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Drug resistance in chronic myeloid leukaemia (CML) may occur via mutations in the causative BCR::ABL1 fusion or BCR::ABL1-independent mechanisms. We analysed 48 patients with BCR::ABL1-independent resistance for the presence of secondary fusion genes by RNA sequencing. We identified 10 of the most frequently detected secondary fusions in 21 patients. Validation studies, cell line models, gene expression analysis and drug screening revealed differences with respect to proliferation rate, differentiation and drug sensitivity. Notably, expression of RUNX1::MECOM led to resistance to ABL1 tyrosine kinase inhibitors in vitro. These results suggest secondary fusions contribute to BCR::ABL1-independent resistance and may be amenable to combined therapies.

Supplementary Figures S1-S6, and Table S1-S3

To understand mechanisms of response to BET inhibitors (BETi), we mined the Beat AML functional genomic data set and performed genome-wide CRISPR screens on BETi-sensitive and BETi-resistant acute myeloid leukemia (AML) cells. Both strategies revealed regulators of monocytic differentiation—SPI1, JUNB, FOS, and aryl-hydrocarbon receptor signaling (AHR/ARNT)—as determinants of BETi response. AHR activation synergized with BETi, whereas inhibition antagonized BETi-mediated cytotoxicity. Consistent with BETi sensitivity dependence on monocytic differentiation, ex vivo sensitivity to BETi in primary AML patient samples correlated with higher expression of the monocytic markers CSF1R, LILRs, and VCAN. In addition, HL-60 cell line differentiation enhanced its sensitivity to BETi. Further, screens to rescue BETi sensitivity identified BCL2 and CDK6 as druggable vulnerabilities. Finally, monocytic AML patient samples refractory to venetoclax ex vivo were significantly more sensitive to combined BETi + venetoclax. Together, our work highlights mechanisms that could predict BETi response and identifies combination strategies to overcome resistance.Significance:Drug resistance remains a challenge for AML, and new therapies, such as BETi, will require combination approaches to boost single-agent responses. We conducted genome-wide CRISPR screens and functional genomics on AML patient samples to identify leukemic differentiation state and AHR signaling as primary mediators of BETi response.This article is highlighted in the In This Issue feature, p. 403

Supplemental Figure 2. CLL-CI correlates with OS across CLL patient subgroups (derivation dataset).

Supplemental Figure 1. CIRS correlates with outcomes in CLL (derivation dataset).

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Purpose:Comorbid medical conditions define a subset of patients with chronic lymphocytic leukemia (CLL) with poor outcomes. However, which comorbidities are most predictive remains understudied.Experimental Design:We conducted a retrospective analysis from 10 academic centers to ascertain the relative importance of comorbidities assessed by the cumulative illness rating scale (CIRS). The influence of specific comorbidities on event-free survival (EFS) was assessed in this derivation dataset using random survival forests to construct a CLL-specific comorbidity index (CLL-CI). Cox models were then fit to this dataset and to a single-center, independent validation dataset.Results:The derivation and validation sets comprised 570 patients (59% receiving Bruton tyrosine kinase inhibitor, BTKi) and 167 patients (50% receiving BTKi), respectively. Of the 14 CIRS organ systems, three had a strong and stable influence on EFS: any vascular, moderate/severe endocrine, moderate/severe upper gastrointestinal comorbidity. These were combined to create the CLL-CI score, which was categorized into 3 risk groups. In the derivation dataset, the median EFS values were 58, 33, and 20 months in the low, intermediate, and high-risk groups, correspondingly. Two-year overall survival (OS) rates were 96%, 91%, and 82%. In the validation dataset, median EFS values were 81, 40, and 23 months (two-year OS rates 97%/92%/88%), correspondingly. Adjusting for prognostic factors, CLL-CI was significantly associated with EFS in patients treated with either chemo-immunotherapy or with BTKi in each of our 2 datasets.Conclusions:The CLL-CI is a simplified, CLL-specific comorbidity index that can be easily applied in clinical practice and correlates with survival in CLL.

Supplemental Fig. 3. CLL-CI correlates with EFS in patient subgroups (validation set).

BackgroundCyclin-dependent kinase 4/6 inhibitors (CDKi) have changed the landscape for treatment of patients with hormone receptor positive, human epidermal growth factor receptor 2-negative (HR+/HER−) metastatic breast cancer (MBC). However, next-line treatment strategies after CDKi progression are not yet optimized. We report here the impact of clinical and genomic factors on post-CDKi outcomes in a single institution cohort of HR+/HER2− patients with MBC.MethodsWe retrospectively reviewed the medical records of patients with HR+/HER2− MBC that received a CDKi between April 1, 2014 and December 1, 2019 at our institution. Data were summarized using descriptive statistics, the Kaplan-Meier method, and regression models.ResultsWe identified 140 patients with HR+/HER2− MBC that received a CDKi. Eighty percent of patients discontinued treatment due to disease progression, with a median progression-free survival (PFS) of 6.0 months (95% CI, 5.0-7.1), whereas those that discontinued CDKi for other reasons had a PFS of 11.3 months (95% CI, 4.6-19.4) (hazard ratio (HR) 2.53, 95% CI, 1.50-4.26 [P = .001]). The 6-month cumulative incidence of post-CDKi progression or death was 51% for the 112 patients who progressed on CDKi. Patients harboring PTEN mutations pre-CDKi treatment had poorer clinical outcomes compared to those with wild-type PTEN.ConclusionThis study highlights post-CDKi outcomes and the need for further molecular characterization and novel therapies to improve treatments for patients with HR+/HER2− MBC.

Using ex vivo drug screening of primary patient specimens, we identified the combination of the p38 MAPK inhibitor doramapimod (DORA) with the BCL2 inhibitor venetoclax (VEN) as demonstrating broad, enhanced efficacy compared with each single agent across 335 acute myeloid leukemia (AML) patient samples while sparing primary stromal cells. Single-agent DORA and VEN sensitivity was associated with distinct, nonoverlapping tumor cell differentiation states. In particular, increased monocytes, M4/M5 French-American-British classification, and CD14+ immunophenotype tracked with sensitivity to DORA and resistance to VEN but were mitigated with the combination. Increased expression of MAPK14 and BCL2, the respective primary targets of DORA and VEN, were observed in monocytic and undifferentiated leukemias, respectively. Enrichment for DORA and VEN sensitivities was observed in AML with monocyte-like and progenitor-like transcriptomic signatures, respectively, and these associations diminished with the combination. The mechanism underlying the combination’s enhanced efficacy may result from inhibition of p38 MAPK-mediated phosphorylation of BCL2, which in turn enhances sensitivity to VEN. These findings suggest exploiting complementary drug sensitivity profiles with respect to leukemic differentiation state, such as dual targeting of p38 MAPK and BCL2, offers opportunity for broad, enhanced efficacy across the clinically challenging heterogeneous landscape of AML.

Chimeric antigen receptor T-cell therapy (CART) has extended survival of patients with relapsed/refractory diffuse large B-cell lymphoma (DLBCL). However, limited durability of response and prevalent toxicities remain problematic. Identifying patients at high risk of disease progression, toxicity, and death would inform treatment decisions. Although the cumulative illness rating scale (CIRS) has been shown to correlate with survival in B-cell malignancies, no prognostic score has been independently validated in CART recipients. We retrospectively identified 577 patients with relapsed/refractory DLBCL indicated for CART at 9 academic centers to form a learning cohort (LC). Random survival forest modeling of overall survival (OS) and progression-free survival (PFS) was performed to determine the most influential CIRS organ systems and severity grades. The presence of a severe comorbidity (CIRS score ≥3) in the respiratory, upper gastrointestinal, hepatic, or renal system - herein termed "Severe4" - had the greatest impact on post-CART survival. Controlling for other prognostic factors (number of prior therapies, Eastern Cooperative Oncology Group performance status, BCL6 translocation, molecular subtype), Severe4 was strongly associated with shorter PFS and OS in the LC (hazards ratio [HR]=2.15 and 1.94, respectively; p

To understand mechanisms of response to BET inhibitors (BETi), we mined the Beat AML functional genomic data set and performed genome-wide CRISPR screens on BETi-sensitive and BETi-resistant acute myeloid leukemia (AML) cells. Both strategies revealed regulators of monocytic differentiation—SPI1, JUNB, FOS, and aryl-hydrocarbon receptor signaling (AHR/ARNT)—as determinants of BETi response. AHR activation synergized with BETi, whereas inhibition antagonized BETi-mediated cytotoxicity. Consistent with BETi sensitivity dependence on monocytic differentiation, ex vivo sensitivity to BETi in primary AML patient samples correlated with higher expression of the monocytic markers CSF1R, LILRs, and VCAN. In addition, HL-60 cell line differentiation enhanced its sensitivity to BETi. Further, screens to rescue BETi sensitivity identified BCL2 and CDK6 as druggable vulnerabilities. Finally, monocytic AML patient samples refractory to venetoclax ex vivo were significantly more sensitive to combined BETi + venetoclax. Together, our work highlights mechanisms that could predict BETi response and identifies combination strategies to overcome resistance. Significance: Drug resistance remains a challenge for AML, and new therapies, such as BETi, will require combination approaches to boost single-agent responses. We conducted genome-wide CRISPR screens and functional genomics on AML patient samples to identify leukemic differentiation state and AHR signaling as primary mediators of BETi response. This article is highlighted in the In This Issue feature, p. 403

The characterization of cancer genomes has provided insight into somatically altered genes across tumors, transformed our understanding of cancer biology, and enabled tailoring of therapeutic strategies. However, the function of most cancer alleles remains mysterious, and many cancer features transcend their genomes. Consequently, tumor genomic characterization does not influence therapy for most patients. Approaches to understand the function and circuitry of cancer genes provide complementary approaches to elucidate both oncogene and non-oncogene dependencies. Emerging work indicates that the diversity of therapeutic targets engendered by non-oncogene dependencies is much larger than the list of recurrently mutated genes. Here we describe a framework for this expanded list of cancer targets, providing novel opportunities for clinical translation.

NPM1 mutations are among the most common mutations in CN-AML (Cytogenetically Normal-AML) and are known to enhance chemosensitivity in AML patients. Across multiple co-operative group studies, in the absence of deleterious mutations, patients with NPM1 mutations show superior overall and relapse-free survival. The FOXM1 regulatory network is a major predictor of poor prognosis in human cancers of different origin, including AML. Early AML cell line and patient sample data from our lab demonstrated that nuclear NPM colocalizes with nuclear FOXM1 while mutant NPM sequesters FOXM1 in the cytoplasm. Applying RNAseq differential expression results from published FOXM1 knockout experiments on KG-1 cells and MLL-AF9-transformed mice to Beat AML RNAseq data, we developed a ~200-gene FOXM1 signature that was significantly associated with inferior CR/CRi rate (OR=0.44 [95% CI: 0.21–0.95], p=0.037) in wild type NPM1, wild type FLT3 de novo AML patients. Notably, this FOXM1 signature was not significantly associated with CR/CRi in mutated NPM1, wild type FLT3 patients (OR=1.78 [95% CI: 0.15–21.51], p=0.651). These data suggest that inactivation of FOXM1 may underlie the chemosensitivity conferred by the highly prevalent NPM1 mutation in favorable risk AML. Using our novel network-centric approach, we found a first-in-class small molecule (STL427944) that affects a pathway that limits FOXM1 expression and significantly disrupts the associated regulatory network. To directly address the role of FOXM1 in chemosensitivity of AML cells, we tested two FOXM1-specific inhibitory compounds: STL427944 and its derivative, STL001 (their structures will be disclosed). From a medicinal chemistry standpoint, STL427944 has two main metabolic liabilities, a hydrolytically labile furane-carboxylic acid phenolic ester and a hydrazone. We applied several paths of structural activity relationship (SAR) optimization to overcome these issues from removal of liabilities to bio-isosteric replacement to cyclizations. The ring replacement is likely to have significantly improved the overall stability observed in STL001, resulting in at least !0-fold more active compound with better ‘drug-like’ properties and thus enhanced potency. We showed that 500nM of STL001 partially suppressed expression of FOXM1 protein in AML, while 5mM of STL001 (versus 50mM of STL427944) fully suppressed FOXM1 and sensitized AML cells to venetoclax and cytarabine treatment. Ex vivo treatment of AML patient samples with STL427944 and STL001 resulted in inhibition of nuclear FOXM1 expression according to IHC and downregulation of canonical FOXM1 transcriptional targets as indicated by real-time PCR. This treatment potentiated the efficacy of venetoclax in inhibiting colony forming activity in AML patient samples. Our data suggest that targeting FOXM1 in AML patients with wild type NPM1 may recapitulate NPM1 mutations and will have a robust therapeutic effect leading to more favorable outcome for AML patients. Citation Format: Irum Khan, Sandjeev Raghuwanshi, Andy Kaempf, Jeff Tyner, Alex Domling, Carlos Camacho, Andrei Gartel. Targeting of FOXM1 recapitulates NPM1 mutations in chemo-sensitization of AML [abstract]. In: Proceedings of the AACR Special Conference: Acute Myeloid Leukemia and Myelodysplastic Syndrome; 2023 Jan 23-25; Austin, TX. Philadelphia (PA): AACR; Blood Cancer Discov 2023;4(3_Suppl):Abstract nr A09.

Significance Acute myeloid leukemia (AML) is the most common acute leukemia in adults, with a 5-y survival of 29%. Immunotherapy is based on the premise that tumors suppress the immune system. We investigated the status of T cell immunity in AML at the time of diagnosis. We found a significant association between T cell percentage in the bone marrow and overall survival in newly diagnosed AML patients. When we evaluated the T cells from the bone marrow of patients with AML, one-third displayed profound functional impairment. Most of these compromised T cells, however, could be rescued using checkpoint inhibitors. Our results support the development of immune checkpoint therapy to combat this deadly disease., Acute myeloid leukemia (AML) is the most common acute leukemia in adults, with approximately four new cases per 100,000 persons per year. Standard treatment for AML consists of induction chemotherapy with remission achieved in 50 to 75% of cases. Unfortunately, most patients will relapse and die from their disease, as 5-y survival is roughly 29%. Therefore, other treatment options are urgently needed. In recent years, immune-based therapies have led to unprecedented rates of survival among patients with some advanced cancers. Suppression of T cell function in the tumor microenvironment is commonly observed and may play a role in AML. We found that there is a significant association between T cell infiltration in the bone marrow microenvironment of newly diagnosed patients with AML and increased overall survival. Functional studies aimed at establishing the degree of T cell suppression in patients with AML revealed impaired T cell function in many patients. In most cases, T cell proliferation could be restored by blocking the immune checkpoint molecules PD-1, CTLA-4, or TIM3. Our data demonstrate that AML establishes an immune suppressive environment in the bone marrow, in part through T cell checkpoint function.

External beam radiation therapy (EBRT) is a safe and emerging bridging liver-directed therapy (LDT) to liver transplant (LT) for patients with hepatocellular carcinoma (HCC). The prevalence and clinical characteristics of patients receiving EBRT as an LDT for LT have not been evaluated. Our aim was to describe the utilization of EBRT in patients with HCC evaluated for LT in the United States.We analyzed United Network for Organ Sharing data from October 2013 to June 2020 and identified patients with HCC who applied for model for end-stage liver disease (MELD) exceptions for LT wait list prioritization. The primary outcome was the period prevalence of EBRT. We examined associations between clinical variables and EBRT and fit survival models with EBRT as a time-varying predictor.We identified 18,543 patients with HCC with MELD exception applications. EBRT was used in 658 patients (3.5%) either alone (1.2%) or combined with other LDT (2.3%). Transarterial chemoembolization was the most used LDT (59.3%), followed by thermal ablation (27.9%) and radioembolization (15.2%). EBRT prevalence rose by an average of 12.2% per year (P = .001). Use of EBRT differed by geographic region, ranging from 2% to 8% (P.001). EBRT and no EBRT groups had similar initial MELD score, portal vein thrombosis, tumor diameter, number of tumors, bilirubin, and α-fetoprotein (P.05). Median time-to-transplant from wait list registration for EBRT versus no EBRT groups was 10 months (95% confidence interval, 9.4-10.9) versus 11.9 months (95% confidence interval, 11.7-12.2; P.001). Evaluated as a time-varying predictor, EBRT increased the risk of LT by 30% (sub-hazard ratio, 1.30; P.001), while the effect of EBRT on the risk of wait list removal due to clinical deterioration or death (sub-hazard ratio, 1.07; P = .489) was nonsignificant.In the United States, EBRT is rarely used compared with other LDTs and exhibits geographic variation. Low EBRT utilization highlights a gap in the treatment armamentarium for HCC.

Introduction: CAR T-cell therapy (CART) has dramatically improved outcomes for patients (pts) with relapsed/refractory (r/r) DLBCL, but the majority of pts still have poor outcomes due to progressive disease and toxicities. Here we used a machine learning algorithm to rank the prognostic impact of specific comorbidities measured by the Cumulative Illness Rating Scale (CIRS) in DLBCL pts indicated for CART in a multicenter learning cohort (LC) and a separate validation cohort (VC). Methods: pts with r/r DLBCL included in RWE analysis underwent leukapheresis for CART at 10 academic centers. CIRS was assessed at the time of T-cell collection (Salvi et al, 2008). Progression-free survival (PFS) and overall survival (OS) were measured from T-cell collection. Random survival forest (RSF) modeling of PFS and OS was repeatedly applied to random subsets of the LC to determine the most important CIRS categories and comorbidity levels in the presence of known prognostic factors. Cox proportional hazards models of PFS and OS were fit to both cohorts. Associations between comorbidities and CART adverse events were evaluated with Fisher’s exact test. Results: The LC comprised 577 pts, median age of 63 (range, 19-90); 90% had ECOG 0-1. Median number of prior therapies was 3 (range, 1-11). GCB subtype was found in 54% of pts, with 38% non-GCB and 8% unknown. Twenty-seven pts (4.7%) died before CART infusion. Of the 550 pts who received CART, 71% got axicabtagene ciloleucel, 22% tisagenlecleucel, and 7% lisocabtagene maraleucel. The median CIRS score was 7 (range, 0-25) with 54% having CIRS ≥7. The median PFS was 11 months (95% CI: 8 – 15) and OS 30 months (95% CI: 23 – NA), with a median follow-up time of 20 months. Although CIRS ≥7 was significantly associated with PFS (HR=1.26) and OS (HR=1.35) in univariable analysis, it did not remain significant in multivariable models. According to a RSF tree depth-weighted nodal split score, severe comorbidities in the following CIRS categories had the greatest impact on PFS: respiratory, upper GI, renal, and hepatic (denoted Severe4, 9% of pts). When accounting for other significant variables, Severe4 was independently associated with inferior PFS (HR=2.45, p Citation Format: Geoffrey Shouse, Andy Kaempf, Max Gordon, David Yashar, Audrey M Sigmund, Gordon Smilnak, Steven M Bair, Agrima Mian, Lindsey Fitzgerald, Amneet Bajwa, Samantha Jaglowski, Neil Bailey, Mayzar Shadman, Krish Patel, Deborah M Stephens, Manali Kamdar, Brian T Hill, Jordon Gauthier, Reem Karmali, Loretta Nastoupil, Adam S Kittai, Alexey V Danilov. Impact of a validated composite comorbidity score on outcomes in patients treated with CAR T-cell therapy for diffuse large B cell lymphoma (DLBCL): A multicenter real-world evidence (RWE) study [abstract]. In: Proceedings of the Third AACR International Meeting: Advances in Malignant Lymphoma: Maximizing the Basic-Translational Interface for Clinical Application; 2022 Jun 23-26; Boston, MA. Philadelphia (PA): AACR; Blood Cancer Discov 2022;3(5_Suppl):Abstract nr A27.

Introduction: Medical comorbidities influence survival in CLL. We previously reported on a simplified CLL-specific comorbidity scale, the CLL-CI (Gordon et al. 2019), which required assessment of only three organ systems and was predictive of outcomes in a heterogeneous retrospective patient cohort. Herein we analyzed CLL-CI among patients treated with ibrutinib. Methods: This retrospective study included 339 CLL patients treated with ibrutinib at 9 academic centers between 2014-2019. Vascular, endocrine and upper-gastrointestinal organ systems were assessed at the time of ibrutinib initiation. Each was scored from 0 to 3, in order of increasing severity of dysfunction to generate the CLL-CI score (range, 0-9; Figure A). As established previously, CLL-CI≥2 was deemed high-risk. Event free survival (EFS) was measured from start of ibrutinib to development of new CLL-related symptoms, disease progression, start of a new therapy or death. Overall survival (OS) was measured from treatment initiation to death. Patients with no EFS or OS events were censored at last follow up. The Kaplan-Meier method and log-rank test were used to estimate and compare survival. Multivariable Cox regression was utilized to model EFS and OS. Differences between CLL-CI groups were evaluated with Wilcoxon rank sum and Fisher's exact tests. Results: Median age was 68 years (range, 30-91), 240 (71%) were treated in the relapsed/refractory setting (range of prior therapies, 0-10). Advanced Rai stage (3-4) was present in 206 (61%) and TP53 aberrancy was present in 93 (27%) patients at the start of ibrutinib therapy. Median follow up was 23 months (range, 1-71). CLL-CI score was high (≥2) in 202/339 patients (60%). Patient characteristics were well balanced between subgroups (CLL-CI In multivariate models adjusted for age, del(17p) and relapsed disease, high CLL-CI was associated with shortened EFS (HR=1.65; p=0.014, Figure) and OS (HR=1.73; p=0.1). CIRS score≥7 also correlated with EFS (HR=1.91; p=0.002) and OS (HR=2.78; p=0.006). Ibrutinib discontinuation rates due to adverse events were more frequent in patients with CLL-CI ≥2 (25% vs 14%; p=0.014). However, dose reduction rates were similar (24% vs 20%; p=0.51). Fifty-two deaths occurred: 40 in the high CLL-CI subgroup and 12 in the low CLL-CI subgroup. Cause of death was known in 31 patients. Death due to disease progression was more frequent in the high CLL-CI subgroup (28% vs 8%; p Since some of the key ibrutinib toxicities (atrial fibrillation, hypertension) may not have been accounted for in the CLL-CI we further elucidated their possible impact. Cardiac disease was significantly more prevalent among patients with high CLL-CI (37% vs 16%, p Conclusions: Here we present the largest cohort of CLL patients treated with ibrutinib in whom comorbidities have been systematically assessed. We find that the CLL-CI (which assesses endocrine, vascular and upper gastrointestinal conditions) correlates with survival and tolerance of therapy in this population. Unexpectedly, we found that hypertension and cardiac comorbidities did not improve CLL-CI's discriminatory power. This result combined with the simplicity of scoring the CLL-CI makes it an attractive tool for clinical practice. CLL-CI needs to be explored prospectively in patients treated with ibrutinib and other targeted therapies. Disclosures Patel: Genentech: Consultancy, Speakers Bureau; BeiGene: Consultancy; Celgene/BMS: Consultancy, Membership on an entity's Board of Directors or advisory committees; Janssen: Consultancy, Speakers Bureau; Adaptive Biotechnologies: Consultancy; AstraZeneca: Consultancy, Research Funding, Speakers Bureau; Pharmacyclics: Consultancy, Speakers Bureau; Kite: Consultancy. Cohen:Janssen, Adicet, Astra Zeneca, Genentech, Aptitude Health, Cellectar, Kite/Gilead, Loxo: Consultancy; Genentech, BMS, Novartis, LAM, BioInvent, LRF, ASH, Astra Zeneca, Seattle Genetics: Research Funding. Choi:Pharmacyclics/Abbvie: Research Funding; Genentech: Consultancy. Hill:BMS: Consultancy, Honoraria, Research Funding; Celgene: Consultancy, Honoraria, Research Funding; Kite, a Gilead Company: Consultancy, Honoraria, Research Funding; Novartis: Consultancy, Honoraria; Abbvie: Consultancy, Honoraria, Research Funding; Takeda: Research Funding; Beigene: Consultancy, Honoraria, Research Funding; AstraZenica: Consultancy, Honoraria, Research Funding; Genentech: Consultancy, Honoraria, Research Funding; Karyopharm: Consultancy, Honoraria, Research Funding; Pharmacyclics: Consultancy, Honoraria, Research Funding. Shadman:Mustang Bio, Celgene, Pharmacyclics, Gilead, Genentech, Abbvie, TG therapeutics, Beigene, Astra Zeneca, Sunesis, Beigene: Research Funding; Abbvie, Genentech, Astra Zeneca, Sound Biologics , Pharmacyclics, Verastem, ADC therapeutics, Beigene, Cellectar, BMS, Morphosys and Atara Biotherapeutics: Consultancy. Stephens:Innate: Consultancy; Gilead: Research Funding; Verastem: Research Funding; Janssen: Consultancy; Acerta: Research Funding; Pharmacyclics: Consultancy; MingSight: Research Funding; Beigene: Consultancy; Arqule: Research Funding; Juno: Research Funding; Karyopharm: Consultancy, Research Funding. Brander:Novartis: Consultancy, Other; NCCN: Other; Tolero: Research Funding; AstraZeneca: Consultancy, Honoraria, Other, Research Funding; Ascentage: Other, Research Funding; ArQule: Consultancy, Other, Research Funding; AbbVie: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other, Research Funding; Tolero: Research Funding; Teva: Consultancy, Honoraria; Novartis: Consultancy, Other; NCCN: Other; Verastem: Consultancy, Honoraria, Other, Research Funding; TG Therapeutics: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other, Research Funding; Pfizer: Consultancy, Other; Pharmacyclics LLC, an AbbVie Company: Consultancy, Honoraria, Other, Research Funding; MEI Pharma: Other, Research Funding; Juno/Celgene/BMS: Other, Research Funding; Genentech: Consultancy, Honoraria, Other, Research Funding; DTRM: Other, Research Funding; BeiGene: Other, Research Funding; Teva: Consultancy, Honoraria. Danilov:BeiGene: Consultancy; Pharmacyclics: Consultancy; Abbvie: Consultancy; Bristol-Myers Squibb: Research Funding; Rigel Pharmaceuticals: Consultancy; Astra Zeneca: Consultancy, Research Funding; Aptose Biosciences: Research Funding; Verastem Oncology: Consultancy, Research Funding; Takeda Oncology: Research Funding; Gilead Sciences: Research Funding; Bayer Oncology: Consultancy, Research Funding; Genentech: Consultancy, Research Funding; TG Therapeutics: Consultancy; Nurix: Consultancy; Celgene: Consultancy; Karyopharm: Consultancy.

1054 Background: Combination of CDKi with endocrine therapy is a key treatment for mHRBC due to survival benefit and favorable safety profile. However, progressive disease inevitably develops and outcomes after CDKi discontinuation (dc) are not well-described. Within our institution, we previously reported clinical characteristics and outcomes for a cohort of 140 mHRBC patients who received CDKi therapy. Median progression-free survival (PFS) and overall survival (OS) post-CDKi dc were 7.0 and 15.4 months, respectively. However, 29% experienced rapid progression or death within 4 months following CDKi dc. Molecular predictors of rapid progression after CDKi are unknown and may help define therapies to improve outcomes. In this study, we sought to identify molecular predictors for rapid disease progression after CDKi dc in mHRBC. Methods: We identified within our cohort 34 patients with mHRBC who progressed on CDKi with next-generation sequencing (NGS) performed on pre-CDKi tissue samples. PFS and OS, measured from CDKi dc, were analyzed with the Kaplan-Meier estimator and log-rank test. Rapid progression was analyzed with logistic regression and Fisher’s exact test to evaluate association between pre-CDKi tumor mutation and rapid progression post-CDKi. Results: NGS of pre-CDKi tumor biopsies found 12 genes ( FGF3, FGF4, FGFR, PIK3CA, PTEN, AKT, RB1, CDKN2A, MYC, CCND1, ESR1, TP53) that were altered in ≥3 of the 34 patients. The six patients (18%) with a PTEN mutation (mut) had a median PFS of 3 months and median OS of 4 mo. In comparison, median PFS and OS of PTEN wild-type (wt) patients were 7 mo. (log-rank p=0.008) and 21 mo. (log-rank p

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Our study details the stepwise evolution of gilteritinib resistance in FLT3-mutated acute myeloid leukemia (AML). Early resistance is mediated by the bone marrow microenvironment, which protects residual leukemia cells. Removing these supportive extrinsic ligands drives evolution of late, intrinsic resistance. Whole exome sequencing, CRISPR/Cas, metabolomics, proteomics, and pharmacologic approaches were used to mechanistically define both early and late resistance. Early resistant cells undergo metabolic reprogramming, grow more slowly, and are dependent upon Aurora kinase B (AURKB). Late resistant cells are characterized byexpansion of pre-existing NRAS mutant subclones and continued metabolic reprogramming. Our model closely mirrors the timing and mutations of AML patients treated with gilteritinib.Pharmacological inhibition of AURKB resensitized both early resistant cell cultures and primary leukemia cells from gilteritinib-treated AML patients. These findings support a combinatorial strategy to target early resistant AML cells with AURKB inhibitors and gilteritinib before the expansion of pre-existing resistance mutations occurs.

Background The combination of everolimus (EVE) and exemestane (EXE) is approved for the treatment of patients with metastatic hormone receptor-positive breast cancer (mHRBC) who progress on nonsteroidal aromatase inhibitor (NSAI) therapy. However, none of the patients enrolled in the trial that led to this approval (BOLERO-2) had previously received CDK4/6 inhibitors (CDK4/6is), which have since become a frontline standard of care for mHRBC. As such, the clinical benefit of EVE plus EXE in patients who have previously received CDK4/6is remains unknown. Materials and Methods Adult patients with mHRBC at our institution who progressed on an NSAI plus CDK4/6i or NSAI therapy alone and were treated with at least one cycle of EVE plus EXE between 2012 and 2018 were analyzed. Collected data included patient demographics, treatment history, adverse events, and clinical outcomes. Primary objectives were to compare progression-free survival (PFS) and overall survival (OS) between patients who received prior NSAI plus CDK4/6i therapy versus an NSAI alone. Results Among 43 patients, 17 had prior CDK4/6i exposure. With the exception of de novo metastatic disease, patient and disease characteristics were comparable across treatment cohorts. There was no significant difference in PFS (median, 3.6 vs. 4.2 months) or OS (median, 15.6 vs. 11.3 months) between patients who had received prior CDK4/6is and those who had not, respectively. Conclusion Prior exposure to CDK4/6i therapy did not impact survival outcomes for patients with mHRBC taking EVE plus EXE. However, there was a trend toward improved OS in the CDK4/6i cohort that should be evaluated in larger cohorts. Implications for Practice The use of CDK4/6 inhibitors in combination with a nonsteroidal aromatase inhibitor has become a standard frontline therapy in metastatic hormone receptor-positive breast cancer. An approved subsequent line of therapy is everolimus plus exemestane; however, the original data supporting this therapy predated approval of CDK4/6 inhibitors. As such, the clinical benefit of everolimus and exemestane in patients previously treated with a CDK4/6 inhibitor was unknown. This retrospective cohort study offers real-world data demonstrating prior CDK4/6 inhibitor exposure does not impact survival outcomes for everolimus plus exemestane.

Purpose Cyclin-dependent kinase (CDK) 4/6 inhibitors are integral treatment for advanced hormone receptor positive breast cancer; however, venous thromboembolic events (VTE) occurred in 1%-5% of clinical trial patients. Thrombosis rates in the real-world setting remain unclear. We aimed to define the rate of thromboembolic events, risk factors for thrombosis on CDK 4/6 inhibitors and evaluate the Khorana VTE risk score as a predictive tool for VTE in patients on CDK 4/6 therapy. Methods Multicenter retrospective analysis of adult breast cancer patients prescribed palbociclib, ribociclib, or abemaciclib. The primary endpoint was thrombosis during treatment or within 30 days of CDK inhibitor discontinuation. Cox regression was used to model time-to-thrombosis, starting from a patient's initiation of CDK 4/6 therapy. The extended Kaplan-Meier method and Cox modeling were used to assess the effect of time-varying thrombosis status on overall survival. Results Two hundred and sixty-six patients were included (89% on palbociclib, 14% on abemaciclib, 7% on ribociclib). Twenty-nine thrombotic events occurred in 26 (9.8%) women. Of these events, 72% were venous and 34% were arterial. The 1-year incidence of thrombosis was 10.4% overall, 10.9% on palbociclib, 8.3% on ribociclib, and 4.8% on abemaciclib. Hemoglobin less than 10 g/dL was a statistically significant predictor of thrombosis (HR 3.53, P: .014). Khorana score ranged from 0-3, with the majority between 0 and 1 and was not predictive of VTE. Thrombosis was associated with reduced overall survival (HR 1.28, P: .128, median 7.3 months) compared to not having a CDK-associated clot (median 35.7 months). Discussion VTE in our analysis is higher than reported in clinical trials and arterial thrombosis comprised over one-third of events. The highest incidence was with palbociclib, followed by ribociclib. Khorana score did not predict VTE risk. Larger, real-world studies are needed. The role for prophylactic anticoagulation is yet to be defined in this patient population.

Purpose: Comorbid medical conditions define a subset of patients with chronic lymphocytic leukemia (CLL) with poor outcomes. However, which comorbidities are most predictive remains understudied. Experimental Design: We conducted a retrospective analysis from 10 academic centers to ascertain the relative importance of comorbidities assessed by the cumulative illness rating scale (CIRS). The influence of specific comorbidities on event-free survival (EFS) was assessed in this derivation dataset using random survival forests to construct a CLL-specific comorbidity index (CLL-CI). Cox models were then fit to this dataset and to a single-center, independent validation dataset. Results: The derivation and validation sets comprised 570 patients (59% receiving Bruton tyrosine kinase inhibitor, BTKi) and 167 patients (50% receiving BTKi), respectively. Of the 14 CIRS organ systems, three had a strong and stable influence on EFS: any vascular, moderate/severe endocrine, moderate/severe upper gastrointestinal comorbidity. These were combined to create the CLL-CI score, which was categorized into 3 risk groups. In the derivation dataset, the median EFS values were 58, 33, and 20 months in the low, intermediate, and high-risk groups, correspondingly. Two-year overall survival (OS) rates were 96%, 91%, and 82%. In the validation dataset, median EFS values were 81, 40, and 23 months (two-year OS rates 97%/92%/88%), correspondingly. Adjusting for prognostic factors, CLL-CI was significantly associated with EFS in patients treated with either chemo-immunotherapy or with BTKi in each of our 2 datasets. Conclusions: The CLL-CI is a simplified, CLL-specific comorbidity index that can be easily applied in clinical practice and correlates with survival in CLL.

Mechanisms of immune suppression in peripheral tissues counteract protective immunity to prevent immunopathology and are coopted by tumors for immune evasion. While lymphatic vessels facilitate T cell priming, they also exert immune suppressive effects in lymph nodes at steady-state. Therefore, we hypothesized that peripheral lymphatic vessels acquire suppressive mechanisms to limit local effector CD8+ T cell accumulation in murine skin. We demonstrate that nonhematopoietic PD-L1 is largely expressed by lymphatic and blood endothelial cells and limits CD8+ T cell accumulation in tumor microenvironments. IFNγ produced by tissue-infiltrating, antigen-specific CD8+ T cells, which are in close proximity to tumor-associated lymphatic vessels, is sufficient to induce lymphatic vessel PD-L1 expression. Disruption of IFNγ-dependent crosstalk through lymphatic-specific loss of IFNγR boosts T cell accumulation in infected and malignant skin leading to increased viral pathology and tumor control, respectively. Consequently, we identify IFNγR as an immunological switch in lymphatic vessels that balances protective immunity and immunopathology leading to adaptive immune resistance in melanoma.

The implementation of targeted therapies for acute myeloid leukaemia (AML) has been challenging because of the complex mutational patterns within and across patients as well as a dearth of pharmacologic agents for most mutational events. Here we report initial findings from the Beat AML programme on a cohort of 672 tumour specimens collected from 562 patients. We assessed these specimens using whole-exome sequencing, RNA sequencing and analyses of ex vivo drug sensitivity. Our data reveal mutational events that have not previously been detected in AML. We show that the response to drugs is associated with mutational status, including instances of drug sensitivity that are specific to combinatorial mutational events. Integration with RNA sequencing also revealed gene expression signatures, which predict a role for specific gene networks in the drug response. Collectively, we have generated a dataset—accessible through the Beat AML data viewer (Vizome)—that can be leveraged to address clinical, genomic, transcriptomic and functional analyses of the biology of AML. Analyses of samples from patients with acute myeloid leukaemia reveal that drug response is associated with mutational status and gene expression; the generated dataset provides a basis for future clinical and functional studies of this disease.

The biological response of normal tissue to high-dose radiation treatment remains poorly understood. Alterations to the microenvironment, specifically the microvasculature, have been implicated as a significant contributor to tumoral cytotoxicity. We used contrast-enhanced ultrasound (CEU) perfusion imaging, which is uniquely suited to assess functional status of the microcirculation, to measure microvascular blood flow after high-dose irradiation to normal skeletal muscle tissue in a murine model. Proximal hindlimbs of wild-type C57Bl/6 mice were irradiated with a single fraction using 6 MV photons, 1 cm bolus and a dynamic wedge. Quantitative perfusion CEU imaging of the skeletal muscle was performed at days 1 and 8 postirradiation in three different regions of interest (ROIs): 1. 15 Gy external-beam irradiated leg; 2. 12 Gy irradiated 5 mm proximal area; 3. single ROI in the nonirradiated contralateral (CL) hindlimb. Perfusion imaging was also performed in the hindlimb of nonirradiated mice. CEU time-intensity data were analyzed to measure microvascular blood flow (MBF, also referred to as perfusion), and its parametric components of microvascular flux rate and functional microvascular blood volume (MBV). Plasma measurements of two potent vasoconstrictors, endothelin-1 and angiotensin II, were also performed to assess systemic response. CEU perfusion imaging values for the 12 and 15 Gy irradiated limb regions were pooled. At day 1, MBF in the irradiated limb was significantly lower than in the CL limb (P = 0.016) but quite similar to the nonirradiated mice. At day 8, both limbs of irradiated mice exhibited a trend towards lower MBF than the limbs of nonirradiated mice (28% decrease in mean MBF, P = 0.149 for CL; 39% decrease, P = 0.065 for irradiated limb). Compared to nonirradiated animals, the reduction in perfusion in irradiated limbs at day 8 may have been more influenced by the microvascular flux rate (25% decrease in the mean, P = 0.079) than the MBV (12% decrease in the mean, P = 0.328). Examination of vasoactive compounds revealed that the average plasma concentration for endothelin-1 at day 8 postirradiation was significantly higher in 14 irradiated animals than in 4 nonirradiated animals (3.07 pg/ ml vs. 2.51 pg/ml; P = 0.011). Up to day 8 after high-dose irradiation, flow deficits in irradiated muscle appear to be a consequence of increased vascular resistance more so than loss or functional de-recruitment of microvascular units.

Introduction Approximately 25-30% of adult acute myeloid leukemias (AMLs) have FLT3 mutation. Among the two main types, internal tandem duplication (ITD) and tyrosine kinase domain (TKD) point mutations, FLT3-ITD is associated with poorer clinical outcomes. Recently, midostaurin was found to improve survival when combined with 7+3 induction for FLT3-ITD or FLT3-TKD AML. Additionally, increased CD33 expression in AML patient samples has been linked to FLT3-ITD+ blasts. This suggests the combined targeting of CD33 and FLT3 in AML patients as a novel treatment approach. Hence, we introduce a combination therapy of gemtuzumab ozogamicin (GO) - a humanized CD33 antibody conjugated to a calicheamicin derivative - and midostaurin with 7+3 (cytarabine + daunorubicin) for newly diagnosed FLT3-mutated AML. Methods This Phase I open-label, dose-finding study was designed to determine the maximum tolerated dose schedule of GO administered with midostaurin and 7+3 induction and to assess the safety and preliminary efficacy of this combination. Patients needed to be >18 years old, have newly diagnosed AML, be fit for 7+3 induction, and harbor a FLT3-ITD or -TKD mutation per next-generation sequencing or PCR testing. In cohorts of 3, patients were assigned to 1 of 4 dose levels according to the "keyboard" Bayesian toxicity probability interval design (Table 1) applied to a target dose-limiting toxicity (DLT) rate of 20%. Non-hematologic DLTs were defined as study drug-related grade ≥3 toxicities, with exceptions for infections, elevated liver enzymes (that resolve in ≤5 days), GI symptoms (≤3 days), and electrolyte abnormalities (≤1 day). Hematologic DLTs were grade 4 neutropenia or grade ≥3 thrombocytopenia at 6 weeks after the start of the induction cycle and in the absence of AML. The DLT evaluation period covered the induction and, if applicable, re-induction cycles. Clinical responses were determined by 2017 ELN criteria. CD33 expression on AML blasts was quantified before and after treatment. Upon completion of 1 or 2 induction cycles, patients with a CR or CRi could receive up to 2 cycles of consolidation therapy, consisting of cytarabine (HiDAC or MiDAC depending on age Results Eight patients have been enrolled out of the planned accrual of 24. The median age was 59 (range: 35-72) years, and every patient identified as White and Non-Hispanic. At screening, all patients had FLT3-ITD mutations. The median bone marrow blast percentage was 64% (range 17%-91%), with the majority of blasts expressing CD33 (median 96%, range 80-100%, 2 patients missing data due to recent enrollment). The current numbers of patients treated by dose level (DL) were: 3 on DL1 (IV GO 3 mg/m 2 Day 1), 3 on DL2 (IV GO 3 mg/m 2 Day 1 and 4), and 2 on DL3 (IV GO 3 mg/m 2 Day 1, 4, and 7). No DLTs were observed among the six patients in DL1 or DL2. Likewise, no DLTs were observed in the two patients in DL3. The most common treatment-emergent adverse events included grade 3 febrile neutropenia (75% of patients), grade 3/4 mucositis (25%), grade 3 sepsis (25%), and grade 3 esophagitis (12.5%). One patient had a serious adverse event: a GO-related Grade 4 sinusoidal obstruction syndrome occurring ~4 months into treatment and lasting 23 days. The median duration of study therapy was 62 days (range 20-95 days); reasons for discontinuation were disease progression (n=2), non-compliance (n=1), and allogeneic HCT (n=5). The overall response rate and composite complete remission rate were both 75% (95% CI: 34.9% - 96.8%). There were no treatment-related deaths in the first 30 days. Due to the early nature of this study, survival outcomes have not been calculated and post-induction CD33 expression data are not available. Conclusions In newly diagnosed FLT3-mutated AML patients, induction chemo/immunotherapy with GO, midostaurin, and 7+3 has yielded promising responses and has been well tolerated with no DLTs thus far. Additional patients and correlative analyses of CD33 expression will provide further insight into the safety and efficacy of this regimen for a patient population with historically poor prognosis. Figure 1 Figure 1. Disclosures Borate: Pfizer: Membership on an entity's Board of Directors or advisory committees, Research Funding; Blueprint Medicine: Membership on an entity's Board of Directors or advisory committees; Jazz Pharma: Research Funding; Genentech: Membership on an entity's Board of Directors or advisory committees, Other: Advisory Board; AbbVie: Membership on an entity's Board of Directors or advisory committees, Research Funding; Daiichi-Sankyo: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding; Takeda: Membership on an entity's Board of Directors or advisory committees; Astellas: Membership on an entity's Board of Directors or advisory committees; incyte: Membership on an entity's Board of Directors or advisory committees, Research Funding; Rampal: Membership on an entity's Board of Directors or advisory committees; Galecto, Inc.: Consultancy; Promedior: Consultancy. Saultz: IKENA: Research Funding. Traer: ImmunoGen: Membership on an entity's Board of Directors or advisory committees; Servier/Agios: Membership on an entity's Board of Directors or advisory committees; Incyte: Research Funding; Abbvie: Consultancy, Membership on an entity's Board of Directors or advisory committees; Schrodinger: Research Funding; Genentech: Membership on an entity's Board of Directors or advisory committees; Astellas: Consultancy, Membership on an entity's Board of Directors or advisory committees. Walter: Amgen: Research Funding; Aptevo: Consultancy, Research Funding; Celgene: Consultancy, Research Funding; Immunogen: Research Funding; Macrogenics: Consultancy, Research Funding; Jazz: Research Funding; Pfizer: Consultancy, Research Funding; Selvita: Research Funding; Amphivena: Consultancy, Other: ownership interests; Agios: Consultancy; Astellas: Consultancy; BMS: Consultancy; Genentech: Consultancy; Janssen: Consultancy; Kite: Consultancy.

Introduction: Comorbidities are common in CLL and associated with shorter overall survival (OS) and disease specific survival (Rotbain, Leukemia, 2021). Using an ensemble machine learning approach, we identified the comorbidities most strongly associated with event free survival (EFS) in CLL in order to construct the CLL-CI (Gordon et al, 2021). Our initial report included heterogeneously-treated derivation (N=570) and validation (N=167) cohorts of patients with CLL. Median EFS in the derivation cohort was 58, 33 and 20 months for low, intermediate and high-risk CLL-CI groups, respectively (p Methods: In our multicenter CLL cohort, we retrospectively analyzed patients from 10 academic centers who received targeted therapies, e.g. ibrutinib, acalabrutinib, idelalisib and venetoclax. Secondly, we now compiled a validation cohort which included patients from the Danish National CLL registry. CLL-CI score was assigned as previously reported (Gordon et al, 2021). Briefly, one point was assigned, if present, for any cumulative illness rating scale (CIRS) vascular comorbidity, moderate/severe upper gastrointestinal CIRS comorbidity and moderate/severe endocrine CIRS comorbidity, for a total maximum score of 3 (Fig. 1A). A score of 0 signifies low risk disease, 1-intermediated risk and 2-3-high risk. The Kaplan-Meier method and Cox models were used to estimate the association between CLL-CI, EFS (defined as death or next therapy) and OS. Results: CLL patients treated with targeted therapies (N=448) had a median age of 68 years (range, 26-91), 124 pts (28%) had TP53 aberrancy, 268 (60%) were treated in the relapsed/refractory (R/R) setting and 398 (89%) received ibrutinib. By CLL-CI group, 194 (43%) were low risk, 173 (39%) intermediate risk and 81 (18%) high risk. Median EFS was 57, 35 and 17 months for low, intermediate and high-risk groups, respectively (p The validation cohort (N=4975) included patients from the Danish CLL registry. Median age was 71 years, 1630 (33%) had low risk CLL-IPI and 425 (8.5%) had high or very-high risk CLL-IPI; 87% received chemoimmunotherapy. From time of diagnosis, in low, intermediate and high risk CLL-CI groups the median OS was not reached, 8.5 and 6 years, respectively (p Conclusions: Here we report that a novel comorbidity index, CLL-CI, predicts outcomes in patients with CLL treated with targeted therapies. Furthermore, we validate this index in a large National Registry. Thus, CLL-CI is a validated measure of comorbidity in CLL which provides important prognostic information for patients with early stage disease treated with a watch and wait approach and in patients with advanced stage disease receiving targeted therapies or chemoimmunotherapy. Figure 1 Figure 1. Disclosures Rotbain: Abbvie: Other: travel grants; AstraZeneca: Consultancy, Other: travel grants; Janssen: Other: travel grants . Shouse: Kite Pharma: Speakers Bureau; Beigene: Honoraria. Mei: Morphosys: Research Funding; BMS: Research Funding; Epizyme: Research Funding; TG Therapeutics: Research Funding; EUSA: Honoraria; Janssen: Honoraria; Beigene: Research Funding. Brander: Ascentage: Research Funding; Pfizer: Consultancy, Other: Biosimilars outcomes research panel; ArQule: Research Funding; DTRM: Research Funding; Genentech: Consultancy, Research Funding; BeiGene: Research Funding; Juno Therapeutics/Celgene/Bristol Myers Squibb: Research Funding; LOXO: Research Funding; MEI Pharma: Research Funding; TG Therapeutics: Consultancy, Research Funding; AstraZeneca: Research Funding; Verastem: Consultancy; NCCN: Other: panel member; Novartis: Research Funding; Pharmacyclics LLC, an AbbVie Company: Consultancy, Research Funding; AbbVie: Consultancy, Other: informCLL registry steering committee, Research Funding; ArQule/Merck: Consultancy. Hill: Incyte/Morphysis: Consultancy, Honoraria, Research Funding; Gentenech: Consultancy, Honoraria, Research Funding; Kite, a Gilead Company: Consultancy, Honoraria, Other: Travel Support, Research Funding; Celgene (BMS): Consultancy, Honoraria, Research Funding; Pfizer: Consultancy, Honoraria; Novartis: Consultancy, Honoraria, Research Funding; Karyopharm: Consultancy, Honoraria, Research Funding; AbbVie: Consultancy, Honoraria, Research Funding; AstraZenica: Consultancy, Honoraria; Epizyme: Consultancy, Honoraria; Beigene: Consultancy, Honoraria, Research Funding. Choi: Abbvie: Other: Institution: Research Grant/Funding; Pharmacyclics: Other: Institution: Research Grant/Funding; Oncternal: Other: Institution: Research Grant/Funding; Velosbio: Other: Institution: Research Grant/Funding; Merck: Other: Institution: Research Grant/Funding; Geron: Other: Institution: Research Grant/Funding. Cohen: Genentech, BMS/Celgene, LAM, BioINvent, LOXO, Astra Zeneca, Novartis, M2Gen, Takeda: Research Funding; Janssen, Adicet, Astra Zeneca, Genentech, Aptitude Health, Cellectar, Kite/Gilead, Loxo, BeiGene, Adaptive: Consultancy. Stephens: Abbvie: Consultancy; Novartis: Research Funding; JUNO: Research Funding; Mingsight: Research Funding; Beigene: Membership on an entity's Board of Directors or advisory committees; Arqule: Research Funding; Adaptive: Membership on an entity's Board of Directors or advisory committees; TG Therapeutics: Membership on an entity's Board of Directors or advisory committees; Epizyme: Membership on an entity's Board of Directors or advisory committees; Innate Pharma: Membership on an entity's Board of Directors or advisory committees; AstraZeneca: Consultancy; CSL Behring: Consultancy; Celgene: Consultancy; Karyopharm: Membership on an entity's Board of Directors or advisory committees, Research Funding. Patel: Kite Pharma: Consultancy, Speakers Bureau; MEI Pharma: Consultancy; TG Therapeutics: Consultancy, Speakers Bureau; Abbvie: Consultancy; Janssen: Consultancy; Genentech: Consultancy; Bristol Myers Squibb: Consultancy, Speakers Bureau; BeiGene: Consultancy; Morphosys: Consultancy; Pharmacyclics: Consultancy; AstraZeneca: Consultancy, Research Funding, Speakers Bureau; ADC Therapeutics: Consultancy; Lilly: Consultancy. Shadman: Mustang Bio, Celgene, Bristol Myers Squibb, Pharmacyclics, Gilead, Genentech, Abbvie, TG Therapeutics, Beigene, AstraZeneca, Sunesis, Atara Biotherapeutics, GenMab: Research Funding; Abbvie, Genentech, AstraZeneca, Sound Biologics, Pharmacyclics, Beigene, Bristol Myers Squibb, Morphosys, TG Therapeutics, Innate Pharma, Kite Pharma, Adaptive Biotechnologies, Epizyme, Eli Lilly, Adaptimmune , Mustang Bio and Atara Biotherapeutics: Consultancy. Niemann: Novo Nordisk Foundation: Research Funding; CSL Behring, Genmab, Takeda, Octapharma: Consultancy; Abbvie, AstraZeneca, Janssen: Consultancy, Research Funding. Danilov: Abbvie: Consultancy, Honoraria; Beigene: Consultancy, Honoraria; Pharmacyclics: Consultancy, Honoraria; Rigel Pharm: Honoraria; Gilead Sciences: Research Funding; Bristol-Meyers-Squibb: Honoraria, Research Funding; Astra Zeneca: Consultancy, Honoraria, Research Funding; TG Therapeutics: Consultancy, Research Funding; Genentech: Consultancy, Honoraria, Research Funding; Takeda Oncology: Research Funding; Bayer Oncology: Consultancy, Honoraria, Research Funding; SecuraBio: Research Funding.

Introduction: CAR T-cell therapy has dramatically improved outcomes for patients (pts) with relapsed/refractory (r/r) DLBCL, with some achieving durable response. Still, the majority of pts have poor outcomes with CAR-T due to progressive disease, while inherent characteristics may predispose pts to CAR-T toxicities. Tools quantifying frailty and comorbidities have not been verified in large patient cohorts. The Cumulative Illness Rating Scale (CIRS) is a comprehensive tool that has been found to predict outcomes in various B cell malignancies. Here we use a machine learning algorithm to rank the prognostic impact of specific comorbidities, as measured by CIRS, and then assess survival and toxicities in pts treated with commercially available CAR T-cell products for DLBCL. Methods: We conducted a retrospective RWE analysis of pts with r/r DLBCL who underwent leukapheresis for CART at 9 academic centers. CIRS was assessed at the time of T-cell collection and calculated as in Salvi et al, 2008. High comorbidity burden was defined using a published cutoff of CIRS score ≥7. Progression-free survival (PFS) and overall survival (OS) were measured from T-cell collection. Repeated subsampling and random survival forest (RSF) modeling of PFS were used to determine the most prognostic CIRS categories in the presence of covariates. Cox proportional hazards models were fit to quantify the association between survival and the following patient features: IPI at diagnosis, concurrent indolent lymphoma, age, ECOG performance status, number of prior therapies, prior transplant, number of medications, cell of origin subtype, complex karyotype, MYC rearrangement by FISH, and MYC+BCL2/BCL6 rearrangement (double-hit). Associations between comorbidities and CAR-T adverse events were evaluated with Fisher's exact test. Results: We analyzed data from 577 pts, with a median age of 63 (range, 19-90); 90% had ECOG 0-1. Median number of prior therapies was 3 (range, 1-11) and 25% of pts (n=143) had prior autologous stem cell transplant. GCB subtype was found in 54% of pts (n=312), with 38% (n=218) non-GCB and 8% (n=47) unknown (Hahn's algorithm). MYC gene rearrangement was present in 21% and double-hit in 16% with 9% lacking FISH. Twenty-seven pts (4.7%) died before CAR-T infusion (progressive disease in 24 and infection in 3). Of the 550 pts who received CAR-T, 71% (n=393) got axicabtagene ciloleucel, 22% (n=120) tisagenlecleucel, and 7% (n=37) lisocabtagene maraleucel. The median CIRS score was 7 (range, 0-25) with 54% (n=309) having CIRS ≥7. The most frequent comorbidities were seen in the vascular (51%), endocrine (45%), and hypertension (43%) organ system categories. PFS event was observed in 56% of pts and 41% died, while survivors had a median follow-up time of 20 months. Median survival estimates were 11 months (95% CI: 8 - 15) for PFS and 30 months (95% CI: 23 - NA) for OS. CIRS ≥7 was significantly associated with PFS (HR=1.26, Fig 1A) and OS (HR=1.35, Fig 1B) in univariable analysis. According to RSF variable importance and node splits, key CIRS categories were respiratory, upper GI, renal, and hepatic (Fig 1C). Multivariable analysis showed higher ECOG (2/3 vs 1 vs 0) and 3 or more prior lines of treatment predicted shorter PFS and OS. Non-GCB subtype was related to reduced PFS but not OS, while double-hit status did not correlate with either outcome. Interestingly, CIRS ≥7 was not a significant predictor of outcomes in multivariable models. However, a severe comorbidity in any of the above four systems (denoted "Severe4", 9% of pts) was independently associated with inferior PFS (HR=2.45, Fig 1D) and OS (HR=2.30, Fig 1E). Although CIRS ≥7 was not associated with the development of CRS, pts with "Severe4" had a higher rate of grade ≥3 CRS (16% vs 6%; p=0.013). In addition, development of grade ≥3 ICANS was associated with CIRS ≥7 (26% vs 12%; p Conclusions: In this large RWE study, we demonstrate that CIRS is predictive of outcomes and identify a composite index comprising four CIRS organ systems (respiratory, upper GI, renal and hepatic; "Severe4") that had prognostic significance in CAR-T recipients for r/r DLBCL. "Severe4" is predictive of severe CRS and CIRS ≥7 is predictive of severe ICANS. The underlying mechanism leading to this observation is an area of active ongoing investigation. Given these results, CIRS evaluation and "Severe4" should be considered prior to CAR-T in DLBCL. GS & AK contributed equally Figure 1 Figure 1. Disclosures Shouse: Beigene Pharmaceuticals: Honoraria; Kite Pharmaceuticals: Speakers Bureau. Jaglowski: Novartis: Consultancy, Research Funding; Kite, a Gilead Company: Consultancy, Research Funding; Juno: Consultancy; Takeda: Consultancy; CRISPR Therapeutics: Consultancy. Shadman: Abbvie, Genentech, AstraZeneca, Sound Biologics, Pharmacyclics, Beigene, Bristol Myers Squibb, Morphosys, TG Therapeutics, Innate Pharma, Kite Pharma, Adaptive Biotechnologies, Epizyme, Eli Lilly, Adaptimmune , Mustang Bio and Atara Biotherapeutics: Consultancy; Mustang Bio, Celgene, Bristol Myers Squibb, Pharmacyclics, Gilead, Genentech, Abbvie, TG Therapeutics, Beigene, AstraZeneca, Sunesis, Atara Biotherapeutics, GenMab: Research Funding. Patel: Kite Pharma: Consultancy, Speakers Bureau; Morphosys: Consultancy; Janssen: Consultancy; Genentech: Consultancy; Bristol Myers Squibb: Consultancy, Speakers Bureau; Pharmacyclics: Consultancy; Abbvie: Consultancy; TG Therapeutics: Consultancy, Speakers Bureau; MEI Pharma: Consultancy; BeiGene: Consultancy; AstraZeneca: Consultancy, Research Funding, Speakers Bureau; ADC Therapeutics: Consultancy; Lilly: Consultancy. Stephens: CSL Behring: Consultancy; Celgene: Consultancy; AstraZeneca: Consultancy; Abbvie: Consultancy; Novartis: Research Funding; JUNO: Research Funding; Mingsight: Research Funding; Arqule: Research Funding; Adaptive: Membership on an entity's Board of Directors or advisory committees; TG Therapeutics: Membership on an entity's Board of Directors or advisory committees; Epizyme: Membership on an entity's Board of Directors or advisory committees; Beigene: Membership on an entity's Board of Directors or advisory committees; Innate Pharma: Membership on an entity's Board of Directors or advisory committees; Karyopharm: Membership on an entity's Board of Directors or advisory committees, Research Funding. Kamdar: TG Therapeutics: Research Funding; Adaptive Biotechnologies: Consultancy; ADC Therapeutics: Consultancy; AstraZeneca: Consultancy; Genentech: Research Funding; Genetech: Other; Celgene: Other; SeaGen: Speakers Bureau; Celgene (BMS): Consultancy; Kite: Consultancy; KaryoPharm: Consultancy; AbbVie: Consultancy. Hill: Novartis: Consultancy, Honoraria, Research Funding; Kite, a Gilead Company: Consultancy, Honoraria, Other: Travel Support, Research Funding; Pfizer: Consultancy, Honoraria; AbbVie: Consultancy, Honoraria, Research Funding; Epizyme: Consultancy, Honoraria; Karyopharm: Consultancy, Honoraria, Research Funding; Celgene (BMS): Consultancy, Honoraria, Research Funding; Gentenech: Consultancy, Honoraria, Research Funding; AstraZenica: Consultancy, Honoraria; Incyte/Morphysis: Consultancy, Honoraria, Research Funding; Beigene: Consultancy, Honoraria, Research Funding. Karmali: Genentech: Consultancy; BeiGene: Consultancy, Speakers Bureau; Kite, a Gilead Company: Consultancy, Research Funding, Speakers Bureau; Karyopharm: Consultancy; Takeda: Research Funding; EUSA: Consultancy; Epizyme: Consultancy; Roche: Consultancy; Morphosys: Consultancy, Speakers Bureau; AstraZeneca: Speakers Bureau; BMS/Celgene/Juno: Consultancy, Research Funding; Janssen/Pharmacyclics: Consultancy. Kittai: Abbvie: Consultancy; Bristol-Meyers Squibb: Consultancy; Janssen: Consultancy. Danilov: Abbvie: Consultancy, Honoraria; TG Therapeutics: Consultancy, Research Funding; Takeda Oncology: Research Funding; Genentech: Consultancy, Honoraria, Research Funding; Pharmacyclics: Consultancy, Honoraria; Beigene: Consultancy, Honoraria; Bayer Oncology: Consultancy, Honoraria, Research Funding; Gilead Sciences: Research Funding; Bristol-Meyers-Squibb: Honoraria, Research Funding; Rigel Pharm: Honoraria; Astra Zeneca: Consultancy, Honoraria, Research Funding; SecuraBio: Research Funding.

The development of molecularly-targeted therapies to improve outcomes relative to chemotherapy for acute myeloid leukemia (AML) is impeded by the heterogeneity of genetic aberrations that contribute to disease. Among the multitude of biological mechanisms that lead to AML disease initiation and progression is dysregulation of cytokine signaling pathways, a hallmark of chronic inflammation, which contribute to the growth, survival, and differentiation state of AML cells. We have previously shown that IL-1β, a pro-inflammatory cytokine expressed by many cell types including macrophages and monocytes, stimulates proliferation of leukemic blasts independent of mutational status in primary AML samples via enhanced phosphorylation of p38α MAPK, an effect that can be blocked by IL-1 receptor knockdown or by pharmacologic inhibition (Carey 2017). Additionally, recent studies have shown sensitivity to the approved BCL2 inhibitor venetoclax in AML associates with undifferentiated leukemic cells (Pei 2020; Zhang 2018; Majumder 2020). Based on these associations, we evaluated the combination of doramapimod (DORA), a p38 MAPK inhibitor, with venetoclax (VEN) for potential enhanced sensitivity on primary AML cells. Ex vivo drug screening of primary AML patient samples (n=335) revealed significantly enhanced efficacy of VEN+DORA compared to either single agent (Nemenyi test; p Disclosures Druker: Novartis Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees, Patents & Royalties; Recludix Pharma, Inc.: Consultancy; EnLiven: Consultancy, Research Funding; Pfizer: Research Funding; The RUNX1 Research Program: Membership on an entity's Board of Directors or advisory committees; Merck & Co: Patents & Royalties; Aileron: Membership on an entity's Board of Directors or advisory committees; ALLCRON: Consultancy, Membership on an entity's Board of Directors or advisory committees; Amgen: Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Aptose Therapeutics: Consultancy, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Blueprint Medicines: Consultancy, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Bristol-Myers Squibb: Research Funding; Cepheid: Consultancy, Membership on an entity's Board of Directors or advisory committees; GRAIL: Current equity holder in publicly-traded company; VB Therapeutics: Membership on an entity's Board of Directors or advisory committees; Iterion Therapeutics: Membership on an entity's Board of Directors or advisory committees; Nemucore Medical Innovations, Inc.: Consultancy; Third Coast Therapeutics: Membership on an entity's Board of Directors or advisory committees; Vincerx Pharma: Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Vivid Biosciences: Membership on an entity's Board of Directors or advisory committees. Tyner: Genentech: Research Funding; Takeda: Research Funding; Astrazeneca: Research Funding; Constellation: Research Funding; Agios: Research Funding; Petra: Research Funding; Incyte: Research Funding; Array: Research Funding; Gilead: Research Funding; Janssen: Research Funding; Seattle Genetics: Research Funding; Schrodinger: Research Funding.

Introduction Despite impressive response rates of >70% in frontline therapy with Azacitidine and Venetoclax, relapsed/refractory (R/R) acute myeloid leukemia (AML) remains a clinical challenge with up-front resistance and relapse developing through mutations of TP53, RAS pathway genes and expression/utilization of alternative BCL2 family members. Through integrative functional genomic analyses performed on a large cohort of primary AML patient samples, we found that the combination of Ruxolitinib (Rux) and Venetoclax (Ven) exhibited broad ex vivo efficacy and synergy. Based on the significant activity seen in both newly diagnosed and R/R AML, we conducted a Phase I study to evaluate the safety and efficacy of Rux+Ven in R/R AML. Methods This Phase I multi-center study will identify a recommended Phase II dose and evaluate the overall safety and preliminary efficacy of Rux+Ven. Eligible patients were >18 years old with R/R AML. There was no restriction on the number of previous therapies or prior exposure to Ven or Rux. We included patients with prior allogeneic transplant if GVHD was controlled and prior MDS patients >75 years old who progressed on a hypomethylating agent (HMA) but had no AML therapy. In cohorts of 3 to 4, patients were assigned to receive Rux+Ven at 1 of 6 dose levels according to the "keyboard" Bayesian toxicity probability interval design applied to a pre-specified target dose-limiting toxicity (DLT) rate of 30%. DLTs were defined as non-disease-related Grade ≥3 non-hematologic toxicities, with exceptions for nausea, vomiting, febrile neutropenia due to disease-related cytopenias, and electrolyte abnormalities that resolve within 48 hours. Hematologic DLT was specified as Grade 4 neutropenia by 42 days post-therapy in the absence of disease. The DLT evaluation period covered the first cycle. Clinical responses were determined using 2017 ELN criteria. Patients could continue study therapy after two cycles if they had a response or were deriving clinical benefit. Results Twenty patients have been enrolled out of the planned accrual of 30. Median age was 73 (range 29-87), 60% of patients had refractory disease, 50% had ≥3 prior therapies, and 35% had prior Ven exposure. At screening, 40% of patients had a complex karyotype and 65% had adverse ELN risk, including 20% with mutated TP53. The numbers of patients treated by dose level (DL) are: 3 on DL0 (200 mg qd Ven and 10 mg bid Rux), 4 on DL1 (400 mg qd Ven and 10 mg bid Rux), 3 on DL2 (400 mg qd Ven and 20 mg bid Rux), and 10 on DL3 (400 mg qd Ven and 30 mg bid Rux), the highest level of the study. Per prescribing guidelines, the cycle 1 Rux doses of 10 patients and Ven doses of 14 patients were reduced because of concomitant CYP3A inhibitor therapy. All 20 patients were DLT evaluable (defined as receiving both study drugs on ≥50% of cycle 1 days) and no DLTs were observed. The median duration of therapy was 55 days (95% CI: 51 - 113 days). The most common hematological AEs, regardless of attribution, were grade 3/4 thrombocytopenia (40%), febrile neutropenia (40%), neutropenia (35%), anemia (35%), and leukopenia (30%). Of the serious AEs recorded in 7 patients (35%), 4 were deemed related to Rux+Ven (3 for febrile neutropenia, 1 for sepsis). There were 2 treatment-emergent deaths, with one ( lung infection) potentially related to study drug. The Clinical Benefit Rate (CBR) was 40% (95% CI: 19.1% - 63.9%) over the first 2 cycles of therapy with five patients (25%) establishing MLFS or better and three achieving a CR or CRi during this time period (Composite Complete Remission rate 15% (95% CI: 3.2% - 37.9%)). For the 5 responders, median number of prior lines was 2, 1 patient had received Ven previously, and none had a TP53 mutation. Ongoing analyses involving ex vivo drug sensitivity, genomic sequencing, CyTOF, and single cell imaging are looking at associations with clinical response. The protocol is being amended to include PK/PD analysis for the final 10 patients. Conclusion The novel, all oral combination of Ven and Rux in heavily pretreated R/R AML patients was well tolerated with no DLTs, promising initial responses, and encouraging duration of therapy. Additional analyses including PK/PD and genomic and immune profiling will give further insight into which patients benefit the most in this poor prognostic group. The well tolerated safety profile makes it a promising combination to advance in the front line setting in combination with HMAs. Figure 1 Figure 1. Disclosures Borate: Astellas: Membership on an entity's Board of Directors or advisory committees; Pfizer: Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding; Daiichi-Sankyo: Membership on an entity's Board of Directors or advisory committees; Genentech: Membership on an entity's Board of Directors or advisory committees, Other: Advisory Board; Jazz Pharma: Research Funding; Blueprint Medicine: Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees; AbbVie: Membership on an entity's Board of Directors or advisory committees, Research Funding; incyte: Membership on an entity's Board of Directors or advisory committees, Research Funding; Rampal: Membership on an entity's Board of Directors or advisory committees; Galecto, Inc.: Consultancy; Promedior: Consultancy. Saultz: IKENA: Research Funding. Madanat: Onc Live: Honoraria; Blue Print Pharmaceutical: Honoraria; Stem line pharmaceutical: Honoraria; Geron Pharmaceutical: Consultancy. Druker: VB Therapeutics: Membership on an entity's Board of Directors or advisory committees; Aptose Therapeutics: Consultancy, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Bristol-Myers Squibb: Research Funding; Recludix Pharma, Inc.: Consultancy; The RUNX1 Research Program: Membership on an entity's Board of Directors or advisory committees; Merck & Co: Patents & Royalties; Third Coast Therapeutics: Membership on an entity's Board of Directors or advisory committees; Blueprint Medicines: Consultancy, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Amgen: Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Pfizer: Research Funding; Iterion Therapeutics: Membership on an entity's Board of Directors or advisory committees; ALLCRON: Consultancy, Membership on an entity's Board of Directors or advisory committees; EnLiven: Consultancy, Research Funding; GRAIL: Current equity holder in publicly-traded company; Novartis Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees, Patents & Royalties; Nemucore Medical Innovations, Inc.: Consultancy; Cepheid: Consultancy, Membership on an entity's Board of Directors or advisory committees; Aileron: Membership on an entity's Board of Directors or advisory committees; Vincerx Pharma: Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Vivid Biosciences: Membership on an entity's Board of Directors or advisory committees. Tyner: Array: Research Funding; Takeda: Research Funding; Genentech: Research Funding; Constellation: Research Funding; Schrodinger: Research Funding; Petra: Research Funding; Agios: Research Funding; Seattle Genetics: Research Funding; Incyte: Research Funding; Gilead: Research Funding; Janssen: Research Funding; Astrazeneca: Research Funding.

Activating mutations in the tyrosine kinase receptor FLT3 are observed in ~35% of all acute myeloid leukemia (AML) cases. Multiple FLT3 inhibitors are currently in clinical development and while most patients initially respond well to FLT3 inhibition, resistance inevitably develops in a period of months. During the initial response to FLT3 inhibitors, residual leukemia cells are able to survive and persist in the marrow microenvironment, which facilitates early resistance. Over time, leukemia cells develop intrinsic mechanisms of resistance, that are not dependent upon the microenvironment, which leads to late resistance and disease relapse. In this study, we utilized a two-step model to study the temporal evolution of gilteritinib resistance. To model initial extrinsic early resistance, we cultured the FLT3-ITD+ AML cell lines, MOLM-14 and MV4;11, with ligands secreted by the marrow microenvironment. We previously reported that fibroblast growth factor 2 (FGF2) is secreted from marrow stromal cells and protects FLT3-ITD+ AML cells from quizartinib, a FLT3 inhibitor. FGF2 binds receptor FGFR1, activates MAPK signaling, and promotes ligand-dependent growth (Traer et al. Cancer Res. 2016). We also used FLT3 ligand (FL), which has been shown to reactivate the FLT3 receptor despite the presence of inhibitor. MOLM14 cells were cultured with 100 nM gilteritinib in media alone, or supplemented with 10 ng/ml FGF2 or FL. After 7 weeks, all cultures supplemented with ligand eventually resumed growth (early resistance), whereas MOLM-14 cells without ligand were unable to resume growth.We then removed ligand, which transiently restored sensitivity to gilteritinib. However, after 2 months the cells resumed exponential growth in the absence of extrinsic factors (late resistance). A similar pattern was observed with MV4;11 cells. We analyzed the mechanisms of early and late resistance using a number of orthogonal tools: whole exome sequencing (WES), genome-wide CRISPR/Cas9, proteomics, metabolomics, and small-molecule inhibitor screening. WES identified NRAS mutations in the majority of late resistant cultures (13/15) and a gatekeeper FLT3 mutation was found in 1, consistent with mutations found in patients treated with gilteritinib on the Admiral trial (McMahon et al., Cancer Discov., 2019). To identify if NRAS mutations were pre-existing, ddPCR was used. NRAS mutations were detected at low level ( We further analyzed both early and late resistant lines with genome-wide resensitization CRISPR/Cas screening and global and phospho-proteomics. As expected, CRISPR/Cas screening revealed that NRAS was important for late resistance, which was confirmed by proteomics. In contrast, early resistance revealed multiple hits in metabolic and cell cycle pathways by CRISPR/Cas and global and phospho-proteomics. The unique metabolic signatures (lipid signaling in particular) were verified by metabolomic analyses. CDK proteins were significantly decreased in early resistance, as was the cell cycle. Aurora Kinase B protein (AURKB) was also increased in early resistance, and cells were uniquely sensitive to AURKB small molecule inhibitors and genetic deletion. We then evaluated primary leukemia cells from 11 patients before and after 1-2 months of gilteritinib to evaluate early resistance. Leukemia cells were isolated by CD33 and CD34 beads and subjected to a targeted proteomic analysis (Figure 1). In agreement with our cell line model, early resistant cells had a distinct protein profile, with significant alteration of cell cycle and lipid metabolism proteins after gilteritinib treatment. Primary patient samples also demonstrated robust sensitivity to AURKB inhibitors only after gilteritinib exposure. Our results suggest that developing drug combinations that selectively target early resistance can improve the depth of initial response and may block development of later resistance mutations, thus improving the durability of response to gilteritinib. Figure 1 Disclosures Tyner: Incyte: Research Funding; Janssen: Research Funding; Syros: Research Funding; Seattle Genetics: Research Funding; Petra: Research Funding; Agios: Research Funding; AstraZeneca: Research Funding; Gilead: Research Funding; Aptose: Research Funding; Array: Research Funding; Genentech: Research Funding; Constellation: Research Funding; Takeda: Research Funding. Druker:Henry Stewart Talks: Patents & Royalties; Third Coast Therapeutics: Membership on an entity's Board of Directors or advisory committees; Novartis Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees, Patents & Royalties, Research Funding; McGraw Hill: Patents & Royalties; MolecularMD (acquired by ICON): Consultancy, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Millipore (formerly Upstate Biotechnology): Patents & Royalties; Merck & Co: Patents & Royalties; Cepheid: Consultancy, Membership on an entity's Board of Directors or advisory committees; GRAIL: Consultancy, Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; Dana-Farber Cancer Institute: Patents & Royalties; EnLiven: Consultancy, Research Funding; Gilead Sciences: Consultancy, Membership on an entity's Board of Directors or advisory committees; Vivid Biosciences: Membership on an entity's Board of Directors or advisory committees; Aileron Therapeutics: Membership on an entity's Board of Directors or advisory committees; ALLCRON: Consultancy, Membership on an entity's Board of Directors or advisory committees; Amgen: Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Aptose Therapeutics Inc. (formerly Lorus): Consultancy, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; ARIAD: Research Funding; Blueprint Medicines: Consultancy, Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; Bristol-Myers Squibb: Research Funding; Oregon Health & Science University: Patents & Royalties; Patient True Talks: Consultancy; Pfizer: Research Funding; The RUNX1 Research Program: Membership on an entity's Board of Directors or advisory committees; Iterion Therapeutics (formerly Beta Cat Pharmaceuticals): Membership on an entity's Board of Directors or advisory committees; VB Therapeutics: Membership on an entity's Board of Directors or advisory committees; Leukemia & Lymphoma Society: Research Funding. Traer:Daiichi Sankyo: Membership on an entity's Board of Directors or advisory committees; Genentech: Membership on an entity's Board of Directors or advisory committees; Abbvie: Consultancy, Membership on an entity's Board of Directors or advisory committees; Agios: Membership on an entity's Board of Directors or advisory committees; Incyte: Research Funding; Notable Labs: Consultancy, Current equity holder in private company; Astellas: Membership on an entity's Board of Directors or advisory committees.