These results demonstrate the lack of MetAP1 aminopeptidase inhibition by SDX-7539 and TNP-470, and supports the findings regarding selectivity of these compounds for MetAP2.

Contains methods and results from additional experiments

Levels of small molecule MetAP2 inhibitors SDX-7539 and TNP-470 in plasma and CSF, used to assess CNS penetration.

Results of enzyme assays that assessed the potential inhibitory activity of the MetPA2 inhibitor against various enzymes.

Variance in the expression of genes between control and SDX-7320-treated tumor samples is displayed in this graph.

Shows binding assay performance and differential MetAP2 binding of test compounds.

Detailed information from the x-ray crystallography of SDX-7539 bound to MetAP2 that enabled determination of the co-crystal’s 3D structure.

Compares the 3D crystal structures of MetAP2 with SDX-7539 bound to that of MetAP2 with TNP-470

Results of competition for radioligand binding by SDX-9402 at different receptors, to assess the selectivity of the MetAP2 inhibitor against known drug targets.

This table provides details on where each cathepsin enzyme was obtained.

Shows the structures of the PDC, the benzenesulfonate salt form of the physiologically active small molecule, and three test compounds, along with the numbering system utilized for NMR assignment of evexomostat.

This graph displays the exposure-time profile of SDX-7539 in rats after IV administration of SDX-7320 or SDX-7539.

Abstract Methionine aminopeptidase type 2 (METAP2) is a ubiquitous, evolutionarily conserved metalloprotease fundamental to protein biosynthesis which catalyzes removal of the N-terminal methionine residue from nascent polypeptides. METAP2 is an attractive target for cancer therapeutics based upon its over-expression in multiple human cancers, the importance of METAP2-specific substrates whose biological activity may be altered following METAP2 inhibition, and additionally, that METAP2 was identified as the target for the anti-angiogenic natural product, fumagillin. Irreversible inhibition of METAP2 using fumagillin analogues has established the anti-angiogenic and anti-tumor characteristics of these derivatives; however, their full clinical potential has not been realized due to a combination of poor drug-like properties and dose-limiting central nervous system (CNS) toxicity. This report describes the physicochemical and pharmacological characterization of SDX-7320 (evexomostat), a polymer–drug conjugate of the novel METAP2 inhibitor (METAP2i) SDX-7539. In vitro binding, enzyme, and cell-based assays demonstrated that SDX-7539 is a potent and selective METAP2 inhibitor. In utilizing a high molecular weight, water-soluble polymer to conjugate the novel fumagillol-derived, cathepsin-released, METAP2i SDX-7539, limitations observed with prior generation, small molecule fumagillol derivatives were ameliorated including reduced CNS exposure of the METAP2i, and prolonged half-life enabling convenient administration. Multiple xenograft and syngeneic cancer models were utilized to demonstrate the anti-tumor and anti-metastatic profile of SDX-7320. Unlike polymer–drug conjugates in general, reductions in small molecule–equivalent efficacious doses following polymer conjugation were observed. SDX-7320 has completed a phase I clinical safety study in patients with late-stage cancer and is currently being evaluated in multiple phase Ib/II clinical studies in patients with advanced solid tumors.

TPS1129 Background: Alpelisib, a selective PI3K p110a inhibitor, was approved for breast cancer patients with PIK3CA mutations. An on-target toxicity of alpelisib is hyperglycemia and patients with baseline insulin resistance/elevated HbA1c are at greater risk of developing grade 3,4 hyperglycemia after receiving alpelisib. Restoring insulin sensitivity and reducing systemic insulin levels improved the efficacy of alpelisib in animal models of breast cancer. Evexomostat is a polymer-drug conjugate of a novel small molecule methionine aminopeptidase 2 (MetAP2) inhibitor. In normal mice, it reduced alpelisib-induced hyperglycemia/hyperinsulinemia and in an animal model of breast cancer synergized with alpelisib to regress tumor growth. In a Phase 1 trial, evexomostat improved insulin resistance in patients with elevated baseline insulin, reduced angiogenic factors in patients with elevated baseline levels, increased the insulin-sensitizing adipokine adiponectin and showed anti-metastatic effects in patients with advanced solid tumors. Methods: This is a phase 1b/2, open-label, single-arm pilot study (NCT05455619) in postmenopausal women with HR+, HER2- metastatic breast cancer with a PIK3CA mutation who progressed following treatment with endocrine therapy plus a CDK4/6 inhibitor and who are at risk for hyperglycemia (HbA1c between 5.5 and 6.4%inclusive). The primary goal is to determine the safety of evexomostat plus alpelisib and fulvestrant (the ‘triplet therapy’), including the severity and number of hyperglycemic events, as well as anti-tumor benefit. The starting dose of evexomostat in the triplet therapy is 36 mg/m 2 (n=6; one dose below the monotherapy MTD of 49 mg/m 2 ). Once the MTD of evexomostat in the triplet therapy has been defined, up to 20 patients will be enrolled at that dose. An additional 20 patients may be enrolled to further characterize the safety and anti-tumor effect of the triplet therapy (up to 52 patients). This trial is open to accrual at multiple sites in the USA. Safety analysis includes the type, frequency, and severity of treatment-emergent adverse events (TEAEs) per NCI CTCAE, v5, and the number and proportion of patients with grade 3 or 4 hyperglycemia during the first 2 cycles of therapy, with an estimate of the exact upper one-sided 97.5% confidence bound. Efficacy analyses include objective response rate (ORR, consisting of complete response (CR) and partial response (PR)). The number of patients without disease progression six months from the start of the triplet therapy will be assessed. The CBR of CRs, PRs plus stable disease ≥24 weeks from C1D1 will be calculated. Overall survival data will be summarized as available. QoL will be analyzed according to functional scores and recommendations in the EORTC scoring manual. ECOG performance status and change from baseline will be summarized. Clinical trial information: NCT05455619 .

Background: Breast cancer patients with mutation(s) in the PIK3CA gene have more aggressive disease and worse outcomes relative to patients without PIK3CA mutations. Alpelisib (Piqray), an inhibitor of PIK3CA, was approved for breast cancer patients with PIK3CA mutations. An on-target toxicity of alpelisib is hyperglycemia leading to hyperinsulinemia which may limit effectiveness of this drug. Patients with baseline metabolic dysfunction, insulin resistance, and/or elevated HbA1c are at greater risk of developing grade 3,4 hyperglycemia after receiving alpelisib (Piqray) than patients without metabolic dysfunction. Restoring insulin sensitivity and reducing systemic insulin levels improved the efficacy of alpelisib in preclinical models of breast cancer. Evexomostat is a polymer-drug conjugate of a novel small molecule methionine aminopeptidase 2 (MetAP2) inhibitor that in normal mice reduced alpelisib-induced hyperglycemia/hyperinsulinemia and in the MCF-7 model of HR+/PIK3CA-mutant breast cancer showed synergistic anti-tumor activity with alpelisib (Piqray). Evexomostat was well-tolerated in a phase 1 monotherapy safety study in late-stage cancer patients and improved insulin resistance in patients with elevated insulin at baseline, among other metabolic and angiogenic markers. Methods: This is a phase 1b/2, open-label, single-arm pilot study (NCT05455619) in postmenopausal women with PIK3CA-mutated, HR+, HER2- metastatic breast cancer with disease progression following treatment with endocrine therapy plus a CDK4/6 inhibitor who are at risk for hyperglycemia, with risk factors defined as HbA1c between 5.7 and 6.4% and/or HOMA-IR ≥1.8. The primary objective is to determine the safety of evexomostat plus standard of care treatment alpelisib (Piqray) and fulvestrant (combined, the ‘triplet therapy’), to measure the severity and number of hyperglycemic events, and to assess clinical, anti-tumor benefit of the triplet therapy. The trial will begin with a dose-escalation cohort (n=6) at an evexomostat dose of 36 mg/m2 (one dose below the monotherapy MTD of 49 mg/m2) in combination with alpelisib and fulvestrant given in accordance with their respective labels. Based on safety data from the first 6 patients (two cycles), the safety review committee may increase the evexomostat dose for the next cohort of six patients to 49 mg/m2 or may decrease the evexomostat dose to 27 mg/m2 and may adjust the dose of alpelisib if warranted. Once the MTD of the triplet therapy has been defined, additional enrollment will occur until a total of up to 20 patients have completed at least two cycles of triplet therapy at that dose. If warranted, an additional 20 patients may be enrolled to further characterize the safety profile and/or anti-tumor effect of the triplet therapy (total of up to 52 patients). This trial will open to accrual in August, 2022. Primary safety analysis consists of the type, frequency, and severity of treatment-emergent adverse events (TEAEs) per the NCI CTCAE, v5.0, the number of patients with grade 3 or 4 hyperglycemia during the first 2 cycles of therapy plus an estimate of the proportion and its exact upper one-sided 97.5% confidence bound will be analyzed. Efficacy analyses include calculation of the ORR, consisting of complete response (CR) and partial response (PR). The number of patients alive without disease progression six months from the start of the triplet therapy will be assessed. The CBR of CRs, PRs plus stable disease ≥24 weeks from C1D1 will be calculated. Overall survival data will be summarized as available or appropriate. QoL will be analyzed according to functional scores and recommendations in the EORTC scoring manual. ECOG performance status and change from baseline will be summarized. Citation Format: Peter Cornelius, Neal Salomon, David Browning, Sakirat Gill, Ben Mayes, Pierre Dufour, James Shanahan, Bradley Carver, Hope Rugo. The Amelia-1 Study: A phase 1b/2 trial of evexomostat (SDX-7320) plus fulvestrant (Faslodex®) and alpelisib (Piqray®) in patients with advanced breast cancer at risk for alpelisib (Piqray)-induced hyperglycemia [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr OT3-31-01.

MetAP2 inhibitors have shown clinical anti-tumor activity, but CNS side effects and poor drug-like properties have generally limited their development. SDX-7320 is a polymer-drug conjugate of a novel fumagillin-derived MetAP2 inhibitor (SDX-7539) attached via a cleavable linker to a hydroxypropylmethacrylamide (HPMA) polymer backbone. This conjugation approach is intended to improve safety (i.e., limit CNS penetration) while also improving biodistribution and pharmacokinetics relative to small molecule MetAP2 inhibitors. SDX-7320 recently completed a phase I safety trial in late-stage cancer patients (NCT02743637) and was well-tolerated with no apparent CNS side effects. In prior models of breast cancer, SDX-7320 significantly inhibited the growth of syngeneic EO771 triple-negative breast cancers (TNBC) accelerated by obesity/metabolic dysfunction (i.e., “metabo-oncology”) and also synergized with the PI3Kα inhibitor alpelisib/Piqray® in ER+/Her2- MCF-7 xenografts. The objectives of these experiments were to determine the effect of SDX-7320 on the growth of Her2+ xenografts alone or in combination with the Akt/mTOR inhibitor capivasertib/AZD-5363 and to determine if SDX-7320 could attenuate hyperglycemia induced by capivasertib/AZD-5363. Female athymic, nude mice (9 weeks of age) were surgically implanted with slow-release estrogen pellets (90-day, 0.72 mg, inter-scapular) and three days later 2 x 107 BT474 cells (in Matrigel/RPMI-1640, 50/50, v/v) were injected into the fourth mammary gland. When the group mean tumor volume exceeded 100 mm3, treatment with test agents commenced. SDX-7320 (sc/q4d, 6, 12 mg/kg) exerted significant, dose-dependent inhibition of BT-474 tumor growth (TGI of 61 and 88% respectively), whereas capivasertib/AZD-5363 (po, qd, 100, 200 mg/kg) exhibited significant anti-tumor activity only at 200 mg/kg (54% TGI). The combination of SDX-7320 (12 mg/kg) plus capivasertib/AZD-5363 (200 mg/kg) produced additive effects in that a greater number of mice had tumor regression relative to mice treated with either SDX-7320 or capivasertib/AZD-5363 alone. Tumor tissue from a subset of treatment groups (Vehicle, SDX-7320-12 mg/kg, SDX-7320-12 mg/kg + AZD-5363-200 mg/kg, and AZD-5363-200 mg/kg) was snap frozen and stored at -80oC. PolyA+ RNA was isolated, cDNA libraries were constructed and RNASeq was conducted (range of 20-30 reads per million (RPM) base pairs). Analysis of RNASeq data showed that SDX-7320 alone significantly attenuated the expression of hypoxia-induced genes as well as genes related to the innate immune system. Based on these results, we conclude that SDX-7320 affected tumor growth by limiting the ability of tumors to adapt to a hypoxic microenvironment and by altering the innate tumor-immune microenvironment. In normal male, C57Bl/6 mice, capivasertib/AZD-5363 (200 mg/kg, po) elevated blood glucose after a single dose. Pretreatment with either one dose (24 hours before the Akt/mTOR inhibitor) or multiple doses (Q4D beginning 14 days prior to Akt/mTOR inhibitor) of SDX-7320 (8 mg/kg) significantly inhibited hyperglycemia induced by capivasertib/AZD-5363. In summary SDX-7320 significantly inhibited the growth of BT-474 xenografts, alone and in combination with capivasertib/AZD-5363, which was associated with suppression of hypoxia-induced genes, as well as genes of the innate immune system. In addition, SDX-7320 improved the safety profile of capivasertib/AZD-5363 by attenuating Akt/mTOR inhibitor-induced hyperglycemia, providing further support for the clinical exploration of SDX-7320 in combination with Akt/mTOR inhibitors in Her2+ breast cancer. Citation Format: Peter Cornelius, Benjamin Mayes, Pierre Dufour, Mark Kalinich, Jonathan Wang, Sara Little, Bradley Carver, James Shanahan. Inhibition of HER2+ tumor growth with SDX-7320, a novel MetAP2 inhibitor, alone and in combination with capivasertib/AZD-5363: Reduced expression of hypoxia-inducible and innate-immune system genes [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P5-05-04.

Abstract Small molecule MetAP2 inhibitor TNP-470 previously showed clinical anti-tumor activity, however CNS side effects and poor drug-like properties limited its development. SDX-7320 is a polymer-drug conjugate of a novel MetAP2 inhibitor (SDX-7539), intended to improve biodistribution (limit CNS penetration) and pharmacokinetics relative to small molecule MetAP2 inhibitors. SDX-7320 has recently completed a phase I trial in late-stage cancer patients (NCT02743637). SDX-7320 was previously shown to inhibit the growth of syngeneic EO771 triple-negative breast cancers (TNBC) accelerated by obesity/metabolic dysfunction (“metabo-oncology”). In addition SDX-7320 synergized with the PI3Kα inhibitor alpelisib (Piqray®) to block the growth of ER+/Her2- MCF-7 xenografts. Here, we show that SDX-7320 inhibits the growth of MCF-7 xenografts alone and in combination with the CDK4/6 inhibitor Palbociclib (Ibrance®). Nude mice were implanted with slow release estrogen pellets and then injected with MCF-7 cells in the fourth mammary gland. Treatment began (n=10/group) when tumors exceeded 100 mm³: SDX-7320 (sc/q4d, 8 mg/kg), palbociclib (po, qd, 20 or 40 mg/kg), SDX-7320 plus palbociclib (20 mg/kg), SDX-7320 plus palbociclib (40 mg/kg). MCF-7 tumor growth inhibition at day 31 relative to vehicle-treated mice: SDX-7320, 29% (NS); palbociclib, 24% and 54% at 20 and 40 mg/kg respectively (NS); SDX-7320 plus palbociclib (20 mg/kg), 64% (p<0.05); and SDX-7320 plus palbociclib (40 mg/kg), 82%, (p<0.01). Tumor samples were homogenized in RIPA buffer containing protease and phosphatase inhibitors for analysis of proteins (by WES). Cell-cycle proteins (cyclin E1, E2, cdk2, cdk4) were decreased in tumor tissue from SDX-7320-treated mice relative to vehicle, and were typically lower in tumors from mice treated with SDX-7320 plus palbociclib (40 mg/kg) compared to either vehicle-treated or SDX-7320-treated mice. A key growth factor-signaling protein Akt was reduced in SDX-7320-treated groups (alone and in combination) as was estrogen receptor alpha (ERα). pAkt (S473) was lowest in SDX-7320-treated tumors and in tumors treated with SDX-7320 plus palbociclib (40 mg/kg). The autophagy marker LC3B was elevated in response to palbociclib (20 and 40 mg/kg) but in combination with SDX-7320, the increase was significantly attenuated. Intracellular pathways and proteins linked to emergence of resistance to palbociclib include cell cycle (cyclin E1), growth factor and hormone signaling (Akt, ERα) and autophagy (LC3B). The protein changes observed (representative of these pathways) suggest that combining SDX-7320 with palbociclib (Ibrance®) may extend the period of progression-free survival in patients with ER+/Her2- breast cancer relative to treatment with palbociclib (Ibrance®) alone. Citation Format: Peter Cornelius, Benjamin Mayes, Pierre Dufour, Sara Little, Andrew Slee, Raphael Nir, Adam Nir, Bradley J. Carver, James Shanahan. SDX-7320, a novel inhibitor of methionine aminopeptidase 2 (MetAP2), inhibits MCF-7 tumor growth in combination with palbociclib (Ibrance®) [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 1068.