Combination Of Azacitidine Research Articles

A Novel Histone Deacetylase Inhibitor MCT-3 Augments Inhibition of Leukemia Cell Growth and Suppresses Induction of MMP-9 Expression by Azacitidine in HL-60 Cells.

Azacitidine (AZA), a demethylating agent, has recently been demonstrated to have efficacy in the treatment of myelodysplasia and acute myeloid leukemia. A potential concern when considering the use of this agent is a recent report demonstrating AZA-mediated re-activation of matrix metalloproteinase-9 (MMP-9) expression facilitating the invasive metastatic phenotype (Sato NM et al J Natl Cancer Inst. 2003 Feb 19;95(4):327–30). Histone deacetylase inhibitors (HDACi) are a recently identified class of agents with considerable in vitro and in vivo activity and early phase clinical efficacy in the treatment of haematological malignancies (Bolden JE Nat Rev Drug Discov. 2006 Sep;5(9):769–84). Pre-clinical in vitro studies suggest that addition of HDACi to 5-aza-2′-deoxycytidine, a derivative of AZA, enhances the efficacy of this agent (Yang H et al Leuk Res. 2005 Jul; 29(7):739–48) whilst early phase clinical trials identify therapeutic activity using a combination of demethylating agents and HDACi (Garcia-Manero G, Blood. 2006 Nov 15;108(10):3271–9). Our current study aimed to determine the in vitro activity and molecular mechanisms of action of the novel HDACi MCT-3, a derivative of Oxamflatin, a hydroxamate analogue, (Dear AE, et al, Org Biomol Chem, 2006, 4, 3778–3784) in the HL-60 cell line alone and combination with AZA. AZA (1.0 microM) and MCT-3 (2.5 microM) alone inhibited HL-60 cell growth over 24hrs by 40%, 30% respectively. The combination of AZA with MCT-3 inhibited HL-60 cell growth up to 50%. Real-time PCR demonstrated that AZA and MCT-3 alone increased p15INK4b and Caspase 3 mRNA expression 2 fold. A Combination of AZA with MCT-3 increased p15INK4b and Caspase 3 mRNA expression up to 2.5 and increased p21WAF1/CIP1 and the orphan nuclear receptor Nur77 expression 2 fold. A combination of AZA and MCT-3 significantly attenuated AZA-induced MMP-9 mRNA expression and proteolytic activity. AZA and MCT-3 alone reduce HL-60 cell growth in vitro. Addition of MCT-3 to AZA increased inhibition of cell growth, suggesting that this HDACi may have the potential for additive activity with demethylating agents. AZA and MCT-3 have similar effects on expression of genes implicated in cell cycle arrest and apoptosis. Increased expression of p21WAF1/CIP1 and the orphan nuclear receptor Nur77 via inhibition of cell cycle progression and enhanced apoptosis may in part be responsible for the enhanced anti-leukaemia activity of the combination of AZA and MCT-3. Importantly MCT-3 is able to inhibit AZA-mediated induction of MMP-9 expression.

Phase I/II Study of MGCD0103, an Oral Isotype-Selective Histone Deacetylase (HDAC) Inhibitor, in Combination with 5-Azacitidine in Higher-Risk Myelodysplastic Syndrome (MDS) and Acute Myelogenous Leukemia (AML).

Epigenetic alterations are common in leukemia. MGCD0103 in an oral isotype-selective HDAC inhibitor that synergizes in vitro with the DNA methyltransferase inhibitor 5-azacitidine (Vidaza, Pharmion). Both agents have single-agent clinical activity in MDS and AML (Garcia-Manero, ASCO, 2006 & Silverman, JCO, 2002). We have developed a Phase I/II study of 5-azacitidine in combination with MGCD0103 in patients with AML and MDS. Patients with MDS (≥10% marrow blasts), relapsed/refractory AML, or untreated elderly patients with AML were eligible. Adequate performance status, renal and hepatic functions were required. 5-azacitidine was administered at its approved dose/schedule: 75 mg/m2 SC daily for the first 7 days of a 28 day cycle. MGCD0103 was administered as a flat dose orally three-times a week starting on the 5th day of 5-azacitidine administration. The phase I portion of the study design followed a classic “3+3” model and only MGCD0103 was dose escalated. The phase II portion targeted a 30% response rate. Final data from the Phase I and II portions of the study will be presented at the Meeting. Five dose levels of MGCD0103 have been evaluated: 35, 60, 90, 110 and 135 mg. At current data cut-off, 37 patients registered in the study were fully evaluable: median age was 67 (range 27–85); 31 patients had AML and 6 MDS. A total of 97 cycles were administered to date, mean = 2.6 (range 1–12). Dose limiting toxicities included nausea, vomiting, anorexia, diarrhea and dehydration which appear similar to dose limiting toxicities for MGCD0103 alone. The MTD of MGCD0103 was initially determined to be 110 mg, however, upon cohort expansion, this dose level was associated with excess toxicity and the starting dose was decreased to 90 mg. Eleven (30%) patients have achieved response: 4 CR, 5 CR-i, and 2 PR. Of these 11 patients, 6 continue on study with mean duration on study of 7 cycles. Of the 5 patients discontinued, 3 discontinued due to SAEs, 1 due to progressive disease and 1 to undergo transplantation. Of the 27 patients at the phase II dose levels of 90 and 110mg, 10 achieved a response (37%; same rate at both doses). Preliminary response data are available at the time of abstract preparation for 13 additional patients, revealing 4 with CR (one of which had 1% residual peripheral blast) and 3 with CR-I for a response rate of 53% in this subset. MGCD0103 pharmacokinetics were not affected by 5-azacitidine. Likewise, co-administration of MGCD0103 had no impact on the pharmacokinetics of 5-azacitidine. A majority of patients exhibited a substantial reduction in PBMC HDAC activity during treatment with the combination. Analysis of DNA methylation is ongoing. In conclusion, the combination of 5-azacitidine with MGCD0103 is safe in patients with advanced AML/MDS and has clinical activity potentially superior to that expected with 5-azacitidine alone in this patient population. These results form the bases of a planned randomized study of 5-azacitidine with or without MGCD0103 in AML and MDS.

Preliminary Results from a Phase I Study of Revlimid® (Lenalidomide) in Combination with Vidaza® (Azacitidine) in Patients with Advanced Myelodysplastic Syndromes (MDS).

Background: Early MDS becomes more advanced as immature myeloid cells proliferate, angiogenesis increases, genetic lesions accumulate, and tumor suppressor genes become inactivated through hypermethylation. Progression to acute myeloid leukemia (AML) may be prevented by targeting these defects through combination therapy, using an immunomodulatory, anti-angiogenic agent, lenalidomide (LEN), and a hypomethylating drug, azacitidine (AZA).Methods: We conducted a multicenter, Phase I trial in patients (pts) with advanced MDS (IPSS score ≥1.5, or FAB or WHO classification with ≥5% myeloblasts) starting in 6/06, with results reported through 7/07. Pts were enrolled using a “3+3” design (See Table), and could not receive LEN or AZA previously. Cycles lasted 28 days, to a maximum of 7 cycles of therapy. The primary endpoint was to determine the maximum tolerated dose (MTD) and dose-limiting toxicities (DLTs, defined as Grade 3/4 non-hematologic toxicity or >50% neutrophil (ANC) or platelet (plt) drop without recovery by Day 56) of the combination. A secondary endpoint was response as defined by the Modified International Working Group.Dose LevelAZA ScheduleLEN Schedule175 mg/m2 SC days 1–55 mg PO days 1–14275 mg/m2 SC days 1–55 mg PO days 1–21375 mg/m2 SC days 1–510 mg PO days 1–21450 mg/m2 SC days 1–5, 8–125 mg PO days 1–14550 mg/m2 SC days 1–5, 8–125 mg PO days 1–21650 mg/m2 SC days 1–5, 8–1210 mg PO days 1–21Results: Seven patients have been enrolled, 6 are evaluable for toxicity data. Median age was 64 years (range 52–70), 1 pt was female, and median follow-up is 5.5 months (range 1.5–13). All pts had RAEB-2; IPSS scores were 1.5 (4), 2.0 (2), and 3.0 (1), with IPSS cytogenetic risk categories of poor (1), intermediate (1), and good (5). No pt had a del (5q) lesion. Median time from MDS diagnosis was 3.5 wks (range 2–106). No DLTs occurred in Dose Levels 1 or 2, and MTD has not yet been reached. Grade 1/2 non-hematologic toxicities (n=6) included fatigue (4), injection site reaction (6), rash (3), pruritis (3), constipation or diarrhea (6), dizziness (1), and mucositis (1). Grade 3/4 non-hematologic toxicities included febrile neutropenia (1). Median ANC drop was 16.4% and plt drop was 10.4%. Although one patient was delayed 1 week in starting cycle 2 for neutropenia, there were no dose-reductions for toxicities. Four pts are evaluable for response: 2 had a complete response, 1 an erythroid response, and 1 progressive disease.Conclusions: The combination of LEN and AZA is well-tolerated and early results suggest efficacy in advanced MDS. Responses and toxicity data from higher Dose Levels will be presented.

Report of a Phase1/2 Study of 5-Azacitidine and Cytarabine in Patients with Relapsed, Refractory Acute Myelogenous Leukemia.

Background: Epigenetic silencing, particularly of tumor suppressor genes, is known to play a significant role in development of leukemia, and is thought to play a role in acquired drug resistance. In vitro, 5-azacitidine (azacitidine) can reverse methylation and activate expression of genes involved in response to chemotherapy, including proapoptotic genes, DNA repair genes and genes that regulate cell cycle. Thus, reversal of DNA methylation can potentially restore chemosensitivity in leukemia and combined use of DNA hypomethylating agent with chemotherapy can be an effective treatment strategy for leukemia.Methods: We developed a phase 1/2 study of azacitidine and cytarabine administered concurrently in patients with relapsed/refractory AML. Newly diagnosed patients deemed ineligible for standard therapy were eligible. Two dose levels of azacitidine were explored, dose level 0= 37.5 mg/m2 intravenously (I.V) daily for 7 days and dose level 1=75 mg/m2 IV daily for 7 days. Similarly two dose levels of cytarabine explored were; low dose- 100 mg/m2 IV continuously (CI) for 7 days and high-dose - 1 gm/m2 CI for 4 days. This resulted in 4 potential groups (Table1). After 2 cycles of induction therapy as in table 1, patients were to receive maintenance therapy upon recovery of counts. In the phase 1 portion of the study groups were opened sequentially in a “3+3” design. The phase 2 part was designed in a 2 by 2 factorial maner with adaptive randomization.Results: Twenty-seven patients with AML (male=13) have been enrolled so far, Group 1= 7, group 2= 6, group 3=11 and group 4= 3. One patient (group 1) did not receive treatment as she withdrew consent. Median age was 61 years (range, 31 to 79) and median number of prior therapies was 2 (range, 0 to 4). Twenty-one patients had received prior cytarabine including 19 who received high-dose cytarbine. Karyotype was complex in 7 patients, deletion 7 in 3 and 11q abnormality in 2. The median number of cycles administered on study was 1 (range, 1–5). One patient (group 2) developed dose-limiting toxicity (DLT) of grade 3 mucositis. No other DLTs were encountered. There was limited efficacy seen, with no patient achieving complete remision (CR). The combination achieved transient control of WBC counts in most patients treated, and one patient had reduction in marrow blasts from 24% to 3% after the 3rd cycle without recovery of blood counts and had progression of disease after 5 cycles.Conclusion: The simultaneous combination of azacitidine with cytarabine as administered in this study is safe. Efficacy evaluation of the combination was compromised by the small number of cycles of azacytidine given, but appears to be limited in a patient population composed predominantly of relapsed/refractory AML.Table 1Low-dose cytarabineHigh-dose cytarabineAzacitidine dose level 0Group 1Group 3Azacitidine dose level 1Group 2Group 4

P151 Azacitidine in combination with EPO+G-CSF and valproic acid rapidly determines hematological improvement in pretreated non responsive IPSS INT-1 MDS patients

P151 Azacitidine in combination with EPO+G-CSF and valproic acid rapidly determines hematological improvement in pretreated non responsive IPSS INT-1 MDS patients

Phase I/II Study of the Oral Isotype-Selective Histone Deacetylase (HDAC) Inhibitor MGCD0103 in Combination with Azacitidine in Patients (pts) with High-Risk Myelodysplastic Syndrome (MDS) or Acute Myelogenous Leukemia (AML).

Aberrant DNA methylation and histone code alterations are common in leukemia. The oral isotype-selective HDAC inhibitor MGCD0103 and the DNA methyltransferase inhibitor azacitidine have been shown to have synergistic antileukemia activity in preclinical models. Both agents have single-agent clinical activity in MDS and AML (Garcia-Manero, ASCO, 2006 & Silverman, JCO, 2002). Based on this, we have developed a Phase I/II study of the combination in leukemia. Patients with relapsed/refractory MDS (10% or more marrow blasts) or AML, or untreated older patients with AML are eligible. Adequate performance status, renal and hepatic functions are required. Azacitidine is administered at its approved dose/schedule: 75 mg/m2 SC daily x 7 every 28 days. MGCD0103 was started on day 5 of azacitidine and was given as an oral dose 3 times a week without adjusting for body weight, without interruption. The phase I portion of the study design followed a classic “3+3” model and only MGCD0103 was dose escalated. Three dose levels of MGCD0103 have been studied so far: 35, 60 and 90 mg. Twelve patients have been registered in this study. The median age is 61 (range 45–85). All, but 1 patient with MDS, had AML. All patients had relapsed or refractory disease. A total of 24 cycles have been administered, mean = 2.0 (range 1–4). Only 1 patient has experienced dose limiting toxicity: grade 3 vomiting at a dose of 90 mg of MGCD0103. Otherwise, the combination has been very well tolerated and the MTD has not yet been defined, with dosing currently ongoing at the approximate MTD of single agent MGCD0103: 110 mg orally three times a week without interruption. Preliminary PK data indicate that the t1/2 for azacitidine is less than 2 h, and does not appear to be affected by MGCD0103. Likewise, MGCD0103 pharmacokinetic characteristics do not appear to be affected by azacitidine. The majority of patients exhibited substantial reduction in HDAC activity during treatment with the combination. Analysis of DNA methylation is ongoing. Two patients have achieved response: 1 complete remission after 4 courses of therapy (in an older patient in first relapsed AML with an initial CR duration of 11 months and multiple cytogenetic abnormalities). A second older patient with refractory AML achieved a complete marrow CR (marrow blasts less than 5%) but died from pneumonia (not drug-related) after the second course of therapy on day 28 before peripheral count recovery. In conclusion, the combination of azacitidine with MGCD0103 is safe in patients with advanced AML/MDS. The combination has encouraging safety, PK, and clinical activity profiles. The study continues at the MTD of single agent MGCD013 in combination with azacitidine. Once the MTD of the combination is documented, the study will continue as a phase II study in this patient population.

A phase I study of a hypomethylating agent azacitidine in combination with carboplatin in patients with platinum resistant epithelial ovarian cancer

5087 Background: Hypermethylation is a common event associated with inactivation of a variety of genes and pathways involved in tumorigenesis and progression as well as drug resistance. Previous evidence displayed that hypomethylating agents could resensitize epithelial ovarian cancer cells to platinum treatment through reexpression of platinum sensitivity-related genes. Therefore, it is of great interest to use a hypomethylating agent to resensitize platinum resistant epithelial ovarian cancer cells to platinum treatment. Methods: This proof-of-concept study used a modified dose-escalation schema (3+3 design). All patients, who have progressed within the 6 months after a platinum-based regimen, are eligible if they have a pathologically confirmed epithelial ovarian cancer. All patients receive subcutaneous injection of azacitidine 75 mg/m2/day for 5 days on days 1–5 and intravenous infusion of carboplatin once on day 2 of each treatment cycle (28 days). Azacitidine is maintained at the same dosage during dose-escalation of carboplatin from AUC4 to AUC5. Results: Six patients have received a total of 16 cycles of treatment. There was no grade 2 or higher toxicity observed. Three patients were evaluable for response: 1 patient, who received 6 cycles of treatment, achieved complete response by CA125 criteria and stable disease by imaging criteria; 1 patient, who received 5 cycles of treatment, displayed stable disease; and the 3rd patient, who received 2 cycles of treatment, showed mixed response by imaging criteria. Conclusions: Preliminary data showed that a regimen of azacitidine 75 mg/m2/day for 5 days plus carboplatin AUC 5 on day 2 was well tolerated in patients with extensive prior treatment. No significant financial relationships to disclose.