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B-cell maturation antigen-targeted chimeric antigen receptor T-cell therapy (BCMA CAR-T) is an effective treatment of relapsed refractory multiple myeloma (MM). However, the pattern of infectious complications is not well elucidated. We performed a single-center retrospective analysis of infection outcomes up to 1 year after BCMA CAR-T for MM from 2018 to 2020. Fifty-five patients with MM were treated with BCMA CAR-T. Before lymphodepletion (LD), 35% of patients had severe hypogammaglobulinemia and 18% had severe lymphopenia. Most patients (68%) received bridging chemotherapy (BC) before LD. In the first month after CAR-T, 98% patients had grade 3 to 4 neutropenia. At 1 year after infusion, 76% patients had hypogammaglobulinemia. With a median follow-up of 6.0 months (95% confidence interval, 4.7-7.4), there were a total of 47 infection events in 29 (53%) patients: 40% bacterial, 53% viral, and 6% fungal. Most (92%) were mild-moderate and of the lower/upper respiratory tract system (68%). Half of the infections (53%) occurred in the first 100 days after CAR-T infusion. Although no statistically significant risk factors for infection were identified, prior lines of therapy, use of BC, recent infections, and post–CAR-T lymphopenia were identified as possible risk factors that need to be further explored. This is the largest study to date to assess infectious complications after BCMA CAR-T. Despite multiple risk factors for severe immunosuppression in this cohort, relatively few life-threatening or severe infections occurred. Further larger studies are needed to better characterize the risk factors for and occurrence of infections after BCMA CAR-T.

Background: Chimeric antigen receptor T-cell (CAR T) immunotherapy is an evolving treatment for relapsed/ refractory non-Hodgkin lymphoma (NHL) and multiple myeloma (MM). There is a growing need to elucidate the infectious complications of BCMA- and CD19-directed CAR T therapy. We performed a single-center retrospective analysis of infection outcomes up to 1-year post BCMA and CD19-directed CAR T treatment. Methods: All patients treated at our institution with a BCMA-directed CAR T therapy for MM or with a CD19-directed CAR T for NHL from 2018-2020 were analyzed for risk factors for infection and infectious complications. Bacterial, viral and fungal infections were recorded if there was a microbiologic or histopathologic diagnosis or if clinical suspicion for infection required empiric treatment. Infection severity was classified as mild, moderate, severe, life-threatening, or fatal as previously described (Young et al. 2016, van Burnik et al. 2007). Infections were identified from the day of the first CAR T-cell infusion up to 1 year after infusion. Observation of infections was terminated at the time of disease progression, initiation of next therapy, or death, whichever occurred first. Fisher's exact test and Wilcoxon rank-sum test were used to compare between cohorts. Poisson mixed effects model with a subject-specific random intercept and an offset to account for the duration at risk was used to identify risk factors for infections. Results: Of the 104 subjects in this study, 55 patients (53%) had MM treated with BCMA CAR T and 49 patients (47%) had NHL treated with CD19 CAR T. Median number of prior therapies was 6 (4 - 9) in the BCMA cohort and 3 (2-4) in the CD19 cohort (Table 1). Prior to starting lymphodepleting (LD) chemotherapy, most patients did not have IgG< 400, ALC< 200, or ANC< 500. Almost all the patients were on antibacterial (99%), antiviral (99%) and antifungal (92%) prophylaxis (ppx), with 18%, 90% and 9% of patients starting this before LD chemotherapy, respectively. Median follow-up time was 5.8 months (95% CI: 4.2-6.4). In total, there were 87 infection events (48 bacterial, 33 viral, and 6 fungal) observed in 56 patients (54%), with 47 infections in 29 (53%) patients in the BCMA CAR T cohort and 40 infections in 27 (55%) patients in the CD19 CAR T cohort (p =0.9). The BCMA cohort had 19 bacterial (40%) vs 29 bacterial (73%) in CD19 cohort (p=0.005), while BCMA cohort had 25 viral (53%) vs 8 viral (20%) in CD19 cohort (p=0.002). Fungal infection rates were comparable between BCMA and CD19 cohorts, 3 (6%) vs 3 (8%) respectively (p=1). Among the infections that occurred, 20 (23%) were high severity occurring in 16 patients (15%) of the overall cohort. Four high severity infections (5%) occurred in 3 patients (5%) in the BCMA cohort while 16 high severity infections (18%) occurred in 13 patients (27%) in the CD19 cohort (p < 0.001) (Figure 1). The BCMA cohort had higher rates of respiratory infections (68% vs 50%, p=0.1), while the CD19 cohort had higher rates of bloodstream infections (15% vs 2%, p=0.05) and gastrointestinal infections (10% vs 0%, p=0.04). The rates of grade (Gr) 3-4 neutropenia and IgG < 300 in both the cohorts at various time points post CAR T were comparable. At 9-12 months, Gr 3-4 neutropenia was 7% vs 8% (p=1) and IgG < 300 was 7% vs 33% (p=0.1) for the BCMA and CD19 cohorts respectively. Adjusting for time periods, risk factors for development of infections were use of steroids (incidence rate ratio [IRR] 1.6, 95% CI: 1.1-2.5, p=0.03) and post CAR T hypogammaglobulinemia (IgG < 600) (IRR =2.1, 95% CI: 1.2-3.9, p=0.02). Conclusions: This retrospective study is one of the largest studies to date comparing the post CAR T infections rates between BCMA and CD19 directed CAR T treated patients. While the incidence of infection, use of antimicrobial ppx, and rates of Gr 3-4 neutropenia and hypogammaglobulinemia were comparable between BCMA and CD19 CAR T cohorts, viral infections were more common post BCMA CAR T while bacterial infections were more frequent post CD19 CAR T. Use of steroids as well as post CAR T hypogammaglobulinemia are possible risk factors for development of infections. Further studies are needed to examine the infectious complications post-CAR T treatment, characterize the underlying risk factors, and to establish appropriate prophylactic approaches in patients undergoing CD19- and BCMA-directed CAR T therapy. Disclosures Fakhri: University of California San Francisco: Current Employment. Ai:Nurix Therapeutics: Membership on an entity's Board of Directors or advisory committees, Research Funding; ADC Therapeutics, Kymera: Membership on an entity's Board of Directors or advisory committees. Martin:Seattle Genetics: Research Funding; GSK: Consultancy; Sanofi: Research Funding; AMGEN: Research Funding; Janssen: Research Funding. Wolf:Adaptive: Consultancy, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Shah:BMS, Janssen, Bluebird Bio, Sutro Biopharma, Teneobio, Poseida, Nektar: Research Funding; GSK, Amgen, Indapta Therapeutics, Sanofi, BMS, CareDx, Kite, Karyopharm: Consultancy. Andreadis:Genentech: Other: Spouse Employee (salary and stock); Novartis: Research Funding; Celgene/Juno: Research Funding; Amgen: Research Funding; Merck: Research Funding; Gilead/Kite: Other: Advisor; Jazz Pharmaceuticals: Other: Advisor; Astellas: Other: Advisor; Seattle Genetics: Other: Advisor; Karyopharm: Other: Advisor; Incyte: Other. Wong:GSK: Research Funding; Amgen: Consultancy; Sanofi: Membership on an entity's Board of Directors or advisory committees; Bristol Myers Squibb: Research Funding; Roche: Research Funding; Janssen: Research Funding; Fortis: Research Funding.

Abstract 763 Studies of the vitamin D3 signaling pathway have revealed a broad role for this hormone in tissue homeostasis and as a result there is great interest in exploring the therapeutic potential of vitamin D3 for the treatment of various diseases including cancer. Studies investigating the role of vitamin D3 in hematological malignancies have shown that in vitro vitamin D3 has anti-proliferative and pro-differentiation effects resulting in differentiation of leukemic cell lines towards the monocytic lineage. Surprisingly, the role of vitamin D3 in HSC homeostasis and leukemia progression in vivo is not well understood. To elucidate the mechanisms of action of vitamin D3 in vivo during HSC self-renewal and differentiation, we utilized the zebrafish (Danio rerio) as vertebrate model due to their evolutionary conserved blood system and their amenability for genetic and chemical manipulation. Vitamin D3 was identified in a chemical screen performed in our laboratory as a regulator of runx1 and c-myb expression in the aorta-gonad mesonephros (AGM), the first site of definitive hematopoiesis. Treatments of zebrafish embryos with active vitamin D3 (1,25OH D3) between 12–36 hours post fertilization (hpf) resulted in increased expression of HSC markers (runx1, c-myb, CD41) as determined by whole mount in situ hybridization (WISH). In contrast, treatment with the non-hydroxylated vitamin D3 precursor cholecalciferol (D3) resulted in decreased runx1 and c-myb expression. D3 treatment during HSC induction and expansion from 24–36 hpf did not affect runx1 and c-myb expression, suggesting that D3 is acting early during the establishment of the vascular niche. To quantify the difference in HSC progenitors observed by WISH, FACS analysis was performed on double positive Tg(Lmo2:dsRed), Tg(c-myb:gfp) embryos. 1,25OH D3 treated embryos had a 20% increase in the double positive cell population corresponding to HSCs, while treatment with D3 resulted in a 25% decrease in the number of HSCs. The differential effect of 1,25OH D3 and D3 suggest that these two compounds act via two distinct mechanisms. To determine if the HSC enhancement observed by 1,25OH D3 was acting through the canonical vitamin D3 receptor (VDR), loss of function experiments were performed by injecting morpholinos targeting the vitamin D receptors. Morpholino knockdown of VDRA and VDRB (zebrafish VDR orthologs) resulted in decreased expression of runx1 c-myb, and CD41 via WISH. The decrease in HSC positive cells was confirmed by fluorescence microscopy using the Tg(Runx1P2:gfp) and Tg(-6.0itga:gfp) reporter lines, further supporting our hypothesis. To address the decrease in HSCs by D3, we postulated that D3 was inhibiting hedgehog signaling, as vitamin D3 has been previously shown to act as a negative regulator of the hedgehog pathway. In support of our hypothesis, treatment with D3 resulted in decreased ptch2 mRNA expression a downstream target of hedgehog signaling. In addition, FACS analysis using the hedgehog reporter Tg(6xGli:mCherry) line showed a 15% reduction in the number of mCherry positive cells in D3 treated embryos compared to controls. Co-treatments of zebrafish embryos with the hedgehog antagonist cyclopamine and D3 resulted in additive loss of reporter activity (45%) and loss of runx1 positive cells in the AGM, revealing synergy between these two compounds. In addition, co-exposure with the hedgehog agonist SAG rescued the HSC defect in D3-treated embryos. Hedgehog signaling is known to regulate vein and artery specification through the activation of notch signaling. Consistent with our hypothesis that D3 reduces hedgehog signaling, an expansion of the venous marker flt4 and a reduction in the expression of the arterial marker ephrinb2a was observed by WISH on D3-treated embryos. Furthermore, treatment of the notch reporter line Tg(Notch:gfp) with D3 resulted in lowered notch activity. In summary, these studies reveal that the active 1,25OH D3 positively regulates HSC progenitors in vivo during the onset of definitive hematopoiesis. In contrast, the non-hydroxylated vitamin D3 precursor acts as negative regulator of hematopoiesis by inhibiting hedgehog signaling and affecting vascular niche formation. Based on our studies, the differential effect between D3 and 1,25OH D3 should be considered when investigating the therapeutic potential of vitamin D3 in the context of hematological malignancies and other cancers. Disclosures: Goessling: Fate Therapeutics: Consultancy. North:Fate Therapeutics: Consultancy.