ABSTRACT
Chronic Myeloid Leukemia is a clonal disorder characterized by the presence of the Ph-chromosome and the BCR-ABL tyrosine-kinase (TK). Target-therapy with Imatinib has greatly improved its outcome. Deeper and faster responses are reported with the second-generation TKI Nilotinib. Sustained responses may enable TKI discontinuation. However, even in a complete molecular response, some patients experience disease recurrence possibly due to persistence of quiescent leukemic CD34+/lin-Ph+ stem cells (LSCs). Degree and mechanisms of LSCs clearance during TKI treatment are not clearly established. The PhilosoPhi34 study was designed to verify the in-vivo activity and timecourse of first-line Nilotinib therapy on BM CD34+/lin-Ph+ cells clearance. Eighty-seven CP-CML patients were enrolled. BM cells were collected and tested for Ph+ residual cells, at diagnosis, 3, 6 and 12 months of treatment. FISH analysis of unstimulated CD34+/lin- cells in CCyR patients were positive in 8/65 (12.3%), 5/71 (7%), 0/69 (0%) evaluable tests, respectively. Per-Protocol analysis response rates were as follows: CCyR 95% at 12 months, MR4.5 31% and 46% at 12 and 36 months, respectively. An exploratory Gene Expression Profiling (GEP) study of CD34+/lin- cells was performed on 30 patients at diagnosis and after, on 79 patients at diagnosis vs 12 months of nilotinib treatment vs 10 healthy subjects. Data demonstrated some genes significantly different expressed: NFKBIA, many cell cycle genes, ABC transporters, JAK-STAT signaling pathway (JAK2). In addition, a correlation between different expression of some genes (JAK2, OLFM4, ICAM1, NFKBIA) among patients at diagnosis and their achievement of an early and deeper MR was observed.
Subject(s)
Antineoplastic Agents/therapeutic use , Gene Expression Regulation, Leukemic/drug effects , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/drug therapy , Neoplastic Stem Cells/drug effects , Protein Kinase Inhibitors/therapeutic use , Pyrimidines/therapeutic use , ATP-Binding Cassette Transporters/genetics , ATP-Binding Cassette Transporters/metabolism , Adolescent , Adult , Aged , Aged, 80 and over , Biomarkers, Pharmacological , Bone Marrow/drug effects , Bone Marrow/metabolism , Bone Marrow/pathology , Case-Control Studies , Cell Cycle Proteins/genetics , Cell Cycle Proteins/metabolism , Female , Gene Expression Profiling , Granulocyte Colony-Stimulating Factor/genetics , Granulocyte Colony-Stimulating Factor/metabolism , Humans , Intercellular Adhesion Molecule-1/genetics , Intercellular Adhesion Molecule-1/metabolism , Janus Kinase 2/genetics , Janus Kinase 2/metabolism , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/diagnosis , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/genetics , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/pathology , Male , Middle Aged , NF-KappaB Inhibitor alpha/genetics , NF-KappaB Inhibitor alpha/metabolism , Neoplastic Stem Cells/metabolism , Neoplastic Stem Cells/pathology , Philadelphia Chromosome , Prospective Studies , RecurrenceABSTRACT
Chronic myeloid leukemia (CML) is characterized by the constitutive tyrosine kinase activity of the oncoprotein BCR-ABL1 in myeloid progenitor cells that activates multiple signal transduction pathways leading to the leukemic phenotype. The tyrosine-kinase inhibitor (TKI) nilotinib inhibits the tyrosine kinase activity of BCR-ABL1 in CML patients. Despite the success of nilotinib treatment in patients with chronic-phase (CP) CML, a population of Philadelphia-positive (Ph+) quiescent stem cells escapes the drug activity and can lead to drug resistance. The molecular mechanism by which these quiescent cells remain insensitive is poorly understood. The aim of this study was to compare the gene expression profiling (GEP) of bone marrow (BM) CD34+/lin- cells from CP-CML patients at diagnosis and after 12 months of nilotinib treatment by microarray, in order to identify gene expression changes and the dysregulation of pathways due to nilotinib action. We selected BM CD34+/lin- cells from 78 CP-CML patients at diagnosis and after 12 months of first-line nilotinib therapy and microarray analysis was performed. GEP bioinformatic analyses identified 2,959 differently expressed probes and functional clustering determined some significantly enriched pathways between diagnosis and 12 months of nilotinib treatment. Among these pathways, we observed the under expression of 26 genes encoding proteins belonging to the cell cycle after 12 months of nilotinib treatment which led to the up-regulation of chromosome replication, cell proliferation, DNA replication, and DNA damage checkpoint at diagnosis. We demonstrated the under expression of the ATP-binding cassette (ABC) transporters ABCC4, ABCC5, and ABCD3 encoding proteins which pumped drugs out of the cells after 12 months of nilotinib. Moreover, GEP data demonstrated the deregulation of genes involved in the JAK-STAT signaling pathway. The down-regulation of JAK2, IL7, STAM, PIK3CA, PTPN11, RAF1, and SOS1 key genes after 12 months of nilotinib could demonstrate the up-regulation of cell cycle, proliferation and differentiation via MAPK and PI3K-AKT signaling pathways at diagnosis.
Subject(s)
ATP-Binding Cassette Transporters/blood , Cell Cycle/drug effects , Gene Expression Regulation, Leukemic/drug effects , Janus Kinases/blood , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/blood , Neoplasm Proteins/blood , Pyrimidines/administration & dosage , STAT Transcription Factors/blood , Signal Transduction/drug effects , Female , Humans , Male , Middle Aged , Time FactorsSubject(s)
Bone Marrow Cells/drug effects , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/pathology , Pyrimidines/pharmacology , Adolescent , Adult , Aged , Aged, 80 and over , Antigens, CD34/analysis , Bone Marrow Cells/pathology , Cell Count , Female , Humans , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/drug therapy , Male , Middle Aged , Neoplastic Stem Cells/pathology , Prospective Studies , Protein Kinase Inhibitors/therapeutic use , Young AdultABSTRACT
BACKGROUND: Chronic myeloid leukemia (CML) is a clonal myeloproliferative disorder with heterogeneous biological and clinical features. The biomolecular mechanisms of CML response to tyrosine-kinase inhibitors are not fully defined. OBJECTIVE: We undertook a gene expression profiling (GEP) study of selected bone marrow (BM) CD34+/lin- cells of chronic-phase CML patients at diagnosis and after 12 months of TKI nilotinib to investigate molecular signatures characterizing both conditions.
