Cytidine Deaminase Inhibitor Research Articles

Pharmacokinetic assessment of low dose decitabine in combination therapies: Development and validation of a sensitive UHPLC-MS/MS method for murine plasma analysis

Decitabine is a DNA methyltransferase inhibitor used in the treatment of acute myeloid leukemia and myelodysplastic syndrome. The notion that ongoing trials are presently exploring the combined use of decitabine, with or without the cytidine deaminase inhibitor cedazuridine, and other antileukemic drugs necessitates a comprehensive understanding of pharmacokinetic properties and an evaluation of drug-drug interaction liabilities. We report here the development and validation of a sensitive UHPLC-MS/MS method for quantifying decitabine in mouse plasma, which should be useful for such studies. The method involved a one-step protein precipitation extraction, and chromatographic separation on an XBridge HILIC column using gradient elution. The method was found to be robust, accurate, precise, and sufficiently sensitive (lower limit of quantitation, 0.4 ng/mL) to determine decitabine concentrations in microvolumes of plasma from mice receiving the agent orally or intravenously in the presence or absence of cedazuridine.

Synthesis of 1,4-azaphosphinine nucleosides and evaluation as inhibitors of human cytidine deaminase and APOBEC3A.

Nucleoside and polynucleotide cytidine deaminases (CDAs), such as CDA and APOBEC3, share a similar mechanism of cytosine to uracil conversion. In 1984, phosphapyrimidine riboside was characterised as the most potent inhibitor of human CDA, but the quick degradation in water limited the applicability as a potential therapeutic. To improve stability in water, we synthesised derivatives of phosphapyrimidine nucleoside having a CH2 group instead of the N3 atom in the nucleobase. A charge-neutral phosphinamide and a negatively charged phosphinic acid derivative had excellent stability in water at pH 7.4, but only the charge-neutral compound inhibited human CDA, similar to previously described 2'-deoxyzebularine (Ki = 8.0 ± 1.9 and 10.7 ± 0.5 µM, respectively). However, under basic conditions, the charge-neutral phosphinamide was unstable, which prevented the incorporation into DNA using conventional DNA chemistry. In contrast, the negatively charged phosphinic acid derivative was incorporated into DNA instead of the target 2'-deoxycytidine using an automated DNA synthesiser, but no inhibition of APOBEC3A was observed for modified DNAs. Although this shows that the negative charge is poorly accommodated in the active site of CDA and APOBEC3, the synthetic route reported here provides opportunities for the synthesis of other derivatives of phosphapyrimidine riboside for potential development of more potent CDA and APOBEC3 inhibitors.

Oral decitabine–cedazuridine versus intravenous decitabine for myelodysplastic syndromes and chronic myelomonocytic leukaemia (ASCERTAIN): a registrational, randomised, crossover, pharmacokinetics, phase 3 study

The DNA methyltransferase inhibitors azacitidine and decitabine for individuals with myelodysplastic syndromes or chronic myelomonocytic leukaemia are available in parenteral form. Oral therapy with similar exposure for these diseases would offer potential treatment benefits. We aimed to compare the safety and pharmacokinetics of oral decitabine plus the cytidine deaminase inhibitor cedazuridine versus intravenous decitabine. We did a registrational, multicentre, open-label, crossover, phase 3 trial of individuals with myelodysplastic syndromes or chronic myelomonocytic leukaemia and individuals with acute myeloid leukaemia, enrolled as separate cohorts; results for only participants with myelodysplastic syndromes or chronic myelomonocytic leukaemia are reported here. In 37 academic and community-based clinics in Canada and the USA, we enrolled individuals aged 18 years or older who were candidates to receive intravenous decitabine, with Eastern Cooperative Oncology Group performance status 0 or 1 and a life expectancy of at least 3 months. Participants were randomly assigned (1:1) to receive 5 days of oral decitabine-cedazuridine (one tablet once daily containing 35 mg decitabine and 100 mg cedazuridine as a fixed-dose combination) or intravenous decitabine (20 mg/m2 per day by continuous 1-h intravenous infusion) in a 28-day treatment cycle, followed by 5 days of the other formulation in the next treatment cycle. Thereafter, all participants received oral decitabine-cedazuridine from the third cycle on until treatment discontinuation. The primary endpoint was total decitabine exposure over 5 days with oral decitabine-cedazuridine versus intravenous decitabine for cycles 1 and 2, measured as area under the curve in participants who received the full treatment dose in cycles 1 and 2 and had decitabine daily AUC0-24 for both oral decitabine-cedazuridine and intravenous decitabine (ie, paired cycles). On completion of the study, all patients were rolled over to a maintenance study. This study is registered with ClinicalTrials.gov, NCT03306264. Between Feb 8, 2018, and June 7, 2021, 173 individuals were screened, 138 (80%) participants were randomly assigned to a treatment sequence, and 133 (96%) participants (87 [65%] men and 46 [35%] women; 121 [91%] White, four [3%] Black or African-American, three [2%] Asian, and five [4%] not reported) received treatment. Median follow-up was 966 days (IQR 917-1050). Primary endpoint of total exposure of oral decitabine-cedazuridine versus intravenous decitabine was 98·93% (90% CI 92·66-105·60), indicating equivalent pharmacokinetic exposure on the basis of area under the curve. The safety profiles of oral decitabine-cedazuridine and intravenous decitabine were similar. The most frequent adverse events of grade 3 or worse were thrombocytopenia (81 [61%] of 133 participants), neutropenia (76 [57%] participants), and anaemia (67 [50%] participants). The incidence of serious adverse events in cycles 1-2 was 31% (40 of 130 participants) with oral decitabine-cedazuridine and 18% (24 of 132 participants) with intravenous decitabine. There were five treatment-related deaths; two deemed related to oral therapy (sepsis and pneumonia) and three to intravenous treatment (septic shock [n=2] and pneumonia [n=1]). Oral decitabine-cedazuridine was pharmacologically and pharmacodynamically equivalent to intravenous decitabine. The results support use of oral decitabine-cedazuridine as a safe and effective alternative to intravenous decitabine for treatment of individuals with myelodysplastic syndromes or chronic myelomonocytic leukaemia. Astex Pharmaceuticals.

Development of Oral Azacitidine with Cedazuridine for Myelodysplastic Syndrome (MDS) and Myeloproliferative Neoplasms (MPN) Including CMML (Chronic Myelomonocytic Leukemia) By Targeting Pharmacokinetic AUC Equivalence Vs Subcutaneous Azacitidine

Introduction/ Background: Hypomethylating agents (HMAs) such as azacitidine (AZA) and decitabine (DEC) are FDA-approved for patients with MDS/CMML and acute myeloid leukemia (AML). Treatment is generally administered parenterally as a daily injection or infusion over 5-7 days per four-week cycle. The oral fixed-dose combination of decitabine with cedazuridine, ASTX727, was approved by regulators in the USA and Canada for patients with MDS and CMML based upon the demonstration of biologically equivalent pharmacokinetics (PK) area under the curve (AUC) vs IV decitabine. ASTX030 is a combination of AZA and cedazuridine (CED), a cytidine deaminase inhibitor (CDAi), which enables oral AZA to achieve systemic AUC similar to that of parenteral (subcutaneous) AZA. Development and approval of an oral AZA providing equivalent PK exposure to the parenteral therapy has the potential to markedly reduce the treatment burden associated with parenteral treatment. This Phase I trial is designed to determine the dose combination for oral AZA plus CED to replicate SC AZA AUC exposures in subjects with MDS and MDS/MPN, including CMML. Methods: Adult patients with confirmed MDS, CMML, and other MDS/MPN or AML who are candidates to receive benefit from single agent AZA were enrolled in this open label Phase 1 trial. In addition to assessing safety and efficacy of each combination, the primary PK endpoint was to achieve oral AZA PK AUC 0-24 equivalence versus subcutaneous AZA (at the approved dose of 75 mg/m 2 over the span of 7 days). In the first cycle, patients received oral AZA alone on Day -3, followed by SC AZA at 75 mg/m 2 on Day 1. Subsequently, oral ASTX030 (combination of AZA with CED) was given at varying dose combinations of AZA and CED for each cohort. After cycle 1, patients received oral ASTX030 on Days 1-7 for each 28-day cycle. Dose levels for cohorts in escalation were determined by the data safety and review committee based on safety and PK results. Pharmacodynamic (PD) impact was assessed by changes in LINE-1 DNA methylation in peripheral blood cells. Clinical response assessments were based upon International Working Group (IWG) 2006 criteria for MDS patients and IWG 2015 criteria for MDS/MPN and CMML patients. Results: As of 05 JUL 2023, 65 patients were treated across 8 different dose combinations, with ≥6 patients per cohort. The median age was 72 years old (range 26-87), 36.9% (n=24) were female, 6.2% (n=4), had previously received HMA treatment (unlimited). Of the enrolled patients, 69% (n=45) had MDS, 25%(n=16) had CMML, and 6% (n=4) had MDS/MPN (other). Evaluated dose levels for AZA ranged from 60-136mg and CED dose ranged from 20-100 mg. In cohorts 1-2, tablet formulation was tested first (AZA ranging 80-100mg with CED ranging 80-100mg). Starting with cohort 3, capsule formulation was introduced for AZA to optimize the interaction effect of CDA inhibition by CED. In both tablet and capsule formulation, grade ≥3 AE's (regardless of causality) were observed in 75.9% of patients. The most common Grade ≥3 AE's regardless of relation to drug were leukopenia (25%), thrombocytopenia (20%), and anemia (20%). Gastrointestinal toxicity were similar (for example, nausea 70% ASTX030 vs 71% SC AZA) to those reported for SC AZA (VIDAZA USPI)). No dose limiting toxicity was observed in any of the cohorts. The CED dose of 20 mg resulted in ~100% bioavailability for AZA, suggesting inhibition in a linear range. LINE-1 demethylation results were similar to those reported historically with SC AZA. Although a clinical response comparable to SC AZA was observed, the immature nature of the data requires continued analysis to yield results on treatment efficacy. Conclusion: ASTX030 successfully achieved the primary endpoint of PK equivalence versus SC AZA based on total cycle AUC. The dose combinations evaluated were well tolerated, and the safety profile similar to that of SC AZA, with no unique AE's. LINE-1 demethylation and clinical response confirm the clinical activity of ASTX030. Phase 1B dose expansion for the ASTX030 study at the recommended dose combination of 144mg AZA with 20mg CED will begin soon to confirm PK equivalence for this dose level. An orally bioequivalent AZA based HMA offers significant opportunity for novel combinations and improved convenience for patients with high grade myeloid malignancy.

Oral Decitabine/Cedazuridine Vs Intravenous Decitabine for Acute Myeloid Leukemia: Final Results of a Randomized, Crossover, Registration-Enabling, Pharmacokinetics Study

Background: Parenterally administered hypomethylating agents (HMAs), decitabine (DEC) and azacitidine (AZA), are approved in Europe for adult patients with acute myeloid leukemia (AML) who are not candidates for standard induction chemotherapy as single agent or in combination with venetoclax. Cedazuridine(C) is a novel, potent, and safe inhibitor of cytidine deaminase (CDA) and when given in combination with decitabine, cedazuridine enables efficient oral availability of decitabine.ASTX727 (DEC-C) is a fixed-dose combination (FDC) tablet of 35 mg DEC and 100 mg cedazuridine, DEC-C has been approved for MDS in the US, Canada, and Australia. Here we present final results using DEC-C in an AML population appropriate for single-agent decitabine treatment. Aims: Describe the final clinical outcomes of patients with AML with additional analysis of genetic profiles. Methods: The ASCERTAIN study used a randomized two-period, two-sequence, two-treatment, crossover study design. Patients were randomized 1:1 to either Sequence A: DEC-C (35 mg DEC/100 mg cedazuridine) in Cycle 1 followed by IV-DEC at 20 mg/m 2 in Cycle 2, or Sequence B: receiving IV-DEC in Cycle 1 followed by DEC-C on Cycle 2 to compare pharmacokinetics (PK) [primary endpoint Area Under the Curve (AUC) equivalence over 5 days of dosing]. All patients received DEC-C from Cycle 3 until treatment discontinuation to assess safety and clinical efficacy. Patients were eligible as per the EMA-approved decitabine label (newly diagnosed AML who are not candidates for standard induction chemotherapy). Clinical responses were assessed according to modified International Working Group (IWG) 2003 response criteria. Pre-treatment peripheral blood was used for DNA isolation from leukocytes, and molecular abnormalities identified using a NGS hematologic malignancy panel. Cox-regression analysis on the various factors (eg binary mutational status, CR, etc.) was used for analyses of risk-factors for overall survival (OS). Results: 89 patients were randomized, of whom 87 were treated. The median age of patients was 78.0 years (range, 61 to 92) with 31 (35.6%) males and 56 (64.4%) females. Cytogenetic risk classification was poor-risk in 33 (37.9%), intermediate-risk in 45 (51.7%), and 9 (10.4%) not evaluated patients. For the primary endpoint, preliminary PK data was available from 69 patients who successfully completed PK assessments for both IV-DEC and DEC-C cycles, and the DEC AUC 0-24 (h*ng/mL) 5-Day geometric mean estimate was 904 for DEC-C and 907 for IV-DEC resulting in an oral/IV geometric LSM AUC ratio of 99.64% (90% CI of 91.23-108.8%). Safety findings were consistent with those anticipated for IV-DEC (related Grade ≥ 3 AEs in more than 10% were thrombocytopenia, neutropenia, anaemia, and febrile neutropenia). As of the database lock (25May2023), median follow up was 24.5 (20.2, 25.0) months (25% of 19.7 and 75% of 25.2 months). The best response was complete response (CR) in 19/87 (21.8%, 95% CI: 13.7, 32.0%); CR with incomplete blood cell count recovery (CRi) in 5 patients (5.7%); and CR with incomplete platelet recovery (CRp) in 2 (2.3%); resulting in composite response rate [CR + CRi] of 27.6%. The median overall survival was approximately 8.9 months (95% CI: 6.0, 13.1).These results obtained with DEC-C are consistent with those observed for IV-DEC. Multiple genes, including ASXL1, TP53, RUNX1, and SRSF2 genes were present in more than 20% of the AML patients. TP53 mutation was associated with worse survival. Summary/Conclusion: This randomized phase 3 study in AML patients not candidates for standard induction chemotherapy demonstrates that Daily X 5 dosing of the oral FDC of DEC-C resulted in an equivalent DEC AUC exposure to IV-DEC at 20 mg/m 2 over 5 days. Pharmacodynamic data showed similar demethylation rates (≤1.1% difference). In addition, safety findings and observed clinical activity (response rates and OS) are consistent with published data from IV-DEC, suggesting that DEC-C has the potential to be an oral alternative to the standard IV-DEC Daily×5 regimen. These findings support the use of DEC-C in patients with AML who are not candidates for standard induction chemotherapy.

Reclassification of Ascertain (ASTX727-02) Myelodysplastic Syndrome (MDS) Patients: Outcomes Including Clinical Response, Overall Survival (OS), and Leukemia Free Survival (LFS) Based on IPSS-R and IPSS-M Scoring Systems

Background: Oral decitabine/cedazuridine (ASTX727) is a fixed dose combination of decitabine (35 mg) and the cytidine deaminase inhibitor cedazuridine (100 mg) given once daily X 5 days on a 28-day cycle producing pharmacokinetic (PK) exposure equivalent to the standard intravenous (IV) decitabine regimen of 20 mg/m 2 daily X 5 days on a 28-day cycle. This was demonstrated in the pivotal ASCERTAIN study (Garcia-Manero, et al, ASH 2019) as it provided a PK bridge to existing decitabine data and demonstrated median OS was 31.7 mo. (Savona,et al. Intl. MDSF International Congress on MDS, 2021). Subjects were initially classified by the International Prognosis Scoring System (IPSS) for historical reasons, however IPSS-R and IPSS-M are enhancements to IPSS that provide dynamic risk assessment for predicting clinical outcomes in MDS. Here, the objective was to re-classify the MDS subjects enrolled on the ASCERTAIN study by IPSS-R and IPSS-M and measure the impact of informing patient outcomes based on re-calculated risk assessment. Methods: One hundred thirty-three subjects with MDS/CMML (chronic myelomonocytic leukemia) were enrolled in ASCERTAIN and were randomly assigned either IV decitabine for cycle 1 and oral decitabine/cedazuridine for cycle 2 or the opposite treatment sequence. All subjects continuing beyond cycle 2 received oral decitabine/cedazuridine for all subsequent cycles until treatment discontinuation for disease progression, toxicity, patient's decision, or HSCT. Whole blood collected prior to treatment was used for DNA isolation and molecular abnormalities identified using next-generation sequencing (NGS) hematologic malignancy panel of 179 genes including all genes commonly mutated in MDS. In the initial analysis of clinical outcomes, subjects were classified by IPSS with response assessment by IWG 2006 IPSS low and Int-1 risk levels were categorized as lower-risk MDS (LR-MDS), whereas IPSS Int-2 and high-risk categories were categorized as higher-risk MDS (HR-MDS). Subjects with sufficient data based on (e.g., available heme parameters, cytogenetics, NGS etc.) were reclassified by IPSS-R and IPSS-M. Subjects with IPSS-R score of ≤3.5 or IPSS-M of either very low, low, or moderate low were categorized as LR-MDS. Similarly subjects with IPSS-R score of >3.5 or an IPSS-M categorization of either moderate high, high, or very high were categorized as HR-MDS. Reclassified subjects were reassessed for CR (Complete Response), OS, and LFS and Harrell's concordance index (c-index) was used to describe the level of agreement between each scale and outcomes. Results: Based on the available data, the number of MDS subjects in the different risk classifications were the following: IPSS: 117, IPSS-R: 104, and IPSS-M: 105. Thirteen and 12 subjects on the IPSS could not be reclassified in the IPSS-R and IPSS-M, respectively, including 5 Int-1 and 2 low-risk MDS cases. CMML subjects were excluded from these analyses. Re-classification generally resulted in the upgrade of the subjects from LR-MDS to HR-MDS (Fig. 1). Thirty-one (26.5%) subjects from IPSS were reallocated in the IPSS-R to different risk categories; 3 (9.7%) were downgraded and 28 (90.3%) were upgraded. Similarly on reclassification with IPSS-M 34 (32.4%) of the patients reclassified; 5 (14.8%) were downgraded and 29 (85.3%) were upgraded. For IPSS LR-MDS, 23.2% of patients achieved CR, and 22.9% in the IPSS HR-MDS. Similarly, 21.6% patients achieved CR in the IPSS-R LR-MDS, and 23.9% in the IPSS-R HR-MDS; 26.3% patients achieved CR in the IPSS-M LR-MDS, and 20.9% in the IPSS-M HR-MDS. The c-index for the IPSS was .64 (OS) and .67 (LFS), IPSS-R was .70 (OS) and .71 (LFS), and .75 (OS) and .78 (LFS) for the IPSS-M (Table1). Conclusion: Reclassification from IPSS to IPSS-R or IPSS-M upgraded multiple subjects from a LR to a HR category, describing the ASCERTAIN patient population as a majority higher risk population with worse prognosis than previously assumed based on the IPSS. The efficacy as measured by the CR rates did not change when the LR and HR categories were defined by the different risk stratification systems. In contrast, the c-index improves with migration from IPSS to IPSS-R to IPSS-M, indicating an increased discriminatory ability of IPSS-M score in comparison to IPSS and IPSS-R, to predict patient outcomes.

Development of the Treatment Preference in Myelodysplasia Questionnaire (TPMQ) for Clinicians (mTPMQ), Carers (cTPMQ), and Patients (pTPMQ)

Objective: Myelodysplastic syndrome (MDS) is a disease of the hematopoietic stem cells and leads to an increased risk of developing acute myeloid leukemia (AML).[1] The injectable hypomethylating agents (HMA) azacitdine and decibtabine have been shown to be superior to best supportive care in improving cytopenias in intermediate to high-risk patients.[2, 3] These injectable HMAs regimens are commonly administered in a healthcare facility, with patients traveling to the facility on consecutive days. The recent availability of an oral form of decitabine, ASTX 727 (a fixed-dose combination of decitabine and the cytidine deaminase inhibitor cedazuridine), may significantly impact patient, carer, and clinician choices. To assess preference for an injectable vs. orally administered HMA, three questionnaires were developed based on qualitative input from clinicians, patients, and carers: the Treatment Preference in Myelodysplasia Questionnaire (TPMQ) for clinicians (mTPMQ), carers (cTPMQ), and patients (pTPMQ). Methods: This was a non-interventional, cross-sectional qualitative interview study consisting of interviews with patients with MDS who were eligible for or were being/had been treated with injectable azacitidine, their carers and their treating clinicians. Data were collected from qualitative mixed method interviews composed of concept elicitation and cognitive debriefing. Two sites were identified to enrol participants. During concept elicitation, participants were asked about MDS and treatment options to best understand the constructs of interest relative to their specific circumstance and preferences. Cognitive debriefing occurred after participants reviewed a draft of the questionnaire. Based on the interviews, the questionnaires were revised to ensure the constructs of importance were captured and the questionnaires, response options, and instructions were all well understood. Results: 15 participants completed interviews (5 clinicians for the mTPMQ, 5 patients for the pTPMQ, and 5 carers for the cTPMQ). Concepts identified as important included across groups included: easier to be compliant, less travel time, less time in the clinic, less painful, costs and convenience (Table 1). The mTPMQ underwent 3 revisions. Overall, clinicians indicated the directions, items and response options were understandable and clear. Comments that significantly modified the questionnaire concerned splitting the last question from 1 to 2 questions, depending on which treatment the patient would be receiving. Patient participants thought the pTPMQ captured concepts of interest and was easy to understand. Modifications were made to the questionnaire either as a direct result of patient participant comments (e.g., changed skip directions to “turn to page X, question X”), study team discussions of the interviewer's comments and notes and making items consistent between questionnaires. Carer participants thought the cTPMQ captured the constructs of interest and the questions were clear and the questionnaire was easy to answer. Modifications were made to the questionnaire as a direct result of carer participant comments (e.g., changed “We spend less time in the clinic” to “It means less time in the clinic for the patient”), study team discussions of the interviewer's comments and notes (e.g., added “It makes it easier for the patient to continue treatment”) and making items consistent between questionnaires. Conclusions: The findings of the participant interviews suggest the concepts assessed in the assessments are complete. Respondents could respond to items and talk about all aspects assessed in the questionnaires in meaningful ways and were able to comprehend the instructions and response options on a categorical scale. The TPMQs are based on direct input from the population of interest and assess the constructs of importance to each group. 1. Bell, J.A., et al., Clin Lymphoma Myeloma Leuk, 2018. 18(4): p. e157-e166. 2. Kantarjian, H., et al., Cancer, 2006. 106(8): p. 1794-803. 3. Fenaux, P., et al., Lancet Oncol, 2009. 10(3): p. 223-32.

CTNI-47. DOSE ESCALATION OF ASTX727 IN ADULT PATIENTS WITH RECURRENT/PROGRESSIVE NON-ENHANCING IDH MUTANT GLIOMAS

Abstract BACKGROUND IDH mutant gliomas typically display a hypermethylation pattern known as the glioma CpG island methylator phenotype (G-CIMP) which results in gene activation that promotes gliomagenesis. This study aims to reverse this methylation signature and disrupt tumor growth via treatment with an oral demethylating agent. METHODS This is 2-stage clinical trial investigating oral ASTX727 (decitabine + cedazuridine, a cytidine deaminase inhibitor, as a combination tablet) in patients with recurrent primarily non-enhancing IDH mutant gliomas. The Dose Escalation Stage consisted of a single arm 3 + 3 design. The primary objective was to establish the MTD and assess safety in this patient population. RESULTS Eleven pts (F = 45%, M = 55%) with a median age of 44 yrs (range 26-61 yrs) were enrolled. Two DLTs (grade 4 neutropenia for >72 hrs) were observed in 5 pts enrolled at the starting dose (35 mg decitabine/ 100 mg cedazuridine for 5 consecutive days). Six additional patients were enrolled at dose level -1 (35 mg decitabine/100 mg cedazuridine for 4 consecutive days). There were no DLTs at dose level -1 and this was declared the MTD. Grade ≥ 3 AEs included neutropenia (5 pts), leukopenia (1 pt), lymphopenia (1 pt), hyponatremia (1 pt) and increased ALT (1 pt). Best treatment response was SD in 11 patients. At the time of last follow-up (5/15/23) 10 pts have discontinued treatment (8 for PD, and 2 based on physician decision), 2 pts (18%) have died due to disease progression, 8 pts remain alive and in follow-up, and 1 patient remains on treatment. Median duration on treatment was 212 days (range 36 days – not reached). CONCLUSIONS ASTX727 was well tolerated in patients with recurrent non-enhancing IDH mutant glioma at dose level -1 with no DLTs . A surgical expansion cohort is currently enrolling to confirm drug target engagement and explore pharmacodynamics.

Preliminary Results from a Phase 1b Dose De-Escalation Stage of Abnl-Marro 001: An International MDS/MPN Working Group Study

Background: Myelodysplastic/Myeloproliferative neoplasms (MDS/MPN) represent a group of rare hematologic malignancies, distinguished by their concurrent dysplastic and proliferative features. Patients with these neoplasms have an increased risk of acute leukemia transformation and often present with extramedullary and constitutional symptoms. The only approved therapies for MDS/MPNs are hydroxyurea, decitabine (oral decitabine/cedazuridine and parenteral decitabine) or azacitidine (parenteral) for pts with CMML. Given the rarity and the heterogeneity of MDS/MPNs, prospective studies focusing specifically on MDS/MPN have been uncommon, and refining treatment strategies has been difficult. ABNL-MARRO ( A Basket study of Nove l therapy for untreated MDS/MPN and Relapsed/ Refractory Overlap Syndromes) is an international cooperative effort that leverages the expertise of the MDS/MPN International Working Group (IWG) and provides a framework for collaborative studies around the world to advance treatment of MDS/MPN and to explore clinical and pathologic markers of disease severity, prognosis, and treatment response (Moyo TK et.al BMC Cancer. 2022). Methods: ABNL-MARRO 001 (AM-001) is an open-label, phase 1b/2 study testing the treatment combination of ASTX727 (days 1-5), a fixed dose oral combination pill containing 35 mg of the DNMTi decitabine (DEC) and 100 mg of the cytidine deaminase inhibitor cedazuridine (CED), together with a series of active agents in myeloid disease (see Figure 1a) for MDS/MPN. In arm A, ASTX727 is combined with itacitinib 300 mg (days 1-28), a selective JAK1 inhibitor in newly diagnosed and relapsed/refractory (R/R) pts. Here we report results from the 3+3 dose de-escalation phase 1b portion used to determine the recommended Phase 2 dose (RP2D) in arm A. Dose-limiting toxicities (DLTs) were assessed in cycle 1. Disease response was assessed by the MDS/MPN IWG response criteria (Savona et.al., Blood. 2015). Results: Seven patients were enrolled with a median age 75 (range 32-82) years with newly diagnosed (4 pts) or R/R (3 pts) MDS/MPN. Three patients had CMML, 2 had MDS/MPN-U, and 2 had refractory MDS/MPN RST. One patient was not DLT evaluable given severe disease-induced cytopenia, and was not included in the assessment (patient 4 in Figure 1b). The majority of pts experienced a treatment-emergent adverse event (TEAE) of any grade; the most common (≥30% of pts) were thrombocytopenia (50%), neutropenia (30%), and constipation (30%); 50% of pts experienced a Grade ≥3 TEAE; the most common was neutropenia (30%). No DLTs were reported, and these AEs were consistent with what has been reported in the use of ASTX727 (Savona et al. Blood. 2022). One patient achieved a CR, and two additional patients had the best overall response of optimal marrow response after receiving 6 cycles of treatment. All patients are alive with a median follow-up of 12 months (range 6-17 months). The clinical responses and time-on-treatment of all pts in phase 1b are summarized in Figure 1b. Summary/Conclusion: Based on the preliminary results of the phase 1b study, the dosing schedule of 35 mg DEC / 100 mg CED daily for 5 days in combination with itacitinib 300 mg daily for 28 days was selected as the RP2D as it balanced clinical efficacy with an acceptable and manageable safety profile. This regimen and dose are being utilized in a 28-day cycle in the ongoing global, multi-center phase 2 study (NCT04061421).

Combined inhibition of histone deacetylase and cytidine deaminase improves epigenetic potency of decitabine in colorectal adenocarcinomas

BackgroundTargeting the epigenome of cancerous diseases represents an innovative approach, and the DNA methylation inhibitor decitabine is recommended for the treatment of hematological malignancies. Although epigenetic alterations are also common to solid tumors, the therapeutic efficacy of decitabine in colorectal adenocarcinomas (COAD) is unfavorable. Current research focuses on an identification of combination therapies either with chemotherapeutics or checkpoint inhibitors in modulating the tumor microenvironment. Here we report a series of molecular investigations to evaluate potency of decitabine, the histone deacetylase inhibitor PBA and the cytidine deaminase (CDA) inhibitor tetrahydrouridine (THU) in patient derived functional and p53 null colon cancer cell lines (CCCL). We focused on the inhibition of cell proliferation, the recovery of tumor suppressors and programmed cell death, and established clinical relevance by evaluating drug responsive genes among 270 COAD patients. Furthermore, we evaluated treatment responses based on CpG island density.ResultsDecitabine caused marked repression of the DNMT1 protein. Conversely, PBA treatment of CCCL recovered acetylation of histone 3 lysine residues, and this enabled an open chromatin state. Unlike single decitabine treatment, the combined decitabine/PBA treatment caused > 95% inhibition of cell proliferation, prevented cell cycle progression especially in the S and G2-phase and induced programmed cell death. Decitabine and PBA differed in their ability to facilitate re-expression of genes localized on different chromosomes, and the combined decitabine/PBA treatment was most effective in the re-expression of 40 tumor suppressors and 13 genes typically silenced in cancer-associated genomic regions of COAD patients. Furthermore, this treatment repressed expression of 11 survival (anti-apoptotic) genes and augmented expression of X-chromosome inactivated genes, especially the lncRNA Xist to facilitate p53-mediated apoptosis. Pharmacological inhibition of CDA by THU or its gene knockdown prevented decitabine inactivation. Strikingly, PBA treatment recovered the expression of the decitabine drug-uptake transporter SLC15A1, thus enabling high tumor drug-loads. Finally, for 26 drug responsive genes we demonstrated improved survival in COAD patients.ConclusionThe combined decitabine/PBA/THU drug treatment improved drug potency considerably, and given their existing regulatory approval, our findings merit prospective clinical trials for the triple combination in COAD patients.

Abstract 1678: Overcoming treatment resistance in advanced bladder cancers using a neo-adjuvant combination with a natural product analog

Abstract Urinary bladder cancer (BC) is a common cancer worldwide, with approximately 573,278 new cases and 212,536 deaths arising annually. Tumors detected beyond the transitional zone of the bladder (advanced BC) account for most deaths. In advanced BC, tumor heterogeneity and resistance to successive chemotherapy regimens are major contributors to morbidity and mortality. Toxicity associated with the combination of Gemcitabine and Cisplatin (G+C) treatment limits the efficacy of chemotherapy. We tested the potential antitumor activity of a novel combination therapy involving a semisynthetic derivative of the pentacyclic triterpene (PTA) Ursolic Acid (N-methyl piperazine conjugated Ursolate, UA4) and Gemcitabine (Gem) in two Gem resistant BC cell lines developed in the laboratory. UA4 was synthesized from the PTA Ursolic Acid and Gem was purchased from LC-Labs. We tested the response to Gem and UA4 on naïve (WT) and Gem Resistant (GemR) cells. We generated Gem resistant BC cell lines HTB-4 (T24) and HTB-9 (5637) by repeated selection in increased Gem doses in vitro. The surviving cells were tested for a stable resistant phenotype. We measured the sensitivity to Gem and UA4 alone, and in adjuvant and neo-adjuvant combinations by colorimetric, cell counting, or clonogenic assays. The antitumor efficacy of UA4 and Gem were tested in vivo by xenograft studies on Fox N1-/- athymic mice (both sexes). The expression of markers associated with drug resistance was determined by qPCR, immunoblotting, and immunofluorescence studies. The 50% growth inhibition (IC50) dose in Gem treated cells increased significantly in 5637 and T24 GemR cells - 10-fold higher in GemR (100nM) than in WT cells (10 nM). Both WT and GemR cells showed equal sensitivity to UA4 (~5 µM). GemR cells showed elevated mRNA expression of a pyrimidine salvaging enzyme, cytidine deaminase (CDA), which degrades intracellular Gem, MRP2, and ENT1, all related to limiting intracellular drug availability. Tetrahydrouridine (THU), a competitive inhibitor of CDA, reduced the resistance significantly, indicating CDA as the principal mediator of resistance in GemR cells. Oral administration of UA4 was highly effective in reducing tumor growth of 5637 WT and GemR xenografts, whereas Gem (25mg/kg, 2x per week, i.p.) did not significantly reduce growth of GemR xenografts. Gem antitumor activity on 5637 WT tumors was predominantly due to apoptosis. Increased expression of autophagy markers, but not markers of apoptosis, was evident in vitro and in vivo treatments with UA4. Overall, treatment with UA4 was equally effective in decreasing viability in both WT and GemR advanced BC cells in vitro and reducing their tumor growth in vivo. Unlike Gem, UA4 did not induce significant levels of apoptosis but mainly autophagy-induced cell death. Ongoing studies indicate potential to use UA4 as an adjuvant or neo-adjuvant therapy in the treatment of advanced BC. Citation Format: Juliette R. Seremak, Sunilkanth Bonigala, Kunj B. Gupta, Siva S. Panda, Bal L. Lokeshwar. Overcoming treatment resistance in advanced bladder cancers using a neo-adjuvant combination with a natural product analog [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1678.

Reversible cardiomyopathy in a patient with chronic myelomonocytic leukemia treated with decitabine/cedazuridine: a case report

BackgroundHypomethylating agents (HMAs) have shown efficacy in the treatment of hematological malignancies and are indicated for the treatment of chronic myelomonocytic leukemia (CMML). While the HMA decitabine, in its intravenous formulation, has been used since 2006 for the treatment of CMML, use of its oral formulation has been limited by poor bioavailability due to first-pass metabolism by the enzyme cytidine deaminase. The dose of intravenous decitabine is limited by toxicities such as cardiomyopathy and heart failure. Therefore, cedazuridine was developed as an inhibitor of cytidine deaminase. Cedazuridine decreases the first-pass metabolism of oral decitabine allowing therapeutic levels to be achieved at lower doses, and thus, the novel oral combination of cedazuridine with decitabine was developed. While cardiomyopathy and heart failure are well-established adverse effects associated with intravenous decitabine alone, there to our knowledge there have been no documented incidences of reversible cardiomyopathy in the literature or in patients who participated in the phase 2 and phase 3 clinical trials of oral decitabine-cedazuridine.CaseThis case study presents an 85 year-old Caucasian female with CMML who developed cardiomyopathy and heart failure with reduced ejection fraction after completing 5 cycles of therapy with decitabine/cedazuridine. Furthermore, her symptoms and cardiac function recovered upon discontinuation of the drug.ConclusionsWe present an occurrence of reversible cardiomyopathy in a patient who completed 5 cycles of decitabine/cedazuridine, an oral combination therapy developed to enhance oral bioavailability of decitabine thereby limiting its adverse effects. As the decitabine/cedazuridine combination therapy rises in popularity due to its convenient oral formulation, more trials are needed to understand the prevalence of cardiomyopathy with this drug and to discover preventative strategies for cardiotoxic effects.

Seven-membered ring nucleobases as inhibitors of human cytidine deaminase and APOBEC3A.

The APOBEC3 (APOBEC3A-H) enzyme family as a part of the human innate immune system deaminates cytosine to uracil in single-stranded DNA (ssDNA) and thereby prevents the spread of pathogenic genetic information. However, APOBEC3-induced mutagenesis promotes viral and cancer evolution, thus enabling the progression of diseases and development of drug resistance. Therefore, APOBEC3 inhibition offers a possibility to complement existing antiviral and anticancer therapies and prevent the emergence of drug resistance, thus making such therapies effective for longer periods of time. Here, we synthesised nucleosides containing seven-membered nucleobases based on azepinone and compared their inhibitory potential against human cytidine deaminase (hCDA) and APOBEC3A with previously described 2'-deoxyzebularine (dZ) and 5-fluoro-2'-deoxyzebularine (FdZ). The nanomolar inhibitor of wild-type APOBEC3A was obtained by the incorporation of 1,3,4,7-tetrahydro-2H-1,3-diazepin-2-one in the TTC loop of a DNA hairpin instead of the target 2'-deoxycytidine providing a Ki of 290 ± 40 nM, which is only slightly weaker than the Ki of the FdZ-containing inhibitor (117 ± 15 nM). A less potent but notably different inhibition of human cytidine deaminase (CDA) and engineered C-terminal domain of APOBEC3B was observed for 2'-deoxyribosides of the S and R isomers of hexahydro-5-hydroxy-azepin-2-one: the S-isomer was more active than the R-isomer. The S-isomer shows resemblance in the position of the OH-group observed recently for the hydrated dZ and FdZ in the crystal structures with APOBEC3G and APOBEC3A, respectively. This shows that 7-membered ring analogues of pyrimidine nucleosides can serve as a platform for further development of modified ssDNAs as powerful A3 inhibitors.

A Phase I/II Study of Combination of ASTX727, Gilteritinib and Venetoclax in Patients with Relapsed/Refractory FLT3 Mutated Acute Myeloid Leukemia (AML) and Frontline FLT3 Mutated AML Patients Unfit for Chemotherapy

Background:FLT3 gene mutations occur in approximately 30% of patients (pts) with AML and are associated with worse survival and higher relapse risk. Gilteritinib, a selective potent oral FLT3 tyrosine kinase inhibitor (FLT3i), improves survival in pts with relapsed FLT3 mutated AML. ASTX727 is an oral fixed-dose combination of decitabine (35 mg) and cedazuridine (a cytidine deaminase inhibitor; 100 mg) approved for treating pts with myelodysplastic syndrome (MDS). We are evaluating the safety and efficacy of the combination of ASTX727, gilteritinib, and venetoclax (VEN) in pts with relapsed or refractory FLT3-mutated AML and older/unfit pts unable to receive chemotherapy. Methods: In the phase I, pts with R/R FLT3-mutated AML or high-risk (HR) MDS or chronic myelomonocytic leukemia (CMML) are eligible. FLT3-ITD and/or FLT3-TKD mutations were allowed. In the phase II, the study will also enroll newly diagnosed pts with FLT3-mutated AML ineligible for induction therapy. Pts are required to have an ECOG performance status ≤3, total bilirubin ≤2.5 x upper limit of normal (ULN), ALT/AST ≤3 x ULN, and creatinine clearance ≥30 mL/min. Hydroxyurea or 1 dose of cytarabine up to 1000 mg was allowed in pts with proliferative disease to lower white blood count to ≤ 25 x 109/L before initiation of study therapy. In the phase 1, we evaluated two dose levels of gilteritinib (80 mg and 120 mg daily) continuously with fixed dosages of ASTX727 (35/100 mg daily on days 1-5) and VEN (400 mg daily, dose adjusted for concomitant azoles, for up to 28 days). Cycles are repeated every 28 days for up to 24 cycles, followed by indefinite therapy with gilteritinib as maintenance. During cycle 1, VEN was given at a dose of 100 mg on day 1, 200 mg on day 2, and 400 mg daily onwards. Bone marrow (BM) aspiration was performed on day 21, and VEN was held in pts with BM blasts <5% or aplasia. During cycles 2-24, VEN was given at 400 mg daily (dose adjusted for concomitant azoles) on days 1-21 but dose reductions depending on tolerance were permitted. Gilteritinib continued at the enrolled dose in cycles 2-24. Results: Between 11/2021 and 6/2022, 8 patients (7 AML and 1 HR-CMML) have been enrolled in the phase 1 portion of the trial (Table 1). Their median age was 57 years (range, 38-83). Median number of prior lines of therapy was 1 (range, 1-10). Four pts (50%) had a previous allogeneic hematopoietic stem cell transplant (HSCT), 6 pts (75%) had prior treatment with hypomethylating agent (HMA)+VEN, and 3 pts (27%) received prior FLT3i. None received prior gilteritinib. Cytogenetics was diploid in 4 pts, complex in 3 pts, and miscellaneous in 1 pt. The median FLT3-ITD allelic ratio (AR) was 0.4 (range, 0.11-10.60), with 50% having a high AR (≥0.5). All pts had a FLT3-ITD mutation and no pt had a FLT3-TKD mutation. NPM1 co-mutation was present in 4 pts (50%), followed by DNMT3A (37%) and IDH2 (37%). Six pts have been enrolled on dose level 1 (gilteritinib 80 mg) and 2 pt on dose level 2 (gilteritinib 120 mg). No DLTs have been observed in either dose level. Myelosuppression was frequent, requiring dose adjustment of ASTX727 and/or VEN in cycle 2 onward in 5 pts. Grade 3 non-hematologic adverse events included: sepsis (25%), bacteremia (25%), pneumonia (13%), and other infection (25%). No tumor lysis syndrome, differentiation syndrome, or QTc prolongation was observed. The 30-day and 60-day mortality rates were 0% and 13%, respectively. The overall response rate (ORR) was 87% with 4 (50%) pts achieving CR/CRi (1 CR, 3 CRi) and 3 (37%) pts achieving MLFS. One pt withdrew from the study in cycle 1 and was not evaluable for response. The ORR in pts with prior HMA+VEN was 83% (3 CRi, 2 MLFS). Five of 7 responders (71%) achieved the best response within 1 cycle and 2 (29%) within 2 cycles. Pts received a median of 2 cycles of therapy (range, 1-6). With a median follow-up of 6 months, median overall survival was not reached (range, 1.2-8+ months), and the estimated 6-month overall survival is 70%. One responding pt proceeded to HSCT. Two pts have died, 1 in CRi due to an unknown cause, and 1 due to disease progression and sepsis. Conclusion: The combination therapy with ASTX727, VEN, and gilteritinib appears to be effective and safe in R/R FLT3 mutated AML and HR-MDS. Myelosuppression was frequent but manageable. Continued accrual is anticipated to define the optimal triplet regimen in both R/R and newly diagnosed FLT3-mutated AML. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Prolonged Survival in Bi-Allelic TP53-Mutated (TP53mut) MDS Subjects Treated with Oral Decitabine/Cedazuridine in the Ascertain Trial (ASTX727-02)

Introduction: TP53 mutations (TP53mut) in myelodysplastic syndrome (MDS) patients have been characterized as an independent prognostic factor for poor outcome. These patients may have similar response rates to hypomethylating agents (HMAs) but markedly diminished overall survival (OS) compared to those with wild-type (WT) TP53 status (9.4 vs. 20.7 months [mo.]; Takahashi, K, et al. Oncotarget. 2016). Further analyses have defined monoallelic (MA) and bi allelic (BA)/multi-hit TP53mut populations with very different survival outcomes (8.4 vs. 30 mo.; Bernard, et al. Nat Med. 2020). Oral decitabine/cedazuridine (ASTX727) is a fixed dose combination of decitabine (35 mg) and the cytidine deaminase inhibitor cedazuridine (100 mg) with pharmacokinetic (PK) exposure equivalent to the standard intravenous (IV) decitabine regimen of 20 mg/m2 daily X 5 days on a 28-day cycle. The ASCERTAIN study enrolled MDS and chronic myelomonocytic leukemia (CMML) subjects and the primary endpoint demonstrating PK (AUC) equivalence of oral decitabine/cedazuridine compared with IV decitabine was met (Garcia-Manero, ASH 2019); median overall survival (mOS) was 31.7 mo. (Savona, et al. MDS symposium 2021). Here we present preliminary analysis of the mutation profile of subjects enrolled on ASCERTAIN and evaluate the impact on overall and leukemia-free survival based on the NCCN MDS panel with a focus on the TP53 mutant population. Methods: 133 subjects with MDS/CMML were enrolled to ASCERTAIN and were randomly assigned either IV decitabine for cycle 1 and oral decitabine/cedazuridine for cycle 2 or the opposite treatment sequence. All subjects continuing beyond cycle 2 received oral decitabine/cedazuridine for all subsequent cycles until treatment discontinuation for disease progression, toxicity, patient's decision, or hematopoietic stem cell transplantation. Whole blood collected prior to treatment was used for DNA isolation and molecular abnormalities identified using next generation sequencing (NGS) hematologic malignancy panel of 179 genes including 30 genes from the NCCN MDS panel. Results: Of the 133 treated subjects, NGS analysis was available for 125 subjects. The percentage of subjects with mutations in the following genes were: TET2 (36.8%), TP53 (35.2%), ASXL1 (28%), DNMT3A (25.6%), SRSF2_MFSD11 (17.6%), SF3B1 (15.2%), STAG2 (12.8%), EZH2 (11.2%), RUNX1 (11.2%), U2AF1 (10.4%), BCOR (10.4%), CBL (8.8%). TP53, EZH2, RUNX1, CBL, DNMT3A, SF3B1, and ASXL1 were selected for further analysis based on their reported negative impact on OS and leukemia-free survival (LFS). TP53 and CBL mutations were closely associated with a worse OS (Hazard Ratio[HR] and 95% CI: 1.70 (1.00, 2.87) and 2.54 (1.19, 5.43), respectively) and LFS (HR and 95% CI: 1.63 (0.98, 2.72) and 2.01 (0.95, 4.26), respectively) compared with WT gene status, while subjects with DNMT3A mutation showed a trending advantageous relationship with OS and LFS over WT gene status. The TP53mut population (N=44) was characterized by median age 70.5 years, 63.6% M: 36.4% F, 91% MDS: 9% CMML, IPSS categories: 20% HR, 30% Int-2, 39% Int-1, 2% LR, 9% N/A, Cytogenetics: 27% Better-risk, 18% Intermediate risk, 48% Poor risk, 5% N/A, ECOG 0: 39%, 1: 61%, MA 68%, BA/multi-hit 32%. The median OS and LFS of the TP53mut population were 25.5 and 22.1 mo., respectively, compared to the TP53 WT group with mOS and LFS estimates 33.7 and 31.7 months, respectively (Figure 1). The TP53mut population was further characterized by allelic status and found to have 14 subjects with BA mutations and 30 subjects with MA TP53 mutations without other chromosomal deletions. The respective estimated mOS and 95% CI in the BA vs MA were 13.0 (5.3, 29.1) months vs. 29.2 (19.8, NE) mo. (Figure 2). Conclusion: The NGS mutational profile of MDS and CMML subjects in the ASCERTAIN trial included 35% with TP53mut and this group had a worse survival than those with WT TP53 apparently driven by the poor outcome of those with BA TP53mut. Further LOH studies will help refine this analysis, but in this conservative estimate, treatment with oral decitabine/cedazuridine in the ASCERTAIN study resulted in an estimated survival of 13 months for BA TP53mut which compares favorably with historical results. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Phase 2 Study of ASTX727 (cedazuridine/decitabine) Plus Venetoclax in Patients with Relapsed/Refractory Acute Myeloid Leukemia (AML) or Previously Untreated, Elderly Patients with AML Unfit for Chemotherapy

Background: ASTX727, an oral formulation of the fixed dose combination of decitabine and cytidine deaminase inhibitor cedazuridine (35 mg/100 mg), has been recently approved for the treatment of patients (pts) with myelodysplastic syndrome. Combination of the hypomethylating agents (decitabine or azacytidine) with venetoclax (Ven), an orally bioavailable BCL-2 inhibitor, is the standard of care for treating pts with AML unfit for intensive induction chemotherapy based on age and/or comorbidity. We are investigating whether a total oral therapy regimen of ASTX727+ven is feasible and safe. Methods: Pts aged ≥18 years with relapsed/refractory AML (R/R) or pts with AML (excluding acute promyelocytic leukemia) aged ≥ 75 or 18 -74 with comorbid conditions prohibiting intensive chemotherapy (frontline, FL) are eligible to participate. Other eligibility criteria included adequate renal and hepatic function and absence of uncontrolled infections or other uncontrolled comorbidities and an ECOG performance status (PS) of ≤2. ASTX727 is administered orally daily on days 1-5 of each treatment cycle and Ven on days 1-28 of the first cycle after a dose ramp up of 100-200-400 mg over 3 days (with tumor lysis prophylaxis precautions and with ven dose adjustments as needed based on concurrent CYP3A4 inhibitor therapy such as azole antifungals). A bone marrow exam is performed on day 21±3 days of first cycle and ven is held if blasts cleared to <5% to allow count recovery. Bone marrow exam is repeated on day 28±7 days to document response. Cycles are repeated every 4-8 weeks depending on count recovery and venetoclax is administered for 21 days in subsequent cycles. Dose adjustments of both ASTX727 and ven are permitted depending on tolerance. Results: As of July 11th, 2022, a total of 37 pts (21 FL, 16 R/R) have been enrolled on the study. The median age is 75 years (range, 46-92); 73 in the FL cohort and 75.5 in the R/R cohort. Nine (43%) frontline pts were older than 75 years including 8 pts (38%) ≥80 years. In the R/R cohort, 11 pts (69%) were 70-80 years. The median PS was 1 (range 0-3). The R/R cohort had a median 2 of prior treatments (range, 1-4). In the frontline cohort, 4 pts (19%) had normal cytogenetics, 9 (43%) a complex karyotype; and 7 (33%) other abnormalities including trisomy 8 and miscellaneous. In the R/R cohort, 44% had normal karyotype, 38% complex karyotype and 13% other. Mutations of note in the frontline cohort were TET2 (35%), ASXL1 (30%), RUNX1 (24%), TP53 (16%), and DNMT3A (11%). The overall response including complete response (CR), CR with incomplete count recovery (CRi) and morphological leukemia free state (MLFS) in the frontline cohort is 65% (4 CR, 6 CRi, 1 MLFS and 4 non-responders and 2 not evaluable). 2 pts received only one day of therapy for severe adverse events unrelated to therapy (1 due to ischemic stroke, 1 due to septic shock) and were not evaluable for response. In the R/R cohort, the overall response rate is 53% (6 CR, 1 CRi, 1 MLFS with 7 non-responders). Both cohorts received a median of 3 cycles (range for both cohorts, 1-8). With a median follow-up of 7 months, the median survival for the FL cohort is 9.5 months (range, 0.6 - 11.1) and has not been reached for the R/R cohort (range, 0.6 - 13). Grade 3 or higher adverse events related to study drugs were mainly myelosuppression-related; 4 patients (11%) had infection complications, including 1 pt with life-threatening pneumonia, and 1 pt (3%) who had liver enzyme abnormality. Conclusion: ASTX727 and venetoclax combination is safe and feasible particularly in the advanced elderly population and demonstrates significant efficacy in pts unfit for chemotherapy both in the FL and R/R settings. Total oral therapy of ASTX727 and venetoclax combination appears to be a promising strategy for older and unfit patients with AML. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Upregulation of cytidine deaminase in NAT1 knockout breast cancer cells.

Arylamine N-acetyltransferase 1 (NAT1), a phase II metabolic enzyme, is frequently upregulated in breast cancer. Inhibition or depletion of NAT1 leads to growth retardation in breast cancer cells in vitro and in vivo. A previous metabolomics study of MDA-MB-231 breast cancer cells suggests that NAT1 deletion leads to a defect in de novo pyrimidine biosynthesis. In the present study, we observed that NAT1 deletion results in upregulation of cytidine deaminase (CDA), which is involved in the pyrimidine salvage pathway, in multiple breast cancer cell lines (MDA-MB-231, MCF-7 and ZR-75-1). We hypothesized that NAT1 KO MDA-MB-231 cells show differential sensitivity to drugs that either inhibit cellular pyrimidine homeostasis or are metabolized by CDA. The cells were treated with (1) inhibitors of dihydroorotate dehydrogenase or CDA (e.g., teriflunomide and tetrahydrouridine); (2) pyrimidine/nucleoside analogs (e.g., gemcitabine and 5-azacytidine); and (3) naturally occurring, modified cytidines (e.g., 5-formyl-2'-deoxycytidine; 5fdC). Although NAT1 KO cells failed to show differential sensitivity to nucleoside analogs that are metabolized by CDA, they were markedly more sensitive to 5fdC which induces DNA damage in the presence of high CDA activity. Co-treatment with 5fdC and a CDA inhibitor, tetrahydrouridine, abrogated the increase in 5fdC cytotoxicity in NAT1 KO cells, suggesting that the increased sensitivity of NAT1 KO cells to 5fdC is dependent on their increased CDA activity. The present findings suggest a novel therapeutic strategy to treat breast cancer with elevated NAT1 expression. For instance, NAT1 inhibition may be combined with cytotoxic nucleosides (e.g., 5fdC) for breast cancer treatment.

MDS-520 A Phase I/II Study of Venetoclax in Combination With ASTX727 (Decitabine/Cedazuridine) in Treatment-Naïve High-Risk Myelodysplastic Syndrome (MDS) or Chronic Myelomonocytic Leukemia (CMML)

In MDS, hypomethylating agents (HMA) remain the standard of care. ASTX727, an oral fixed dose combination of HMA decitabine (35mg) and cytidine deaminase inhibitor cedazuridine (100mg), was recently approved in the US for the treatment of MDS and CMML. Venetoclax (Ven), an orally bioavailable BCL-2 inhibitor in combination with azacitidine has shown preliminary clinical activity in treatment-naïve, higher risk MDS. To evaluate a total-oral regimen of Ven+ASTX727 combination in pts with higher risk MDS or CMML. This single arm Phase I/II study of orally administered ASTX727 in combination with Ven (NCT04655755) is enrolling patients ≥18 years with treatment-naïve, higher risk MDS (intermediate-2- or high-risk categories) per IPSS or CMML with excess blasts ≥5%. ASTX727 is administered orally daily on D1-5 and Ven is administered orally daily on D1-14 of 28-d cycles. The primary objective is to determine the safety and tolerability (phase 1) and overall response rate (ORR, defined as CR+mCR) (phase 2) of Ven+ASTX727 combination. 19 pts have been enrolled to date with a median age of 72 years (range 54-94) with 16 pts aged ≥65 years. These patients had a median bone marrow blast count of 12% (range 6-15%) and harbored a median number of 4 (range 1-9) mutations. Most pts had adverse risk mutations such as ASXL1 (58%) and RUNX1 (42%). There were no deaths during the 30-day and 60-day window. No tumor lysis syndrome was observed. The ORR was 95% with 3 pts achieving CR (16%) and 15 pts achieving marrow CR (79%). All pts achieved a response within 1 cycle among which 7 pts, including one with TP53mut, proceeded to hematopoietic stem cell transplant. At a median follow up of 5.9 months, the median duration of response was not reached (range 0.9-11.1 months), and the median overall survival was not reached (range 1.0-12.1 months). Ven+ASTX727 combination appears safe and demonstrates preliminary efficacy in pts with higher risk MDS or CMML with excess blasts. Total-oral regimen of Ven+ASTX727 combination appears to be a promising strategy for high-risk MDS or CMML pts.

P495: PHASE 2 STUDY OF ASTX727 (DECITABINE/CEDAZURIDINE) PLUS VENETOCLAX IN PATIENTS WITH RELAPSED/REFRACTORY ACUTE MYELOID LEUKEMIA (AML) OR PREVIOUSLY UNTREATED, ELDERLY PATIENTS UNFIT FOR CHEMOTHERAPY

Background: ASTX727, is an oral formulation of the fixed dose combination of decitabine and cytidine deaminase inhibitor cedazuridine (35 mg/100 mg). Aims: To investigate whether a total oral therapy regimen of ASTX727+venetoclax (ven) is feasible and safe. Methods: Pts aged ≥18 years (yrs) with relapsed/refractory AML (R/R) or pts with AML aged ≥ 75 or 18 -74 with comorbid conditions prohibiting intensive chemotherapy were eligible to participate (frontline-FL). Other eligibility criteria included adequate renal and hepatic function and an ECOG performance status (PS) of≤2. ASTX727 is administered daily on days 1‐5 of each treatment cycle and ven on days 1‐28 of the 1st cycle after a dose ramp up of 100-200-400 mg over 3 days (with tumor lysis prophylaxis precautions and with ven dose adjustments as needed. A bone marrow exam is performed on day 21±3 of 1st cycle and ven is held if blasts <5% to allow count recovery. Cycles are repeated every 4-8 weeks depending on count recovery and Ven is administered for 21 days in subsequent cycles, with dose adjustments as necessary depending on count recovery. Results: Between March 2021 and January 2022, 28 pts (15 FL and 13 R/R) have been treated on the study. The median age is 75 yrs (range, 47-90) with FL cohort 81 and R/R cohort 72. 9 FL pts (60%) were ≥80 and 5 (30%) 70-80 years. In R/R cohort 9 pts (69%) were 70-80 yrs. The median PS is 2 (range 0-3) and in the R/R cohort, the median number of prior treatments is 2 (range, 1-4). In the FL cohort 5 (33%) had normal and 6 (40%) a complex karyotype; 3 had other. In the R/R cohort, 15% had normal, 46% complex karyotype and 31% other. Mutations of note in the FL cohort were RUNX1 (33%), ASXL1 (33%), DNMT3A (7%), TET2 (40%) and TP53 (20%). The overall response (ORR) including complete response (CR), CR with incomplete count recovery (CRi) and morphological leukemia free state (MLFS) in the FL cohort is 61% (4 CR, 4 CRi, 1 MLFS and 3 non-responders). 3 pts received only one day of therapy for severe adverse events unrelated to therapy (1 due to ischemic stroke, 1 septic shock and 1 debilitation) and were not evaluable for response. In the R/R cohort, the ORR rate was 45% (2 CR, 2 CRi, 2 MLFS with 5 non-responders and 2 not evaluable). The median number of cycles received is 2 (range, 1-5) for both cohorts. With a median follow-up of 5 months, the median survival for the FL cohort has not been reached (range, 0.6 – 7.3) and is 7.2 (range, 0.8-7.3) months for the R/R cohort. Grade 3 or higher adverse events directly attributable to therapy were mainly myelosuppression-related and included neutropenic infections in 3 (11%) and elevation of liver enzymes in 1 (4%) pt. Image:Summary/Conclusion: The combination of ASTX727+ven is feasible, particularly in the advanced elderly population, and demonstrates significant efficacy in pts unfit for chemotherapy both in the FL and R/R settings.

P784: A PHASE I/II STUDY OF VENETOCLAX IN COMBINATION WITH ASTX727 (DECITABINE/CEDAZURIDINE) IN TREATMENT‐NAÏVE HIGH‐RISK MYELODYSPLASTIC SYNDROME (MDS) OR CHRONIC MYELOMONOCYTIC LEUKEMIA (CMML)

Background: In MDS, hypomethylating agents (HMA) remain the standard of care. ASTX727, an oral fixed dose combination of HMA decitabine (35mg) and cytidine deaminase inhibitor cedazuridine (100mg), was recently approved in the US for the treatment of MDS and CMML. Venetoclax (Ven), an orally bioavailable BCL-2 inhibitor in combination with azacitidine has shown preliminary clinical activity in treatment-naïve, higher risk MDS Aims: Based on this data, we designed a study to evaluate a total-oral regimen of Ven+ASTX727 combination in pts with higher risk MDS or CMML Methods: This single arm Phase I/II study of orally administered ASTX727 in combination with Ven (NCT04655755) is enrolling patients ≥18 years with treatment‐naïve, higher risk MDS (intermediate-2- or high-risk categories) per IPSS or CMML with excess blasts ≥5%. The primary objective is to determine the safety and tolerability (phase 1) and overall response rate (ORR, defined as CR+mCR) (phase 2) of Ven+ASTX727 combination. ASTX727 is administered orally daily on D1‐5 and Ven is administered orally daily on D1‐14 of 28-d cycles. 3 dose levels of venetoclax in combination with ASTX727 will be tested. To mitigate tumor lysis syndrome, pretreatment white blood cell count should be less than 10 × 109/L. Cytoreduction is allowed. The safety population includes all patients who received any dose of Ven+ ASTX727, and the efficacy population includes patients who have a valid baseline and post-baseline disease assessment and had received at least one dose of the study drug Results: 15 pts have been enrolled to date (Figure). The median age is 72 years (range 54-94) with 12 pts aged ≥65 years. These patients had a median bone marrow blast count of 12% (range 6-15%) and harbored a median number of 4 (range 1-9) mutations. Most pts had adverse risk mutations such as ASXL1 (67%) and RUNX1 (47%). No DLTs were observed in the initial 6 patient safety lead-in. There were no deaths during the 30-day and 60-day window. No tumor lysis syndrome was observed. The ORR was 87% with 3 pts achieving CR (20%) and 10 pts achieving marrow CR (67%). All pts achieved a response within 1 cycle among which 4 pts, including one with TP53mut,proceeded to hematopoietic stem cell transplant. 2 pts had stable disease at the end of 1 cycle. At a median follow up of 8.8 months, the median duration of response was not reached (range 0.9-11.1 months), and the median overall survival was not reached (range 1.0-12.1 months). Image:Summary/Conclusion: Ven+ASTX727 combination appears safe and demonstrates preliminary efficacy in pts with higher risk MDS or CMML with excess blasts. Total-oral regimen of Ven+ASTX727 combination appears to be a promising strategy for high-risk MDS or CMML pts and may alleviate the burden of chronic, long-term parenteral HMA treatment