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PROTEOME CHANGES IN ACUTE MYELOID LEUKEMIA PATIENTS BEFORE AND AFTER INDUCTION TREATMENT
Author(s):
Luisa Fernanda Restrepo Rodríguez
,
Luisa Fernanda Restrepo Rodríguez
Affiliations:
Biomedical Engineering,Instituto Tecnologico Metropolitano,Medellín,Colombia
Sarah Rothlisberger
Sarah Rothlisberger
Affiliations:
Biomedical Engineering,Instituto Tecnologico Metropolitano,Medellín,Colombia
(Abstract release date: 05/18/17) EHA Library. L. 05/18/17; 182383; PB1669
Luisa
Luisa
Contributions
PDF Abstract
Abstract

Abstract: PB1669

Type: Publication Only

Background

Acute myeloid leukemia (AML) is a malignant disorder of hematopoietic stem and progenitor cells (HSPCs), characterized by the accumulation of immature blasts in the bone marrow and peripheral blood (PB) of affected patients. Standard induction therapy, based on cytarabine and an anthracycline, leads to complete remission in approximately 50% to 75% of patients, depending on prognostic factors, such as age or the presence of certain gene or chromosomal changes. In spite of favorable primary response rates, only approximately 20% to 30% of the patients enjoy long-term disease survival.

Aims

Our aim was to compare the protein expression profile of peripheral blood mononuclear cells (PBMCs) of AML patients at time of diagnosis and after induction therapy.

Methods
PB samples were taken from seven AML patients in Medellin-Colombia at time of diagnosis and after concluding the induction therapy. Informed consent was obtained prior to sample collection. PBMCs were isolated from the 14 blood samples using a Histopaque-1077 solution. Cells were resuspended in lysis buffer (0.5 % Triton x-100, 50 mM Tris-HCL pH 8.0, 150 mM NaCL, 1 mM EDTA, protease inhibitor) and proteins precipitated with trichloroacetic acid. Proteins were separated by 2D SDS-PAGE (pI 3–10 NL), and stained with SYPRO®Ruby. The proteomes were compared using PDQuest™ Advanced 8.0.1 Software. Protein spots of interest were those with a fold change of +/- 1.5 and p < 0.05.

Results

Image analysis revealed an average of 464 protein spots in PB samples taken at time of diagnosis, and an average of 346 spots in PB taken after induction therapy, reflecting changes in protein expression due to treatment. Comparing the proteomes, we found 11 spots that differed significantly (fold change of +/- 1.5 and p < 0.05). Of these, seven proteins were up-regulated and four were down-regulated at time of diagnosis (before treatment) compared to after induction treatment. Nine of these spots correspond to low molecular weight proteins (<40 kDa) and 2 spots have a molecular weight between 40-60 kDa.
Based on the molecular weight and isoelectric point information of these spots we were able to search for proteins reportedly involved in leukemia, in order to propose possible identities (see Table). In terms of biological process, four proteins (eIF5B, HSP27, 14-3-3 protein zeta/delta, and GST-P) are involved in the regulation of apoptosis. The F-actin-capping protein subunit beta could also be of interest, as reorganization of F-actin reflects unique characteristics of the differentiation process in promyelocytic leukemia cells. RuvB-like 2 is a positive regulator of histone acetylation and DNA repair. GRBP2 is a protein involved in the MAPK cascade and regulation of PI3K signaling, pathways regulating diverse cellular functions altered in leukemogenesis such as proliferation, differentiation, and apoptosis. Alpha-enolase is a key glycolytic enzyme; however, it has been shown to be a multifunctional protein involved in cancer. It promotes cell proliferation by also regulating the MAPK and PI3K pathways. Transaldolase is part of the pentose-phosphate pathway. Annexin II acts in angiogenesis and has multifaceted role in human health and disease.

Conclusion
The protein expression profile of AML patients changes after induction treatment. We found 11 spots that differed significantly, and propose possible identities for these. Further analyses are pending in order to experimentally establish the identities and correlate with response to treatment.

Session topic: 3. Acute myeloid leukemia - Biology

Keyword(s): proteomics, Induction chemotherapy, Acute Myeloid Leukemia

Abstract: PB1669

Type: Publication Only

Background

Acute myeloid leukemia (AML) is a malignant disorder of hematopoietic stem and progenitor cells (HSPCs), characterized by the accumulation of immature blasts in the bone marrow and peripheral blood (PB) of affected patients. Standard induction therapy, based on cytarabine and an anthracycline, leads to complete remission in approximately 50% to 75% of patients, depending on prognostic factors, such as age or the presence of certain gene or chromosomal changes. In spite of favorable primary response rates, only approximately 20% to 30% of the patients enjoy long-term disease survival.

Aims

Our aim was to compare the protein expression profile of peripheral blood mononuclear cells (PBMCs) of AML patients at time of diagnosis and after induction therapy.

Methods
PB samples were taken from seven AML patients in Medellin-Colombia at time of diagnosis and after concluding the induction therapy. Informed consent was obtained prior to sample collection. PBMCs were isolated from the 14 blood samples using a Histopaque-1077 solution. Cells were resuspended in lysis buffer (0.5 % Triton x-100, 50 mM Tris-HCL pH 8.0, 150 mM NaCL, 1 mM EDTA, protease inhibitor) and proteins precipitated with trichloroacetic acid. Proteins were separated by 2D SDS-PAGE (pI 3–10 NL), and stained with SYPRO®Ruby. The proteomes were compared using PDQuest™ Advanced 8.0.1 Software. Protein spots of interest were those with a fold change of +/- 1.5 and p < 0.05.

Results

Image analysis revealed an average of 464 protein spots in PB samples taken at time of diagnosis, and an average of 346 spots in PB taken after induction therapy, reflecting changes in protein expression due to treatment. Comparing the proteomes, we found 11 spots that differed significantly (fold change of +/- 1.5 and p < 0.05). Of these, seven proteins were up-regulated and four were down-regulated at time of diagnosis (before treatment) compared to after induction treatment. Nine of these spots correspond to low molecular weight proteins (<40 kDa) and 2 spots have a molecular weight between 40-60 kDa.
Based on the molecular weight and isoelectric point information of these spots we were able to search for proteins reportedly involved in leukemia, in order to propose possible identities (see Table). In terms of biological process, four proteins (eIF5B, HSP27, 14-3-3 protein zeta/delta, and GST-P) are involved in the regulation of apoptosis. The F-actin-capping protein subunit beta could also be of interest, as reorganization of F-actin reflects unique characteristics of the differentiation process in promyelocytic leukemia cells. RuvB-like 2 is a positive regulator of histone acetylation and DNA repair. GRBP2 is a protein involved in the MAPK cascade and regulation of PI3K signaling, pathways regulating diverse cellular functions altered in leukemogenesis such as proliferation, differentiation, and apoptosis. Alpha-enolase is a key glycolytic enzyme; however, it has been shown to be a multifunctional protein involved in cancer. It promotes cell proliferation by also regulating the MAPK and PI3K pathways. Transaldolase is part of the pentose-phosphate pathway. Annexin II acts in angiogenesis and has multifaceted role in human health and disease.

Conclusion
The protein expression profile of AML patients changes after induction treatment. We found 11 spots that differed significantly, and propose possible identities for these. Further analyses are pending in order to experimentally establish the identities and correlate with response to treatment.

Session topic: 3. Acute myeloid leukemia - Biology

Keyword(s): proteomics, Induction chemotherapy, Acute Myeloid Leukemia

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