Alkhateeb, Hassan B.

Abstract The ZRSR2 gene is a member of the spliceosome gene family which are frequently mutated in myeloid neoplasms. ZRSR2 mutations (ZRSR2m) occur in less than 5% of MDS, CMML, and AML. Our study included 164 ZRSR2m patients (98.8% males) and 98 ZRSR2wt MDS control cohort. In the ZRSR2m group, there were 53 MDS (32%), 39 CCUS (24%), 33 MPN (20%), 23 MDS/MPN overlap (14%), 15 AML (9%), and 1 MPAL (0.6%). Most MDS patients were the low blast subtype (n=37, 70%). Twenty-seven MDS patients (51%), and 23 CCUS patients (59%) had absolute monocyte count ≥0.5 x109/L and 18 of 23 MDS/MPN overlap were CMML (78%). Mutations in ZRSR2 were spread across the entire gene. The median number of co-mutations was 2, with TET2 (51%) and ASXL1 (32%) being the most common. U2AF1 and SRSF2, previously described as mutually exclusive with ZRSR2, were found in 10.4% of patients. Median overall survival (OS) was 51 months, and significantly varied among MNs (p=0.004). ZRSR2m MDS patients had better mOS than the MDS control cohort with ZRSR2wt (35 vs 22 months, p=0.013). ZRSR2m patients with isolated TET2 co-mutation and higher hemoglobin showed improved survival, while patients with RUNX1m, higher WBC count showed worse OS.

6513 Background: PTPN11 (Ch12q24) is a proto-oncogene that codes for SHP2, a regulatory protein tyrosine phosphatase that impacts RAS/MAPK signaling. Germline variants have been described in juvenile myelomonocytic leukemia, while somatic variants have been associated with inferior survival in acute myeloid leukemia. PTPN11 variants are infrequent in chronic myelomonocytic leukemia (CMML), and their impact on disease phenotype and survival remains unclear. Methods: After IRB approval, 396 molecularly annotated CMML patients at Mayo Clinic were screened for somatic PTPN11 variants within 12 months of diagnosis while still in chronic phase disease. Overall (OS), acute myeloid leukemia (AML)-free survival (LFS), and predictors of survival were estimated using standard statistical measures. Results: 396 patients (median age 71 years; males 67%) with CMML were considered; pathogenic PTPN11 variants were seen in 12 (3%) with a median VAF of 20.5% (range: 4-50): A72T (n = 2), F285S (n = 2), and A72S (n = 2). Variants were most likely to be seen in the N-terminal SH2 domain (n = 7), followed by the PTP domain (n = 6), and the C-terminal SH2 domain (n = 1). PTPN11 variants clustered with CMML-2 (p = 0.02), higher absolute neutrophil count (p = 0.04), peripheral blood (p = 0.06) and bone marrow (p = 0.02) blast %, older age (p <0.01), and were less likely to have SRSF2 comutations (p = 0.02). After a median follow-up of 18 months (range 0-193), 253 (64%) deaths and 68 (17%) AML transformations were recorded. PTPN11 variants negatively and independently impacted both OS (median 13 vs. 31 months; p <0.01) and LFS (AML events: 42% (5/12) vs. 16% (63/384); p <0.01). In age-adjusted multivariate analysis restricted to genetic risk factors, survival was negatively affected by abnormal karyotype and mutations involving PTPN11, DNMT3A, and SETBP1, and positively by TET2 mutations. All but SETBP1 retained significance in the presence of additional independent risk factors, namely male sex (p <0.01) and leukocyte count (p <0.01); HR in the latter analysis was 4.3 for PTPN11, 2.8 for DNMT3A, and 1.6 for abnormal karyotype. A similar analysis for LFS identified, as risk factors, PTPN11 (p = 0.01), DNMT3A (p <0.01), and ASXL1 (p = 0.03) mutations, as well as bone marrow blast % (p <0.01), absolute monocyte count ≥10 (p = 0.02), and immature circulating myeloid cells (p <0.01). Conclusions: The current study identifies PTPN11 and DNMT3A mutations as independent risk factors for both OS and LFS in CMML. In regard to additional genetic risk factors, OS was positively affected by TET2 mutations and negatively by abnormal karyotype, while LFS was negatively affected by ASXL1 mutations.

7516 Background: Immune effector cell hemophagocytic lymphohistiocytosis IEC-HS is a complication of CAR T cell therapy (CART). This study aims to look at the outcomes of CART patients who develop IEC-HS. Methods: Patients who developed IEC-HS after receiving CART and patients who developed IEC-HS without (nonCART) between December 2020 and March 2022 were included. Results: The 20 CART patients were older than the 25 nonCART patients. Peak ferritin, fibrinogen, CRP, serum creatinine and transaminases were similar between the 2 group. CART patients had lower white blood cell count and platelets and bilirubin than the nonCART patients. CART patients were more like to have markedly elevated IL-6 levels (> 315 mg/ml), higher IL-18, higher MCP-1, and suppressed IL-1β and GMCSF than nonCART patients (Table). Treatment of IEC-HS included tocilizumab (n = 19), anakinra (n = 18) siltuximab (n = 4) and basiliximab (n = 4), dexamethasone (n = 18) methylprednisolone (n = 6), etoposide (n = 1) and ruxolitinib (n = 1). A significant response defined by a 90% reduction or normalization of the cytokine was observed with IL-10 and IFN-ϒ in 92.3% and 84.6% of the CART patients respectively. An intermediate response was noted with TNF, IL-6, MCP-1 and MIP-1 in 38.4%, 30.8% and 61.6% and 38.5% of the patients respectively. No patient had a significant response in IL-2Rα or IL-1β and only 7.7% had a significant response in IL-18. Of the 5 patients who had a MIP-1 response, 4 were on basiliximab +/- high flow continuous venovenous hemofiltration and the only patient that had an IL-18 response was on both modalities. The 100-day mortality after IEC-HS in the CART patients was 40% and was associated with higher peak and lower best ferritin but not the percentage reduction (Table). Higher TNF (90.3 pg/ml vs 37.2 pg/ml, p = 0.05), markedly elevated levels of IL-6 (92% vs 50%, p = 0.03) and IL-10 (33% vs 0%) and an IL-6 response (44.4% vs 0%) were associated with survival at day 100. IL-6 responders had lower peak ferritin vs IL-6 nonresponders (14066 mcg/L vs 45051 mcg/L, p = 0.08). Enterococcus faecium bacteremia developed in 75% of the nonsurvivors vs 16.6% of the 100-day survivors (p = 0.009). Survivors cleared the enterococcus bacteremia within days while nonsurvivors had protracted bacteremia. Conclusions: IEC-HS is devastating complication of CART. Anti-cytokine therapy can reduce some cytokines and ferritin but overimmunosuppression may lead infectious complication, in particular with E. faecium. Better biomarkers are needed to finetune immunosuppressive therapy in order to avoid over-immunosuppression. [Table: see text]

7038 Background: While chimeric antigen receptor T-cell (CAR-T) therapy has transformed treatment for hematological malignancies, severe (≥ grade 3,Gr 3, CTCAE) cytopenias post CAR-T and therapy related myeloid neoplasms (t-MN) are particularly difficult to manage. Survival after diagnosis of t-MN is dismal. Management of ≥Gr 3 cytopenias that do not recover by month 3 after CAR-T require frequent monitoring and transfusion support. Growth factor and thrombopoietin mimetics have variable success, with stem cell boost emerging as having more consistent success. Evaluation for CAR-T eligibility, prior to leukapheresis, is the critical timepoint where factors identified at this time could inform risks and benefits of CAR-T versus alternative treatment and for consideration of stem cell collection in patients who may benefit from stem cell boost to treat severe cytopenia post CAR-T. Methods: We conducted a retrospective analysis comparing clinical data of patients (pts) with lymphoma (NHL) and multiple myeloma (MM) treated with FDA-approved and investigational CAR-T between 01/2016 – 06/2022 at Mayo Clinic. Prolonged cytopenias were defined as hemoglobin (Hg)<8 g/dL, absolute neutrophil count (ANC)<0.5 x 109/L, and/or platelet count (PLT)<50 x 109/L at 3 months after CAR-T. Pts with progressive disease were excluded from cytopenia analysis. Logistic regression to identify factors correlated with cytopenia and t-MN. Results: Among the 186 pts who received CAR-T, 42 (22%) patients developed severe cytopenia (26/102, 25%, in NHL; 16/84, 19% in MM); 15/186 (8%) had more than 1 concurrent cytopenia. Univariate analysis identified that baseline ferritin and CAR-HEMATOTOX score correlated with cytopenia. Multivariate analysis showed that advanced age (≥ 65 years) (HR 2.69, 95%CI 1.17 – 6.40, P = 0.02) and thrombocytopenia (HR 4.01, 95%CI 1.77 – 9.34, P = 0.001) were associated with severe cytopenia. Twenty (10.7%) patients [8 (40%) males] developed t-MN at a median of 9.5 months (IQR 4.8 – 19.3 months) after CAR-T (14/102, 13% in NHL; 6/84, 7% in MM). Univariate analysis identified ferritin and CAR-HEMATOTOX score correlated with t-NM. Multivariate logistic regression showed advanced age (HR 5.03; 95%CI 1.59-18.7, P= 0.009), hemoglobin ≤ 10 g/dl (HR 3.63, 95%CI 1.04-13.30, P = 0.04), and thrombocytopenia (HR 4.06, 95%CI 1.32-14.2, P 0.02) are significantly associated with development of post-CART t-MN. Conclusions: Advanced age, anemia and thrombocytopenia at the time of evaluation for CAR-T eligibility, prior to leukapheresis, are associated with the development of post-treatment severe cytopenia and t-MN. Larger, multi-center studies are needed to validate these findings.

e14535 Background: Cytokine release syndrome (CRS) is the most common complication of chimeric antigen receptor T cells (CAR-T) for hematologic malignancies. We hypothesized that patients undergoing CAR-T therapy who are on metformin will experience a lower incidence or severity of CRS than patients not on this medication. Methods: We evaluated all patients treated with CAR-T at Mayo Clinic Rochester between 1/1/2018 and 1/31/2023, and abstracted relevant clinical and laboratory characteristics to determine the incidence and severity of CRS, PFS as well as OS in patients who were, or were not, on metformin before and during CAR-T therapy for Type 2 diabetes mellitus. We used the Chi-Square test, Mann-Whitney test and the Kaplan-Meier method to analyze data. Results: In the 5-year interval under consideration 237 patients received CAR-T, 168 for non-Hodgkin lymphoma (NHL), and 69 for multiple myeloma (MM). 23 patients (15 NHL and 8 MM) were on metformin at the time of CAR-T. 15 patient developed CRS while on metformin and 175 patients not on metformin (p=0.05). Grade 3 or 4 CRS incidence was 4% in both groups (p=0.41). Of the 153 patients with NHL not on metformin, 121 developed any grade of CRS, while 10 of 15 patients on metformin developed CRS (p=0.268). 4 NHL patients developed CRS Grade ≥3 or higher, while 0 of 15 on metformin developed Grade ≥3 CRS (p=0.237). For the MM patients, the incidence of CRS was 49 patients not on metformin and 5 patients on metformin (p<0.001). 3 MM patients developed CRS Grade ≥3 or higher, while 0 of 8 on metformin developed Grade ≥3 CRS (p=0.346). 12 of 15 patients on metformin with NHL achieved CR compared to 92 of 153 patients not on metformin (p=0.1305). 3 of 8 patients with MM on metformin compared to 24 of 61 patients with MM not on metformin achieved CR or better post CAR-T (p=0.92). Tocilizumab was used in 40% of patients on metformin but in 66% of patients not on metformin (p=0.0016). In NHL patients, PFS from CAR-T infusion was not reached (range: 6.3 to NR) for patients on metformin versus 9.9 months (range: 6.3–19.5) in patients not on metformin (p = 0.3332). In the MM patients, progression-free survival (PFS) from CAR-T infusion was 8.4 months (95% CI: 7.9–not reached [NR]) for patients on metformin and 22.6 months (13.7–NR) for those not on metformin (p = 0.07). Median OS for NHL patients on metformin was not reached (9.766–NR) versus 32.6 months (18.6–NR) for patients not on metformin (p = 0.7022). Median OS was 17 months (95% CI: 8.3 to NR) for patients with MM on metformin vs NR (95% CI: 9.4 to NR) for patients not on metformin (p = 0.2559). Conclusions: In this retrospective study, the co-administration of metformin during CAR-T cell therapy for NHL and MM appears to be associated with a numerically reduced incidence of CRS. The effect appears to be particularly marked in patients with multiple myeloma. Metformin therapy did not seem to affect the response to CAR-T therapy in either NHL or MM.

Abstract Clonal cytopenia of undetermined significance (CCUS) is defined by a myeloid driver mutation in the context of otherwise unexplained cytopenia. CCUS has an inherent risk of progressing to myeloid neoplasm. However, it is unknown how exposure to previous cytotoxic therapy may impact the risk of progression and survival. We stratified patients with CCUS by prior exposure to DNA-damaging therapy. Of 151 patients, 46 (30%) had received cytotoxic therapy and were classified as therapy-related CCUS (t-CCUS), whereas 105 (70%) had de novo CCUS. A lower proportion of t-CCUS had hypercellular marrows (17.8% vs 44.8%, P = .002) but had higher median bone marrow blast percentages. After a median follow-up of 2.2 years, t-CCUS had significantly shorter progression-free survival (PFS, 1.8 vs 6.3 years; hazard ratio [HR], 2.1; P = .007) and median overall survival (OS; 3.6 years vs not reached; HR, 2.3; P = .007) compared with CCUS. Univariable and multivariable time-to-event analyses showed that exposure to cytotoxic therapy independently accounted for inferior PFS and OS. Despite the similarities in clinical presentation between CCUS and t-CCUS, we show that exposure to prior cytotoxic therapies was an independent risk factor for inferior outcomes. This suggests that t-CCUS represents a unique clinical entity that needs more stringent monitoring or earlier intervention strategies.

SummaryAmong 301 newly diagnosed patients with acute myeloid leukaemia receiving venetoclax and a hypomethylating agent, 23 (7.6%) experienced major cardiac complications: 15 cardiomyopathy, 5 non‐ST elevation myocardial infarction and/or 7 pericarditis/effusions. Four patients had more than one cardiac complication. Baseline characteristics included median age ± interquartile range; 73 ± 5 years; 87% males; 96% with cardiovascular risk factors; and 90% with preserved baseline ejection fraction. In multivariate analysis, males were more likely (p = 0.02) and DNMT3A‐mutated cases less likely (p < 0.01) to be affected. Treatment‐emergent cardiac events were associated with a trend towards lower composite remission rates (43% vs. 62%; p = 0.09) and shorter survival (median 7.7 vs. 13.2 months; p < 0.01). These observations were retrospectively retrieved and warrant further prospective examination.

Abstract Heterozygous germline variants in DDX41 are identified in ~5% of patients with Myelodysplastic Syndrome (MDS) or Acute Myeloid Leukemia (AML). Recognition of the hereditary nature of such cancers and a better understanding of the factors that affect progression to malignancy has deep implications for health surveillance of patients and relatives that share a pathogenic DNA variant. DDX41 has a role in ribosome biogenesis and in opposing R-loop accumulation at promoters, but the mechanisms through which DDX41 loss drives cancer development are poorly understood. In a collaboration between Cambridge, UK and Mayo Clinic, USA we identified 85 patients with germline DDX41 variants [73/85 with germline pathogenic variants (GPV) and 12 with variants of uncertain significance (VUS)]. The majority were identified through targeted myeloid NGS upon presentation with AML (30/85) or MDS (44/85). Average age at diagnosis was 65 (35-93) with male predominance (57/85). The most common GPV was the NM_016222.4(DDX41):c.415_418dup; p.D140Gfs*2 (15/85). The majority (53.4% 39/73), also acquired a somatic variants in the other DDX41 allele (referred to as GPV+Som). The most common somatic variant (82% 32/39) was the R525H, a variant predicted to reduce the ATPase function of the helicase domain. We hypothesise that the markedly reduced DDX41 activity in cells with biallelic variants, is a key cause of progression to overt malignancy. In our cohort, we observed that somatic variants occurred at a similar frequency in association with start loss, truncating, splice, in-frame deletion, and missense GPV. However, whereas MDS was the predominant diagnosis within GPV (67.6% 23/34, AML 26.4% 9/34), AML was the most frequent diagnosis within GPV+Som (AML 51%.2 20/39, MDS 38.4% 15/39). Overall, there was a significantly higher likelihood of AML in the presence of a somatic variant (p&lt;0.05), suggesting that there might be a dosage effect on the phenotype. This pattern was observed across all the GPV, with the exception of missense, suggesting that many might be benign polymorphysms. In addition, although somatic variants are typically subclonal (overall mean variant allele fraction VAF=9.87%, 3-27.8%), we observed a higher somatic VAF in AML over MDS cases (12.4% vs 7.7% respectively p&lt;0.05). A lower bone marrow cellularity was also characteristic of GPV+Som vs GPV alone (mean observed/expected cellularity for age 0.9 vs 1.32 respectively p&lt;0.05). In two cases, serial bone marrow morphology and NGS data were available, and we observed marked reduction in bone marrow cellularity upon acquisition of the R525H clone. The mechanisms of AML evolution appear different in DDX41 HMMS compared to sporadic cases. Interestingly, over half of our patients (21/34 GPV and 23/39 GPV+Som) did not exhibit additional somatic drivers. Our work suggests that acquisition of a somatic variant is a critical event in disease progression. Furthermore, the near universally subclonal nature of somatic variants suggest that they may affect disease phenotype through non-cell autonomous mechanisms. Citation Format: Ludovica Marando, Talha Badar, Yael Kusne, Audrey Morris, Terra Lasho, Abhishek Mangaonkar, Christy Finke, Carles Crawley, James M. Foran, Aref Al-Kali, Andrew King, Hassan Alkhateeb, Naseema Gangat, Chi Wong, Rong He, David Viswanatha, Faisal Basheer, Mark Litzow, Alejandro Ferrer, George Vassiliou, Mrinal Patnaik. Investigating the role of the somatic DDX41 variant in the context of DDX41 hereditary myeloid malignancy syndromes (HMMS) [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Translating Cancer Evolution and Data Science: The Next Frontier; 2023 Dec 3-6; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(3 Suppl_2):Abstract nr A037.

AbstractRelapsed and refractory B‐cell acute lymphoblastic leukemia (B‐ALL) is an aggressive B‐cell neoplasm associated with poor outcomes. Conventional multiagent chemotherapy and bispecific antibody therapy may induce remission; however, relapse rates remain high and overall survival is poor. Chimeric antigen receptor T‐cell (CAR‐T) therapy provides durable, deep complete remission, and long‐term cures in relapsed and refractory B‐ALL. However, with this new treatment modality, 10%–30% of patients do not achieve remission, and over 50% experience relapse after therapy. Currently, there are two approved CD19‐specific CAR‐T cell constructs in B‐ALL, Tisagenlecleucel and Brexucabtagene Autoleucel by the United States Food and Drug Administration, and the European Medicines Agency (EMA). In this review, we discuss patients, disease, and CAR‐T predictors of outcomes in B‐ALL. We describe the two approved CD19‐directed CAR‐T cell products, review the current literature, and discuss factors associated with high risks of therapy failure and future direction in CAR‐T cell therapy for B‐ALL.