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Acute myeloid leukemia (AML) with the FLT3 internal tandem duplication (FLT3-ITD) mutation accounts for ~25% of all AMLs, carries a poor prognosis, and is prone to relapse despite targeted therapy. FLT3 mutations are associated with aberrant activation of the Wnt signaling pathway, which itself is implicated in AML initiation/progression and is required for the self-renewal and survival of leukemic stem cells. CLKs regulate the activity of serine/arginine-rich splicing factors (SRSFs) that modulate spliceosome assembly, mRNA splicing, and gene expression. SM09419 is a novel, oral, small-molecule pan-CLK inhibitor that potently inhibits the Wnt pathway. These studies examined the antitumor activity of SM09419 as a single agent and in combination with targeted and standard therapies in preclinical models of FLT3-ITD AML. In MV-4-11 and MOLM-13 AML cells carrying the FLT3-ITD mutation, SM09419 dose-dependently inhibited SRSF6 phosphorylation and potently suppressed expression of Wnt pathway-related genes (CCND1, MYC, TCF7, DVL2). The effect on cell proliferation was tested in 8 AML cell lines with varying mutation profiles as well as 26 different leukapheresis-derived primary human AML cells. Proliferation was strongly impaired by SM09419 across all tested cell lines (average EC50=0.2 + 0.048 µM]); MV-4-11 and MOLM-13 cells had EC50 of 0.049 and 0.144 µM, respectively. SM09419 also potently inhibited proliferation in all primary AML cells (average EC50=0.048 + 0.0097 µM) regardless of FLT3 mutation status, cytogenetics, or AML diagnosis (de novo or relapsed/refractory). SM09419 also induced apoptosis in MV-4-11 and MOLM-13 cells, increasing caspase 3/7 activation and PARP cleavage while reducing survivin and MCL-1 expression relative to vehicle. In vivo antitumor effects and tolerability of oral SM09419 (QD) alone or combined with either midostaurin (FLT3 inhibitor) or venetoclax (BCL2 inhibitor) and/or azacitidine were assessed in FLT3-ITD xenograft models (n=5-6/group). In MOLM-13 xenografts, SM09419 (12.5 and 25 mg/kg) induced strong tumor growth inhibition (TGI) vs. vehicle at Day 14 (TGI 52% [p<0.05] and 74% [p<0.001], respectively). Midostaurin (50 mg/kg) induced significant TGI vs. vehicle (50%, p<0.05), which was increased when administered in combination with 12.5 mg/kg SM09419 (81%, p<0.001). In MV-4-11 xenografts, single-agent SM09419 (6.25, 12.5, and 25 mg/kg) induced significant TGI vs. vehicle (56% [p<0.05], 94%, and 95% [p<0.001], respectively) at Day 26 with tumor regression in all mice dosed at 12.5 mg/kg and 25 mg/kg. In a subsequent experiment, midostaurin (50 mg/kg) alone and combined with 6.25 mg/kg SM09419 for 23 days induced tumor regression in MV-4-11 xenografts (100% TGI vs. vehicle, p<0.0001). After treatment discontinuation, tumor regression was maintained in all mice (6/6) treated with the combination for 26 days, whereas tumor regrowth was immediately observed in midostaurin-treated mice. In another MV-4-11 xenograft study, the combination of 6.25mg/kg SM09419 with azacitidine (0.8 mg/kg QD) and/or venetoclax (25 mg/kg QD) induced significant TGI (95-98% vs. vehicle, p<0.001) with tumor regression at Day 26. Azacitidine + venetoclax induced 79% TGI (p<0.001), but no tumor regression was observed. The triple combination induced tumor regression in all mice and complete regressions in 4/6 mice (67%); it had a greater effect on slowing tumor regrowth after treatment discontinuation vs. a single agent or doublet. SM09419 alone or in combination was well tolerated in these xenograft models based on body weight measurements. In summary, SM09419 potently inhibited SRSF6 phosphorylation and Wnt signaling pathway activity and induced apoptosis in FLT3-ITD cell lines. It also inhibited proliferation in cell lines and primary AML cells regardless of FLT3 status. The strong in vivo antitumor effects observed as combination treatment suggest that SM09419 combined with standard therapies may provide a clinical benefit by slowing or preventing relapse in AML with a marker of poor prognosis such as FLT3-ITD. A Phase 1 study assessing safety, tolerability, and pharmacokinetics of SM09419 in subjects with advanced hematologic malignancies is being initiated. Disclosures Chung: Samumed, LLC: Employment, Equity Ownership. Creger:Samumed, LLC: Employment, Equity Ownership. Sitts:Samumed, LLC: Employment, Equity Ownership. Chiu:Samumed, LLC: Employment, Equity Ownership. Mak:Samumed, LLC: Employment, Equity Ownership. KC:Samumed, LLC: Employment, Equity Ownership. Tam:Samumed, LLC: Employment, Equity Ownership. Bucci:Samumed, LLC: Employment, Equity Ownership. Stewart:Samumed, LLC: Employment, Equity Ownership. Phalen:Samumed, LLC: Employment, Equity Ownership. Cha:Samumed, LLC: Employment, Equity Ownership.

Mantle cell lymphoma (MCL) is a B-cell non-Hodgkin lymphoma (NHL) that accounts for ~7% of all NHL in the U.S. MCL is associated with aberrant activation of the Wnt signaling pathway, which plays a key role in the survival and maintenance of MCL-initiating cells. Many MCL patients experience relapse and subsequent disease progression due to chemoresistance following initial therapy; hence, novel therapies are needed. CLKs regulate the activity of serine/arginine-rich splicing factors (SRSFs) that modulate spliceosome assembly, mRNA splicing, and gene expression. SM09419 is a novel, oral, small-molecule pan-CLK inhibitor that potently inhibits the Wnt pathway. The purpose of these studies was to examine the antitumor activity of SM09419 in preclinical models of MCL. SM09419 potently inhibited both CLK1-CLK4 (IC50 for all <0.02 µM) and Wnt signaling pathway (average EC50=0.068 µM) activities. In REC-1 and GRANTA-519 MCL cells, SM09419 dose-dependently inhibited SRSF6 phosphorylation and potently suppressed expression of Wnt-related genes (CCND1, LEF1, TCF7) and proteins vs. vehicle. In tests on 5 MCL cell lines, cell proliferation was strongly impaired by SM09419 across all lines (average EC50=0.102 µM [0.021-0.236]). SM09419 also induced apoptosis in REC-1 and GRANTA-519 cells, increasing caspase 3/7 activation and PARP cleavage while reducing survivin and MCL-1 expression vs. vehicle. In vivo antitumor effects and tolerability of oral SM09419 (QD 20-21 days) were assessed in mice bearing REC-1 and JeKo-1 flank xenografts (n=5/group). In REC-1 xenografts, strong tumor growth inhibition (TGI) vs. vehicle occurred in mice treated with 12.5, 25, and 50 mg/kg SM09419 (TGI 88% [p<0.01], 100%, and 100% [p<0.001], respectively), and the two highest doses induced complete tumor regression in all mice from D14. Similarly, in JeKo-1 xenografts, SM09419 (12.5 and 25 mg/kg) induced significant TGI vs. vehicle (71% and 100%, respectively; p<0.0001) with complete tumor regression at 25 mg/kg, whereas acalabrutinib (50 mg/kg BID) was not efficacious (27% TGI) when tested in parallel. SM09419 25mg/kg induced reversible suppression of phospho-SRSF6 protein and inhibited Wnt pathway-related gene expression (TCF7 and DVL2) in JeKo-1 tumors in a single-dose PD study, demonstrating downstream target engagement in vivo. SM09419 was also assessed in 2 patient-derived xenograft (PDX) mouse models of MCL. PDX cells were injected intravenously and treatment was initiated upon 8-12% engraftment of human CD45+CD19+ cells in peripheral blood. In the first model, derived from a patient who was progressive after 8 modalities including ibrutinib, SM09419 (25 mg/kg QD) increased survival vs. vehicle (100% through D26 vs. 0% by D12, respectively; n=6/group) and suppressed MCL engraftment in the blood (12% at D26 vs. 69% at D8 and D12, respectively; p=0.002) and bone marrow (30% at D26 vs. 91% at D8 and D12, respectively; p=0.002). In the second model, derived from a patient refractory after ibrutinib and anti-PDL1 treatment, SM09419 (25 mg/kg QD) significantly suppressed MCL engraftment vs. vehicle in the blood (8% vs. 72%), bone marrow (20% vs. 57%), and spleen (15% vs. 96%) at D28 (study end; p<0.001 for all; n=4/group). In addition, SM09419 greatly inhibited splenomegaly vs. vehicle (0.04 g vs. 0.4 g, respectively; p<0.001). In a subsequent experiment in the same model, mice (n=7/group) were treated with 12.5 or 25 mg/kg SM09419 or vehicle for 12 weeks (to D85). Blood MCL engraftment at D41 was significantly lower in mice treated with SM09419 (40% at 12.5 mg/kg and 23% at 25 mg/kg) vs. vehicle (88%; p<0.01 and p<0.001, respectively). SM09419 dose-dependently increased survival (28.6% at 12.5 mg/kg and 85.7% at 25 mg/kg at D85) vs. vehicle (0% at D63); survival was maintained in both dose groups during post-treatment monitoring (to D99). SM09419 was well tolerated in all tested mouse models based on body weight measurements. In summary, SM09419 potently inhibited SRSF6 phosphorylation Wnt signaling pathway activity, and cell proliferation and induced apoptosis in MCL cell lines. The strong in vivo antitumor effects observed as a single agent suggest that SM09419 may provide a clinical benefit for patients with treatment-resistant or refractory MCL. A Phase 1 study assessing safety, tolerability, and pharmacokinetics of SM09419 in subjects with advanced hematologic malignancies is being initiated. Disclosures Chung: Samumed, LLC: Employment, Equity Ownership. Creger:Samumed, LLC: Employment, Equity Ownership. Sitts:Samumed, LLC: Employment, Equity Ownership. Chiu:Samumed, LLC: Employment, Equity Ownership. Mak:Samumed, LLC: Employment, Equity Ownership. KC:Samumed, LLC: Employment, Equity Ownership. Tam:Samumed, LLC: Employment, Equity Ownership. Bucci:Samumed, LLC: Employment, Equity Ownership. Stewart:Samumed, LLC: Employment, Equity Ownership. Phalen:Samumed, LLC: Employment, Equity Ownership. Cha:Samumed, LLC: Employment, Equity Ownership.

Acute myeloid leukemia (AML) with TP53 mutation makes up ~13% of AML cases and is an aggressive, treatment-resistant subtype with dismal prognosis and limited therapeutic options. Aberrant activation of the Wnt signaling pathway is associated with AML initiation/progression and is required for the self-renewal and survival of leukemic stem cells, making Wnt signaling inhibition a potential therapeutic modality for adverse AML. CLKs regulate the activity of serine/arginine-rich splicing factors (SRSFs) that modulate spliceosome assembly, mRNA splicing, and gene expression. SM09419 is a novel, oral, small-molecule pan-CLK inhibitor that potently inhibits the Wnt pathway in a Wnt signaling reporter assay. The purpose of these studies was to examine the antitumor activity of SM09419 as a single agent and in combination with standard therapies in preclinical models of TP53 mutant (TP53mut) AML. In TF-1a and KG-1a AML cells with TP53 mutations, SM09419 dose-dependently inhibited SRSF6 phosphorylation and potently suppressed expression of Wnt-related genes (LEF1, MYC, DVL2) and proteins vs. vehicle. The effect of SM09419 on cell proliferation was tested in 6 TP53mut AML cell lines. Proliferation was strongly impaired by SM09419 across all cell lines (EC50=0.23 + 0.056 µM). SM09419 significantly induced apoptosis in TF-1a and KG-1a cells, increasing caspase 3/7 activation and PARP cleavage while reducing survivin and MCL-1 expression relative to vehicle. In addition, SM09419 potently inhibited cell proliferation when tested in 27 leukapheresis-derived human primary AML cell lines (EC50=0.046 + 0.0061 µM) regardless of TP53 status, cytogenetics, or AML diagnosis (de novo or relapsed/refractory). In vivo antitumor effects and tolerability of oral SM09419 (QD) alone or combined with cytarabine (Ara-C), venetoclax (VEN), or azacytidine (AZA) were assessed in mice bearing TP53mut flank xenografts (n=5-15/group). In TF-1a xenografts, SM09419 (12.5 and 25 mg/kg) induced significant tumor growth inhibition (TGI) vs. vehicle at D20 (TGI 55-56% [p<0.01]). VEN (50mg/kg) was not effective (3% TGI) and combining VEN with SM09419 had no additional benefit (52%-60% TGI). In Kasumi-1 xenografts, SM09419 (12.5 and 25 mg/kg), AZA (0.8 mg/kg), and VEN (25 mg/kg) induced TGI vs. vehicle of 87%, 95% (both p<0.0001), 72% (p<0.001), and 48% (NS), respectively at D18. SM09419 25 mg/kg alone induced tumor regression in 40% (2/5) of the mice. SM09419 (12.5 mg/kg) + VEN induced greater TGI vs. vehicle (96%, p<0.0001) with tumor regression in 80% (4/5) of the mice, while AZA + VEN induced 79% TGI (p<0.001) with no tumor regression. In KG-1a xenografts, single-agent SM09419 (12.5 and 25 mg/kg) and Ara-C (10mg/kg) induced significant TGI vs. vehicle (53%, 98%, and 80% [p<0.001], respectively) at D28 but VEN (12.5mg/kg) did not (35% TGI). The combination of SM09419 (12.5 mg/kg) + VEN (12.5 mg/kg) improved TGI (98%) vs. vehicle. Tumor regression was seen in all mice with single-agent SM09419 (25 mg/kg) and 12.5 mg/kg + VEN. In another KG-1a xenograft study, mice were treated with combinations of SM09419 (12.5 mg/kg), AZA (0.8 mg/kg), and VEN (25 mg/kg) for 20 days followed by 21 days of SM09419 (25 mg/kg) or vehicle maintenance in some groups. SM09419 + VEN, SM09419 + AZA, and AZA + VEN induced TGI of 95%, 64%, and 58%, respectively (all p<0.0001), with 80% (12/15) regression in SM09419 + VEN. The triplet induced 91% TGI but was not well tolerated due to GI toxicity. In the maintenance phase, SM09419 given QD or QOD greatly slowed tumor regrowth vs. vehicle at D41 in mice previously treated with SM09419 + VEN (80% and 72% TGI [p<0.001], respectively). SM09419 QD maintenance therapy also slowed tumor regrowth following AZA + VEN (p<0.0001). SM09419 alone and in combination (except with AZA + VEN) was well tolerated in all tested xenografts. In summary, SM09419 potently inhibited SRSF phosphorylation and Wnt pathway signaling and induced apoptosis in TP53mut AML cell lines. It also inhibited proliferation in cell lines and primary AML cells regardless of TP53 status. Strong in vivo antileukemic effects were observed with SM09419 as a single agent or in combination with other AML therapies, suggesting that it is a potential treatment for hard-to-treat AML subtypes such as TP53mut AML. A Phase 1 study assessing safety, tolerability, and pharmacokinetics of SM09419 in subjects with advanced hematologic malignancies is being initiated. Disclosures Chung: Samumed, LLC: Employment, Equity Ownership. Creger:Samumed, LLC: Employment, Equity Ownership. Sitts:Samumed, LLC: Employment, Equity Ownership. Chiu:Samumed, LLC: Employment, Equity Ownership. Mak:Samumed, LLC: Employment, Equity Ownership. KC:Samumed, LLC: Employment, Equity Ownership. Tam:Samumed, LLC: Employment, Equity Ownership. Bucci:Samumed, LLC: Employment, Equity Ownership. Stewart:Samumed, LLC: Employment, Equity Ownership. Phalen:Samumed, LLC: Employment, Equity Ownership. Cha:Samumed, LLC: Employment, Equity Ownership.