Mahesh Seetharam

Treatment of higher risk myelodysplastic syndrome patients unresponsive to hypomethylating agents with ON 01910.Na

In a Phase I/II clinical trial, 13 higher risk red blood cell-dependent myelodysplastic syndrome (MDS) patients unresponsive to hypomethylating therapy were treated with the multikinase inhibitor ON 01910.Na. Responses occurred in all morphologic, prognostic risk and cytogenetic subgroups, including four patients with marrow complete responses among eight with stable disease, associated with good drug tolerance. In a subset of patients, a novel nanoscale immunoassay showed substantially decreased AKT2 phosphorylation in CD34+ marrow cells from patients responding to therapy but not those who progressed on therapy. These data demonstrate encouraging efficacy and drug tolerance with ON 01910.Na treatment of higher risk MDS patients.

Mixed Phenotype Acute Leukemia

OBJECTIVES The 2008 World Health Organization (WHO) classification system grouped bilineal and biphenotypic acute leukemias together under a new heading of mixed phenotype acute leukemia (MPAL). The lineage-specific marker criteria have also changed for a diagnosis of MPAL. The goal of this study was to characterize clinical significance of this new group. METHODS Sixty-one patients diagnosed with MPAL using either European Group for the Immunological Classification of Leukaemias (EGIL) criteria or 2008 WHO criteria were included in this study. RESULTS Sixteen patients (26%) diagnosed with acute biphenotypic leukemia using EGIL criteria did not fulfill 2008 WHO criteria for MPAL. Cytogenetic data were available for 32 patients, and the most common abnormality was t(9;22) (five of 32 cases). Clinical outcome data suggested that younger patients with MPAL (≤21 years) had better overall survival (OS) in both the EGIL and WHO groups (EGIL, P = .0403; WHO, P = .0601). Compared with 177 patients with acute myeloid leukemia (AML), MPAL patients had better OS (P = .0003) and progression-free survival (P = .0001). However, no difference in OS between MPAL and 387 patients with acute lymphoblastic leukemia was present (P = .599). CONCLUSIONS As defined by the 2008 WHO classification, fewer patients are now classified as having MPAL than with the EGIL criteria. In this study, patients with MPAL have a better clinical outcome compared with patients with AML.

DNA methylation analysis of ALOX12 and GSTM1 in acute myeloid leukaemia identifies prognostically significant groups

To determine the role of DNA methylation in the progression of acute myeloid leukaemia (AML), we analysed the methylation status of ALOX12, GSTM1, HS3ST2 and FZD9 in 127 AML patients. Aberrant methylation of ALOX12 was associated with the subcategory AML with myelodysplasia‐related changes (P = 0·0439) and specifically with megakaryocytic dysplasia (P = 0·0003). An association between HS3ST2 and AML patients with favourable cytogenetic risk was identified (P = 0·0469). In univariate and multivariate analysis, methylation of GSTM1 was associated with worse overall survival (OS) and disease‐free survival (DFS), with hazard ratios of 2·57 and 1·86, respectively. Furthermore, the significance of methylation of GSTM1 in predicting poor prognosis was maintained within the subcategories of AML not otherwise specified (NOS), AML with intermediate cytogenetic risk and normal karyotype AML. Finally, patients with both GSTM1 and ALOX12 methylated, demonstrated worse outcomes when all AML patients were assessed (OS; P = 0·000411) as well as within AML NOS (DFS; P = 0·0023), AML with intermediate cytogenetic risk (OS; P = 0·0104) and normal karyotype AML (OS; P = 0·00636). This study implicates methylation of specific genes in the classification and prognostication of AML and suggests that the morphological feature of multilineage dysplasia may be a surrogate marker of gene methylation in at least a subset of AML cases.

Acute Myeloid Leukemia With Monosomal Karyotype Morphologic, Immunophenotypic, and Molecular Findings

OBJECTIVES Acute myeloid leukemia (AML) with monosomal karyotype (MK) recently has been reported to be associated with worse outcome than the traditional complex karyotype. METHODS In this retrospective study of 111 patients with AML, we identified 14 patients with MK (13% of all patients with AML) using the definition proposed by Breems et al. RESULTS Five (36%) of these 14 patients had a loss of a single chromosome in the presence of other structural abnormalities, and nine (64%) had a loss of two or more autosomal chromosomes. Patients with AML-MK presented at an older age, with lower bone marrow blasts, and their blasts less frequently expressed CD34. Most patients with AML-MK had morphologic multilineage dysplasia and were predominantly subclassified as having AML with myelodysplasia-related changes (AML-MRC). Molecular analysis showed a significant absence of NPM1 and FLT3 in patients with AML-MK. CONCLUSIONS Outcome data showed that patients with AML-MK had significantly worse overall survival, disease-free survival, and complete response compared with the rest of the patients with AML as well as within the AML-MRC group.

Acute Myeloid Leukemia With Monosomal Karyotype

OBJECTIVES Acute myeloid leukemia (AML) with monosomal karyotype (MK) recently has been reported to be associated with worse outcome than the traditional complex karyotype. METHODS In this retrospective study of 111 patients with AML, we identified 14 patients with MK (13% of all patients with AML) using the definition proposed by Breems et al. RESULTS Five (36%) of these 14 patients had a loss of a single chromosome in the presence of other structural abnormalities, and nine (64%) had a loss of two or more autosomal chromosomes. Patients with AML-MK presented at an older age, with lower bone marrow blasts, and their blasts less frequently expressed CD34. Most patients with AML-MK had morphologic multilineage dysplasia and were predominantly subclassified as having AML with myelodysplasia-related changes (AML-MRC). Molecular analysis showed a significant absence of NPM1 and FLT3 in patients with AML-MK. CONCLUSIONS Outcome data showed that patients with AML-MK had significantly worse overall survival, disease-free survival, and complete response compared with the rest of the patients with AML as well as within the AML-MRC group.

Acute Myeloid Leukemia With Monosomal KaryotypeMorphologic, Immunophenotypic, and Molecular Findings

OBJECTIVES Acute myeloid leukemia (AML) with monosomal karyotype (MK) recently has been reported to be associated with worse outcome than the traditional complex karyotype. METHODS In this retrospective study of 111 patients with AML, we identified 14 patients with MK (13% of all patients with AML) using the definition proposed by Breems et al. RESULTS Five (36%) of these 14 patients had a loss of a single chromosome in the presence of other structural abnormalities, and nine (64%) had a loss of two or more autosomal chromosomes. Patients with AML-MK presented at an older age, with lower bone marrow blasts, and their blasts less frequently expressed CD34. Most patients with AML-MK had morphologic multilineage dysplasia and were predominantly subclassified as having AML with myelodysplasia-related changes (AML-MRC). Molecular analysis showed a significant absence of NPM1 and FLT3 in patients with AML-MK. CONCLUSIONS Outcome data showed that patients with AML-MK had significantly worse overall survival, disease-free survival, and complete response compared with the rest of the patients with AML as well as within the AML-MRC group.

Abstract 4136: Single-cell network profiling (SCNP) to evaluate the proteomic profiles associated with ON 01910.Na treatment of MDS patients (Pts)

Proceedings: AACR 102nd Annual Meeting 2011‐‐ Apr 2‐6, 2011; Orlando, FL Background: Single Cell Network Profiling (SCNP) is used to measure simultaneously the effects of multiple modulators (including drugs) on intracellular signaling cascades at the single cell level. ON 01910.Na has been reported to inhibit polo-like kinase 1, PI3-kinase and Akt pathways. In an ongoing phase II study in Int-1, 2 or high risk MDS pts refractory to hypomethylating agents, biomarker assays are being performed to assess CD34+ cell functional signaling profiles associated with biological activity of ON 01910.Na. Objectives: The objectives were to simultaneously compare the functional effects of a panel of modulators on different signaling pathways (such as the PI3K and the Janus Kinases (Jak) signal transducers and activators of transcription (Stat) pathway) to identify specific proteomic profiles associated with the biological activity of and response to ON 01910.Na in MDS pts. Methods: MDS pt bone marrow samples were collected at baseline and after treatment cycles 1, 3, 5, & 7. Bone marrow mononuclear cells (BMMCs) were isolated and cryopreserved for longitudinal analysis. Activation of signaling pathways was measured with fluorochrome-conjugated antibodies that recognize p-Erk1/1 (T202/Y204), p-Akt (S473), p-S6 (S235/236), p-Stat1 (Y701), p-Stat3 (S727), and p-Stat5 (Y694). BMMCs were modulated with FMS-like tyrosine kinase 3 ligand (FLTL3), stem cell factor (SCF), granulocyte colony stimulating factor (G-CSF), or granulocyte-monocyte colony stimulating factor (GM-CSF) for 15 minutes. Cells were processed for SCNP by fixation, permeabilization, and incubation with fluorochrome-conjugated antibodies. Results: SCNP analyses in Pt ON103 (progressed to AML after completion of trial) showed that frequency of CD34+ cells increased during the course of the clinical trial; when modulated with either FLT3L or SCF, compared to baseline findings, CD34+ cells exhibited increased p-S6 and p-Akt responsiveness with treatment; and interestingly, CD34+ cell responsiveness to G-CSF decreased (p-Stat1 and p-Stat5) while no signaling was observed in response to GM-CSF. In contrast, SCNP analyses in Pt ON104 (stable disease), showed that frequency of CD34+ cells was maintained throughout treatment; when modulated with either FLT3L or SCF, CD34+ cells exhibited decreased p-S6, p-Akt, p-Erk (slight); and while CD34+ cell responsiveness to G-CSF decreased (p-Stat1 and p-Stat5), a robust p-Stat5 response was induced by GM-CSF which increased during the course of the clinical trial.Conclusions: The data here suggest that measurement of intracellular signaling responses using SCNP is feasible using BMMCs from pts with MDS. Interrogating key signaling pathways thought to be involved in the action of ON 01910.Na provides a tool for developing functional proteomic signaling profiles associated with biological activity of this novel cell cycle active agent. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4136. doi:10.1158/1538-7445.AM2011-4136