E-selectin Antagonist Research Articles

Combining 5-azacitidine with the E-selectin antagonist uproleselan is an effective strategy to augment responses in myelodysplasia and acute myeloid leukaemia.

The interaction of acute myeloid leukaemic (AML) blasts with the bone marrow (BM) microenvironment is a major determinant governing disease progression and resistance to treatment. The constitutive expression of E-selectin in the vascular compartment of BM, a key endothelial cell factor, directly mediates chemoresistance via E-selectin ligand/receptors. Despite the success of hypomethylating agent (HMA)-containing regimens to induce remissions in older AML patients, the development of primary or secondary resistance is common. We report that following treatment with 5-azacitidine, promoter regions regulating the biosynthesis of the E-selectin ligands, sialyl Lewis X, become further hypomethylated. The resultant upregulation of these gene products, in particular α(1,3)-fucosyltransferase VII (FUT7) and α(2,3)-sialyltransferase IV (ST3GAL4), likely causes functional E-selectin binding. When combined with the E-selectin antagonist uproleselan, the adhesion to E-selectin is reversed and the survival of mice transplanted with AML cells is prolonged. Finally, we present clinical evidence showing that BM myeloid cells from higher risk MDS and AML patients have the potential to bind E-selectin, and these cells are more abundant in 5-azacitidine-non-responsive patients. The collective data provide a strong rationale to evaluate 5-azacitidine in combination with the E-selectin antagonist, uproleselan, in this patient population.

Patient Recorded Outcomes from a Randomized, Controlled Phase 2 Trial of E-Selectin Inhibition with Uproleselan Vs Placebo to Reduce GI Toxicity during Melphalan-Conditioned Autologous Hematopoietic Cell Transplantation for Multiple Myeloma

Background: The use of high-dose melphalan conditioning followed by autologous hematopoietic cell transplant (AHCT) continues to be a key component in the treatment of patients with multiple myeloma (MM). However, high-dose melphalan is associated with a high rate of GI toxicity (>90%), which adversely impacts clinical outcomes and negatively influences patient quality of life (QoL). Indeed, it has been reported that AHCT leads to short-term deterioration of health-related QoL in patients with MM, and QoL typically improves by 2-3 months post-AHCT. Pre-clinical and clinical data suggest that E-selectin inhibition may play a role in reducing immune-mediated GI epithelial injury following cytotoxic chemotherapy. We hypothesized that uproleselan (upro), a synthetic, competitive E-selectin antagonist, may improve health-related QoL by reducing risk of chemotherapy-induced diarrhea in patients undergoing AHCT. Methods: We conducted a Phase 2, single-center, randomized, double-blind, placebo-controlled trial in patients with MM receiving high-dose melphalan (200 mg/m2) as conditioning for AHCT for MM. Patients were randomized 1:1 to receive prophylactic upro+standard of care (SoC) vs placebo+SoC. Uproleselan was given in 6 doses of 800 mg IV starting on day-3 through day 0. The primary endpoint was diarrhea severity as assessed by CTCAE v5.0 (secondary endpoint - Bristol Stool Scale). In addition, a key secondary endpoint was patient reported outcomes (PRO) of GI-related QoL using the National Cancer Institute (NCI) PRO-CTCAE Measurement System, a validated PRO tool developed to evaluate symptomatic toxicities in patients with cancer in clinical trials. Significance level of p<0.2 was pre-specified for all endpoints, per protocol. Patients were surveyed during the peri-transplant period at day-3 (baseline), day+8, and day+14 or day of discharge (DoD). Responses were graded on a scale of 1-5, with 1 indicating absence of symptoms or zero interference with daily activities and 5 indicating very severe symptoms or very significant interference with daily activities. GI-specific domains assessed included swallowing, mouth sores, appetite, nausea, vomiting, heartburn, bloating, abdominal pain, flatulence, and incontinence. Results: Fifty adult patients with MM were enrolled from 5/2021-10/2022. Baseline PRO symptoms were similar between the upro+SoC and placebo+SoC arms at baseline (day-3). By day+8 post-AHCT patients in the upro+SoC arm reported a significant improvement in severity of GI-related symptoms across 30% (6/20) of domains surveyed vs placebo+SoC (p=0.07-0.14). Furthermore, improvement in mouth sore-related symptoms continued to day+14 post-AHCT in the upro+SoC arm vs placebo+SoC (p=0.17). The remaining GI-related symptoms returned to baseline by D+14 or DoD (p>0.20). Also of note, a numerically lower mean diarrhea severity score and lower incidence of Grade 3 diarrhea (by CTCAE v5.0) was observed favoring upro+SOC which did not meet the pre-specified significance threshold (p=0.34, p=0.26, respectively). However, a significant reduction in severe diarrhea (by Bristol Stool Scale) was observed favoring upro+SoC (p=0.10) vs placebo+SoC, with improvement in the upro+SoC arm to mild diarrhea (p=0.02). Conclusions: Upro+SoC resulted in a clinically significant improvement in GI-toxicity related symptoms as assessed via PRO-CTCAE in patients with MM undergoing melphalan-conditioned AHCT, when compared to placebo+SoC. These improvements in patient reported outcomes are further supported by data from the primary and additional secondary endpoints of the study. Taken together, these results suggest prophylactic E-selectin inhibition may represent a promising strategy to mitigate chemotherapy-associated GI-toxicity. Further studies are needed to verify these findings and to further define the optimal dosing of upro in the peri-AHCT period.

Uproleselan Added to Cladribine Plus Low Dose Cytarabine (LDAC) in Patients with Treated Secondary Acute Myeloid Leukemia (TS-AML)

AML arising from HMA-treated myeloid disorders, also known as “treated-secondary AML” (ts-AML) is associated with an adverse prognosis (Complete Remission [CR] rates 15-30% and median Overall Survival [OS] 6-10 months). E-selectin, a marker of cell survival and resistance to chemotherapy, is highly expressed in the leukemic microenvironment. Exposure of leukemic blasts to HMAs has been shown to increase its expression. Uproleselan is an e-selectin antagonist that has demonstrated antileukemic activity in AML. We sought to study the combination of low-intensity chemotherapy with Cladribine + LDAC (CLAD/LDAC) with uproleselan to overcome local and microenvironmental resistance. This Phase Ib/II clinical trial (NCT04848974) evaluate the safety, tolerability, and efficacy of Uproleselan added to Cladribine and LDAC. A 3+3 dose-escalation approach was implemented to evaluate 2 different dose levels for Cladribine (CLAD)+ LDAC; each 4-week cycle consists of Uproleselan (at fixed dose of 800mg intravenously [IV]). Induction and reinduction if required, included IV uproleselan twice a day from day 1-12; consolidation was given same days but once a day if patients(pts) achieved response (complete response[CR] with incomplete hematological recovery [CRi] or morphologic leukemia-free state [MLFS]). CLAD was IV for 5 days (3.75mg/m2 and 5mg/m2; level -1 and 1 respectively) and subcutaneous LDAC twice daily 10 days (15mg, and 20mg; level -1 and 1 respectively) during induction; consolidation was similar except it was with 3-days of CLAD, for up to 6 cycles, pts aged ≥18 years with a diagnosis of ts-AML with adequate organ function, who have not received therapy for their AML were enrolled. ts-AML is defined as AML arising from previously treated myeloid neoplasm. Presence of the e-selectin was assessed as exploratory analysis. 20 pts have been treated, 18 pts evaluable: 14 (70%) male, median age was 72 yrs (range, 58-86); at the start of therapy, the median bone marrow blasts were 26.5% (20-82%), median WBC was 2.1x10 9/L (0.6-26.4), and median platelets were 26x10 9/L (4-667). Pts had received a median of 1 (1-6) treatments prior to AML transformation. Prior diagnoses were: Chronic Myelomonocytic Leukemia (CMML), MDS, and MDS/MPN in 3 (25%), 14 (70%), and 1 (5%) respectively; all had received HMA, 11 (55%) additionally had received Ven, and 5 (25%) had stem cell transplantation (SCT) prior to enrolling. All pts had unfavorable cytogenetics by ELN 2022. The most frequent mutations were: TET2, TP53, RUNX1 and SRSF2 in 9 (45%), 7 (35%), 6 (30%) and 6 (30%) pts respectively. 18 pts were evaluable for e-selectin; median plasmatic concentration was 26.2ng/mL (15.3-131 ng/mL). Most common AEs were grade(gr) ≥3 neutropenic fever (13 pts, 65%; including 2 gr 5 events), gr 3 bleeding (10%) and gr 2 thrombosis (5%). There were no dose-limiting toxicities observed on dose levels -1 or 1. The chosen dose level was 1 as described above. 2/3 pts who received dose level -1 died during the study follow-up due to sepsis in the first 4-week after induction. Median time to 0.5x10 9/L neutrophil and 50x10 9/L platelets recovery was 30 (17-52) and 38 (20-69) days, respectively. The response rates were 2 (11%) CRi, 1 (6%) CRp, and 4 (22%) MLFS. There was a reduction in BM blasts in 13 pts (72%) (Figure 1). The ORR was 50%(5/10) ( p=0.60) and 20% (1/5) ( p=0.06) among pts who had prior Ven exposure or prior SCT, respectively. The median cycles received was 1.5 (1-4), median cycles at which the best response was achieved was 1 (1-2). 12 pts were taken off protocol due to progression, 3 for death, 3 for allogeneic SCT and 1 continued in remission. The pt who achieved negative MRD-negative by flow cytometry underwent SCT and is still alive. The median follow-up is 8.1 months. Median OS and EFS were 5.3 and 1.4 months respectively; 4-month RFS (CRi, CRp and MLFS) was 30% (Fig 2). In the univariate analysis by Cox regression for plasmatic E-selectin concentration the HR for OS was 1.016 (95% CI 1.002-1.031). Ts-AML, progressing from previously treated antecedent myeloid disorders, is associated with low rates of remission and poor OS. Uproleselan combined with Cladribine + LDAC was well-tolerated with minimal therapy-related AEs - allowing a safe approach to marrow blast reduction and disease control in preparation for a potential allogeneic SCT. We are currently determining the relationship of plasmatic e-selectin concentration and response to treatment.

A Phase I Study of Uproleselan Combined with Azacitidine and Venetoclax for the Treatment of Older or Unfit Patients with Treatment Naïve Acute Myeloid Leukemia

Introduction: VIALE-A established azacitidine (AZA) with venetoclax (VEN) as a standard of care for AML ineligible for induction chemotherapy (IC). Patients (Pts) who achieved a measurable residual disease (MRD) negative complete remission (CR) or CR with incomplete count recovery (CRi) by multiparameter flow cytometry (MFC) had improved outcomes; however, only 41% of CR/CRi responders achieved MRD negativity. Uproleselan (UPRO) is a novel E-selectin antagonist that disrupts cell survival pathways and improves chemotherapy responses in preclinical models, including in combination with AZA +/- VEN. A Phase I/II study of UPRO combining with IC in AML pts established a recommended Phase II dose (RP2D). UPRO was associated with higher CR/CRi and MRD negative (MRD-ve) rates, lower induction mortality, and less severe mucositis compared to historical controls. We are exploring the combination of UPRO with AZA/VEN to improve outcomes in pts with untreated AML ineligible for standard IC. Methods: This ongoing single center Phase I trial of frontline UPRO in combination with AZA/VEN in older or unfit AML pts (NCT04964505) consists of a dose optimization portion, using a modified 3+3 dose optimization design to confirm the RP2D UPRO dose level (800 mg IV q12h for 7 days), and a dose expansion cohort. We report results of the dose optimization portion and initial expansion phase subjects. Key eligibility criteria included age ≥18 years, AML diagnosis by WHO criteria, and eligibility for frontline AZA/VEN. Pts received UPRO 800 mg IV q12h and AZA 75 mg/m2 IV/SC q24h for 7 days, and VEN 400 mg PO daily for 28 days in 28-day cycles. Treatment continued until progression, intolerance, or patient decision to stop. Pts had a VEN ramp-up in cycle 1 including tumor lysis syndrome monitoring and prophylaxis. A bone marrow biopsy was done after every cycle until morphologic leukemia-free state or better response (MLFS+). UPRO was given for up to 6 cycles with dosing decreased to daily after achieving MLFS+. The primary objective was to determine safety and tolerability. Adverse events (AE) were monitored throughout treatment, and dose-limited toxicities (DLT) assessed in cycle 1. The secondary objective was to evaluate preliminary efficacy, notably MRD-ve CR/CRi rate, as measured by MFC. Results: As of June 26, 2022, 8 pts [75% female, median age 78 (range 70-81)] were enrolled in the dose optimization (n=6) and dose expansion (n=2) portions of the study. Four (50%) had de novo AML and four (50%) had secondary AML, including 3 (38%) with therapy-related AML. Six (75%) had ELN 2017 adverse risk disease; 3 (38%) had complex cytogenetics. The most common mutations identified were RUNX1 (n=4), BCOR (n=2) and TET2 (n=2), and one patient each had TP53, NPM1, FLT3-ITD, or IDH2 mutations. All pts completed cycle 1. Median time on study is currently 126 days (range 32-179). Pts received a median of 3 treatment cycles (range 1-6). No DLTs were observed. There were no deaths in the first 30 days from treatment initiation; there was one death from sepsis in the first 60 days. All pts had at least one treatment-emergent AE (TEAE). The most common TEAE regardless of grade and attribution included anemia (n=6), thrombocytopenia (n=6), anorexia (n=4), nausea (n=4), neutropenia (n=4), fatigue (n=3), hypocalcemia (n=3), and hyponatremia (n=3). All pts experienced at least one ≥grade 3 TEAE, with anemia (n=6), thrombocytopenia (n=6), neutropenia (n=4), and febrile neutropenia (n=2) most common. Four pts (50%) experienced nine total serious AEs (SAEs), including thrombocytopenia (n=2). There were two grade 5 events, sepsis and AML disease progression, both unrelated to UPRO. Two pts remain on study treatment, and six have discontinued study therapy [patient decision (n=4) and death (n=2)]. All pts with an MLFS+ had dose modifications and/or cycle delays. All 8 pts had a MLFS+ response. Five (63%) achieved CR and one CRi, for a total CR/CRi rate of 75%, and two achieved MLFS. Five CR/CRi responses occurred with cycle 1. Four of the CR/CRi responses were MFC MRD-ve, for an overall MRD-ve CR/CRi rate of 50% and 67% among the CR/CRi responders. Conclusions: Preliminary results from this Phase I study reveals a tolerable safety profile of UPRO with AZA/VEN in pts with untreated AML ineligible for IC. No DLTs were observed, and the most common Grade 3-4 AE and SAE were hematologic. The combination shows promising preliminary efficacy, including a 50% rate of MRD-ve CR/CRi.

Uproleselan Added to Cladribine Plus Low Dose Cytarabine (LDAC) in Patients with Treated Secondary Acute Myeloid Leukemia (TS-AML)

Background Treated secondary AML (TS-AML), arising after prior HMA-treated MDS, is associated with very poor prognosis (Complete Remission [CR] rates 15-30% and median Overall Survival [OS] 6-8 months). E-selectin ligand is highly expressed on AML blasts in the leukemic microenvironment and may be a marker of cell survival and resistance to chemotherapy. Exposure of leukemic blasts to HMAs has been shown to increase their expression of E-selectin ligand. Uproleselan is an E-selectin antagonist that overcomes resistance to chemotherapy in AML (Barbier, Nat Commun 2020). We sought to study the combination of low-intensity chemotherapy with Cladribine + LDAC (CLAD/LDAC) with uproleselan to overcome local and microenvironmental resistance and improve outcomes in this difficult subset. Methods This is Phase Ib/II clinical trial (NCT04848974) to evaluate the safety, tolerability, and explore the efficacy of Uproleselan added to Cladribine and LDAC. A 3+3 dose-escalation approach was implemented to evaluate 2 different dose levels for Cladribine (CLAD)+ LDAC; each 4-week cycle consists of Uproleselan (at a fixed dose of 800mg intravenously [IV]) added to IV CLAD 5 days (3.75mg/m2 and 5mg/m2; level -1 and 1, respectively) and subcutaneous LDAC twice daily 10 days (15mg, and 20mg; level -1 and 1, respectively) during induction; consolidation was similar except it was with 3-days of CLAD, for up to 6 cycles. Pts aged ≥18 years with a diagnosis of TS-AML with adequate organ function, who have not received therapy for their AML were enrolled. TS-AML is defined as AML arising from a previously treated myeloid neoplasm. Presence of the E-selectin ligand was assessed using Flow Cytometry (FC). Results 10 pts have been treated, with 9 pts currently evaluable: 6 (67%) were male and the median age was 68 years (range, 58-80); at the start of therapy, the median bone marrow blasts were 33% (1-78%), median WBC was 2.2x109/L (0.6-20.1), median platelets were 18x109/L (4-305), and median creatinine was 0.98mg/dL (0.67-1.52). Pts had received a median of 1 (1-2) treatments prior to AML transformation. Prior diagnoses were: therapy-related Myelodysplastic Syndrome (t-MDS), Chronic Myelomonocytic Leukemia (CMML), MDS and MDS/MPN in 3 (33%), 3 (33%), 2 (22%) and 1 (11%) respectively; all had received HMA, 5 (56%) additionally had Ven and 3 (33%) had stem cell transplantation (SCT) prior to enrolling. All pts had unfavorable features by ELN 2017. The most frequent mutations were:SXL1, TP53 and TET2 in 4 pts each (44%), SRSF2 and NRAS in 3 patients each (33%) and SETBP1, RUNX1 and EZH2 in 2 patients each (22%). 6 pts were evaluable for E-selectin ligand expression; the median expression was 59% (42%-95%) and median MFI was 20.5 (13-262). The most common SAEs were ≥ grade 3 neutropenic fever (70%), (including 2 grade 5 events), grade 3 bleeding (10%), and grade 2 thrombosis (5%) (Table 1). There were no dose-limiting toxicities observed on dose levels -1 or 1. Two pts treated on dose level -1 die during the study follow-up due to sepsis within the first 4-weeks during induction. Median time to 0.5x109/L neutrophil and 50x109/L platelets recovery was 29 (17-39) and 38 (33-48) days respectively. The median follow-up is 4+ months. 8 pts were evaluable for response at the time of analysis. The ORR was 62% (5/8), including 2 (25%) PR, 1 (13%) CRi, 1 (13%) CRp and 1 (13%) MLFS. There was a reduction in BM blasts in 6 pts (75%) (Figure 1). 5 pts were taken off protocol due to progression, 2 for death, 1 for allogeneic SCT and 1 continued onto maintenance in remission. The one pt who achieved negative MRD, underwent SCT and is still alive. Median OS and EFS were 4.7 and 1.3 months respectively; 4-month RFS (CRi, CRp, and MLFS) was 67%. The median cycles received was 1 (1-3), median cycles at which the best response was achieved was 1 (1-2). The 4-month OS were 100% and 60% among responders vs. non-responders, respectively (p=0.27), and the 4-month EFS were 50% and 0% respectively (p=0.01). The ORR was 40% (2/5) (p=0.85) and 33% (1/3) (p=0.85) among pts who had prior Ven exposure or prior SCT, respectively. Conclusions The combination of Cladribine + LDAC with Uproleselan was overall well tolerated with few treatment-related AEs. The combination produced an ORR of 62% in a high-risk, refractory population whose prognosis is very dismal. The relationship of E-selectin ligand expression, response to treatment, and outcomes is being analyzed. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

A randomized clinical trial of the efficacy and safety of rivipansel for sickle cell vaso-occlusive crisis

AbstractThe efficacy and safety of rivipansel, a predominantly E-selectin antagonist, were studied in a phase 3, randomized, controlled trial for vaso-occlusive crisis (VOC) requiring hospitalization (RESET). A total of 345 subjects (204 adults and 141 children) were randomized and 320 were treated (162 with rivipansel, 158 with placebo) with an IV loading dose, followed by up to 14 additional 12-hourly maintenance doses of rivipansel or placebo, in addition to standard care. Rivipansel was similarly administered during subsequent VOCs in the Open-label Extension (OLE) study. In the full analysis population, the median time to readiness for discharge (TTRFD), the primary end point, was not different between rivipansel and placebo (−5.7 hours, P = .79; hazard ratio, 0.97), nor were differences seen in secondary end points of time to discharge (TTD), time to discontinuation of IV opioids (TTDIVO), and cumulative IV opioid use. Mean soluble E-selectin decreased 61% from baseline after the loading dose in the rivipansel group, while remaining unchanged in the placebo group. In a post hoc analysis, early rivipansel treatment within 26.4 hours of VOC pain onset (earliest quartile of time from VOC onset to treatment) reduced median TTRFD by 56.3 hours, reduced median TTD by 41.5 hours, and reduced median TTDIVO by 50.5 hours, compared with placebo (all P < .05). A similar subgroup analysis comparing OLE early-treatment with early-treatment RESET placebo showed a reduction in TTD of 23.1 hours (P = .062) and in TTDIVO of 30.1 hours (P = .087). Timing of rivipansel administration after pain onset may be critical to achieving accelerated resolution of acute VOC. Trial Registration: Clinicaltrials.gov, NCT02187003 (RESET), NCT02433158 (OLE).

Uproleselan sodium. E-selectin antagonist, Treatment of acute myeloid leukemia

Uproleselan sodium. E-selectin antagonist, Treatment of acute myeloid leukemia

Co-Targeting E-Selectin/CXCR4 with GMI-1359 Facilitates AML Stem Cell Mobilization and Protects BM Niches from Anti-Leukemia Therapy

Acute myeloid leukemia (AML) is characterized by the heterogeneous clonal expansion of undifferentiated myeloid cells in the bone marrow (BM). AML cells compete with normal hematopoietic cells and rewire the BM microenvironment into niches that selectively support leukemia stem cells (LSC). The leukemic niche produces soluble factors that facilitate the retention of LSC and provide protection from cytotoxic and targeted agents. The vascular adhesion molecule, E-selectin is expressed on endothelial cells (EC) in the perivascular niche where therapy-resistant AML cells have an increased affinity to E-selectin compared to normal hematopoietic stem cells (HSC) (Winkler et al., 2020). We previously demonstrated (Chang et al., ASH 2020) that E-selectin blockade by the pharmacological antagonist, GMI-1271 (uproleselan; GlycoMimetics, Inc) sensitized therapy-resistant LSC to Bcl-2 targeted therapy. Efficacious eradication of LSC in the BM however requires blocking multiple receptors and/or associated signaling pathways. A more optimal dislodgement of LSC from the BM could be attained by combining an E-selectin antagonism with blockade of the CXCR4/SDF-1α axis. The dual antagonist of E-selectin and CXCR4, GMI-1359 (GlycoMimetics, Inc.), has been tested in a phase 1 clinical trial (NCT02931214). Previously, we showed that GMI-1359 in combination with a FLT3-ITD inhibitor, improved survival in a xenograft model of FLT3-ITD + AML (Zhang et al., 2016). Hence, we hypothesized that co-targeting E-selectin/CXCR4 more efficiently mobilizes AML cells from BM niches and synergizes with the anti-leukemia activity of venetoclax/hypomethylating agent (Ven/HMA).Intra-vital 2-photon imaging and tracking of individual leukemia cells in triple reporter mice (Blood: dextran-TRITC; Host T-cells: DsRed; Host myeloid CD11 cells: EYFP) injected with AML cells carrying a turquoise fluorescent protein reporter gene suggested that dual inhibition of E-selectin/CXCR4 with GMI-1359 significantly enhanced AML cell motility (Fig 1. from 2.2 um/min to 5.4 um/min, p<0.001). Individual cells were dislodged from the niche and traveled long-distance. The combined inhibition of E-selectin and CXCR4 depleted BM leukemia cells in vascular niche areas. In a patient-derived primary AML xenograft (PDX) model (harboring mutations in JAK2 and c-Kit), combinatorial treatment of GMI-1359 with Ven/HMA significantly reduced BM retention of LSC compared to control cohorts or to Ven/HMA alone (p = 0.02 and p=0.003, respectively).In order to better understand how the augmented AML mobilization improves the efficacy of AML therapy, BM cells from PDX mice treated for 2 weeks with GMI-1271, GMI-1359, Ven/HMA, and their combinations were analyzed by single-cell proteomics (CyTOF). Blockade of E-selectin alone or dual E-selectin/CXCR4 inhibition in combination with Ven/HMA diminished levels of E-selectin ligand, mTOR, pFAK, pRb, cMyc, while increasing p21 and cleaved caspase3, which was associated with significant reduction of BM-resident LSC compared to Ven/HMA alone (CD45+34+CD38-CD123+, p= 0.03). AML blasts from the BM of the combinatorial treatment groups showed altered signaling including decreased Ki67, pRb, pNFkB, pPI3K, and E-selectin ligand, and increased levels of cleaved caspase 3. We further found that Ven/HMA significantly diminished CD31+ EC in the BM compared to control cohorts (p= 0.009). However, pharmacological antagonists of E-selectin or E-selectin/CXCR4 protected EC from Ven/HMA-induced detrimental insults through upregulation of survival signaling cascades including pAKT, pERK, pMAPK and decreased eNOS expression in EC compared to Ven/HMA treatment alone. Both EC and MSC were protected by dual inhibition of E-selectin/CXCR4 with GMI-1359. We also observed upregulated pro-survival signaling pathways such as phosphorylation of AKT-MAPK-ERK along with increased Bcl-xL, Bcl-2, and Idu expression in MSC from the GMI-1359 + Ven/HMA treated PDX mice compared to Ven/HMA single treatment cohorts.Collectively, our results provide strong evidence that co-targeting E-selectin/CXCR4 with GMI-1359 profoundly reduces BM retention of LSC as well as protects BM niche component cells from apoptosis induced by targeted therapy, resulting in improving the anti-leukemia activity of Ven/HMA therapy in AML. [Display omitted] DisclosuresFogler: GlycoMimetics Inc.: Current Employment, Current equity holder in publicly-traded company, Patents & Royalties. Magnani: GlycoMimetics Inc.: Current Employment, Current equity holder in publicly-traded company, Patents & Royalties. Carter: Ascentage: Research Funding; Syndax: Research Funding. Andreeff: Oxford Biomedica UK: Research Funding; ONO Pharmaceuticals: Research Funding; AstraZeneca: Research Funding; Reata, Aptose, Eutropics, SentiBio; Chimerix, Oncolyze: Current holder of individual stocks in a privately-held company; Karyopharm: Research Funding; Breast Cancer Research Foundation: Research Funding; Syndax: Consultancy; Daiichi-Sankyo: Consultancy, Research Funding; Novartis, Cancer UK; Leukemia & Lymphoma Society (LLS), German Research Council; NCI-RDCRN (Rare Disease Clin Network), CLL Foundation; Novartis: Membership on an entity's Board of Directors or advisory committees; Amgen: Research Funding; Aptose: Consultancy; Glycomimetics: Consultancy; Medicxi: Consultancy; Senti-Bio: Consultancy.

FLT3 Inhibitors Upregulate CXCR4 and E-Selectin Ligands and CD44 Via ERK Suppression in AML Cells, and Blockade of CXCR4 and E-Selectin Signaling with GMI-1359 Overcomes AML Resistance to Quizartinib in Vitro and In Vivo

FLT3 Inhibitors Upregulate CXCR4 and E-Selectin Ligands and CD44 Via ERK Suppression in AML Cells, and Blockade of CXCR4 and E-Selectin Signaling with GMI-1359 Overcomes AML Resistance to Quizartinib in Vitro and In Vivo

Phase 1/2 study of uproleselan added to chemotherapy in patients with relapsed or refractory acute myeloid leukemia

Uproleselan (GMI-1271) is a novel E-selectin antagonist that disrupts cell survival pathways, enhances chemotherapy response, improves survival in mouse xenograft and syngeneic models, and decreases chemotherapy toxicity in vivo. A phase 1/2 study evaluated the safety, tolerability, and antileukemic activity of uproleselan (5-20 mg/kg) with MEC (mitoxantrone, etoposide, and cytarabine) among patients with relapsed/refractory (R/R) acute myeloid leukemia (AML). Among the first 19 patients, no dose-limiting toxicities were observed. The recommended phase 2 dose (RP2D) was 10 mg/kg twice daily. An additional 47 patients with R/R AML were treated with uproleselan at the RP2D plus MEC. At the RP2D, the remission rate (complete response [CR]/CR with incomplete count recovery [CRi]) was 41% (CR, 35%), and the median overall survival (OS) was 8.8 months. In a separate cohort, 25 newly diagnosed patients age ≥60 years received uproleselan at the RP2D plus cytarabine and idarubicin (7 + 3). In these frontline patients, the CR/CRi rate was 72% (CR, 52%), and the median OS was 12.6 months. The addition of uproleselan was associated with low rates of oral mucositis. E-selectin ligand expression on leukemic blasts was higher in patients with relapsed vs primary refractory AML and in newly diagnosed older patients with high-risk cytogenetics and secondary AML. In the R/R cohort, E-selectin expression >10% was associated with a higher response rate and improved survival. The addition of uproleselan to chemotherapy was well tolerated, with high remission rates, low induction mortality, and low rates of mucositis, providing a strong rationale for phase 3 randomized confirmatory studies. This trial was registered at www.clinicaltrials.gov as #NCT02306291.

Abstract LB164: A novel glycomimetic compound (GMI-1757) with dual functional antagonism to E-selectin and galectin-3 attenuates fibrosis, facilitates mononuclear cell infiltration and optimizes anti-PD-L1 therapeutic activity in a pancreatic adenocarcinoma model

Abstract The inability to generate robust anti-tumor responses with therapeutics affecting T cell function has been ascribed to multiple parameters such as the tumor fibrotic compartment and the degree/makeup of host infiltrative cells. These parameters are affected through pathways involving the interaction of galectin-3 and E-selectin with cognate ligands. In the current investigations we assessed the impact of a dual E-selectin and galectin-3 glycomimetic antagonist, GMI-1757, in combination with anti-PD-L1 on tumor progression using an orthotopic pancreatic carcinoma model. Eight days following the orthotopic injection of 5x105 luciferase-enabled Pan02 carcinoma cells into the pancreas of female C57BL/6 mice, mice began daily IP treatment for 11 days with control diluent (C) or 50 mg/kg GMI-1757. Beginning on study day 15 C- or GMI-1757-treated mice were given 10 mg/kg anti-PD-L1 or an isotype control antibody (LTF-2). Sixteen mice were allocated into each of 4 groups: group 1 = C + LTF-2; group 2 = GMI-1757 + LTF-2; group 3 = C + anti-PD-L1; and group 4 GMI-1757 + PD-L1. Bioluminescence imaging (BLI) intensities were determined over the course of the study (Day 0 to 26) to assess anti-tumor activity, and tissue was excised at study conclusion to assess fibrosis and immune infiltration. Tumor growth progressed in mice treated with LTF-2 in combination with C or GMI-1757 (final %T/C of 100 and 60.7, respectively), and appeared arrested in mice treated with C and anti-PD-L1 antibody (%T/C = 18.3). In contrast, treatment of tumor-bearing mice with GMI-1757 in combination with anti-PD-L1 antibody resulted in tumor regressions in 5/8 mice with a final %T/C = 0.60. Using a histopathologic scoring system, the extent of fibrosis within the primary pancreatic tumors was dependent on treatment with GMI-1757. Tumor fibrosis was moderate to marked in groups 1 and 3, and minimal to mild in groups 2 and 4. These results were confirmed by morphometric analysis of Masson's Trichrome stained sections where the percent tumor fibrosis area was approximately 29 and 19% in groups 1 and 3, respectively, and 10 and 5% in groups 2 and 4, respectively. Notably, tumor infiltrating immune cells were enhanced with GMI-1757 treatment as compared to C (moderate to marked vs. minimal). The immune phenotypes comprising the increased mononuclear cell infiltration into orthotopic Pan02 tumors continues to be investigated. In summary, we report on the use of a dual E-selectin and galectin-3 antagonist to attenuate fibrosis and enhance mononuclear cell infiltration in an orthotopic model of pancreatic carcinoma. The shifts in fibrotic response and cellular infiltration obtained with GMI-1757 treatment in this model creates a more robust antitumor effect when combined with anti-PD-L1 treatment compared to anti-PD-L1 treatment alone. The impact of GMI-1757 to combine with immune modulators where fibrosis and restricted host cell infiltration negatively impact tumor response continues to be investigated. Citation Format: William E. Fogler, Theodore A. Smith, Ji-Won Lee, Silvia Locatelli-Hoops, David Stewart, John Peterson, John L. Magnani. A novel glycomimetic compound (GMI-1757) with dual functional antagonism to E-selectin and galectin-3 attenuates fibrosis, facilitates mononuclear cell infiltration and optimizes anti-PD-L1 therapeutic activity in a pancreatic adenocarcinoma model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr LB164.

Glycomimetic Antagonist of E-Selectin, GMI-1271, Enhances Therapeutic Activity of the Hypomethylating Agent, 5-Azacitidine, in the KG1 Model of AML

Components of the bone marrow stroma afford a permissive environment for the malignant progression of AML. These stromal components include E-selectin, an adhesion molecule expressed in the bone marrow vasculature. Through the recognition and binding of E-selectin by specific ligands on AML blasts, survival pathways are activated within the blasts leading to the development of/increase in chemoresistance. GMI-1271 is a novel E-selectin antagonist that disrupts this process, down-regulating survival pathways and enhancing chemotherapy response with improved survival in mouse xenograft and syngeneic AML tumor models. GMI-1271 is currently in clinical trials for the treatment of AML in combination with intensive chemotherapy; the FDA has granted Breakthrough Therapy Designation to GMI-1271 for the treatment of adults with relapse/refractory AML. Azacitidine (5-AZC) is a DNA-methyltransferase inhibiting cytosine nucleoside analog that at low doses induces DNA hypomethylation and transcriptional activation, while at higher doses is directly cytotoxic to neoplastic cells including AML blasts. Vidaza (5-AZC, Celgene) is approved in Europe for the treatment of limited populations with AML. In these studies we evaluated the anti-tumor activity of GMI-1271 in combination with 5-AZC in the KG1 AML tumor model to identify if there was potential for therapeutic benefit of the combination.Seven days post i.v. injection of KG1 cells, female NSG mice (n=10/group) were treated with saline; GMI-1271 (40 mg/kg IP QDx14) alone; 5-AZC (5 mg/kg IP Q3Dx5) alone, or the combination of GMI-1271 and 5-AZC. All treatments were well tolerated. The median survival time (MST) of mice treated with 5-AZC was 88 days and statistically different (P<0.005) to groups treated with saline (MST=69.5 days) or GMI-1271 alone (MST=69 days). At study conclusion (Day 110 post tumor injection), 20% of mice treated with 5-AZC remained alive; as expected all mice treated with saline or GMI-1271 alone succumbed to progressive tumor growth. Importantly, the therapeutic activity of 5-AZC was significantly enhanced when combined with GMI-1271 (MST>110 days, P=0.0217 compared to 5-AZC alone) with 70% of mice surviving to study conclusion. These results indicate that E-selectin/AML blast interaction in the KG1 model protects from the anti-tumor activity of 5-AZC and that GMI-1271 can attenuate this protection.To investigate the nature of the observed in vivo activity of GM-1271 and 5-AZC in the KG1 model, in vitro assays of cytotoxicity and E-selectin ligand expression in KG1 and the KG1a variant were assessed. The IC50 of 5-AZC alone or in combination with a saturating dose of GMI-1271 on KG1 cells was 518 and 528 nM, respectively. The IC50 of 5-AZC alone or in combination with GMI-1271 on KG1a cells was 637 and 711 nM, respectively. These results imply that the enhanced anti-tumor activity observed in vivo was not due to a simple shift in the cytotoxicity profile of 5-AZC. KGI and KG1a cells were subsequently cultured for 96 h in a noncytotoxic concentration (100 nM) of 5-AZC and the reactivity of the cells to HECA-452, a surrogate marker of E-selectin ligand, or binding E-selectin was determined by flow cytometry. Treatment with 5-AZC resulted in an increased reactivity of KG1 and KG1a cells to HECA-452 (29 and 27%, respectively compared to nontreated cells) and an increased binding of E-selectin to KG1 cells (32% compared to nontreated cells). Collectively, these results demonstrate that noncytotoxic concentrations of 5-AZC lead to an increased expression of E-selectin ligands on KG1 and KG1a cells.In summary, these studies show that even though the hypomethylating agent 5-AZC improves outcomes with AML, therapeutic potential could be improved. In this regard, 5-AZC at noncytotoxic concentrations promotes the increased expression of E-selectin ligands on AML blasts which may dampen 5-AZC activity. However, attenuation or disruption of E-selectin ligand/E-selectin interactions with GMI-1271 in combination with 5-AZC overcomes this tumor survival pathway with the potential to maximize therapeutic outcome. This work has relevance to patients with AML who do not achieve optimal or long-term response with 5-AZC and other hypomethylating agents. Clinical assessment of GMI-1271 with hypomethylating agents in AML is planned. DisclosuresFogler:GlycoMimetics: Employment, Equity Ownership. Smith:GlycoMimetics: Employment, Equity Ownership. Lee:GlycoMimetics: Employment, Equity Ownership. Magnani:GlycoMimetics, Inc.: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees.

Targeting E-selectin to Tackle Cancer Using Uproleselan.

Simple SummaryThis review focuses on eradicating cancer by targeting a surface protein expressed on the endothelium—E-selectin—with a novel drug, uproleselan (GMI-1271). Blocking E-selectin in the tumor microenvironment acts on multiple levels; uproleselan was shown (i) to inhibit cancer cell tethering, rolling and extravasating, i.e., cancer dissemination, (ii) to reduce adhesion and lose stem cell-like properties, (iii) to mobilize cancer cells to circulation where they are more susceptible to chemotherapy, which altogether contributes (iv) to overcome drug resistance. Uproleselan has been tested effective in leukemia, myeloma, pancreatic, colon and breast cancer cells, all of which can be found in the bone marrow as a primary or as a metastatic tumor site. In addition, uproleselan has a good safety profile in patients. It improves the efficacy of chemotherapy, reduces side effects such as neutropenia, intestinal mucositis and infections, and extends overall survival.E-selectin is a vascular adhesion molecule expressed mainly on endothelium, and its primary role is to facilitate leukocyte cell trafficking by recognizing ligand surface proteins. E-selectin gained a new role since it was demonstrated to be involved in cancer cell trafficking, stem-like properties and therapy resistance. Therefore, being expressed in the tumor microenvironment, E-selectin can potentially be used to eradicate cancer. Uproleselan (also known as GMI-1271), a specific E-selectin antagonist, has been tested on leukemia, myeloma, pancreatic, colon and breast cancer cells, most of which involve the bone marrow as a primary or as a metastatic tumor site. This novel therapy disrupts the tumor microenvironment by affecting the two main steps of metastasis—extravasation and adhesion—thus blocking E-selectin reduces tumor dissemination. Additionally, uproleselan mobilized cancer cells from the protective vascular niche into the circulation, making them more susceptible to chemotherapy. Several preclinical and clinical studies summarized herein demonstrate that uproleselan has favorable safety and pharmacokinetics and is a tumor microenvironment-disrupting agent that improves the efficacy of chemotherapy, reduces side effects such as neutropenia, intestinal mucositis and infections, and extends overall survival. This review highlights the critical contribution of E-selectin and its specific antagonist, uproleselan, in the regulation of cancer growth, dissemination, and drug resistance in the context of the bone marrow microenvironment.

Restoration of Normal Blood Flow in Mouse Models of Sickle Cell Vaso-Occlusion Following Intravenous or Subcutaneous Administration of a Highly Potent E-Selectin Specific Inhibitor

Background: In sickle cell disease (SCD), recurrent episodes of vaso-occlusive crisis (VOC) contribute to morbidity and accelerated mortality. Analysis of the RESET Phase 3 study demonstrated that administration of the pan-selectin inhibitor, rivipansel, early in the course of VOC was associated with efficacy outcomes in favor of rivipansel, suggesting that reversal of vaso-occlusion may be achieved by treatment early in VOC. While both P- and L-selectin likely contribute to the underlying inflammatory cascade leading to VOC, previous studies have shown that E-selectin expression on the vascular luminal surface appears to be crucial for adhesion of sickled (SS)-RBC and leukocytes to the endothelium. GMI-1687 is an innovative potent E-selectin specific antagonist that demonstrates high bioavailability following subcutaneous (SC) administration. In the current investigation, we report on in vitro and in vivo studies evaluating the activity of GMI-1687 in SCD mouse models of VOC. Methods: Townes transgenic mice and nude mice transfused with human SS-RBC were used as in vivo models of SCD. VOC was initiated by TNF injection in both models. Adhesion to inflamed venules and development of VOC was recorded using intravital microscopy at sites of window chamber implants. The in vivo activity of E-selectin inhibition with GMI-1687 was assessed following IV injection in the nude mouse and Townes transgenic models, and SC injection in the nude mouse model of SCD. Results: In an initial study, IV treatment with saline or GMI-1687 was initiated 30 min post infusion of SS-RBC (t30) into TNF-treated mice, a time during which sickle cells are adherent to inflamed venules and VOC has initiated, and repeated 30 minutes later (t60). As expected, mice treated with saline alone showed marked adhesion of human SS-RBC to venules (61% occlusion of the total vessels recorded at t30) with evident blood stasis. Under these conditions, GMI-1687 at 40 μg/kg significantly reversed SS-RBC adhesion and VOC at both t30 and t60 (p=0.002 and 0.0073, respectively, compared to saline treatment). This led to restoration of blood flow in 85% of the venules in GMI-1687 treated mice vs. 18% of vessels with normal blood flow in the control saline treated group. Because of reduced sickle cell adhesion and VOC, the number of circulating SS-RBC increased by approximately 6-fold in mice treated with GMI-1687. Similar anti-adhesive and anti-VOC activities of GMI-1687 treatment were obtained in the Townes transgenic model of SCD where following IV administration of the E-selectin antagonist, adhesion of sickled RBCs was significantly diminished and as a result, the percentage of the total occluded vessel segments was reduced by 80-90%. We next assessed the activity of SC administration with 40 μg/kg GMI-1687 on human SS-RBC adhesion and development of VOC in the nude mouse model. Similar to the IV experiments, GMI-1687 initiated at t30 and t60 post SS-RBC infusion significantly reduced SS-RBC adhesion in inflamed vessels (69% reduction at t30, and 63% at t60, p=0.0001). The reduction in SS-RBC adhesion led to a lower incidence of VOC (2% compared to 30% of vessels in mice injected with saline), and a 5.1-fold improvement (p=0.0315) in the number of circulating human SS-RBC. It is noteworthy that the effective dose of GMI-1687 used in these IV and SC studies resulting in the restoration of normal blood flow was approximately 500-fold less than that reported for rivipansel. Conclusions: These data demonstrate that the administration of the potent and E-selectin specific compound GMI-1687 was effective in restoring blood flow in two mouse models of SCD, and support the development of GMI-1687 formulated for SC use and self-administration with the potential for early intervention of VOC. Disclosures Thamilarasan: GlycoMimetics: Research Funding. Fogler:GlycoMimetics: Current Employment, Current equity holder in publicly-traded company, Patents &amp; Royalties. Magnani:GlycoMimetics, Inc.: Current Employment, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees, Patents &amp; Royalties. Zennadi:GlycoMimetics: Research Funding.

Abstract 486: Enhanced survival of lethally-irradiated mice with HSC reconstitution in combination with the E-selectin antagonist, GMI-1271 (uproleselan)

Abstract The involvement of the adhesion molecule E-selectin and its interactions with E-selectin ligands as pertains to hematopoietic stem cells (HSC) and transplantation has been investigated (Winkler et al. 2012; Winkler et al. 2014). These preclinical studies focused on the role of E-selectin and the use of uproleselan, an E-selectin antagonist, during HSC mobilization in harvesting procedures of donors to accelerate recovery in transplant recipients. However, the impact of E-selectin and uproleselan administration on transplant recipients was less clear. In the current investigations we assessed the survival outcome of bone marrow depleted mice when reconstituted with HSC in combination with uproleselan. Lethally-irradiated, bone marrow depleted C57BL/6 mice were reconstituted with bone-marrow harvested from a congenic strain. Twenty-four hours post irradiation (6Gy x2), cohorts of mice (n=10/group) were injected i.v. with 1 × 106 cells (study day 0) from congenic donors using three i.p. dosing regimens with 40 mg/kg uproleselan. These regimens were: (a) bid on study days 0 and1; (b) bid on study days 1 and 2; and (c) bid on study day 1 only. Control groups in this study included irradiated mice alone (expected survival = 0%), non-irradiated mice alone (expected survival = 100%), and irradiated, reconstituted mice (no uproleselan). The survival of mice was determined over the course of the study (Day 0 to 30). Additional parameters of evaluation included sinusoidal obstructive syndrome such as hepatic veno-occlusive disease known to be E-selectin dependent and a complication of HSC transplantation. Treatment with uproleselan as part of the transplant regimen significantly increased the median survival time (MST) of mice compared with the control group – the MST of mice treated with uproleselan and HSC was &amp;gt;30 days with 80-90% of mice alive at study completion. In contrast, the MST of irradiated mice (no transplant) was 11.5 days with no survivors at study conclusion. The MST of mice irradiated and transplanted with congenic HSC was 9 days with 40% survival on day 30. The impact of uproleselan on survival represented a &amp;gt;233.3% increase in life span. Flow cytometric analysis in all surviving mice on day 30 showed that the mean percentage of CD45.1+ cells from donor congenic mice was approximately 90% (blood and bone marrow) indicating that all surviving mice were successfully reconstituted. In summary, we report on a novel therapeutic use of inhibitors of E-selectin, such as uproleselan, which results in the increased survival of mice when combined with HSC transplantation for reconstitution of depleted and compromised bone marrow. The impact on increased host survival could extend to the use of peripheral blood and stem cell transplantations as a therapeutic option in various malignancies where curative intent is intended. Citation Format: William E. Fogler, Dylan Daniel, Sheri Barnes, Alden Wong, David Draper, John L. Magnani. Enhanced survival of lethally-irradiated mice with HSC reconstitution in combination with the E-selectin antagonist, GMI-1271 (uproleselan) [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 486.

Abstract 5867: Transcriptome profiling of ST3GAL4 and FUT7 in multiple tumor types and prognostic value in adult acute myeloid leukemia

Abstract E-selectin is a cell adhesion glycoprotein expressed on endothelial cells and participates in the development of environmental-mediated drug resistance and poor clinical outcome in cancer. The E-selectin antagonist, uproleselan, interrupts leukemic cell homing to the vascular niche and increases susceptibility to cytotoxic therapies. Recent clinical data demonstrate a correlation between leukemic cell surface E-selectin ligands and response to uproleselan, linking E-selectin ligand expression to response. Multiple genes involved in the glycan synthesis of E-selectin ligands are highly expressed in pediatric AML. Expression levels of two of these genes, ST3GAL4 and FUT7 are associated with poor outcome and are associated with cell surface E-selectin ligand expression. In the current studies we extend transcriptome profiling of E-selectin ligand forming glycosylation genes with an emphasis on ST3GAL4 and FUT7 in different cancers. Initially, we investigated expression levels of ST3GAL4 and FUT7 in 10,258 samples covering 33 cancer types from the TCGA PanCanAtlas. ST3GAL4 and FUT7 were consistently expressed in the majority of cancers evaluated. The cancer types that expressed ST3GAL4 most highly were melanoma (uveal and skin), kidney chromophobe, adrenocortical carcinoma and bladder urothelial carcinoma, while for FUT7, it was AML, diffuse large B-cell lymphoma, thymoma, testicular germ cell tumors and head and neck squamous cell carcinoma. Of particular interest was the identification of adult AML for the highest expression of FUT7 and high expression of ST3GAL4 (mean log2 gene expression = 8.1 and 9.4, respectively). Augmented expression of FUT7 and ST3GAL4 were also observed in an analysis of 39 AML cell lines among the 1,457 cell lines comprising the Cancer Cell Line Encyclopedia RNAseq data set. The prognostic significance of FUT7 and ST3GAL4 in adult AML was further assessed using the TCGA-LAML RNAseq dataset for differential expression and associations with overall survival (OS). The data set included 151 RNAseq profiles of bone marrow samples from adult patients with AML, and within this data set the status of the FMS-like tyrosine kinase 3 (FLT3) proto-oncogene was considered. Mutational alterations of FLT3 are associated with higher risk of relapse and shorter OS compared with wild-type FLT3. ST3GAL4 and FUT7 were both identified as being upregulated (fold-change = 1.73 and 1.40, respectively) in the mutated FLT3 subset (n=46) as compared to wild type FLT-3 (p=0.000033 and 0.046, respectively). Notably in the mutated FLT-3 subset expression of FUT7 was significantly associated with a poor prognosis and decreased OS (HR = 4.56, p= 0.015). Collectively, these studies extend the prognostic importance of the E-selectin ligand glycosylation genes, ST3GAL4 and FUT7, to adult AML where these genes may be useful as predictive biomarkers. In addition, these studies suggest potential additional tumor types beyond AML where treatment protocols with uproleselan may have therapeutic benefits. Citation Format: William E. Fogler, Vince Deng, David Stewart, Michael Jarman, Simone Daminelli, Adrian Carr, John L. Magnani. Transcriptome profiling of ST3GAL4 and FUT7 in multiple tumor types and prognostic value in adult acute myeloid leukemia [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 5867.

Alliance A041701 - a Randomized Phase 2/3 Study of Conventional Chemotherapy +/- Uproleselan (GMI-1271) in Older Adults with Acute Myeloid Leukemia (AML) Receiving Intensive Induction Chemotherapy

Uproleselan (GMI-1271) is a novel antagonist of E-selectin, an adhesion molecule expressed on endothelial cells. E-selectin is expressed transiently in the normal vasculature during an inflammatory response and constitutively in the bone marrow (BM). Binding of E-selectin (E-sel) to sialyl Lex, the E-sel ligand (E-sel-L), on the leukemic cell surface activates cell survival pathways and promotes chemotherapy resistance in AML. In preclinical models, blockade of E-selectin with uproleselan disrupts the activation of cell survival pathways and enhanced the efficacy of chemotherapy across multiple AML tumor models. Furthermore, uproleselan protected against chemotherapy-induced mucositis by regulating macrophage trafficking to the site of injury in the gut lining. A previous phase I/II study of uproleselan added to chemotherapy in patients with untreated AML (older adults ≥60 yrs) and relapsed/refractory AML (≥ 18yrs) showed promising remission rates (CR/CRi) and survival outcomes, and reduced rates of mucositis. In the newly diagnosed older patients, high remission rates were achieved overall (CR/CRi 72%) and for the high risk subgroup with sAML (CR/CRi 69%). In this phase 2/3 study, we will test the addition of uproleselan to a standard daunorubicin/cytarabine regimen in older adults with previously untreated AML. The study will enroll patients age ≥ 60 yrs with untreated acute myeloid leukemia. Patients with acute promyelocytic leukemia, activating mutations in FLT3, or evidence of central nervous system involvement are excluded. Subjects are randomized to 7+3 induction (cytarabine + daunorubicin) +/- uproleselan. Subjects achieving a CR/CRi may receive up to 3 cycles of consolidation with intermediate dose cytarabine 2 gm/m2 IV d1-5 +/- uproleselan. The primary phase II objective is to compare the event-free survival (EFS). A sample size of 262 patients was selected for the phase II to detect an improvement in median EFS from 7 months to 11 months (HR= 0.64) with &amp;gt; 95% power, using a log rank test. The phase III primary objective will compare overall survival (OS). For the phase III, a sample size of 335 evaluable patients per arm (670 total inclusive of patients enrolled in phase II) will provide &amp;gt;90% power to detect an improvement in median OS from 12 months to 16 months (HR= 0.75), using a log-rank test. Correlative studies will measure E-selectin ligand on AML blasts and soluble E-selectin and will also measure minimal residual disease after remission induction by multiparameter flow cytometry. A comprehensive geriatric assessment will identify baseline measures associated with EFS and develop a risk model to predict OS among older adults receiving intensive AML therapy. The study is endorsed by SWOG and ECOG-ACRIN and opened to enrollment on 1/16/2019. Figure Disclosures Uy: Astellas: Consultancy; Pfizer: Consultancy; Curis: Consultancy; GlycoMimetics: Consultancy. Dinner:Agios: Consultancy; Pfizer: Consultancy; AstraZeneca: Consultancy. Strickland:Jazz: Consultancy; Sunesis Pharmaceuticals: Research Funding; Pfizer: Consultancy; Kite: Consultancy; Astellas Pharma: Consultancy; AbbVie: Consultancy. Liesveld:Onconova: Other: Data safety monitoring board; Abbvie: Membership on an entity's Board of Directors or advisory committees. Byrd:Acerta: Research Funding; Acerta: Research Funding; Genentech: Research Funding; Janssen: Consultancy, Other: Travel Expenses, Research Funding, Speakers Bureau; Ohio State University: Patents &amp; Royalties: OSU-2S; BeiGene: Research Funding; Janssen: Consultancy, Other: Travel Expenses, Research Funding, Speakers Bureau; BeiGene: Research Funding; Gilead: Other: Travel Expenses, Research Funding, Speakers Bureau; Gilead: Other: Travel Expenses, Research Funding, Speakers Bureau; BeiGene: Research Funding; Novartis: Other: Travel Expenses, Speakers Bureau; Novartis: Other: Travel Expenses, Speakers Bureau; Gilead: Other: Travel Expenses, Research Funding, Speakers Bureau; Pharmacyclics LLC, an AbbVie Company: Other: Travel Expenses, Research Funding, Speakers Bureau; Pharmacyclics LLC, an AbbVie Company: Other: Travel Expenses, Research Funding, Speakers Bureau; Novartis: Other: Travel Expenses, Speakers Bureau; TG Therapeutics: Other: Travel Expenses, Research Funding, Speakers Bureau; Genentech: Research Funding; Genentech: Research Funding; Pharmacyclics LLC, an AbbVie Company: Other: Travel Expenses, Research Funding, Speakers Bureau; Acerta: Research Funding; TG Therapeutics: Other: Travel Expenses, Research Funding, Speakers Bureau; Ohio State University: Patents &amp; Royalties: OSU-2S; Ohio State University: Patents &amp; Royalties: OSU-2S; TG Therapeutics: Other: Travel Expenses, Research Funding, Speakers Bureau; Janssen: Consultancy, Other: Travel Expenses, Research Funding, Speakers Bureau. Stone:Argenx, Celgene, Takeda Oncology: Other: Data and Safety Monitoring Board/Committee: ; AbbVie, Actinium, Agios, Argenx, Arog, Astellas, AstraZeneca, Biolinerx, Celgene, Cornerstone Biopharma, Fujifilm, Jazz Pharmaceuticals, Amgen, Ono, Orsenix, Otsuka, Merck, Novartis, Pfizer, Sumitomo, Trovagene: Consultancy; Novartis, Agios, Arog: Research Funding. OffLabel Disclosure: Uproleselan in AML

A Double-Blind, Placebo-Controlled, Phase 3 Registration Trial to Evaluate the Efficacy of Uproleselan (GMI-1271) with Standard Salvage Chemotherapy in Patients with Relapsed/Refractory (R/R) Acute Myeloid Leukemia

A Double-Blind, Placebo-Controlled, Phase 3 Registration Trial to Evaluate the Efficacy of Uproleselan (GMI-1271) with Standard Salvage Chemotherapy in Patients with Relapsed/Refractory (R/R) Acute Myeloid Leukemia

A Novel and Potent Inhibitor of E-Selectin, GMI-1687, Attenuates Thrombus Formation and Augments Chemotherapeutic Intervention of AML in Preclinical Models Following Subcutaneous Administration

Abstract Uproleselan (GMI-1271), an E-selectin antagonist, has been shown in preclinical models to disrupt activation of cell survival pathways in acute myeloid leukemia (AML), enhance chemotherapy efficacy, and improve survival. Uproleselan received FDA breakthrough therapy designation for adult relapsed/refractory AML in 2017 and Phase III studies are ongoing. In the present studies we report on the in vitro and in vivo comparative activities of an innovative high potency E-selectin antagonist, GMI-1687, a potential subcutaneously administered follow-on drug candidate to Uproleselan. The binding constant, association and dissociation rates of GMI-1687 to immobilized recombinant human (rh) E-selectin were determined by surface plasmon resonance (SPR) at 25oC. The KD of GMI-1687 was 2.4 nM, with Kon = 3 x 106 M-1s-1 and Koff = 1 x10-2 s-1. Under similar experimental conditions the KD of Uproleselan was 520 nM with Kon = 0.02 x 106 M-1s-1 and Koff = 1 x10-2 s-1. GMI-1687 was evaluated for its ability to inhibit binding of sialyl Lea to immobilized rh E-selectin. The median IC50 (n=6 independent assays) of GMI-1687 and Uproleselan in this assay was 15 and 550 nM, respectively. The in vitro activity of GMI-1687 to release adherent KG1a AML cells from E-selectin coated wells was also determined. GMI-1687 at 100 nM detached approximately 55% of adherent AML cells and was significantly different from Uproleselan at an identical concentration (38% detachment, P=0.0216). The percent bioavailability (%F) of GMI-1687 was evaluated in male Sprague-Dawley rats following intravenous (IV) and subcutaneous (SC) routes of administration at 5 mg/kg. The mean (+/- SD) SC %F for GMI-1687 was 126 +/- 3.8%. GMI-1687 also showed high bioavailability in CD-1 mice after SC administration of 0.58 mg/kg with %F = 132 +/-38. The in vivo therapeutic activity of GMI-1687 following SC administration was assessed in an acute model of inferior vena cava (IVC) thrombosis and a tumor model of AML.Immediately following the induction of a non-occlusive thrombosis via electrical stimulation (250 mAmp) of the IVC, cohorts of male C57BL/6J mice (n=5/group) were given a single SC injection of saline (0.1 mL); Uproleselan (40 mg/kg); or GMI-1687 (4 mg/kg, 0.4 mg/kg or 0.04 mg/kg), and twenty-four hrs post thrombus induction the IVC was harvested from all mice and thrombus weights were determined. Treatment with GMI-1687 decreased thrombus formation with significant inhibition at 0.04 mg/kg (92%, P&lt;0.001 compared to saline control). The inhibition of thrombus formation with GMI-1687 dosed at 0.04 mg/kg was statistically indistinguishable from Uproleselan administered SC at 40 mg/kg (97% inhibition). The therapeutic activity of SC GMI-1687 was also observed in combination with chemotherapy in a U937 tumor model. Three days post IV injection of U937 tumor cells, bone marrow ablated, female NOD/SCID mice (n=10/group) were treated with saline (0.1 mL SC QDx14); GMI-1687 (0.04 mg/kg SC QDx14) alone; cytarabine (AraC 300 mg/kg IP QDx3) + daunorubicin (DNR 3 mg/kg IV QDx1), or the combination of GMI-1687 and AraC + DNR. All treatments were well tolerated. The median survival time (MST) of mice treated with AraC + DNR was 36 days and statistically different (P&lt;0.001) to groups treated with saline (MST=22 days) or GMI-1687 alone (MST=23 days). Importantly, the therapeutic activity of AraC+DNR was significantly enhanced when combined with GMI-1687 (MST&gt;47.5 days, P=0.0153 compared to AraC+DNR alone). In summary, a highly potent innovative antagonist of E-selectin, GMI-1687, has been produced that demonstrates high bioavailability following SC administration. SC injection of GMI-1687 shows significant activity in preclinical models previously reported for parenteral administration of Uproleselan, but at approximately 250-fold lower dose. GMI-1687 is therefore well-positioned for potential use in outpatient treatment settings where an E-selectin antagonist has therapeutic relevance. IND-enabling studies with GMI-1687 are currently underway. Disclosures Peterson: GlycoMimetics: Employment, Equity Ownership. Baek:GlycoMimetics: Employment, Equity Ownership. Locatelli-Hoops:GlycoMimetics: Employment, Equity Ownership. Lee:GlycoMimetics: Employment, Equity Ownership. Deng:GlycoMimetics: Employment, Equity Ownership. Stewart:GlycoMimetics: Employment, Equity Ownership. Smith:GlycoMimetics: Employment, Equity Ownership. Fogler:GlycoMimetics: Employment, Equity Ownership. Magnani:GlycoMimetics: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees.

Uproleselan (GMI-1271), an E-Selectin Antagonist, Improves the Efficacy and Safety of Chemotherapy in Relapsed/Refractory (R/R) and Newly Diagnosed Older Patients with Acute Myeloid Leukemia: Final, Correlative, and Subgroup Analyses

Background Binding of E-selectin (E-sel) to sialyl Lex on the leukemic cell surface activates cell survival pathways and promotes chemotherapy resistance in AML. Expression of the E-sel ligand (E-sel-L) is associated with increased relapse and poor survival. Uproleselan (GMI-1271), a novel E-sel antagonist, disrupts cell survival pathway activation, enhances chemotherapy response and protects from toxicity with improved survival in vivo. We added uproleselan to mitoxantrone, etoposide, cytarabine (MEC) chemotherapy for R/R AML patients (pts) and to cytarabine and idarubicin (7+3) induction for older, treatment naive (TN) AML pts. Here we report on final outcomes and correlative studies. Methods A Phase (Ph) 1/2 trial evaluated safety and efficacy of escalating doses of upro (5-20 mg/kg) combined with MEC in pts with R/R AML. The recommended Ph 2 dose (RP2D) was 10 mg/kg. Ph 2 added pts ≥60 yrs with TN AML treated with upro and 7+3. Uproleselan was given 24 hrs prior, every 12 hrs during and 48 hrs post chemotherapy. Responders could receive consolidation therapy (1 cycle MEC or 1-3 cycles IDAC) with uproleselan. Baseline E-sel-L expression on AML blasts (CD45+,SSC) and leukemic stem cells (LSC, CD34+CD38-CD123+) in blood and bone marrow (BM) was assessed by flow cytometry, for percentage of blasts binding to E-sel-Fc chimeric protein and HECA452 (antibody to sialyl Lex). Post-induction measurable residual disease (MRD-/MRD+) was assessed locally. Results 91 pts were enrolled (Ph 1 R/R=19; Ph 2 R/R=47. TN=25). Median age in R/R pts was 59 yrs (26-84) with 22/66 (33%) primary refractory, 22 (33%) CR1 Median age in TN pts was 67 yrs (60-79). 48% had adverse risk (ELN) and 52% secondary AML (sAML). Uproleselan was well tolerated, with no increase in adverse events, no Grade 3/4 mucositis, and mortality 8% (30d) and 12% (60d). CR/CRi was 72% (all), and 69% (sAML). 5/9 (56%) evaluable pts were MRD-. 19/25 (76%) proceeded to further anti-leukemic therapy; 11 (44%) proceeded to SCT. Median (95% CI) EFS, OS, and remission duration were 9.2 m (3.0-12.6), 12.6 m (9.9-NR), and 10.4 m (7.1-17.8) respectively; 1-year OS was 52%. For sAML, median (95% CI) EFS and OS were 7.7 m (1.1-9.5) and 10.5m (4.4-NR), respectively. For TN/MRD-, 1-year OS was 60%. E-sel-L was detectable on BM blasts in all 24 evaluable pts: median expression 31% (2-92) of blasts. In a subset of MRD evaluable pts (N=8), E-sel-L expression was higher in those who were MRD+ (35% vs 8%). In BM blasts (Figure N=24), LSC expression of E-sel-L correlated with blast E-sel-L (R2=0.87, p Conclusion The addition of uproleselan to chemotherapy was well tolerated, with low oral mucositis rates, high remission rates, high MRD- and transplant rates, and promising survival outcomes in pts with R/R and TN AML. High E-sel-L expression is associated with improved remission and survival with uproleselan treatment in R/R AML. Phase III studies in pts with R/R and (older) TN AML are underway. Disclosures DeAngelo:Shire: Honoraria; Amgen: Consultancy; BMS: Consultancy; ARIAD: Consultancy, Research Funding; Blueprint Medicines: Honoraria, Research Funding; Takeda: Honoraria; Glycomimetics: Research Funding; Novartis Pharmaceuticals Corporation: Consultancy, Honoraria; Amgen: Consultancy; Novartis Pharmaceuticals Corporation: Consultancy, Honoraria; Incyte: Consultancy, Honoraria; Glycomimetics: Research Funding; Blueprint Medicines: Honoraria, Research Funding; Pfizer Inc: Consultancy, Honoraria; Shire: Honoraria; BMS: Consultancy; Takeda: Honoraria; Incyte: Consultancy, Honoraria; Pfizer Inc: Consultancy, Honoraria; ARIAD: Consultancy, Research Funding. Jonas:Glycomimetics: Research Funding; Genentech/Roche: Research Funding; Celgene: Consultancy, Research Funding; Accelerated Medical Diagnostics: Research Funding; Daiichi Sankyo: Research Funding; Incyte: Research Funding; Esanex: Research Funding; Tolero: Consultancy; Kalobios: Research Funding; LP Therapeutics: Research Funding; Amgen: Consultancy; Pharmacyclics: Research Funding; Forma: Research Funding; AbbVie: Consultancy, Research Funding. Liesveld:Onconova: Other: DSMB; Abbvie: Honoraria. Bixby:GlycoMimetics: Research Funding. Advani:Glycomimetics: Consultancy; Novartis: Consultancy; Amgen: Research Funding; Pfizer: Honoraria, Research Funding. Marlton:GlycoMimetics: Research Funding; Novartis: Honoraria, Membership on an entity9s Board of Directors or advisory committees; Roche: Honoraria, Membership on an entity9s Board of Directors or advisory committees; Janssen: Honoraria, Membership on an entity9s Board of Directors or advisory committees; AbbVie: Honoraria, Membership on an entity9s Board of Directors or advisory committees; Astellas: Honoraria, Membership on an entity9s Board of Directors or advisory committees; Celgene: Membership on an entity9s Board of Directors or advisory committees; Pfizer: Membership on an entity9s Board of Directors or advisory committees. O9Dwyer:Janssen: Membership on an entity9s Board of Directors or advisory committees, Research Funding; BMS: Research Funding; Onkimmune: Equity Ownership, Membership on an entity9s Board of Directors or advisory committees, Research Funding; Celgene: Research Funding; Glycomimetics: Research Funding; Abbvie: Membership on an entity9s Board of Directors or advisory committees. Fogler:GlycoMimetics: Employment, Equity Ownership. Wolfgang:GlycoMimetics: Employment, Equity Ownership. Magnani:GlycoMimetics: Employment, Equity Ownership, Membership on an entity9s Board of Directors or advisory committees. Thackray:GlycoMimetics: Employment, Equity Ownership. Becker:GlycoMimetics: Research Funding.