exchange. The mean HbS% at mobilization was 10.8% (range 2.9–21.5%). Overall, the preparative transfusion regimen reduced PB reticulocyte percentage by 50% to a mean of 7.2% (range 3.7–12.3), indicating decreased stress erythropoiesis. Resting PB CD34+ counts before and after 3-months of transfusion were 10.8 cells/μL (range 2–21) and 6.2 cells/μL (range 1–13), respectively. A total of 39 mobilization/collection procedures were performed in 16 patients with a mean post-plerixafor CD34+ count of 43 cells/μL (range 13–85) with collections performed between the two darkest color preferences of the manufacturer. Cell collection preference was monitored using real-time intraprocedural CD34+ cell sampling. A mean of 4.3 total blood volumes was processed (range 2.5–6.4), with mean collection efficiency (CE) (CD34+ cells collected ÷ [pre-apheresis CD34+ count × total blood processed]) of 48% (range 8.2–107.1). The mean reticulocyte percent in patients with CE <30% (11.4%, range 2.5 –18.9) was significantly higher than in those with CE >30% (6.2%, range 1.8–11.9, p=0.0026). Two patients had a history of delayed hemolytic transfusion reactions and/or multiple RBC alloantibodies before study entry that limited preparative transfusions to reach a HbS of 30% for 3 months. They were successfully collected, and products were generated after simple transfusions with a single limited-volume red cell exchange immediately prior to collection. The success of this study was driven by optimizing apheresis strategies, including suppression of stress erythropoiesis through transfusion, real-time instrument adjustments, and efficient manufacturing that allowed a collection target of nearly half the CD34+ cells recommended in the FDA-approved gene product. Strikingly, the highly efficient ex vivo manipulation platform described here enables successful mobilization and manufacturing of gene therapy products for SCD in several months, easing the path to autologous gene therapy for SCD.

Pancreatic ductal adenocarcinoma (PDAC) remains resistant to immunotherapy due to a highly immunosuppressive tumor microenvironment. Lahusen et al identify transglutaminase 2 (TGM2) as a critical tumor cell-intrinsic regulator of immune suppression in PDAC. TGM2 enhances the secretion of suppressive cytokines such as granulocyte colony-stimulating factor (G-CSF) and granulocyte-macrophage-CSF (GM-CSF) via a microtubule-dependent pathway that secondarily impairs T-cell activation and cytotoxicity. Targeting TGM2, either genetically or with microtubule-disrupting agents, restores T cell function and promotes tumor control in preclinical PDAC models. These findings are discussed in the context of our recent work on vasoactive intestinal peptide (VIP) signaling in PDAC, which independently drives immune evasion through VIP receptor (VPAC)-mediated induction of TGF-β1 secretion from tumor cells, upregulation of programmed death-ligand 1 on myeloid cells, and inhibition of effector T cells. Here, we have explored the mechanistic intersections between TGM2 and VIP signaling—including cytokine secretion, vesicle trafficking, and immune checkpoint regulation—and propose combination strategies that integrate TGM2 or VIP inhibition with checkpoint blockade. Together, these insights suggest a new framework for overcoming immune resistance in PDAC by targeting cancer cell-intrinsic secretory programs and offer a rationale for biomarker-guided, multimodal immunotherapeutic approaches.

Despite significant advancements in understanding the pancreatic ductal adenocarcinoma (PDAC) tumor microenvironment (TME) through single-cell RNA sequencing (scRNAseq) and the development of novel therapeutic combinations, PDAC remains one of the deadliest cancers, with a five-year survival rate of ∼3%. Notably, survival outcomes are particularly worse in males compared to females. The limited efficacy of immune checkpoint inhibitors (ICI), such as anti-CTLA-4 and anti-PD-1/PD-L1 therapies, in PDAC has been attributed to a profoundly immunosuppressive TME driven by M2-like tumor-associated macrophages (TAM). Our translational studies demonstrated that dual blockade of PD-1 and VPAC stimulates CD8+ T cell activity and eradicates tumors in ∼50% of mice across various PDAC models. We hypothesized that a combinatorial ICI strategy is targeting TAM-driven immunosuppression in the TME and could enhance T cell-mediated anti-tumor immunity in PDAC. To test this hypothesis, we performed in vivo tumor growth and survival experiments in syngeneic PDAC models. We also conducted RNA sequencing (RNAseq) of bone marrow-derived macrophages (BMDM) polarized to M1/M2 phenotypes to investigate immune checkpoint expressions, supplemented by scRNAseq analysis of TAM from PDAC patients using public datasets. Using VIP-knockout mice, we explored the mechanisms underlying VPAC blockade-mediated tumor control. As anticipated, syngeneic PDAC tumors (e.g., KPC-luc and MT5) grew significantly slower in VIP-knockout mice compared to wildtype controls. Notably, the anti-tumor response was markedly superior in female VIP-knockout mice compared to males. To dissect the cellular basis of this sexual dimorphism, we depleted specific immune subsets, including T cells and TAM. Phagocytic TAM depletion accelerated tumor growth in females but had no effect on males, while the depletion of other immune subsets increased tumor growth in both sexes. In vitro, androgen receptor (AR) inhibition (mimicking the female phenotype) reversed the suppressive state of BMDM, marked by reduced Arg1, CD206, and PD-L1 expression. In vivo, AR inhibition or PD-L1 loss significantly enhanced the anti-tumor efficacy of VIP blockade in male mice. Mechanistically, RNAseq and Western blotting revealed increased VPAC1 expression on IC21 and BMDM cells following AR inhibition, reflecting the female-like phenotype. These findings suggest that androgen and VIP pathways mediate sexual dimorphism in TAM phagocytosis, which limits ICI efficacy in PDAC. VPAC1 and PD-L1 are identified as potential sex-specific phagocytosis checkpoints, providing promising therapeutic targets to overcome sex-based differences in PDAC immunotherapy outcomes. Zihan Chen, Tenzin Passang, Fanyuan Zeng, Yuou Wang, JianMing Li, Cynthia Giver, Kiranj Chaudagar, Edmund Waller. Sexual dimorphism in macrophage phagocytosis limits immune checkpoint inhibitor efficacy in pancreatic ductal adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 1211.

Background: Impaired endogenous vascular regenerative capacity, reflected by reduced levels of circulating progenitor cells (CPC), has been linked to age-related diseases, especially adverse cardiovascular outcomes. We have previously reported that CPC are associated with cognitive aging, but their impact on cognitive impairment and related brain phenotypes is unclear. Here we report the impact of CPC on cognitive and neuroimaging markers of cognitive impairment. Methods: We analyzed data from 283 community-dwelling participants (59% female, 39% Black) enrolled in the Brain Stress, Hemodynamics and Risk Prediction (B-SHARP) program. Participants underwent (a) cognitive assessments (including Montreal Cognitive Assessment [MoCA]); (b) brain magnetic resonance imaging (MRI) to derive white matter hyperintensity (WMH) volumes, whole-brain cortical thickness and hippocampal volumes; and (c) flow cytometry for enumerating CPCs as CD45med mononuclear cells expressing CD34 with co-expression with either CD133, chemokine CXC motif receptor 4 (CXCR4), or vascular endothelial growth factor receptor-2 (VEGF2R). Linear regression models were adjusted for demographic and vascular risk factors. Results: In fully adjusted models, lower levels of CD34+/CD133+ CPCs were associated with worse global cognition (MoCA: β = 0.59, p = 0.01), reduced cortical thickness (β = 0.01, p = 0.01), and greater WMH burden (β = -0.15, p = 0.01). Lower levels of CD34+ and CD34+/CXCR4+ CPCs were significantly associated with greater WMH volume burden (CD34+: β = -0.27, p < 0.01; CD34+/CXCR4+: β = -0.14, p = 0.03). Higher CD34+/VEGFR2+ CPCs were associated with higher MoCA scores ( β = 0.37, p < 0.01) and language performance on the Boston Naming Test( β = 0.01, p = 0.03) but not with brain phenotypes. Conclusions: Reduced regenerative capacity is associated with worse cognitive performance on global tests and with vascular brain injury, including WMH volume and cortical thinning. If validated in future studies, these findings may highlight regenerative capacity as a promising therapeutic target for mitigating cognitive decline. ### Competing Interest Statement The authors have declared no competing interest. ### Funding Statement This work was supported by the National Institute on Aging through grants AG051633 and AG057470 (VAScular ContribUtors to prodromaL AlzheimeR?s disease [VASCULAR] study) and R01AG042127 (CAndesartan vs LIsinopril effects on the BRain and Endothelial function in eXecutive MCI [CALIBREX] study). No additional external funding was received for the present analyses. The authors and their institutions did not receive payment or services from any third party, including for study design, data collection, management, analysis, interpretation, manuscript preparation, or statistical support. ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: The VASCULAR (VAScular ContribUtors to prodromaL AlzheimeR's disease) and CALIBREX (CAndesartan vs LIsinopril effects on the BRain and Endothelial function in eXecutive MCI) studies were approved by the Emory University Institutional Review Board (Atlanta, GA). All participants provided written informed consent prior to enrollment. I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable. Yes The data that support the findings of this study are not publicly available due to privacy and institutional restrictions. Deidentified participant data may be available from the corresponding author upon reasonable request and with appropriate institutional approvals.