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2 results on '"Oksana Montecchini"'

1. A Novel ELISA-Based Peptide Biosensor Assay for Screening ABL1 Activity in vitro: A Challenge for Precision Therapy in BCR-ABL1 and BCR-ABL1 Like Leukemias

Author

Oksana Montecchini, Stefania Braidotti, Raffaella Franca, Giulia Zudeh, Christian Boni, Claudio Sorio, Eleonora Toffoletti, Marco Rabusin, Alberto Tommasini, Giuliana Decorti, and Gabriele Stocco

Subjects
ABL1-class BCR-ABL1 like acute lymphoblastic leukemia, ABL1 peptide biosensor, in vitro ELISA assay, ABL1 tyrosin kinase inhibitors, precision therapy, Therapeutics. Pharmacology, RM1-950
Abstract

The pathogenic role of the overactivated ABL1 tyrosine kinase (TK) pathway is well recognized in some forms of BCR-ABL1 like acute lymphoblastic leukemia (ALL); TK inhibitors represent a useful therapeutic choice in these patients who respond poorly to conventional chemotherapy. Here we report a novel peptide biosensor (PABL)-ELISA assay to investigate ABL1 activity in four immortalized leukemic cell lines with different genetic background. The PABL sequence comprises an ABL1 tyrosine (Y) phosphorylation site and a targeting sequence that increases the specificity for ABL1; additional peptides (Y-site-mutated (PABL-F) and fully-phosphorylated (PPHOSPHO-ABL) biosensors) were included in the assay. After incubation with whole cell lysates, average PABL phosphorylation was significantly increased (basal vs. PABL phosphorylation: 6.84 ± 1.46% vs. 32.44 ± 3.25%, p-value < 0.0001, two-way ANOVA, Bonferroni post-test, percentages relative to PPHOSPHO-ABL in each cell line). Cell lines expressing ABL1-chimeric proteins (K562, ALL-SIL) presented the higher TK activity on PABL; a lower signal was instead observed for NALM6 and REH (p < 0.001 and p < 0.05 vs. K562, respectively). Phosphorylation was ABL1-mediated, as demonstrated by the specific inhibition of imatinib (p < 0.001 for K562, NALM6, ALL-SIL and p < 0.01 for REH) in contrast to ruxolitinib (JAK2-inhibitor), and occurred on the ABL1 Y-site, as demonstrated by PABL-F whose phosphorylation was comparable to basal levels. In order to validate this novel PABL-ELISA assay on leukemic cells isolated from patient’s bone marrow aspirates, preliminary analysis on blasts derived from an adult affected by chronic myeloid leukaemia (BCR-ABL1 positive) and a child affected by ALL (BCR-ABL1 negative) were performed. Phosphorylation of PABL was specifically inhibited after the incubation of BCR-ABL1 positive cell lysates with imatinib, but not with ruxolitinib. While requiring further optimization and validation in leukemic blasts to be of clinical interest, the PABL-based ELISA assay provides a novel in vitro tool for screening both the aberrant ABL1 activity in BCR-ABL1 like ALL leukemic cells and their potential response to TK inhibitors.

Published
2021
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2. Targeting kinase-activating genetic lesions to improve therapy of pediatric acute lymphoblastic leukemia

Author

Alice Poropat, Claudio Sorio, Debora Curci, Marco Rabusin, Giuliana Decorti, Natasa Karas Kuzelicki, Raffaella Franca, Dino Paladin, Oksana Montecchini, Eleonora Toffoletti, Gabriele Stocco, Franca, Raffaella, Kuzelicki, Natasa K., Sorio, Claudio, Toffoletti, Eleonora, Montecchini, Oksana, Poropat, Alice, Rabusin, Marco, Curci, Debora, Paladin, Dino, Stocco, Gabriele, and Decorti, Giuliana

Subjects
0301 basic medicine, Pathology, medicine.medical_treatment, bcr-abl, Fusion Proteins, bcr-abl, Chromosomal translocation, Biochemistry, Targeted therapy, Fusion gene, hemic and lymphatic diseases, Protein-Tyrosine Kinase, Drug Discovery, Medicine, Sirolimu, ALL, Chemotherapy, Hematologic malignancies, Kinase-activating genetic lesions, Lymphoblastic leukemia, Molecular Medicine, Pharmacology, Child, TOR Serine-Threonine Kinase, Lymphoblast, TOR Serine-Threonine Kinases, Combination chemotherapy, Protein-Tyrosine Kinases, Precursor Cell Lymphoblastic Leukemia-Lymphoma, targeted therapy, kinase-activating genetic lesions, pediatric acute lymphoblatic leukemia, Hematologic malignancie, Tyrosine kinase, medicine.drug, Human, medicine.medical_specialty, Protein Kinase Inhibitor, 03 medical and health sciences, Nitriles, Humans, Protein Kinase Inhibitors, Kinase-activating genetic lesion, Janus Kinases, Sirolimus, 030102 biochemistry & molecular biology, business.industry, Organic Chemistry, Fusion Proteins, Imatinib, Pyrazoles, Pyrimidines, Pyrazole, Cancer research, Janus Kinase, Janus kinase, business
Abstract

Acute lymphoblastic leukemia (ALL) is the most common hematologic malignancy in children, characterized by an abnormal proliferation of immature lymphoid cells. Thanks to risk-adapted combination chemotherapy treatments currently used, survival at 5 years has reached 90%. ALL is a heterogeneous disease from a genetic point of view: patients’ lymphoblasts may harbor in fact several chromosomal alterations, some of which have prognostic and therapeutic value. Of particular importance is the translocation t(9;22)(q34;q11.2) that leads to the formation of the BCR-ABL1 fusion gene, encoding a constitutively active chimeric tyrosine kinase (TK): BCR-ABL1 that is present in ~3% of pediatric ALL patients with B-immunophenotype and is associated with a poor outcome. This type of ALL is potentially treatable with specific TK inhibitors, such as imatinib. Recent studies have demonstrated the existence of a subset of BCR-ABL1 like leukemias (~10-15% of Bimmunophenotype ALL), whose blast cells have a gene expression profile similar to that of BCR-ABL1 despite the absence of t(9;22)(q34;q11.2). The precise pathogenesis of BCR-ABL1 like ALL is still to be defined, but they are mainly characterized by the activation of constitutive signal transduction pathways due to chimeric TKs different from BCR-ABL1. BCR-ABL1 like ALL patients represent a group with unfavorable outcome and are not identified by current risk criteria. In this review, we will discuss the design of targeted therapy for patients with BCR-ABL1 like ALL, which could consider TK inhibitors, and discuss innovative approaches suitable to identify the presence of patient’s specific chimeric TK fusion genes, such as targeted locus amplification or proteomic biosensors.

Published
2018

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