Abstract: INTRO: Immune profiles and immune reconstitution are increasingly studied as important contributors to the prognosis and treatment responses of hematologic malignancy patients. Data on epidemiologic factors influencing immune profiles in hematologic malignancy patients are lacking. METHODS: We performed flow cytometric analyses of immune panels including T-cell, B-cell, NK cell and dendritic cell (DC) subsets in 1,025 consecutive adult hematologic malignancy patients (N=873) and controls/donors (N=152) between 2006-2016. Immune panels were analyzed on fresh peripheral blood samples drawn during workup before autologous or allogeneic hematopoietic cell transplant (HCT). Hematologic malignancy diagnoses were AML (N=235), MM (N=228), NHL (N=197), MDS/MPN (N=85), ALL (N=60), HL (N=33) and other leukemias (N=35). Patients were 58% male, median age 58 years (range 18-77), 91% non-Hispanic white (NHW), 6% African-American (AA), 1.3% Hispanic, 1.5% other. The HCT-comorbidity index (HCT-CI) in patients showed: 63% with & gt; 1 co-morbidity; 26% moderate pulmonary, 22% psychologic (requiring therapy/counseling), 16% severe pulmonary, 12% cardiac (CHF/CAD/MI), 11% diabetes requiring insulin, 9% obese (BMI & gt;35mg/kg 2), 9% prior cancer. Controls were related apheresis or marrow donors of allogeneic HCT patients: 51% male, median age 49 years (range 19-73), 92% NHW, 4% AA, 1.3% Hispanic, 2.6% other. HCT-CI scoring of controls: 45% with & gt; 1 comorbidity, 20% obese, 20% psychologic, 10% mild liver disease, 7% diabetes. Due to inter-individual cell count variability, immune cells were normalized as percent gated of lymphocytes except DC populations which were gated on mononuclear cells, both on forward and side scatter. To control for multiple comparisons, Bonferroni corrected statistically significant P was set at & lt;0.001. RESULTS: In controls, males had a significantly lower proportion of CD3+ cells/µl than females (68% vs 73%, P=0.001), NHW had a higher proportion of CD8+ central memory (CM) cells than other race/ethnicities (4.6 vs 2.7%, P=0.004) with no other significant differences by sex or race, although some of the race groups were low in our cohort. In contrast, among hematologic malignancy patients, males had significantly lower CD4+, CD4+ naïve, CD4+ recent thymic emigrants (RTEs), total T-regulatory cells (Treg), and CD19+ naïve cells but significantly higher CD8+ CM and effector memory (EM) cells than females. In addition, NHW had higher CD8+ CM than AA, Hispanic and Other races. Significant differences by age in patients and controls are shown in the Table. Across all age groups, patients had higher proportions of CD3+ cells, lower proportions of B-cells and no difference in NK or DCs than controls. As controls increased in age, CD4+ total significantly increased, while CD8+total, CD8+naive, T-γδ cells decreased, and myeloid DCs were highest at each end of the age spectrum. As patients increased in age, activated HLA-DR T-cells significantly increased, while T-γδ, CD8+naïve, and RTEs significantly decreased. The CD4:8 ratio increased while the CD4+ and CD8+ naïve:EM ratio decreased with age in both controls and patients, however patients had lower CD4:CD8 and naïve:EM ratios than controls. Immunophenotypes by patient disease are shown in the Figure. In general, patients with lymphoid diseases had lower CD3+CD4+ but higher CD3+CD8+, NK and DCs. In controls/donors, there were no significant differences in immune cell profiles by the presence or absence of comorbidities: obesity, diabetes, psychologic and mild liver disease. In patients, T-γδ were significantly lower in patients with diabetes, with no other significant differences for cardiac, psychologic, prior cancer, obesity or pulmonary co-morbidities. CONCLUSIONS: Additional analyses are ongoing to investigate the influence of prior therapies (chemotherapy, hypomethylating agents, monoclonal antibodies, etc.) and cytogenetic risk groups within each disease (AML, ALL, MDS/MPN, MM, NHL) on immune cell profiles. Studies of immunophenotyping in hematologic malignancies should include adjustment for confounders such as age, sex and race as biologic variables, as well as consideration of the diseases and treatments given. Interestingly, common co-morbidities did not broadly influence immune cell profiles in our cohort of hematologic malignancy patients. Figure 1 Figure 1. Disclosures Chen: Actinium Pharmaceuticals: Other: Principal Investigator, SIERRA Trial, Actinium. Hillengass: Bristol Myers Squibb: Membership on an entity's Board of Directors or advisory committees; Beijing Medical Award Foundation: Speakers Bureau; Oncopeptides: Membership on an entity's Board of Directors or advisory committees; Sanofi: Membership on an entity's Board of Directors or advisory committees; Beijing Life Oasis Public Service Center: Speakers Bureau; Skyline: Membership on an entity's Board of Directors or advisory committees; Curio Science: Speakers Bureau; Adaptive: Membership on an entity's Board of Directors or advisory committees; GlaxoSmithKline: Membership on an entity's Board of Directors or advisory committees; Oncotracker: Membership on an entity's Board of Directors or advisory committees; Axxess Network: Membership on an entity's Board of Directors or advisory committees. Wang: Genentech: Membership on an entity's Board of Directors or advisory committees; DAVA Oncology: Consultancy, Speakers Bureau; BMS/Celgene: Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Honoraria, Other: Advisory Board; Mana Therapeutics: Consultancy, Honoraria; Astellas: Consultancy, Membership on an entity's Board of Directors or advisory committees; AbbVie: Consultancy, Membership on an entity's Board of Directors or advisory committees; Kura Oncology: Consultancy, Honoraria, Other: Advisory board, steering committee, Speakers Bureau; GlaxoSmithKline: Consultancy, Honoraria, Other: Advisory Board; Jazz Pharmaceuticals: Consultancy, Honoraria, Other: Advisory Board; Kite Pharmaceuticals: Consultancy, Honoraria, Other: Advisory Board; Pfizer: Consultancy, Honoraria, Other: Advisory Board, Speakers Bureau; Stemline Therapeutics: Consultancy, Honoraria, Other: Advisory board, Speakers Bureau; Takeda: Consultancy, Honoraria, Other: Advisory board; Rafael Pharmaceuticals: Other: Data safety monitoring committee; Gilead: Consultancy, Honoraria, Other: Advisory board; Daiichi Sankyo: Consultancy, Honoraria, Other: Advisory board; PTC Therapeutics: Consultancy, Honoraria, Other: Advisory board; Genentech: Consultancy; MacroGenics: Consultancy. Griffiths: Takeda Oncology: Consultancy, Honoraria; Alexion Pharmaceuticals: Consultancy, Research Funding; Apellis Pharmaceuticals: Research Funding; Abbvie: Consultancy, Honoraria; Astex Pharmaceuticals: Honoraria, Research Funding; Genentech: Research Funding; Taiho Oncology: Consultancy, Honoraria; Celgene/Bristol-Myers Squibb: Consultancy, Honoraria, Research Funding; Novartis: Honoraria; Boston Biomedical: Consultancy. Torka: TG Therapeutics: Membership on an entity's Board of Directors or advisory committees. McCarthy: Bluebird: Honoraria, Membership on an entity's Board of Directors or advisory committees; Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees; Takeda: Honoraria, Membership on an entity's Board of Directors or advisory committees; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Karyopharm: Honoraria, Membership on an entity's Board of Directors or advisory committees; Bristol Myers Squibb: Honoraria, Membership on an entity's Board of Directors or advisory committees; Magenta Therapeutics: Honoraria, Membership on an entity's Board of Directors or advisory committees; Oncopeptides: Honoraria, Membership on an entity's Board of Directors or advisory committees; Juno: Honoraria, Membership on an entity's Board of Directors or advisory committees.

Abstract: β2 adrenergic receptor signaling is a key regulator of various immune cells, including T cells; however, its role in T cell function in the context of graft versus host disease (GvHD) is poorly understood. We previously showed that housing mice at thermoneutral temperature (TT; 30°C), which reduces systemic adrenergic stress, increased the incidence and severity of GvHD after allogeneic hematopoietic cell transplant (allo-HCT) compared to mice housed at standard temperature (ST; 22°C) which exerts a mild but chronic adrenergic stress (Leigh et al J Immunol 2015). The increased incidence and severity of GvHD in TT mice can be reversed by the administration of a β2-adrenergic receptor (β2-AR) agonist, suggesting an important role of epinephrine and norepinephrine in allo-HCT outcome (Leigh et al., J. Immunol 2015; Mohammadpour et al J Immunol 2018). We investigated the mechanisms and downstream events of β2-AR signaling in donor T cells after allo-HCT by using β2-AR knockout (β2-AR-/-) mice and commercially available β2-AR agonists. The main goal here was to explore whether signaling through β2-AR in donor T cells could control GvHD incidence and severity without minimizing the graft-versus leukemia (GvL) effect. We utilized both a major MHC-mismatch C57B6 (H-2kb) into BALB/c (H-2kd) model and a MHC-matched, multiple minor histocompatibility antigen (miHA) mismatched B6 (H-2kb) into C3H/SW (H-2kb) model. Recipient BALB/c and C3H/SW WT mice were lethally irradiated with 850 and 1100 cGy respectively and injected by tail vein with T cell depleted bone marrow (TCD-BM) alone (3 ×106) or TCD-BM and splenic T cells derived from allogeneic WT or β2-AR-/- B6 donors (0.7 × 106 T cells in B6 → BALB/c and 1.5 × 106 in B6 → C3H/SW). We found that donor T cells express β2-AR after allo-HCT and that β2-AR expression on WT T cells plays an important role in controlling GvHD, as evidenced by less severe weight loss, and increased survival compared to mice receiving β2-AR-/- donor T cells (Figure 1A). Histopathologic examination showed that β2-AR-/- T cells induced more damage in the small and large intestine. To explore further the mechanism(s) by which β2-AR signaling controls the severity of GvHD, we used NanoString analysis and discovered that β2-AR-/- T cells have the Th1 phenotype with an increase in Tbx21, Ifng, Irf8 and Emoes genes, while WT CD4+ T cells had higher levels of Th2 and Treg associated genes, including Foxp3, Ptgs5, Tgfb2, Il10, Il21 and Il22. We also observed a significant increase in the inflammatory cytokines IFN-γ and IL-17 in β2-AR-/- CD4+ T cells from the spleen and liver on days 7 and 14 after allo-HCT as compared to WT T cells (Figure 1B), while the expression of IL-10 was significantly higher in WT T cells compared to β2-AR-/- T cells (P 〈 0.01). We next sought to determine whether GvL may be affected by use of long acting β2-AR agonist (Bambuterol) to control GvHD. Bambuterol was administered daily at a dose of 1mg/kg from day 0. We observed that Bambuterol controlled the severity and mortality of GvHD after allo-HCT in both major and minor mismatch mouse models, as evidenced by reduced weight loss and an improved clinical score and survival rate in mice receiving Bambuterol compared to vehicle (P 〈 0.001). We showed that treatment increased the expression of IL-10 and decreased the expression of IFN-γ and IL-17 in CD4+ T cells. Interestingly, we found that β2-AR agonist treatment significantly increased the generation of myeloid derived suppressor cells (MDSCs) from WT BM without any effect on β2-AR-/- BM both in vitro and in vivo, suggesting an important role of β2-AR signaling in the generation of MDSCs. To investigate the effect of Bambuterol on GvL, the A20 lymphoma cell line was injected 4 hours before allo-HCT. Using two different doses of T cells (0.5 × 106 and 0.2 × 106) in B6 → BALB/c model, we found that Bambuterol preserved GvL by inducing CD44+ CD62L- NKG2D+ effector cells and CD44+ CD62L+ central memory cells. Since β2-AR agonists can affect cardiac function, we measured heart rate (HR) and blood pressure (BP) using a tail-cuff. There was no difference in BP and HR at day 21 and 28 after allo-HCT between mice receiving Bambuterol compared to mice receiving vehicle. In conclusion, these data reveal how β-AR signaling can influence donor T cell differentiation and function in murine GvHD models without decreasing GvL effect pointing to the feasibility of manipulation of β2-AR signaling to ameliorate clinical GvHD. Disclosures No relevant conflicts of interest to declare.