Exercise-induced ß2-adrenergic Receptor Activation Enhances the Antileukemic Activity of Expanded γδ T-Cells via DNAM-1 Upregulation and PVR/Nectin-2 Recognition.

Exercise mobilizes cytotoxic lymphocytes to blood which may allow superior cell products to be harvested and manufactured for cancer therapy. Gamma-Delta (γδ) T-cells have shown promise for treating solid tumors, but there is a need to increase their potency against hematologic malignancies. Here, we show that human γδ T-cells mobilized to blood in response to just 20 minutes of graded exercise have surface phenotypes and transcriptomic profiles associated with cytotoxicity, adhesion, migration, and cytokine signaling. Following 14 days ex vivo expansion with zoledronic acid and IL2, exercise mobilized γδ T-cells had surface phenotypes and transcriptomic profiles associated with enhanced effector functions and demonstrated superior cytotoxic activity against multiple hematologic tumors in vitro and in vivo in leukemia-bearing xenogeneic mice. Infusing humans with the ß1+ß2-agonist isoproterenol and administering ß1 or ß1+ß2 antagonists prior to exercise revealed these effects to be ß2-adrenergic receptor (AR) dependent. Antibody blocking of DNAM-1 on expanded γδ T-cells, as well as the DNAM-1 ligands PVR and Nectin-2 on leukemic targets, abolished the enhanced antileukemic effects of exercise. These findings provide a mechanistic link between exercise, ß2-AR activation, and the manufacture of superior γδ T-cell products for adoptive cell therapy against hematologic malignancies. SIGNIFICANCE: Exercise mobilizes effector γδ T-cells to blood via ß2-adrenergic signaling which allows for generation of a potent expanded γδ T-cell product that is highly cytotoxic against hematologic malignancies.

Comparable Efficacy of Oral Bendamustine versus Intravenous Administration in Treating Hematologic Malignancies.

Purpose: The purpose of this study was to analyze potential differences in antitumor efficacy and pharmacokinetics between intravenous (IV) bendamustine (BEN) and a novel orally administered bendamustine agent (PO) that is utilizing the beneficial properties of superstaturated solid dispersions formulated in nanoparticles. Methods: Pharmacokinetics of IV versus PO BEN were determined by analysis of plasma samples collected from NSG mice treated with either IV or PO BEN. Plasma samples were analyzed using liquid chromatography-mass spectrometry (LC/MS/MS) following a liquid-liquid extraction to determine peak BEN concentration (Cmax), area under the concentration-time curve (AUC) and the half-life (t1/2) in-vivo. in-vitro cytotoxicity of BEN against human non-Hodgkin's Burkitt's Lymphoma (Raji), multiple myeloma (MM.1s), and B-cell acute lymphoblastic leukemia (RS4;11) cell lines was determined over time using MTS assays. Luciferase-tagged versions of the aforementioned cell lines were used to determine in-vivo BEN cytotoxicity of IV versus PO BEN at two different doses. Results: Bendamustine at a high dose in-vitro causes cell death. There was no significant difference in antitumor efficacy between IV and novel PO BEN at a physiologically relevant concentration in all three xenograft models. In-vivo pharmacokinetics showed the oral bioavailability of BEN in mice to be 51.4%. Conclusions: The novel oral BEN agent tested exhibits good oral bioavailability and systemic exposure for in-vivo antitumor efficacy comparable to IV BEN. An oral BEN formulation offers exciting clinical potential as an additional method of administration for bendamustine and warrants further evaluation in clinical studies.

COVID-19 vaccination produces exercise-responsive SARS-CoV-2 specific T-cells regardless of infection history.

BACKGROUND: The mobilization and redistribution of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) specific T-cells and neutralizing antibodies (nAbs) during exercise is purported to increase immune surveillance and protect against severe coronavirus disease 2019 (COVID-19). We sought to determine if COVID-19 vaccination would elicit exercise-responsive SARS-CoV-2 T-cells and transiently alter nAb titers. METHODS: Eighteen healthy participants completed a 20-min bout of graded cycling exercise before and/or after receiving a COVID-19 vaccine. All major leukocyte subtypes were enumerated before, during, and after exercise by flow cytometry, and immune responses to SARS-CoV-2 were determined using whole blood peptide stimulation assays, T-cell receptor (TCR)-ß sequencing, and SARS-CoV-2 nAb serology. RESULTS: COVID-19 vaccination had no effect on the mobilization or egress of major leukocyte subsets in response to intensity-controlled graded exercise. However, non-infected participants had a significantly reduced mobilization of CD4+ and CD8+ naive T-cells, as well as CD4+ central memory T-cells, after vaccination (synthetic immunity group); this was not seen after vaccination in those with prior SARS-CoV-2 infection (hybrid immunity group). Acute exercise after vaccination robustly mobilized SARS-CoV-2 specific T-cells to blood in an intensity-dependent manner. Both groups mobilized T-cells that reacted to spike protein; however, only the hybrid immunity group mobilized T-cells that reacted to membrane and nucleocapsid antigens. nAbs increased significantly during exercise only in the hybrid immunity group. CONCLUSION: These data indicate that acute exercise mobilizes SARS-CoV-2 specific T-cells that recognize spike protein and increases the redistribution of nAbs in individuals with hybrid immunity.

Has the shortage of fludarabine altered the current paradigm of lymphodepletion in favor of bendamustine?

The most common lymphodepletion regimen used prior to infusion of chimeric antigen receptor-T cells (CAR-T) is cyclophosphamide (CY) in combination with fludarabine (Flu) (CY-FLU). While cyclophosphamide (CY) possesses lymphotoxic effects, it concurrently preserves regulatory T cell activity, potentially affecting the efficacy of CAR-T cells. Moreover, the use of fludarabine (FLU) has been linked to neurotoxicity, which could complicate the early detection of immune effector cell-associated neurotoxicity syndrome (ICANS) observed in CAR-T cell therapy. Given the ongoing shortage of FLU, alternative lymphodepleting agents have become necessary. To date, only a limited number of studies have directly compared different lymphodepleting regimens, and most of these comparisons have been retrospective in nature. Herein, we review the current literature on lymphodepletion preceding CAR-T cell therapies for lymphoid hematologic malignancies, with a specific focus on the use of bendamustine (BEN). Recent evidence suggests that administering BEN before CAR-T cell infusion yields comparable efficacy, possibly with a more favorable toxicity profile when compared to CY-FLU. This warrants further investigation through randomized prospective studies.

Mediport use as an acceptable standard for CAR T cell infusion.

Introduction: Mediport use as a clinical option for the administration of chimeric antigen receptor T cell (CAR T cell) therapy in patients with B-cell malignancies has yet to be standardized. Concern for mediport dislodgement, cell infiltration, and ineffective therapy delivery to systemic circulation has resulted in variable practice with intravenous administration of CAR T cell therapy. With CAR T cell commercialization, it is important to establish practice standards for CAR T cell delivery. We conducted a study to establish usage patterns of mediports in the clinical setting and provide a standard of care recommendation for mediport use as an acceptable form of access for CAR T cell infusions. Methods: In this retrospective cohort study, data on mediport use and infiltration rate was collected from a survey across 34 medical centers in the Pediatric Real-World CAR Consortium, capturing 504 CAR T cell infusion routes across 489 patients. Data represents the largest, and to our knowledge sole, report on clinical CAR T cell infusion practice patterns since FDA approval and CAR T cell commercialization in 2017. Results: Across 34 sites, all reported tunneled central venous catheters, including Broviac® and Hickman® catheters, as accepted standard venous options for CAR T cell infusion. Use of mediports as a standard clinical practice was reported in 29 of 34 sites (85%). Of 489 evaluable patients with reported route of CAR T cell infusion, 184 patients were infused using mediports, with no reported incidences of CAR T cell infiltration. Discussion/Conclusion: Based on current clinical practice, mediports are a commonly utilized form of access for CAR T cell therapy administration. These findings support the safe practice of mediport usage as an accepted standard line option for CAR T cell infusion.

Social isolation and social connectedness among young adult cancer survivors: A systematic review.

BACKGROUND: Social isolation and connectedness are social determinants of health that have demonstrated effects on cancer-related outcomes. These constructs have been systematically evaluated among pediatric and older adult cancer populations. In this review, the authors evaluated the prevalence, correlates, and psychosocial implications of social isolation and connectedness among young adult (YA) cancer survivors aged 18-39 years. METHODS: Peer-reviewed articles published in English before June 2021 were identified from database searches and included articles' reference lists according to PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-Analyses) guidelines. Included articles described studies that assessed social isolation and/or connectedness among YA cancer survivors. RESULTS: In total, 5094 unique records were identified; 4143 were excluded after title/abstract screening, and 907 were excluded after full-text review. Forty-four articles were included. Few studies used validated measures or directly assessed social isolation or connectedness. Social isolation was similarly prevalent among YAs and older cancer survivors and noncancer populations. Demographic, clinical, and behavioral risk and protective factors for social isolation were identified. Social isolation was related to worse psychological well-being, whereas social connectedness was often, but not always, related to better psychological well-being. CONCLUSIONS: This growing literature underscores the relevance of social isolation and connectedness as important health determinants among YA cancer survivors. The identified risk and protective factors can identify YAs who especially may benefit from screening for social isolation. Future studies are needed that directly, reliably, and validly evaluate social isolation and connectedness to inform the development of interventions to decrease isolation and increase connectedness.

Human lymphocytes mobilized with exercise have an anti-tumor transcriptomic profile and exert enhanced graft-versus-leukemia effects in xenogeneic mice.

Background: Every bout of exercise mobilizes and redistributes large numbers of effector lymphocytes with a cytotoxic and tissue migration phenotype. The frequent redistribution of these cells is purported to increase immune surveillance and play a mechanistic role in reducing cancer risk and slowing tumor progression in physically active cancer survivors. Our aim was to provide the first detailed single cell transcriptomic analysis of exercise-mobilized lymphocytes and test their effectiveness as a donor lymphocyte infusion (DLI) in xenogeneic mice engrafted with human leukemia. Methods: Peripheral blood mononuclear cells (PBMCs) were collected from healthy volunteers at rest and at the end of an acute bout of cycling exercise. Flow cytometry and single-cell RNA sequencing was performed to identify phenotypic and transcriptomic differences between resting and exercise-mobilized cells using a targeted gene expression panel curated for human immunology. PBMCs were injected into the tail vein of xenogeneic NSG-IL-15 mice and subsequently challenged with a luciferase tagged chronic myelogenous leukemia cell line (K562). Tumor growth (bioluminescence) and xenogeneic graft-versus-host disease (GvHD) were monitored bi-weekly for 40-days. Results: Exercise preferentially mobilized NK-cell, CD8+ T-cell and monocyte subtypes with a differentiated and effector phenotype, without significantly mobilizing CD4+ regulatory T-cells. Mobilized effector lymphocytes, particularly effector-memory CD8+ T-cells and NK-cells, displayed differentially expressed genes and enriched gene sets associated with anti-tumor activity, including cytotoxicity, migration/chemotaxis, antigen binding, cytokine responsiveness and alloreactivity (e.g. graft-versus-host/leukemia). Mice receiving exercise-mobilized PBMCs had lower tumor burden and higher overall survival (4.14E+08 photons/s and 47%, respectively) at day 40 compared to mice receiving resting PBMCs (12.1E+08 photons/s and 22%, respectively) from the same donors (p<0.05). Human immune cell engraftment was similar for resting and exercise-mobilized DLI. However, when compared to non-tumor bearing mice, K562 increased the expansion of NK-cell and CD3+/CD4-/CD8- T-cells in mice receiving exercise-mobilized but not resting lymphocytes, 1-2 weeks after DLI. No differences in GvHD or GvHD-free survival was observed between groups either with or without K562 challenge. Conclusion: Exercise in humans mobilizes effector lymphocytes with an anti-tumor transcriptomic profile and their use as DLI extends survival and enhances the graft-versus-leukemia (GvL) effect without exacerbating GvHD in human leukemia bearing xenogeneic mice. Exercise may serve as an effective and economical adjuvant to increase the GvL effects of allogeneic cell therapies without intensifying GvHD.

Multiagent Intratumoral Immunotherapy Can Be Effective in A20 Lymphoma Clearance and Generation of Systemic T Cell Immunity.

The use of immunotherapies has shown promise against selective human cancers. Identifying novel combinations of innate and adaptive immune cell-activating agents that can work synergistically to suppress tumor growth and provide additional protection against resistance or recurrence is critical. The A20 murine lymphoma model was used to evaluate the effect of various combination immunotherapies administered intratumorally. We show that single-modality treatment with Poly(I:C) or GM-CSF-secreting allogeneic cells only modestly controls tumor growth, whereas when given together there is an improved benefit, with 50% of animals clearing tumors and surviving long-term. Neither heat nor irradiation of GM-CSF-secreting cells enhanced the response over use of live cells. The use of a TIM-3 inhibitory antibody and an OX40 agonist in combination with Poly(I:C) allowed for improved tumor control, with 90% of animals clearing tumors with or without a combination of GM-CSF-secreting cells. Across all treatment groups, mice rejecting their primary A20 tumors were immune to subsequent challenge with A20, and this longstanding immunity was T-cell dependent. The results herein support the use of combinations of innate and adaptive immune activating agents for immunotherapy against lymphoma and should be investigated in other cancer types.

Exercise mobilizes diverse antigen specific T-cells and elevates neutralizing antibodies in humans with natural immunity to SARS CoV-2.

Epidemiological data suggest that physical activity protects against severe COVID-19 and improves clinical outcomes, but how exercise augments the SARS-CoV-2 viral immune response has yet to be elucidated. Here we determine the antigen-specific CD4 and CD8 T-cell and humoral immunity to exercise in non-vaccinated individuals with natural immunity to SARS CoV-2, using whole-blood SARS-CoV-2 peptide stimulation assays, IFN-γ ELISPOT assays, 8-color flow cytometry, deep T-cell receptor (TCR) ß sequencing, and anti-RBD-1 neutralizing antibody serology. We found that acute exercise reliably mobilized (∼2.5-fold increase) highly functional SARS-CoV-2-specific T-cells to the blood compartment in those with natural immunity to the virus. The mobilized cells reacted with spike protein (including alpha (α) and delta (δ)-variants), membrane, and nucleocapsid peptides in those previously infected but not in controls. Both groups reliably mobilized T-cells reacting with Epstein-Barr viral peptides. Exercise mobilized SARS-CoV-2 specific T-cells maintained broad TCR-ß diversity with no impact on CDR3 length or V and J family gene usage. Exercise predominantly mobilized MHC I restricted (i.e. CD8+) SARS-CoV-2 specific T-cells that recognized ORF1ab, surface, ORF7b, nucleocapsid, and membrane proteins. SARS-CoV-2 neutralizing antibodies were transiently elevated ∼1.5-fold during exercise after infection. In conclusion, we provide novel data on a potential mechanism by which exercise could increase SARS-CoV-2 immunosurveillance via the mobilization and redistribution of antigen-specific CD8 T-cells and neutralizing antibodies. Further research is needed to define the tissue specific disease protective effects of exercise as SARS-CoV-2 continues to evolve, as well as the impact of COVID-19 vaccination on this response.

Clonal Kinetics and Single-Cell Transcriptional Profiles of T Cells Mobilized to Blood by Acute Exercise.

PURPOSE: Acute exercise redistributes large numbers of memory T cells, which may contribute to enhanced immune surveillance in regular exercisers. It is not known, however, if acute exercise promotes a broad or oligoclonal T-cell receptor (TCR) repertoire or evokes transcriptomic changes in "exercise-responsive" T-cell clones. METHODS: Healthy volunteers completed a graded bout of cycling exercise up to 80% V̇O 2max . DNA was extracted from peripheral blood mononuclear cells collected at rest, during exercise (EX), and 1 h after (+1H) exercise, and processed for deep TCR-ß chain sequencing and tandem single-cell RNA sequencing. RESULTS: The number of unique clones and unique rearrangements was decreased at EX compared with rest ( P < 0.01) and +1H ( P < 0.01). Productive clonality was increased compared with rest ( P < 0.05) and +1H ( P < 0.05), whereas Shannon's Index was decreased compared with rest ( P < 0.05) and +1H ( P < 0.05). The top 10 rearrangements in the repertoire were increased at EX compared with rest ( P < 0.05) and +1H ( P < 0.05). Cross-referencing TCR-ß sequences with a public database (VDJdb) revealed that exercise increased the number of clones specific for the most prevalent motifs, including Epstein-Barr virus, cytomegalovirus, and influenza A. We identified 633 unique exercise-responsive T-cell clones that were mobilized and/or egressed in response to exercise. Among these clones, there was an upregulation in genes related to cell death, cytotoxicity, and activation ( P < 0.05). CONCLUSIONS: Acute exercise promotes an oligoclonal T-cell repertoire by preferentially mobilizing the most dominant clones, several of which are specific to known viral antigens and display differentially expressed genes indicative of cytotoxicity, activation, and apoptosis.

Busulfan, fludarabine, and melphalan are effective conditioning for pediatric and young adult patients with myeloid malignancies underdoing matched sibling or alternative donor transplantation.

BACKGROUND: Allogeneic hematopoietic cell transplantation (allo-HCT) remains a curative option for patients with high-risk myeloid malignancies. PROCEDURE: We present our 10-year experience (October 2012 to October 2021) of consecutive allo-HCT in patients with myeloid malignancies treated on the pediatric HCT service and conditioned with myeloablative targeted dose-busulfan (BU), fludarabine (FLU), and melphalan (MEL). Twenty-three children, adolescents, and young adult patients (CAYA) (median age 15.4 years) with acute myeloid leukemia (AML, n = 17), myelodysplastic syndrome (MDS, n = 4), or chronic myeloid leukemia (CML, n = 2) underwent allo-HCT post-BU-FLU-MEL. Four patients had treatment-related AML/MDS. Donor/stem cell source was matched sibling donor (MSD) PBSC (n = 7), matched unrelated donor (MUD) PBSC (n = 2), umbilical cord blood (UCB) (n = 3), or haploidentical-BMT (n = 11). Risk stratification was low (n = 2), intermediate (n = 15), high (n = 3), and very high risk (n = 1). The two patients with CML had failed tyrosine kinase inhibitor therapies. RESULTS: With a median follow-up of 41.6 months, the relapse rate is only 4.5% with an overall survival (OS) 100%, progression-free survival (PFS) 95.5%, and graft-versus-host-free-relapse-free survival (GRFS) 67.8%. The donor source and the acute graft-versus-host disease (GvHD) prophylaxis regimen significantly impacted grade II-IV aGvHD 66.7% versus 19.2% (p = .039) and chronic graft-versus-host-disease (cGvHD) 66.7% versus 0% (p = .002) in the patients receiving MSD or MUD PBSC compared to haplo-BMT, respectively, resulting in improved GRFS in haplo-BMT, 83.3% compared to 40% matched donor peripheral blood stem cell transplant (PBSCT) (p = .025). CONCLUSIONS: Our results demonstrate that BU-FLU-MEL is efficacious conditioning for disease control in young patients with myeloid malignancies undergoing MSD or alternative donor allo-HCT, but in the setting of PBSC grafts with cyclosporine A-methotrexate (CSA-MTX) GvHD prophylaxis, it results in an unacceptably high incidence of GvHD.

Refractory Pure Red Blood Cell Aplasia Secondary to Major ABO-Incompatible Allogeneic Stem Cell Transplantation Successfully Treated With Daratumumab.

Pure red cell aplasia (PRCA) is a rare hematologic phenomenon that is usually associated with inherited genetic mutations such as in Diamond-Blackfan anemia. However, due to the emergence of allogenic stem cell transplantation in the treatment of various malignant and non-malignant disorders, the incidence of PRCA has increased. PRCA following hematopoietic stem cell transplant (HSCT) is more commonly seen in the setting of a major ABO-incompatible transplant. Treatment of allo-HSCT induced PRCA can be initially supportive as it takes time for the bone marrow to fully recover. However, prolonged and/or failure of the bone marrow to recover, significantly increases patient's risk of iron overload in the setting of frequent transfusions. Iron deposition can potentially lead to severe life-threatening multiorgan involvement which can be fatal. Therefore, earlier recognition and intervention with immunomodulators in patients who undergo frequent transfusions can be beneficial to mitigate this risk. Here, we present a case with severe transfusion-dependent PRCA following major ABO-incompatible allo-HSCT successfully treated with daratumumab.

Development and initial testing of TOGETHER-YA: an eHealth-delivered and group-based psychosocial intervention for young adult cancer survivors.

PURPOSE: This study aimed to (1) develop TOGETHER-YA, an e-Health-delivered and group-based health-related quality of life (HRQOL) intervention for young adult (YA) cancer survivors aged 18-39 (Part 1), and (2) determine its initial feasibility and acceptability in a single-arm pilot trial (Part 2). METHODS: TOGETHER-YA is a manualized, 10-week intervention for YA survivors that includes elements of relaxation training, cognitive-behavioral therapy, and health education. In Part 1, content was adapted from existing evidence-based interventions with feedback from YAs (N = 22) in four iterative focus groups. In Part 2, YA survivors (N = 11) participated in a single-arm pilot trial of TOGETHER-YA. Intervention groups were led by a trained facilitator over videoconference. Primary outcomes were feasibility (i.e., recruitment, session attendance, retention) and acceptability (i.e., participant satisfaction). RESULTS: Focus groups reacted positively to TOGETHER-YA and provided actionable recommendations for enhancing its relevance and acceptability, which were implemented. In initial testing, all feasibility and acceptability benchmarks were met; 58% of eligible YAs were recruited, participants attended M = 6 intervention sessions (SD = 3), and 82% of participants were retained post-intervention. On average, participants "agreed" to "strongly agreed" with positive statements about the weekly sessions and the overall program. CONCLUSION: TOGETHER-YA was developed in collaboration with YA cancer survivors and found to be feasible and acceptable in initial testing. TOGETHER-YA is the first HRQOL intervention for a broad range of YA survivors that is eHealth-delivered for convenience and group-based for peer support. Future large-scale trials should test its efficacy for improving HRQOL. TRIAL REGISTRATION: NCT05048316, September 17, 2021; NCT05054569, September 23, 2021.

Acute exercise mobilizes NKT-like cells with a cytotoxic transcriptomic profile but does not augment the potency of cytokine-induced killer (CIK) cells.

CD3+/CD56+ Natural killer (NK) cell-like T-cells (NKT-like cells) represent <5% of blood lymphocytes, display a cytotoxic phenotype, and can kill various cancers. NKT-like cells can be expanded ex vivo into cytokine-induced killer (CIK) cells, however this therapeutic cell product has had mixed results against hematological malignancies in clinical trials. The aim of this study was to determine if NKT-like cells mobilized during acute cycling exercise could be used to generate more potent anti-tumor CIK cells from healthy donors. An acute exercise bout increased NKT-like cell numbers in blood 2-fold. Single cell RNA sequencing revealed that exercise mobilized NKT-like cells have an upregulation of genes and transcriptomic programs associated with enhanced anti-tumor activity, including cytotoxicity, cytokine responsiveness, and migration. Exercise, however, did not augment the ex vivo expansion of CIK cells or alter their surface phenotypes after 21-days of culture. CIK cells expanded at rest, during exercise (at 60% and 80% VO2max) or after (1h post) were equally capable of killing leukemia, lymphoma, and multiple myeloma target cells with and without cytokine (IL-2) and antibody (OKT3) priming in vitro. We conclude that acute exercise in healthy donors mobilizes NKT-like cells with an upregulation of transcriptomic programs involved in anti-tumor activity, but does not augment the ex vivo expansion of CIK cells.

Murine precursors to type 1 conventional dendritic cells induce tumor cytotoxicity and exhibit activated PD-1/PD-L1 pathway.

The immediate precursor to murine type 1 conventional DCs (cDC1s) has recently been established and named "pre-cDC1s". Mature CD8α+ cDC1s are recognized for suppressing graft-versus-host disease (GvHD) while promoting graft-versus-leukemia (GvL), however pre-cDC1s have not previously been investigated in the context of alloreactivity or anti-tumor responses. Characterization of pre-cDC1s, compared to CD8α+ cDC1s, found that a lower percentage of pre-cDC1s express PD-L1, yet express greater PD-L1 by MFI and a greater percent PIR-B, a GvHD-suppressing molecule. Functional assays were performed ex vivo following in vivo depletion of CD8α+ DCs to examine whether pre-cDC1s play a redundant role in alloreactivity. Proliferation assays revealed less allogeneic T-cell proliferation in the absence of CD8α+ cDC1s, with slightly greater CD8+ T-cell proliferation. Further, in the absence of CD8α+ cDC1s, stimulated CD8+ T-cells exhibited significantly less PD-1 expression compared to CD4+ T-cells, and alloreactive T-cell death was significantly lower, driven by reduced CD4+ T-cell death. Tumor-killing assays revealed that T-cells primed with CD8α-depleted DCs ex vivo induce greater killing of A20 B-cell leukemia cells, particularly when antigen (Ag) is limited. Bulk RNA sequencing revealed distinct transcriptional programs of these DCs, with pre-cDC1s exhibiting activated PD-1/PD-L1 signaling compared to CD8α+ cDC1s. These results indicate distinct T-cell-priming capabilities of murine pre-cDC1s compared to CD8α+ cDC1s ex vivo, with potentially clinically relevant implications in suppressing GvHD while promoting GvL responses, highlighting the need for greater investigation of murine pre-cDC1s.

IL-2 and Zoledronic Acid Therapy Restores the In Vivo Anti-Leukemic Activity of Human Lymphocytes Pre-Exposed to Simulated Microgravity.

BACKGROUND: We have previously shown that the anti-tumor activity of human lymphocytes is diminished in vitro after 12-hours pre-exposure to simulated microgravity (SMG). Here we used an immunocompromised mouse model to determine if this loss of function would extend in vivo, and to also test the efficacy of IL-2 and zoledronic acid (ZOL) therapy as a potential countermeasure against SMG-induced immune dysfunction. We adoptively transferred human lymphocytes that were exposed to either SMG or 1G-control into NSG-Tg (Hu-IL15) mice 1-week after they were injected with a luciferase-tagged human chronic myeloid leukemia (K562) cell line. Tumor growth was monitored 2x weekly with bioluminescence imaging (BLI) for up to 6-weeks. RESULTS: Mice that received lymphocytes exposed to SMG showed greater tumor burden compared to those receiving lymphocytes exposed to 1G (week 6 BLI: 1.8e10 ± 8.07e9 versus 2.22e8 ± 1.39e8 photons/second; p < 0.0001). Peak BLI was also higher in the SMG group compared to 1G-control (2.34e10 ± 1.23e10 versus 3.75e8 ± 1.56e8 photons/second; p = 0.0062). Exposure to SMG did not affect the ability of human lymphocytes to engraft or evoke xeno-graft-versus-host disease in the mice. Additionally, we injected the mice with IL-2 and zoledronic acid (ZOL) to expand and activate the anti-tumor activity of NK cells and γ δ-T cells, respectively. This treatment was found to revive the loss of anti-leukemic function observed in vivo when lymphocytes were pre-exposed to SMG. CONCLUSIONS: Microgravity plays a contributory role in loss of tumor control in vivo. Immuno-stimulating agents like ZOL+IL-2 may offer an important countermeasure for immune dysregulation during prolonged spaceflight.