Fernández-Zapico, Martín E.

AbstractThe modulation of GLI2, an oncogenic transcription factor commonly upregulated in cancer, is in many cases not due to genetic defects, suggesting dysregulation through alternative mechanisms. The identity of these molecular events remains for the most part unknown. Here, we identified TFII-I as a novel repressor of GLI2 expression. Mapping experiments suggest that the INR region of the GLI2 promoter is necessary for GLI2 repression. ChIP studies showed that TFII-I binds to this INR. TFII-I knockdown decreased the binding of NELF-A, a component of the promoter–proximal pausing complex at this site, and enriched phosphorylated RNAPII serine 2 in the GLI2 gene body. Immunoprecipitation studies demonstrate TFII-I interaction with SPT5, another pausing complex component. TFII-I overexpression antagonized GLI2 induction by TGFβ, a known activator of GLI2 in cancer cells. TGFβ reduced endogenous TFII-I binding to the INR and increased RNAPII SerP2 in the gene body. We demonstrate that this regulatory mechanism is not exclusive of GLI2. TGFβ-induced genes CCR7, TGFβ1 and EGR3 showed similar decreased TFII-I and NELF-A INR binding and increased RNAPII SerP2 in the gene body post-TGFβ treatment. Together these results identify TFII-I as a novel repressor of a subset of TGFβ-responsive genes through the regulation of RNAPII pausing.

Abstract Waldenström macroglobulinemia (WM) is a B-cell malignancy characterized by the aberrant production of a monoclonal IgM protein that may lead to hyperviscosity. Although this is a major factor causing significant morbidity in patients, little is known about the mechanisms that regulate monoclonal protein synthesis. Cytokines are protein mediators that are known to be involved in many biological processes, and can profoundly affect tumor cells and the tumor microenvironment. Many cytokines have been shown to have potent therapeutic efficacy in preclinical cancer models; however, the role of cytokine networks in WM is not fully understood and few studies have described the precise functional roles of cytokines in WM. To address this issue we performed a multiplex ELISA analysis to test cytokine levels in sera from patients with WM. We found that Rantes/CCL5 is significantly elevated in WM patients and correlates with disease activity. Elevated expression of RANTES in the serum was confirmed by ELISA and was also detected in the bone marrow of WM patients by ELISA and immunohistochemistry. RANTES expression serves as a marker for recruitment of immune cells and is associated with a wide range of immune-mediated diseases. However, the impact of RANTES in WM is not known. We analyzed the expression of receptors for Rantes by flow cytometry and found that WM B cells and stromal cells express CCR1 and CCR3, but not CCR5. Using a standard chemotaxis assay, we determined that Rantes had no effect on B cell or stromal cell recruitment. Rantes also had no effect on B cell or stromal cell survival, however it did promote stromal cell proliferation (p<0.04). The interaction between Rantes and IL-6 has been described in an autoimmune disease model. Since stromal cells secrete significantly more IL-6 than WM B cells, we treated stromal cells with Rantes for 24 hr and found that Rantes increases IL-6 secretion (p<0.03). To characterize the mechanism of Rantes-mediated IL-6 secretion, we transfected stromal cells with an IL-6 promoter construct and treated with Rantes for 12 hr and found a significant increase in IL-6 promoter activity (p<0.0162) indicating Rantes can regulate IL-6 transcription. Bioinformatics analysis of the IL-6 promoter indicates the presence of multiple candidate binding sites for transcription factors that have been previously shown to play a role in the biology of B cells, including NFkB, AP1, and GLI. Co-transfection of stromal cells with an IL-6 reporter construct and a plasmid expressing GLI1, GLI2 or GLI3 demonstrates that GLI2 and GLI3 proteins can regulate the IL-6 promoter. We then transfected stromal cells with a reporter construct containing 8X-GLI binding sites and demonstrate that Rantes can regulate GLI transcription further supporting a role for the interaction between Rantes and IL-6 through GLI transcription factors. IL-6 rich tumor microenvironment supports malignant cells. Elevated IL-6 levels have no effect on survival of BCWM.1 cells or CD19+ 138+ cells from WM patients, but leads to upregulation of IgM secretion by BCWM.1 cells and CD19+ CD138+ cells from WM patients, and increases their proliferation (p<0.0039). IL-6 activates Erk1/2 and Jak/ Stat in WM and stimulation of the IgM secreting cells BCWM.1 with IL-6 in the presence of PD98059 MAPK inhibitor had no effect on IgM secretion. However, stimulation of BCWM.1 cells with IL-6 in the presence of JakI inhibitor abolished the IL-6 mediated IgM secretion suggesting IL-6 mediated increase in IgM secretion occurs through Jak/ Stat signaling pathway. Analysis of Rantes levels in other B cell malignancies including follicular lymphoma, chronic lymphocytic leukemia, monoclonal gammopathy of undetermined significance and multiple myeloma indicates that Rantes is elevated in other B cell lymphoproliferative disorders and suggest Rantes may play a similar role in other malignancies. In summary, our data identifies a novel Rantes-GLI-IL-6 interplay in the stromal microenvironment that promotes IgM production by malignant B cells. This therefore provides multiple new potential therapeutic avenues, targeting both malignant cells and the microenvironment to control malignant cell growth, and immunoglobulin secretion in WM and Ig-mediated diseases.

Abstract Background: A wide range of cancers, including pancreatic cancer, display altered expression of amino acid transporter LAT-1. LAT-1 overexpression has been shown to be an important prognostic factor and predicts chemo resistance in pancreatic cancer (Kaira K, 2012; Altan B, 2018). Radiolabeled amino acids for PET imaging, including 3-18F-l-α-methyl-tyrosine ([18F] FAMT), have successfully leveraged cancer’s aberrant LAT1 expression and amino acid uptake for decades. SM-88 is a denatured D/L racemic form of α-methyl-tyrosine designed for selective uptake and disruption of protein synthesis in cancer cells, as well as disruption of catecholamine production via inhibition of tyrosine hydroxylase. Catecholamines have been reported to potentially have a role in driving pancreatic cancer progression and changes in the tumor immune status (Renz B, Cancer Cell 2018; Calvani M, Oncotarget 2014). Initial clinical and preclinical results with SM-88 indicate that this agent could pose a novel approach in the treatment of pancreatic cancer. Methods and Results: Initial xenograft models of human gastrointestinal cancers using monotherapy SM-88 showed a >50% decrease in tumor growth at three weeks of treatment (p<0.05). Ongoing preclinical experiments are testing the full pharmacodynamic effects of SM-88 on pancreatic cancer cells, as well as potential effects on the tumor microenvironment due to catecholamine interruption. In results from the ongoing phase 2 trial of SM-88 in heavily pretreated metastatic pancreatic cancer patients, SM-88 has shown monotherapy antitumor activity based on RECIST 1.1 criteria. Additionally, 16/24 patients demonstrated a reduction in circulating tumor cells, with the median nadir decline of 63%. LAT1 has been reported to be important in EMT transition and mesenchymal cell survival (Holldorsson S, Cancer letters 2017); examination of the impact of SM-88 on this important emerging biomarker is ongoing. Conclusion: Altered amino acid (LAT1) metabolism has been shown to be a negative prognosticator in pancreatic cancer outcomes and progression. Targeting LAT1 and amino acid pathways has been identified as a therapeutic approach in multiple cancers (Fuchs BC, Semin Ca Biol 2005). Initial and ongoing studies of SM-88 may clarify the potential utility of this approach in the treatment of pancreatic cancer. Citation Format: Martin Fernandez-Zapico, Dae Won Kim, Philip Philip, Alexander Vandell, Jonathan Eckard, Ron Korn, Giuseppe Del Priore, Diane Simeone. Therapeutic potential of targeting amino acid metabolism in pancreatic cancer [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer: Advances in Science and Clinical Care; 2019 Sept 6-9; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2019;79(24 Suppl):Abstract nr B15.

TPS594 Background: Effective treatment options are very limited for patients with advanced refractory biliary tract cancer (BTC). Fluoropyrimidine-based chemotherapy regimen such as 5-fluorouracil and irinotecan are frequently utilized for these patients after first-line therapy despite lack of FDA approval. Trifluridine/tipiracil (FTD/TPI) is a novel oral nucleoside with antitumor activity in both fluoropyrimidine sensitive and resistant tumors due to its unique mechanisms of action. Given early toxicity and efficacy data from our previous study on single-agent trifluridine/tipiracil (FTD/TPI) in advanced BTC, the clinical evaluation of its combination with irinotecan represents a rational approach for the treatment of advanced refractory BTC. Methods: This is a single-arm phase II trial with a two-stage design to assess the efficacy of trifluridine/tipiracil (FTD/TPI) and irinotecan in advanced refractory BTC. Key eligibility criteria include histologically confirmed advanced, unresectable BTC who have progressed on at least one line of systemic therapy and have measurable disease per RECIST v1.1. Target accrual is 25. Treatment includes trifluridine/tipiracil (FTD/TPI) 25 mg/m2 on days 1-5 and irinotecan 180 mg/m2 on day 1 in 14-day cycles. Patients will be evaluated for response every 4 cycles and in the absence of disease progression, therapy may be given up to 2 years. The primary end point is the progression-free survival rate at 16 weeks. Secondary endpoints include overall response rate, disease control rate, progression-free survival, overall survival, and incidence of adverse events. Correlative biomarker studies include evaluations of circulating tumor DNA and circulating tumor cells at baseline, after 4 cycles and at progression; and development of patient-derived tumor organoids from pre-treatment biopsies for parallel treatments. This study was approved and funded in part by the National Comprehensive Cancer Network (NCCN) Oncology Research Program from general research support provided by Taiho Oncology, Inc. Clinical trial information: NCT 04072445.

Abstract Background : The 2016 revision to the World Health Organization classification of myeloid neoplasms has recommended distinction between "proliferative" (WBC ≥ 13 x 10(9)/L) and "dysplastic" (WBC < 13 X 10(9)/L) subtypes of chronic myelomonocytic leukemia (CMML). In addition, CMML is characterized by a spectrum of cytogenetic and molecular abnormalities (Patnaik Leukemia 2014). In the current study, we looked for correlations between gene expression profiles (GEP) and morphologic or molecular categories of previously untreated patients with CMML. Methods : 35 patients with CMML were included in the study. Targeted capture assays were carried out on BM DNA specimens obtained at diagnosis for 28 myeloid-relevant genes (Patnaik Blood C. J.). RNA sequencing (Stranded Total RNA- Illumina, San Diego, CA), was performed on 9 normal BM controls, and 35 CMML samples (25 peripheral blood (PB) and 10 BM). The RNA-Seq data was analyzed using MAPRSeq v.2.0. Normalization and differential expression analysis was performed using edgeR 2.6.2. We selected genes that were differentially expressed with a False Discovery Rate (FDR) <0.1 and a fold change bigger than 2. Pathway enrichment analysis was performed using Ingenuity Pathway Analysis (IPA). Results: Among the 35 study patients, 75% were males and median age was 71 years (range, 55-87). 21 (60%), 9 (25%) and 5 (15%) patients were classified as CMML-0, 1 and 2, respectively. 16 (45%) had a dysplastic while 19 (55%) had a proliferative phenotype. At a median follow-up of 16 months, 5 (14%) deaths and 4 (14%) leukemic transformations were documented. Mutational frequencies included; TET2 45%, ASXL1 45%, SRSF2 40%, NRAS 14%, SETBP1 13%, CBL 10%, JAK2 7%, RUNX1 6%, DNMT3A 6%, U2AF1 6%, SF3B1 5%, ZRSR2 4%, Tp53 4%, and IDH2 4%. Abnormal cytogenetics (n=9, 35%) included; -Y 8%, trisomy 8 6%, monosomy 7 3%, del(13q) 3% and monosomal karyotype 3%. i) Differential GEP in PB samples: 25 PB CMML samples were analyzed. Unsupervised GEP demonstrated two unique clusters (figure one); cluster one was enriched with dysplastic CMML (90%) and cluster two with proliferative CMML (64%). Consistent with the difference in the distribution of morphologic categories, in comparison to cluster one, patients in cluster two had a higher white blood count (p=0.003), higher monocyte count (p=0.02) and higher risk prognostication by the Mayo Model (p=0.016); there was no significant difference between the two clusters in terms of age and gender distribution, hemoglobin level, neutrophil and platelet counts, PB or BM blast content, cytogenetic risk stratification, ASXL1, TET2, RAS, CBL and SETBP1 mutational status. Genes significantly upregulated (FDR<1e-5 and fold change ≥ 16x); in cluster two (proliferative CMML) in comparison to cluster one included; PRG2, CTSG, BEX1, CD34, CRISP2, while those significantly down-regulated included (FDR<1e-15 and fold change ≥ 5x); IL2RB, PRF1, GNLY, PDGFRB. IPA analysis identified the following pathways preferentially expressed in cluster two (proliferative CMML) 1) mitotic roles of polo-like kinase (KIF23, WEE-1, PLK1, PLK4), 2) Cell cycle: G2/M checkpoint regulation (CDC25C, WEE-1, AURKA, CDK1), 3) Cell cycle: chromosomal replication (CDC45, RPA3, MCM2, CDC7), and 4) BRCA1 in DNA damage response (BARD1, RBBP8, PLK1, RAD51). Whereas in the cluster one (dysplastic CMML), preferentially expressed pathways included; 1) T-cell receptor signalling (CD247 PRKCQ, CD4, PLCG1, CD28) and 2) STAT3 signalling (SOCS3, MAP3K9). ii) Differential GEP in BM samples: 10 BM CMML samples were analyzed along with 10 normal controls. Samples and controls clustered separately and within the CMML samples two clusters were identified (figure two). These clusters had equal representations of dysplastic (n=5) and proliferative (n=5) CMML sub-types. Additional samples are being processed. Conclusions: Differential gene expression profiling in patients with CMML identifies two unique clusters, clinically demarcated, at least in PB samples, by "proliferative" and "dysplastic" CMML subtypes. The proliferative cluster is enriched in genes involved in cell cycle regulation and DNA damage response; whereas the dysplastic cluster is enriched in T-cell receptor and STAT3 signalling. The prognostic and therapeutic impact of these gene clusters remains to be assessed in a larger group of informative patients. Figure 1. Figure 1. Figure 2. Figure 2. Disclosures Al-Kali: Novartis: Research Funding; Celgene: Research Funding.

Introduction: Chronic myelomonocytic leukemia (CMML) is a clonal hematopoietic neoplasm characterized by sustained peripheral blood (PB) monocytosis and an inherent risk for leukemic transformation. Clonal origins of the disease can be detected in hematopoietic progenitor cells (CD34+/CD38-), while the complete spectrum of mutational evolution can be seen in circulating monocytes (CD14+). Cell sorting strategies have been employed to select cells in CMML, and while there are adequate monocyte numbers in the PB, there are very few circulating progenitor cells. In addition, attrition related to the selection process significantly depletes primary cells available for biological experiments and multiomics studies such as RNA-seq, ChIP-seq, ATAC-seq, and DIP-seq. While single-cell methods may be able to overcome this challenge, bulk sequencing methods remain a robust and cost-effective approach. We hypothesized that, secondary to the stem cell origin of this disease and significant myeloproliferation, PB mononuclear cells (MNC) would provide comparable results with regards to transcriptomic analysis, in comparison to cell selection procedures. Methods: Peripheral blood obtained from 15 molecularly annotated patients with WHO-defined CMML was ACK-lysed and subjected to a Ficoll procedure for collection of MNC. MNC were left unsorted (n=5) or further selected for CD34+/CD38- (n=5) and CD14+ (n=5) using a fully automated RoboSep-S (StemCell Technologies) protocol. All samples were then subjected to bulk whole transcriptome shotgun sequencing (using Illumina TruSeq and an Illumina HiSeq 4000). After data quality control, counts of detectable transcripts were log2-normalized and Pearson's product-moment correlation coefficients were calculated to evaluate the correlation between the two cell-sorting strategies and unsorted cells in terms of detectable transcripts. To visualize sample differences log2-normalized transcripts counts were centered and scaled per gene for a select number of genes relevant to myeloid biology as well as a number of housekeeping genes. Results: Fifteen patients with WHO-defined CMML, median age 69 years (55-73 years), 66% male, were included. Next generation sequencing for somatic mutations was performed on PB MNC obtained at CMML diagnosis (Figure 1, top heatmap). Considering the small sample size, mutations were evenly distributed among groups with the exception of ASXL1 (higher frequency in CD14+ and CD34+/CD38- cells), ZRSR2 (higher frequency in unsorted cells), and TET2 (lower frequency in CD14+ cells). The three groups were also well matched with regards to other CMML-related variables such as WHO and FAB morphological subtypes, cytogenetic abnormalities, and risk stratification by the Mayo Molecular Model. Transcriptomic analysis revealed a strong positive correlation between the median number of log2-normalized detectable transcripts in unsorted cells and CD34+/CD38- cells (ρ = 0.96, p < 0.001, top scatterplot). Likewise, there was a strong positive correlation between the median number of log2-normalized detectable transcripts in unsorted cells and CD14+ cells (ρ = 0.91, p < 0.001, bottom scatterplot). The latter correlation was marginally lower, which was explained by increased global gene expression in 3 of the 5 CD14+ samples (bottom heatmap). Increased gene expression in these 3 samples involved key myeloid genes and housekeeping genes known to have stable expression across human tissues alike. In comparison to PB MNC, both cell sorting strategies resulted in significant depletion of primary cells required for other experiments, and for procedures such as ChIP-seq, DIP-seq and ATAC-seq (CD34+/CD38- had greater depletion than CD14+). Additional experiments to assess this strategy for the above mentioned epigenetic studies are currently being planned. Conclusions: Accounting for sample differences, different cell sorting strategies (unsorted, CD34+/CD38- selection, and CD14+ selection) yielded similar results when performing bulk transcriptomic assessments on PB MNC from patients with CMML. For the purpose of gene expression profiling there was no clear advantage with CD34+/CD38- or CD14+ selection. These results support the use of unsorted cells for bulk transcriptomic analysis in CMML. Figure 1 Disclosures Patnaik: Stem Line Pharmaceuticals.: Membership on an entity's Board of Directors or advisory committees.

Background: Myelodysplastic syndrome (MDS)/myeloproliferative neoplasm (MPN) with ring sideroblasts and thrombocytosis (MDS/MPN-RS-T) was formally included as a unique WHO-defined entity in the 2016, iteration of the classification of hematological neoplasms (under MDS/MPN overlap syndromes). Patients with MDS/MPN-RS-T have features of MDS-RS-single lineage dysplasia (SLD), persistent thrombocytosis (platelet count ≥450x109/L), with proliferation of large atypical megakaryocytes. Anemia, including red blood cell transfusion dependency (RBC-TD) is a major problem. While extensive studies have documented erythroid response rates (ERR) to erythropoiesis stimulating agents (ESA) in MDS and MDS-RS-SLD, the data in MDS/MPN-RS-T is less clear. We carried out this study to asses ERR in ESA treated patients with MDS/MPN-RS-T. Methods: After approval by the IRB, patients with 2016, WHO-defined MDS/MPN-RS-T were included in the study. The bone marrow (BM) morphology, cytogenetics and BM RS% were retrospectively reviewed. All study patients underwent next generation sequencing for 29 myeloid-relevant genes, obtained on BM mononuclear cells, at diagnosis, or at first referral, by previously described methods (Patnaik et al., BCJ 2016). Treatment details, including the type of ESA used, dose administered, side effects and reason for discontinuation were retrospectively abstracted. Erythroid responses were assessed using the 2006, international working group (IWG) MDS and the 2015, IWG MDS/MPN response criteria. Results: Forty seven patients with MDS/MPN-RS-T were identified, of which 37 (79%) patients received ESA treatment at any time point during their disease course; median age 73 years (range, 52-93), 46% male.The hemoglobin at diagnosis (HB) and baseline erythropoietin (EPO) levels were 9.1 gm/dL (range, 6.6-12.1) and 39 IU/L (range, 8-500), respectively. Ten (29%) patients were RBC-TD at baseline. None of the 37, ESA-treated patients were on concomitant cyto-reductive therapy for thrombocytosis. Twenty (54%) patients were treated with erythropoietin-a, 13 (35%) with darbepoetin, and 4 (11%) with both sequentially. Median doses were 40,000 IU weekly (range, 20,000-80,000) for erythropoietin-a and 200 mcg every 2 weeks (range, 100-500), for darbepoetin. Median treatment duration was 14 months (range, 1-173) and causes for discontinuation included; ESA failure/loss of response in 20 (54%), sustained HB rise in 1 (3%) and other causes, including adverse events in 4 (11%) patients. With regards to safety, the following events were considered potentially ESA related; treatment emergent hypertension in 2 (5%), venous thromboembolism, ischemic stroke and splenic infarction in 1 (3%) patient each, respectively. All patients with thrombotic/ischemic events had been on low dose aspirin prophylaxis. Erythroid responses by the IWG MDS and MDS/MPN response criteria were assessable in 35 (95%) patients: with 16 (46%) meeting criteria for an erythroid response, by both assessment systems. The median duration of ER was 20 months (range, 2-172). In comparison to ESA non-responders, those that responded were more likely to have baseline EPO levels ≤44 IU/L (ROC analysis; median, 28 vs 112 IU/L; p=0.0080), while there was a trend for inferior responses associated with RBC-TD (p=0.0857). Age, gender, baseline white blood cell and platelet counts, type of ESA used, BM RS %, JAK2V617F, SF3B1, DNMT3A, TET2 and ASXL1 mutational status did not impact ESA response (Table 1). On a univariate survival analysis (OS), survival was not significantly different between ESA responders vs non-responder (median OS, 76 vs 45 months; p=0.2929). After ESA failure, 7 (19%) patients received hypomethylating agents (HMA), while 5 (14%) received lenalidomide. There were no erythroid responses to HMA, while 1 of 3 (33%) assessable lenalidomide treated patients achieved a morphological complete response. Conclusion: Erythropoiesis stimulating agents are effective first line therapies for the management of anemia in patients with MDS/MPN-RS-T, with 46% achieving an erythroid response (median duration of response of 20 months). Low baseline endogenous EPO level (<44 IU/L) was the best predictor of response, with a trend towards an inferior response in the presence of RBC-TD. In the context of clonal thrombocytosis, potentially related thrombotic/ischemic events were documented in 8% of patients. Disclosures Al-Kali: Astex Pharmaceuticals, Inc.: Research Funding. Patnaik:Stem Line Pharmaceuticals.: Membership on an entity's Board of Directors or advisory committees.

Waldenström macroglobulinemia (WM) is a low-grade B-cell lymphoma associated with the accumulation of lymphoplasmacytic cells in the bone marrow (BM) and a high level of monoclonal IgM protein in the serum. Genome-wide studies revealed genomic alternations in WM, with the most prevalent being a point mutation in the myeloid differentiation primary response gene 88 (MyD88) resulting in an amino acid substitution (L265P) that is present in over 90% of WM patients. MyD88 is an effector of Toll-like receptor (TLR) signaling pathway that mediates inflammatory cytokine expression and secretion upon activation. Here, we found that activation of the TLR-MyD88 pathway in WM cells carrying the MyD88 L265P somatic mutation increases the expression of IL-6 and CCL2, two cytokines known to participate in the pathobiology of several neoplasms including B cell malignancies. Furthermore, IL-6 has been shown to modulate IgM secretion and malignant B cell growth in WM. We demonstrate that an active ERK1/2 pathway, which is elevated in WM patient samples, is required to maintain IL-6 and CCL2 expression in BCWM.1 (p=0.0339) and RPCI-WM1 (p=0.0005) cells. Further characterization identified a change in the chromatin landscape in response to TLR-MyD88 activation in WM cells. We found increased levels of trimethylation of histone 3 lysine 4 (H3K4me3) at the promoters of the inflammatory cytokines IL-6 and CCL2 by chromatin immunoprecipitation (ChIP) assay followed by qPCR (ChIP-qPCR). The H3K4me3 modification is catalyzed by six related homologs of the yeast histone methyltransferase (HMT) family. Further analysis identified the Mixed-lineage leukemia 1 (MLL1) as the enzyme bound to these promoters in response to TLR-MyD88 stimulation of WM cells. Analysis of WM cell lines and primary WM patient cells showed that MLL1 and its binding partner, menin, are expressed at significantly higher levels in CD19+CD138+cells from WM patients compared to CD19+B cells from peripheral blood of healthy donors (p<0.001). We also found an increase in H3K4me3 deposition on IL-6 and CCL2 promoters during early (1-3 hr) and late (12-24 hr) kinetics following TLR-MyD88 stimulation. This also coincided with increased deposition of MLL1 on these cytokine promoters by ChIP-qPCR. Disruption of menin-MLL1 using the MI-2 or MI-503 menin-MLL1 inhibitors significantly reduced IL-6 and CCL2 expression in WM cell lines (BCWM.1 p=0.039; RPCI-WM1 p=0.0041). This also results in significantly reduced IgM expression (p<0.0001) and secretion at inhibitor dose (5 μM) that has no effect on cell proliferation. Finally, MLL1 knockdown using RNAi significantly reduces IgM expression in BCWM.1 (p<0.0001), MWCL (p=0.0016) and RPCI-WM1 (p<0.0001) and IgM secretion in BCWM.1 (p<0.0001) and MWCL-1 (p<0.0001). Taken together, these results identify a novel role for menin-MLL1 in regulating inflammatory cytokines and IgM expression and secretion in WM and provide the rationale for targeting these molecules in WM patients. Disclosures Ansell: Mayo Clinic Rochester: Employment; Mayo Clinic Rochester: Employment; Regeneron: Research Funding; Seattle Genetics: Research Funding; Trillium: Research Funding; Seattle Genetics: Research Funding; Regeneron: Research Funding; Trillium: Research Funding; Bristol-Myers Squibb: Research Funding; Mayo Clinic Rochester: Employment; Trillium: Research Funding; Affimed: Research Funding; Affimed: Research Funding; Regeneron: Research Funding; Trillium: Research Funding; Mayo Clinic Rochester: Employment; Bristol-Myers Squibb: Research Funding; Mayo Clinic Rochester: Employment; LAM Therapeutics: Research Funding; Bristol-Myers Squibb: Research Funding; Seattle Genetics: Research Funding; LAM Therapeutics: Research Funding; LAM Therapeutics: Research Funding.