A Multiplexed Screening Assay to Evaluate Chemotherapy-Induced Myelosuppression Using Healthy Peripheral Blood and Bone Marrow

Abstract

Myelosuppression is a major side effect of chemotherapy in cancer patients and can result in infections, bleeding complications, and increased risk of morbidity and mortality, as well as limit the drug dose and frequency of administration. Chemotherapy-induced myelosuppression is caused by the disruption of normal hematopoiesis. Thus, prior understanding of the adverse effects of chemotherapies on hematopoietic cells is essential to minimize the side effects of cancer treatment. Traditional methods such as colony-forming assays for studying chemotherapy-induced myelosuppression are time-consuming and labor intensive. High-throughput flow cytometry technologies and methods to detect rare hematopoietic cell populations are critical in advancing our understanding of how different blood cell types in complex biological samples respond to chemotherapeutic drugs. In the present study, hematopoietic progenitor cells were induced to differentiate into megakaryocytes and myeloid lineage cells. The expanded cells were then used in a multiplexed assay to monitor the dose-response effects of multiple chemotherapies on different stages of megakaryocyte differentiation and myeloid cell populations in a 96-well plate format. The assay offers an alternative method to evaluate the myelosuppressive potential of novel chemotherapeutic drugs compared to traditional lower throughput and labor-intensive assays.

Guidelines for the use of flow cytometry and cell sorting in immunological studies (second edition)

“…HTFC can be performed using living cells, fixed cells, suspension cells, or even adherent cells and bead based assays. Examples for applications are Ab development , toxicology and apoptosis studies , analysis of protein phosphorylation , identification of stem cell expanding compounds , identification of immune enhancing or immune inhibiting compounds , monitoring of chemotherapy side effects , and characterization and cytotoxicity monitoring of engineered T cells . In addition, secreted molecules such as chemokines or cytokines as well as gene expression can be quantified using barcoded bead based multiplex assays .…”

Chemotherapy-induced anemia: etiology, pathophysiology, and implications for contemporary practice

“…The development of screening assays utilizing hematopoietic stem and progenitor cells (HPCs CD34+) from peripheral blood and bone marrow help to evaluate the effects of chemotherapy-induced myelosuppression and may identify particularly cytotoxic pharmacologic therapies. 106 In addition to novel screening mechanisms, various hormones and pharmacologic agents influence the prevention and management of CIA. Administration of FLT3 tyrosine kinase inhibitors following chemotherapy may selectively induce myelosuppression of neoplastic cells, but not bone marrow progenitor cells.…”

High-Throughput Flow Cytometry in Drug Discovery

“…The authors identify several clinically approved tyrosine kinase inhibitors targeting T-ALL in a hypoxic environment similar to that in bone marrow. Javarappa et al 28 describe a multiplexed moderate-throughput flow cytometry assay for evaluating myelosuppressive effects induced by chemotherapeutic drugs. A nine-color antibody panel is used to evaluate the effects of a panel of 15 chemotherapeutic drugs on the differentiation and maturation of megakaryocytes and myeloid cells differentiated from CD34-positive hematopoietic progenitor cells, purified from peripheral blood and bone marrow samples from healthy donors.…”