Reduction In Bone Marrow Blasts Research Articles

Phase IIB Trial of Oral Midostaurin (PKC412), the FMS-Like Tyrosine Kinase 3 Receptor (FLT3) and Multi-Targeted Kinase Inhibitor, in Patients With Acute Myeloid Leukemia and High-Risk Myelodysplastic Syndrome With Either Wild-Type or Mutated FLT3

Mutations leading to constitutive activation of the FMS-like tyrosine kinase 3 receptor (FLT3) occur in blasts of 30% of patients with acute myeloid leukemia (AML). Midostaurin (PKC412; N-benzoylstaurosporin) is a multitargeted tyrosine kinase inhibitor, with demonstrated activity in patients with AML/myelodysplastic syndrome (MDS) with FLT3 mutations. Ninety-five patients with AML or MDS with either wild-type (n = 60) or mutated (n = 35) FLT3 were randomly assigned to receive oral midostaurin at 50 or 100 mg twice daily. The drug was discontinued in the absence of response at 2 months, disease progression, or unacceptable toxicity. Response was defined as complete response, partial response (PR), hematologic improvement, or reduction in peripheral blood or bone marrow blasts by ≥ 50% (BR). The rate of BR for the population in whom efficacy could be assessed (n = 92) was 71% in patients with FLT3-mutant and 42% in patients with FLT3 wild-type. One PR occurred in a patient with FLT3-mutant receiving the 100-mg dose regimen. Both doses were well-tolerated; there were no differences in toxicity or response rate according to the dose of midostaurin. These results suggest that midostaurin has hematologic activity in both patients with FLT3-mutant and wild-type. The degree of clinical activity observed supports additional studies that combine midostaurin and other agents such as chemotherapy especially in FLT3-mutant AML.

The Activity of AT9283 in Acute Myeloid Leukemia in Vitro and in the Clinic Is Karyotype Dependent.

Abstract 4159Aurora kinases (AK) A and B are overexpressed in a proportion of patients with acute myeloid leukemia (AML) and the level of overexpression correlates with their sensitivity to AK inhibition in vitro. We recently reported data from a dose escalation study of AT9238, a small molecule inhibitor of AKs in patients with refractory leukemias in which eight of 24 AML patients with relapsed/refractory AML achieved a 3 33% reduction in bone marrow blasts and haematological improvement. All patients had received at least one line of previous therapy. Further analysis has revealed that of the eight patients with relapsed/refractory AML that benefited from treatment with AT9283 five had a normal karyotype and the remaining three patients showed evidence of isolated abnormalities of chromosome 7, including 7q loss.Separately, we have reported that AT9283 inhibits the proliferation and survival of AML cell lines in vitro and suggested that those cell lines with complex karyotypic abnormalities responded differently from normal diploid lines.In these experiments AML cell lines exhibit one of two phenotypes following exposure to AT9283; rapid induction of cell death at low nM concentrations (Phenotype 1) or endo-reduplication followed by cell death at a later time point (Phenotype 2). In both scenarios treatment with AT9283 results ultimately in cell death. Cell lines with a normal karyotype tended to undergo rapid apoptosis without evidence of endoreduplication at low concentrations of AT9283. These findings provide further support for the potential importance of karyotype as a determinant of outcome in the clinical study. This is the first indication that cytogenetics might be used to predict responsiveness to Aurora kinase inhibitors in the clinic. Disclosures:Lyons:astex therapeutics: Employment. Squires:Astex Therapeutics, Ldt: Employment. Goodall:astex therapeutics: Employment. Yule:Astex Therapeutics Ldt: Employment. Ravandi:BMS: Consultancy, Honoraria, Research Funding.

A Phase 1 Dose-Escalation Study of the Novel KSP Inhibitor ARRY-520 in Advanced Leukemias.

Abstract 2047Poster Board II-24 Introduction:The mitotic kinesin spindle protein (KSP) is required for the assembly of a normal bipolar spindle and cell cycle progression through mitosis. ARRY-520 is a potent, selective inhibitor of KSP that arrests cells in mitosis forming an abnormal monopolar spindle with subsequent onset of apoptosis. ARRY 520 has shown potent activity in preclinical models of hematological malignancies which support clinical investigation of this novel targeted antimitotic therapy in patients with leukemias. ARRY-520 was evaluated in a Phase 1 trial, administered as an intravenous (IV) infusion on Day 1 or on Days 1, 3 and 5 in patients with advanced/refractory leukemias. Objectives:The primary objectives of this study were to establish the safety and the maximum tolerated dose (MTD) of ARRY-520 when given as a 1-hour infusion in either a single dose or on a Days 1, 3 and 5 divided-dose schedule per cycle. Secondary objectives were to characterize the pharmacokinetics (PK) of ARRY-520 when given on these schedules, to assess evidence of preliminary clinical activity, and to explore potential biomarkers of KSP inhibition. Methods:ARRY-520 was administered as a 1-hour IV infusion as a single dose per cycle or on a divided dose schedule, in a “3 + 3” Phase 1 design. PK analyses for ARRY-520 were performed on plasma samples collected during Cycle 1 and Cycle 2. Results:A total of 33 patients with acute myelogenous leukemia (AML) and with a median age of 66 years (range 21-88 yrs) were enrolled: 15 in the single-dose schedule (dose levels 2.5, 3.75, 4.5 and 5.6 mg/m2) and 18 in the divided dose schedule (dose levels 0.8, 1.2, 1.5 and 1.8 mg/m2/day). All but one patient had disease refractory to and/or relapsed from prior therapy with a median of 4 prior regimens (range 0-7). The MTD was 4.5 mg/m2 for the single-dose schedule with the dose-limiting toxicity (DLT) of Grade 3 mucositis and was 1.5 mg/m2/day (cumulative dose per cycle of 4.5 mg/m2) for the divided dose schedule, with DLTs of Grade 3 mucositis, hand-foot syndrome and hyperbilirubinemia. ARRY-520 was well tolerated at the MTD and doses below. At the MTD, Grades 3 or 4 reversible drug-related leukopenia were observed in 4/8 evaluable patients (Grade 0 or 1 WBC at baseline) with a median nadir occurring on Day 7. In both schedules ARRY-520 showed promising signs of clinical activity as measured by significant decreases in leukemic blasts in both the peripheral blood and bone marrow. Four of 33 patients (12%) showed at least 50% reduction in bone marrow blasts and 12/33 patients (36%) showed > 1 log reduction in blasts in the peripheral blood. Preliminary plasma PK analyses revealed increasing ARRY-520 concentrations with increasing dose, a mean terminal t1/2 of ∼90 hours, with clearance values ranging from 1.6 to 8.0 L/hr. Conclusions:ARRY-520 showed promising signs of clinical activity and has been well tolerated in both schedules investigated in patients with AML. The most prominent DLTs were mucositis and hand-foot syndrome. The MTD was determined to be the same total dose per cycle for the single-dose and the divided-dose schedule. Data including the safety, PK, pharmacodynamics and preliminary activity of ARRY 520 from this study will be presented. Disclosures:Bethelmie-Bryan:Array BioPharma: Employment. Rush:Array BioPharma: Employment. Freeman:Array BioPharma: Employment. Simmons:Array BioPharma: Employment. Ptaszynski:Array BioPharma: Employment.

Temozolomide and cisplatin in relapsed/refractory acute leukemia

Cisplatin depletes MGMT and increases the sensitivity of leukemia cells to temozolomide. We performed a phase I study of cisplatin and temozolomide in patients with relapsed and refractory acute leukemia. Fifteen patients had AML, 3 had ALL, and 2 had biphenotypic leukemia. The median number of prior chemotherapy regimens was 3 (1–5). Treatment was well tolerated up to the maximal doses of temozolomide 200 mg/m2/d times 7 days and cisplatin 100 mg/m2 on day 1. There was one complete remission in this heavily pretreated patient population. Five of 20 (25%) patients demonstrated a significant reduction in bone marrow blasts.

Phase I dose escalation trial of sorafenib as a single agent for adults with relapsed and refractory acute leukemias

7065 Background: Sorafenib is a multikinase inhibitor with activity against B-raf, VEGF, and FLT3. Based on preclinical activity in FLT3 mutant AML, sorafenib was studied in refractory acute leukemia. Methods: The primary objective was to determine the safety and tolerability of sorafenib in refractory acute leukemias. Secondary objectives included pharmacokinetics (PK) and pharmacodynamic (PD) effects of sorafenib on FLT3 phosphorylation. Dose escalation began at 400 mg BIDx14days per month, and proceeded through 600 mg BID x 21 days per month. Plasma concentration of sorafenib and its primary metabolite sorafenib N-oxide were measured by LC/MS//MS method. The plasma inhibitory assay was used to measure target inhibition of phosphorylated FLT3 and phosphorylated Erk. Results: Fifteen patients (13 = AML, 2 = ALL) were enrolled (ages 37–85) and treated on three dosing schedules (400 mg BID x 14 d, 400 mg BID x 21 days, 600 mg BID x 21days) of single agent sorafenib. The maximally tolerated dose was 400 mg BID x 21 days per month. Grade 3 or greater toxicities were experienced in 55% of cycles, most common grade 3 or greater toxicities being fatigue (16%) and hypokalemia (13%). No patients met criteria for complete or partial response, but 11 of 15 (73%) patients experienced stable disease as best response, with 6 showing a reduction in bone marrow blasts after only one cycle, half of who experienced a >50% reduction in bone marrow blasts. Interestingly, 2 pts with FLT3-ITD mutations both showed marrow blast response (1 pt >50%). Sorafenib resulted in sustained complete inhibition of FLT3 and Erk as demonstrated in all patients assessed (n = 11). Importantly, this inhibition was maintained throughout treatment cycle and 3/5 pts had FLT3 inhibitory activity 7 days post their last dose. Correlative studies suggest sorafenib N-oxide is an active metabolite. Conclusions: Sorafenib is a potent inhibitor of FLT3 with favorable PK and PD properties. Clinical activity as a single agent was limited to transient reductions in bone marrow blast counts and dose escalation was limited due to toxicities. Based on PK data in conjunction with standard curves for inhibition of FLT3 by sorafenib in plasma, the minimum FLT3 inhibitory dose of sorafenib is likely less than 400 mg BID. No significant financial relationships to disclose.

A phase I dose-escalation study of the novel KSP inhibitor ARRY-520 in advanced leukemias

7022 ARRY-520 is a potent, selective KSP inhibitor that arrests cells in mitosis with the subsequent onset of apoptosis. ARRY-520 has shown potent activity in preclinical models of hematological cancers and is being evaluated in a phase I trial in patients with advanced or refractory leukemias. The primary objectives of this study are to establish the safety and the MTD of ARRY-520 given as a single dose each cycle. Secondary objectives are to characterize the pharmacokinetics (PK) of ARRY-520, to assess evidence of preliminary clinical activity, and to explore biomarkers of KSP inhibition. ARRY-520 is administered as a 1-h IV infusion as a single dose per cycle in a “3 + 3 phase I design”. Dose escalations are following a prespecified schema. PK analyses for ARRY-520 are performed on plasma samples collected during cycle 1 and cycle 2. Pretreatment and post-treatment peripheral blood samples are collected for analysis of markers of KSP activity. To date, 15 patients, with a median age of 69 yrs (range 44–84 yrs), have been enrolled in the single dose schedule, 3 patients each at doses of 2.5 mg/m2, 3.75 mg/m2, and 5.6 mg/m2 and 6 patients at 4.5 mg/m2 per cycle. All patients had disease refractory to, and/or relapsed from, 1 or more prior therapies with a median of 3 prior regimens (range 1–10). Two patients at 5.6 mg/m2 experienced a DLT of gr3 mucositis. ARRY-520 was well tolerated at doses below 5.6 mg/m2. 4.5 mg/m2 has been determined to be the MTD. ARRY-520 has shown promising signs of clinical activity at doses of 3.75 mg/m2 and above. 2 patients had a complete reduction in their peripheral blasts on day 5 of cycle 1. Of these, one patient experienced a 70% reduction in bone marrow blasts while on study. One patient without peripheral blood blasts at baseline had a 43% reduction in bone marrow blasts and four additional patients had marked reductions in WBC counts. This is the first reported use of a KSP inhibitor in refractory and/or relapsed leukemias. ARRY-520 has been well tolerated in patients on a single dose schedule. Mucositis was observed as the DLT at 5.6 mg/m2. At doses 3.75 mg/m2 and above, ARRY-520 has shown signs of clinical activity. Based on these data, an alternative dose schedule is being explored. Updated data including safety, PK, PD, and preliminary activity of ARRY 520 from this study will be presented. [Table: see text]

5′ Azacitidine in Combination with Valproic Acid Induces Complete Remissions in Patients with Advanced Acute Myeloid Leukaemia but Does Not Eradicate Clonal Leukaemic Stem/Progenitor Cells.

Demethylating drugs (e.g. 5′ Azacitidine) and histone deacetylation inhibitors are a relatively new class of clinically active agents for acute myeloid leukaemia (AML) and high-risk myelodysplasia (MDS) that have engendered a lot of interest. However, their mechanisms of action (in terms of achieving a clinical response), and how best to use them, both remain unclear. These agents rarely cure patients; in most cases the disease relapses. To better understand the impact of these agents on haemopoiesis, we have treated 45 patients with high risk AML/MDS (30 relapsed/refractory AML, 6 de novo AML, 9 high risk MDS) with azacitidine (75 mg/m2 5 days every 28 days), sodium valproate, all-trans retinoic acid and theophylline and correlated clinical response with immunophenotypic and clonogenic analysis of committed and multi-potent leukemic stem/progenitor cells. The median age of the cohort was 66 years (range 32–85 years). Clinical responses were assessed using Cheson criteria. Responses were observed in 15 patients; 7 achieved a complete remission (CR) or CRi and 8 a partial response (>50% reduction in bone marrow blasts). The median time to achievement of maximal response was 2 cycles (range 1–6). Cytogenetic data in 40/45 patients studied showed abnormalities in 60% (24/40) of patients of which 50% (12/24) were adverse risk. 40% (6/15) patients achieving CR, CRi or PR had adverse risk cytogenetics. The most common grade 3/4 haematological toxicities were thrombocytopenia (51%), leukopenia (42%) and neutropenia (42%). Common non-haematologic toxicities were fatigue (31%), neutropenic sepsis (20%), chest infection (16%) and diarrhoea (13%). Detailed immunophenotypic analysis, clonal assays (methyl cellulose and cobblestone-area forming colony assays, CAFC) and FISH analysis were performed on stem cell enriched (CD34+CD38−), common myeloid progenitor (CMP), granulocyte-monocyte progenitor (GMP) and megakaryocyte erythroid progenitor (MEP) compartments at diagnosis and post-treatment. In all diagnostic samples there were abnormal immunophenotypic CD34+CD38-, CMP, GMP and MEP (myeloid progenitor) compartments. There was an anticipated failure of normal colony and CAFC growth from sorted CD34+CD38- and myeloid progenitor populations. In responding, but not non-responding, patients there was partial or complete restoration of normal immunophenotypic populations accompanied by normal haemopoietic colony output. However, we extended these analyses to look at the kinetics of change in clonally abnormal cells that remain after treatment in total bone marrow mononuclear cells and FACS sorted cells CD34+CD38- and myeloid progenitor populations from one informative AML patient with an abnormal karyotype at diagnosis, who had a complete morphological response. At diagnosis, cytogenetic FISH analysis on sorted cells showed that 90–100% cells were clonally abnormal in the CD34+CD38- and myeloid progenitor compartments. At the beginning of cycle 3, there wascomplete haematological response (normal blood count and bone marrow myelogram and morphology):karyotypic remission in bone marrow mononuclear cells (50 cells examined);restoration of normal colony output; andnormal immunophenotypic pattern of stem/progenitor populations.However, in sorted stem/progenitor populations, clonally abnormal cells were still detected by FISH at ~15% level. These abnormal cells provide a cellular substrate for future loss of complete clinical response. This was seen by cycle 6. Now the patient developedperipheral cytopenias;reduction in normal colony output;increased bone marrow dysplasia but no excess of blasts.The bone marrow mononuclear karyotype was still normal. However, now there was an increase to 100% clonally abnormal cells in the GMP compartment whereas the CD34+CD38- compartment there were still 15% clonally abnormal cells. Though information from additional patients under study will be critical to understand the generality of these findings, these data provide the first insights into the impact of treatment and the cellular kinetics of relapse. In conclusion, this is the first study to document the clinical and biological impact on stem/progenitor and peripheral haemopoiesis of this class of agent, or indeed of any drug in AML and MDS. The findings from studies like this will have general implications on how to assess the clinical and biological impact of therapies in AML and MDS.

A Phase I Dose Escalation Study of KW-2449, An Oral Multi-Kinase Inhibitor against FLT3, Abl, FGFR1 and Aurora in Patients with Relapsed/Refractory AML, ALL and MDS or Resistant/Intolerant CML

Background: Activating mutations of FMS-like tyrosine kinase 3 (FLT3) occur in 30% of patients with de novo AML and confer a poor prognosis. KW-2449 is an oral multi-kinase inhibitor which is highly potent against mutant FLT3 (IC50=0 1–7 nmol/L) and other tyrosine kinases including FGFR1, TrkA, Abl (including T315I) and Aurora A serine threonine kinase. Based on the activity of KW-2449 and its metabolite (M1) in both in vitro and in vivo preclinical leukemia models, KW-2449 was evaluated in patients with leukemia and MDS in this first-in-man study. M1 is formed via monoamine oxidase-B and aldehyde oxidase mediated oxidation of KW-2449.Methods: The objectives were to assess the safety, tolerability, pharmacokinetic (PK) and pharmacodynamic effects of KW-2449 in patients with refractory/relapsed AML, ALL and MDS, or resistant/intolerant CML. A range of daily doses of KW-2449 (12.5–250 mg twice daily, i.e. 25–500 mg/day) on 2 treatment schedules (14 or 28 days) with a recovery period of 7–28 days between cycles were evaluated. Dose limiting toxicity (DLT) and maximum tolerated dose (MTD) were assessed for the 1st cycle. The 28-day schedule was later eliminated. The plasma concentration of KW-2449 and M1 were analyzed by LC-MS/MS method. A plasma inhibitory activity (PIA) assay [Blood 108(10) 3477–83] for P-FLT3 and P-STAT5 was used to measure FLT3 inhibition.Results: 37 patients aged 26–88 years (16 male) were treated at 7 dose levels: 25, 50, 100, 200, 300, 400, and 500 mg daily. Thirty-one patients had AML, 5 CML and 1 ALL. The mean duration of therapy was 2 cycles in AML patients and 4 cycles in CML patients. KW-2449 was rapidly absorbed and metabolized to M1. The half-lives for KW-2449 and M1 were not dose-related and ranged from 2.4 to 4.9 hours and 2.6 to 6.6 hours, respectively. Administration of KW-2449 in a BID regimen led to minor accumulation and was consistent with the short half-life. The PIA assay demonstrated the near complete down-regulation of P-FLT3 and P-STAT5 2 hours post-dose at a dosing level of 400 mg daily. The extent of inhibition was lower at 8 hours and generally absent at 12 hours post-dose. The most frequently reported adverse events (AEs; any grade, regardless of causality) were nausea (70.3%), vomiting (48.6%), fatigue (45.9%), diarrhea (32.4%), dyspnea (29.7%), febrile neutropenia (29.7%), pain in extremity (29.7%), and arthralgia (27.0%). Febrile neutropenia (24.3%), pneumonia (10.8%), and thrombocytopenia (10.8%) were the most frequently reported Grade 3/4 AEs. DLTs occurred in 2 patients: Grade 3 atrial fibrillation (100 mg daily) and Grade 3 nausea and vomiting (500 mg daily). A total of 70 SAEs were reported in 27 patients including 11 on-study deaths; only atrial fibrillation and pleural effusion were considered possibly related to KW-2449. Eight of 31 patients with AML (26%) (FLT3 mutation: 5 positive and 3 negative) and 1 of 5 patients with CML (20%) exhibited a ³ 50% reduction in peripheral blasts and/or bone marrow blasts from baseline to the end of Cycle 1. One patient (500 mg daily) with AML exhibited a > 50% decrease in peripheral blasts, increased platelets, and ANC, and decreased WBC count. A patient with CML (Bcr-Abl T315I +) lost the mutant clone while on KW-2449 treatment.Conclusions: KW-2449 was safe and well tolerated at the dose levels evaluated. There were no complete or partial responses, but transient decreases in peripheral blood and bone marrow blasts were observed, justifying continued investigation of this agent. Sustained inhibition of P-STAT5 and P-FLT3 was not achieved at trough at the highest BID dose evaluated. TID and QID dosing schedules should be evaluated to accommodate the short t1/2 and to achieve sufficient target inhibition.

Expanding the Use of Retinoids in Acute Myeloid Leukemia: Spotlight on Bexarotene

In this issue, Tsai et al. ([1][1]) report on a phase-I clinical trial of bexarotene in 27 non-M3 acute myeloid leukemia (AML). AML is characterized by abnormalities in the myeloid line at various stages of commitment and maturation, leading to an accumulation of granulocyte or monocyte precursors.

A Phase I Study of Bexarotene, a Retinoic X Receptor Agonist, in Non-M3 Acute Myeloid Leukemia

Bexarotene is a retinoic X receptor agonist that has been shown in vitro to inhibit growth and induce differentiation of myeloid leukemic cell lines. We therefore conducted a phase I dose escalation study to assess the maximum tolerated dose, toxicities, and activity of bexarotene in patients with non-M3 acute myeloid leukemia (AML). We enrolled patients with active non-M3 AML who had either relapsed or refractory disease or were not eligible for standard cytotoxic chemotherapy. Cohorts of three to six patients received escalating doses of daily oral bexarotene ranging from 100 to 400 mg/m(2) until evidence of disease progression or unacceptable adverse events occurred. Twenty-seven patients, with median age of 69 years (range, 51-82 years), were treated. Twenty-four (89%) patients had undergone prior chemotherapy. At the highest dose level tested (400 mg/m(2)), three of six patients had to reduce their dose of bexarotene due to grade 3 adverse events. The maximum tolerable dose of bexarotene was determined to be 300 mg/m(2). Clinical activity was manifested by 4 (15%) patients with reduction in bone marrow blasts to <or=5%, 11 (41%) patients with improved platelet counts, and 7 (26%) patients with improved neutrophil counts. Three patients with relapsed AML survived >1 year while taking bexarotene. Leukemic blast differentiation was suggested by the presence of the leukemic cytogenetic abnormality in mature circulating granulocytes and the occurrence of differentiation syndrome. The recommended dose of bexarotene for future studies is 300 mg/m(2)/d. Bexarotene is well tolerated in patients with non-M3 AML and has evidence of antileukemic activity.

Phase 1 study of XL119, a rebeccamycin analog, in patients with refractory hematologic malignancies

XL119 is a water-soluble derivative of rebeccamycin with dose-dependent myelosuppression as dose-limiting toxicity in phase 1 studies of solid tumors. A phase 1 study was conducted to determine the maximum tolerated dose and toxicities of XL119 in patients with advanced myelodysplastic syndrome and relapsed or refractory acute leukemias. Thirty-one patients were treated at 7 dose levels ranging from 140 to 260 mg/m(2)/daily times 5 in a 21-day cycle. Consenting patients had correlative biologic parameters studied. Dose-limiting toxicity was grade 3/4 mucositis. The recommended phase 2 dose in hematologic malignancies is 240 mg/m(2)/daily times 5 in a 21-day cycle. Clinically significant reduction in bone marrow blasts were seen in 5 patients and additional patients had reductions in peripheral blood blasts. However, the responses were transient. Changes of plasma vascular endothelial growth factor levels from Day 1 to Day 7 correlated negatively with changes in peripheral blood blasts from Day 1 to Day 7. Further assessment of XL119 in combination with other agents in patients with acute leukemias and high-risk myelodysplastic syndrome is warranted.

Feasibility Study of Temozolomide Maintenance Therapy in Elderly Patients with AML Following Induction Therapy with High Dose Mitoxantrone and High Dose Ara-C.

With intensive chemotherapy, approximately 50% of elderly patients with AML achieve a complete remission, however essentially all will relapse within 2 years. To date, there is no standard effective postremission therapy for these patients, and most elderly patients tolerate multiple courses of intensive chemotherapy poorly. We previously demonstrated that temozolomide has significant antileukemic activity in patients with relapsed/refractory acute leukemia (Seiter K, et al JCO 2002). In that study 9/20 patients treated had a significant reduction in bone marrow blasts, and 4 patients had a formal complete response. Currently we are evaluating the feasibility and efficacy of temozolomide maintenance therapy administered after one cycle of intensive induction chemotherapy in elderly patients with newly-diagnosed AML. Induction consists of ara-C, 3 gm/m2 daily for 5 days, and mitoxantrone, 80 mg/m2 times one on day 2. Patients achieving complete remission then receive temozolomide 200 mg/m2/d for 7 days every 5 weeks until progression. To date, 34 pts have been treated: median age 69.5 (range 61–84); M/F: 21/13; prior AHD: 47%; cytogenetics: good: 1, intermediate: 24, poor: 7, insufficient: 2. Response to high-dose ara-C/high-dose mitoxantrone induction: CR: 56%, failure: 32%, toxic death: 12%. Two pts expired in remission unrelated to temozolomide: pneumonia, 1 pt, C. difficile colitis: 1 pt. Of 19 patients in CR, 16 received temozolomide maintenance. The median number of temozolomide cycles was 2 (range 1–14). 3 pts stopped temozolomide after 1 cycle (1 refused further therapy, 1 had stem cell transplant, 1 died from C. difficile colitis). Treatment was well tolerated and was administered in the outpatient setting. The median duration of remission is 6.8 months (95% CI: 4.5–9.1 months), and median survival is 5.5 months (95% CI: 3.7–7.3 months) for all patients and 10.1 months (95% CI: 4.9–15.2 months) for patients achieving CR. Temozolomide maintenance is feasible in patients with AML in CR. Further accrual is continuing to assess if remission duration and survival will be improved compared to historical controls.

A Phase 1 Trial of Imatinib Mesylate in Combination with Daunorubicin and Cytarabine for Patients for C-kit Positive Relapsed Acute Myeloid Leukemia (AML).

C-kit is expressed on more than 10% of blasts in 95% of patients with relapsed AML. Imatinib mesylate (IM) inhibits c-kit and is active in relapsed/ refractory AML. We conducted a Phase 1 trial of IM combined with daunorubicin (D) and cytarabine (A) in patients with c-kit+ relapsed AML.Methods: All patients were treated at the Cleveland Clinic from 2003–2007. Cytogenetic (CG) risk was defined by CALGB criteria. Eligibility criteria included: relapsed AML (excluding APL), relapse more than 6 mo from induction therapy, ECOG performance status 0–2, ≥20% c-kit+ blasts (CD117+ by flow cytometry), age ≥18 yrs, not pregnant or lactating, creatinine ≤2 mg/dL, AST and ALT ≤2 x upper limits of normal, bilirubin ≤2 mg/dL. The concurrent use of antimicrobials which significantly interact with IM was prohibited. All patients received D 45 mg/m2 IV for 3 days, and A 100 mg/m2/d continuous infusion for 7 days (7+3). IM dose was escalated using a standard 3 x 3 design. Dose levels were: −1 (300 mg), 1 (400 mg), 2 (600 mg), 3 (800 mg). IM was administered with the first dose of chemotherapy and continued daily until disease progression, intolerance, removal from study for another treatment (including alloBMT), or dose-limiting toxicity (DLT). Initially, patients with persistent leukemia on Day 14 bone marrow were removed from protocol. A later amendment allowed patients with persistent leukemia, but ≥50% reduction in bone marrow blasts to remain on protocol. These patients continued IM and received 5 days of A (100 mg/m2/d) and 2 days of D (45 mg/m2/d).Results: Seventeen patients have been enrolled: median age was 47 yrs (range 30–71) and nine (53%) were male. The average time from CR to relapse was 19.2 mos (range, 7.4–36.2 mos). CG were poor risk in 29%, intermediate risk in 59%, and good risk in 12%. The median percentage of c-kit positive blasts was 86% (range, 52–98%). Nine patients were treated at dose level 1 (400 mg) because 3 patients discontinued IM before DLT could be evaluated (2 due to nausea, 1 due to persistent leukemia on a Day 14 bone marrow). Of the 6 evaluable patients, 2 patients had a DLT. Both of these were ≥Grade 3 hepatotoxicity. One patient had a bilirubin of 6.0 mg/dL (normal range 0–1.5). Therefore, subsequent patients were treated at dose level −1. Eight patients have been treated at dose level -1. Of these, 6 are evaluable for DLT. Only 1 DLT (bilirubin 5.5 mg/dL) was seen, making 300 mg/d the MTD. Fifteen out of 17 patients are evaluable for response, and 9 of these 15 have achieved CR (53%) or CRp (7%). Patients received IM for a median of 35.9 days (range 9–101 days). Three patients discontinued IM when they proceeded to alloBMT. One patient continues on IM and remains in remission on Day 180 of therapy. Enrollment to the expanded cohort at the MTD is ongoing to better evaluate response.Conclusions: The MTD of IM in combination with 7+3 was 300 mg. Since the hepatotoxicity appeared to be reversible with discontinuation of IM, it may be possible to escalate IM doses further and re-challenge. The reason for the elevated bilirubin is unclear but may be due to pharmacokinetic interactions, chemotherapy, and/or the presence of active leukemia. Further pharmacokinetic studies are being performed for further evaluation. Given the preliminary encouraging activity of this Phase 1 combination, a Phase 2 study is planned.

Phase II Study of High Dose Lenalidomide as Initial Treatment for Older Acute Myeloid Leukemia Patients: Early Results Show a Significant Reduction of Bone Marrow Blasts after 14 Days of Therapy.

AML patients over the age of 60 years have a poor prognosis, share disease characteristics with myelodysplastic syndrome (MDS) patients, and warrant novel therapeutic approaches. Lenalidomide has immunomodulatory and anti-neoplastic properties which can induce morphologic and cytogenetic responses in MDS patients, including those with excess blasts. We hypothesized that lenalidomide may be active against AML, and have employed a high dose strategy without dose reductions for hematologic toxicities. Here, we report preliminary results from a phase II study of high dose lenalidomide for front-line treatment of AML ≥ 60 without chromosome 5q deletion or favorable cytogenetics. Treatment included 2 cycles of high dose lenalidomide (50mg/day x 14 days, 30 days of rest, 50mg/day x 21 days), followed by maintenance therapy (10 mg/day) in non-progressing patients. Fifteen patients were enrolled in the first stage between 2/27/07 and 8/3/07. Median age was 71 years (range 60–86 years); ECOG performance status was 0 (4/15, 27%), 1 (10/15, 67%), or 2 (1/15, 7%); 11/15 (73%) patients were male; and 5/15 (33%) patients had prior MDS. Cytogenetics were normal (n=9), loss of chromosome 7 (n=2), loss of chromosome 20 (n=1), trisomy 13 (n=1), or complex (n=2). Overall, the treatment regimen was well tolerated. In the first stage of the study, 12/15 patients are evaluable for day 15 bone marrow and peripheral blood blast changes following the initial high dose lenalidomide cycle (50 mg/day x 14 days). Pre-therapy WBC counts (mean ± SEM) were 13,825 ± 4,447/uL (range 1,100–45,300/uL) and day 15 WBC counts were 4,742 ± 2,136/uL (range 300–24,400/uL). Day 15 bone marrow myeloblast percentages were significantly reduced in 9/12 patients (mean ± SEM decrease of 53 ± 10%, P=0.01, range 18–100%). In addition, the bone marrow blast index (% cellularity x fraction of blasts) decreased significantly after 14 days of high dose lenalidomide (mean ± SEM reduction of 66 ± 11%, P=0.02). Moreover, 5/8 patients with circulating blasts at diagnosis showed clearance of their peripheral blasts at day 15. These findings suggest that lenalidomide is an active agent against acute myeloid leukemia. Results on AML blast changes, response by international working group criteria, and toxicities in this patient cohort with ≥4 months of follow-up will be updated and presented.

A Phase I Trial of AT9283, a Multitargeted Kinase Inhibitor, in Patients with Refractory Hematological Malignancies.

AT9283 is a small molecule inhibitor of aurora kinases A and B, JAK2, and JAK3 (IC50 <5nM in each case). AT9283 was administered as a 72-hour continuous intravenous infusion to patients with refractory hematological malignancies at 6 different dose levels: 3, 6, 12, 24 and 48 mg/m2 daily x 3. Inclusion/exclusion criteria, and definitions of DLT and MTD were standard. A 3+3 dose escalation phase I design was implemented. To date, 20 patients have been treated: 3 at 3, 3 at 6, 7 at 12, 3 at 24 and 4 at 48 mg/m2/daily x 3. Seven were females; median age was 58 years (range 22 to 86); 19 patients had previously treated acute myeloid leukemia (AML); 15 were in second or subsequent salvage; abnormal cytogenetics were present in 15. Two patients had undergone prior matched unrelated allografts. The median number of prior chemotherapy regimens was 2 (range 1 to 5). Pharmacokinetic data is available for patients treated at early dose levels only. Median systemic clearance varied up to two-fold between patients with some suggestion of a non-linear increase in exposure at higher dose levels. At the completion of the infusion, the plasma concentration of AT9283 decreased rapidly exhibiting a multiphasic elimination and a terminal half-life of 6 to 13 hours. At doses of 3 to 48 mg/m2/D x 3, no MTD has been defined; 2 patients treated at 12 mg/m2/D x 3 developed tumor lysis syndrome which resolved spontaneously in one case and required hemodialysis in the second. This dose level was expanded to treat 6 patients. Patients received a median of two cycles of treatment (range 1 to 3). So far, 6 of 20 patients (30%) exhibited evidence of anti-leukemia activity, with significant reductions in bone marrow blasts (≥ 50%) observed at all dose levels: In summary, AT9283 shows good early results and continues to accrue patients to define the DLT and MTD with the 72-hour infusion schedule. In vitro studies suggest that prolonging the duration of infusion will increase the drug's biological effect. This will be explored following the identification of the MTD for a 72-hour infusion.

Lenalidomide in High-Risk Myelodysplastic Syndrome and Acute Myelogenous Leukemia Associated with Chromosome 5 Abnormalities.

Background: Treatment with lenalidomide produces remarkable responses in patients with myelodysplastic syndrome and deletion 5q 31 (5q-) cytogenetic abnormality; transfusion independence in 67% of patients, major cytogenetic responses in 44% and a median duration of response of 115.9 weeks (List et al. Abstract #251; Blood: 108). Responses have been reported in few patients with MDS and complex karyotype as well. Since chromosome 5 abnormality is considered to be an early genetic event, we designed a phase 2 study of lenalidomide in patients with high-risk MDS and relapsed/refractory acute myelogenous leukemia (AML) with any chromosome 5 abnormality.Method: The dose of lenalidomide was 25 mg/day, orally for 21 days with 7 days rest (28 day cycle). For patient with platelet count < 50,000/μl, the starting dose is 15 mg/day orally for 21 days with 7 days rest (28 day cycle). Dose interruptions/reductions are planned for ≥ grade3 neutrpenia with fever or ≥ grade 4 neutropenia (for patients starting with absolute neutrophil count [ANC] >1000/μl) and for grade 3 thrombocytopenia (for patients starting with platelet count ≥ 50,000/μl).Results: Eleven patients out of this 30 patient study have been treated so far; 6 with AML and 5 with MDS. All patients with AML and 2 patients with MDS had previous therapy. Median age is 63 years (range, 44 to 80 years). Nine patients had complex karyotypic abnormality and 2 patients had single additional chromosomal abnormality (+8 in one and 11q abnormality in another) along with 5q deletion. One patient with AML (previously treated with decitabine) and with karyotypic abnormality of trisomy 8 in addition to 5q deletion, achieved complete remission after 3 cycles of therapy. Another patient had reduction in bone marrow blasts from 21 to 5%, but could not continue lenalidomide due to profound thrombocytopenia complicated by gastrointestinal bleeding and eventually had disease progression. Four (4/6) patients with a platelet count of ≥ 30,000/μl and 5 (5/6) patients with ANC of ≥500/μl at study entry experienced worsening cytopenias. No grade 3/4 non-hematological toxicities were seen. One patient in CR currently remains on study. Reasons off study include disease progression (8 patients), stem cell transplant (1 patient) and one patient died while on study from sustained pancytopenia, infection and multiorgan failure.Conclusion: The role of lenalidomide in the treatment of high-risk MDS and AML with chromosome 5 and additional karyotypic abnormalities needs to be defined in the context of clinical trials. Future studies are needed to examine lower doses with or without growth factor support.

Quantitative Molecular Expression of the Immunoregulatory Enzyme Indoleamine 2,3-Dioxygenase in Acute Myeloid Leukemia Cells as a Possible Marker for Minimal Residual Disease Detection.

The expression of the catalytic enzyme of tryptophan, indoleamine 2,3 dioxygenase (IDO) has been identified as a T-cell inhibitor effector pathway in different normal and neoplastic cells. We have recently shown that normal bone marrow (BM) cells, including hematopoietic CD34+ cells, express IDO mRNA only upon IFN-γ stimulation, whereas in a subset of human acute myeloid leukemia cells (AML) IDO is constitutively expressed both at molecular and protein level and induces immunological escape by promoting the generation of T-reg cells. To investigate whether the IDO transcript can be used as a marker for minimal residual disease (MRD) detection, we used a sensitive real-time reverse transcription-polymerase reaction assay (qRT-PCR) to quantify IDO mRNA levels in peripheral blood (PB) and BM samples of newly diagnosed AML patients. The level of IDO transcript was evaluated as IDO copy number/104 ABL copies. As control samples, we used normal PB and BM mononuclear cells (MNCs). Our data showed that normal BM and PB cells expressed minimal amount of IDO mRNA (range 100–1000). Among AML samples, we identified three subsets of patients according to IDO mRNA expression: 1) 33/74 (44.6%) IDO negative (i.e. range < 100), 28/74 (37.84%) IDO low (range 100–1000) and 13/74 (17.6%) IDO high (i.e. range > 1000). BM and PB AML blasts gave similar results. Assessment of protein expression and enzymatic activity was in accordance with molecular results. Some patients were evaluated for IDO mRNA expression before and after induction chemotherapy and the IDO levels were found to correlate with the reduction of BM blasts. Taken together, our qRT-PCR data demonstrate that1.normal PB and BM cells are negative for IDO mRNA expression, which, in turn, is significantly up-regulated in a subset of AML patients (IDO high) and2.IDO mRNA expression correlates with tumor burden,thus suggesting its possible role in the detection of MRD in IDO high patients.

A Phase I Dose Escalation Study of KW-2449, an Oral Multi-Kinase Inhibitor Against FLT3, Abl, FGFR1 and Aurora in Patients with Relapsed/Refractory AML, Treatment Resistant/Intolerant CML, ALL and MDS.

Background: Activating mutations of FMS-like tyrosine kinase 3 (FLT3) occur in 30% of patients (pts) with de novo AML and confer a worse prognosis. KW-2449 is an oral multi-kinase inhibitor highly potent against mutant FLT3 (IC50= 1–7 nmol/L) and other tyrosine kinases including FGFR1, TrkA, Abl (including T315I), JAK2, c-KIT, and c-SRC and Aurora A serine tyrosine kinase. Based on the anti-leukemia activity of KW-2449 demonstrated both in vitro and in vivo preclinical leukemia models, KW-2449 is being evaluated in hematologic conditions in the first-in-man study.Methods: The study objectives were to assess the safety, tolerability, pharmacokinetic (PK) and pharmacodynamic effects of KW-2449 in pts with refractory/relapsed AML or treatment resistant/intolerant CML/ALL and MDS. A range of daily doses of KW-2449 (25–500 mg/day) divided into q 12h dosing on 2 treatment schedules (14 days vs. 28 days) with a recovery period of 7–28 days between cycles. Dose limiting toxicity (DLT) and maximally tolerated dose were assessed for the 1st cycle. Serial samples for PK analysis were collected immediately before and after KW-2449 on treatment days 1, 14 and 28. The 28-day schedule was later eliminated. Plasma concentrations of KW-2449 and its active metabolite (M1) were analyzed by LC-MS/MS. A plasma inhibitory activity (PIA) assay [Blood 108(10) 3477–83] for P-FLT3 and P-STAT5 was used to measure FLT3 inhibition.Results: To date, 29 pts (15 female) have been enrolled (median age 60 years; range 25–82) and treated at 5 dose levels: 25, 50, 100, 200, and 300 mg daily. Twenty-five pts had AML and 4 CML (3 with T315I mutation); 24 pts completed at least 1 cycle. KW-2449 was rapidly absorbed and metabolized to M1. Elimination half-lives were 2.8–3.9 h for KW-2449 and 3.8–5.5 h for M1. Plasma levels of M1 were lower on Days 14 and 28 compared to Day 1, suggesting inhibition of this pathway upon multiple dosing. A single DLT of grade 3 pneumonia was reported on 100 mg but no further DLTs were seen in the expanded cohort at that dose or at 200 or 300 mg daily. A total of 33 SAEs were reported of which 6 were considered possibly related (by the Investigator) to KW-2449: dyspnea, pneumonia, atrial fibrillation, cardiac ischemia, ventricular arrhythmia, and pleural effusion. There have been 5 deaths on study (none drug-related): disease progression (2), neutropenic sepsis, infection with renal failure, and pneumonia. Seven pts had stable disease after 1 cycle. Three pts with AML (2 FLT3+) had ≥ 50% reduction in peripheral and/or bone marrow blasts in the 1st cycle. In vivo FLT3 inhibition, as measured directly in patient blasts, correlated with blast reduction. The extent and duration of FLT3 inhibition increased with increases in dose.Conclusions: KW-2449 appears safe and well tolerated at the dose levels evaluated. Transient decreases in peripheral blood and bone marrow blasts have been observed in a few patients justifying continued investigation with this agent. Accrual is ongoing and a different schedule to accommodate the short t1/2 will be explored.

A phase 2 study of the oral farnesyltransferase inhibitor tipifarnib in patients with refractory or relapsed acute myeloid leukemia

AbstractThis phase 2 study evaluated the efficacy and safety of the oral farnesyltransferase inhibitor tipifarnib in adults with refractory or relapsed acute myeloid leukemia (AML). Patients (n = 252) received tipifarnib 600 mg twice a day for 21 days in 28-day cycles. Median age was 62 years; 99 (39%) patients were 65 years or older. Eleven (4%) of 252 patients achieved complete remission (CR) or complete remission with incomplete platelet recovery (CRp; 9 CR and 2 CRp). Nineteen patients (8%), including those who achieved CR/CRp, achieved a reduction in bone marrow blasts to less than 5% blasts. Bone marrow blasts were reduced more than 50% in an additional 8 patients (total = 27; 11%). Median survival was 369 days for patients who achieved CR/CRp. Myelosuppression was the most common adverse event. The most common nonhematologic toxicities were fever, nausea, and hypokalemia. Single-agent treatment with tipifarnib induced durable CR/CRp, which was associated with prolonged survival, in some patients with refractory or relapsed AML. The response rate observed in this heavily pretreated group of patients suggests the requirement to enhance the response rate either by combining tipifarnib with other active agents or determining factors that are predictive of response.

Feasibility Study of Temozolomide Maintenance Therapy in Elderly Patients with AML Following Induction Therapy with High Dose Mitoxantrone and High Dose Ara-C.

With intensive chemotherapy, approximately 50% of elderly patients with AML achieve a complete remission, however essentially all will relapse within 2 years. To date, there is no standard effective postremission therapy for these patients, and most elderly patients tolerate multiple courses of intensive chemotherapy poorly. We previously demonstrated that temozolomide has significant antileukemic activity in patients with relapsed/refractory acute leukemia (Seiter K, et al JCO 2002). In that study 9/20 patients treated had a significant reduction in bone marrow blasts, and 4 patients had a formal complete response. Currently we are evaluating the feasibility and efficacy of temozolomide maintenance therapy administered after one cycle of intensive induction chemotherapy in elderly patients with newly-diagnosed AML. Induction consists of ara-C, 3 gm/m2 daily for 5 days, and mitoxantrone, 80 mg/m2 times one on day 2. Patients achieving complete remission then receive temozolomide 200 mg/m2/d for 7 days every 5 weeks until progression. To date, 30 pts have been treated: median age 69 (range 60–84); M/F: 17/13; prior AHD: 23%; cytogenetics: good: 1, intermediate: 21, poor: 6, insufficient: 2. Response to high-dose ara-C/high-dose mitoxantrone induction: CR: 59%, progression: 24%, aplastic death: 17%. Two pts expired in remission unrelated to temozolomide: pneumonia, 1 pt, c. difficile: 1 pt. Of 17 patients in CR, 15 received temozolomide maintenance. The median number of temozolomide cycles was 2 (range 1–11). 3 pts stopped temozolomide after 1 cycle (1 refused further therapy, 1 had stem cell transplant, 1 died c. diff). Treatment was well tolerated and was administered in the outpatient setting. The median duration of remission is 5.5 months (95% CI: 3.0–9.8 months), and median survival is 7.6 months (95% CI: 5.7–13.3 months) for all patients and 10.2 months (95% CI: 5.6–15.2 months) for patients achieving CR. Temozolomide maintenance is feasible in patients with AML in CR. Further accrual is continuing to assess if remission duration and survival will be improved compared to historical controls.