SINGLE CELL CYTOKINE ANALYSIS OF CIRCULATING MALIGNANT HEMATOPOIETIC CELLS


Year of Award:
2015
Award Type:
R33
Project Number:
CA196411
RFA Number:
RFA-CA-14-004
Technology Track:
Molecular & Cellular Analysis Technologies
PI/Project Leader:
FAN, RONG
Other PI or Project Leader:
LEVINE, ROSS L
Institution:
YALE UNIVERSITY
Myeloproliferative malignancies are clonal blood diseases in which a dysregulated excessive or dysplastic production of blood cells driven by the gain-of-function mutations in the bone marrow hematopoietic stem/progenitor cells. While the common mutations of these diseases have been identified and the mutation profile is relatively uniform across patients, the clinical outcomes vary substantially, indicating it is insufficient topredict the outcome by detecting the genetic differences. It is known that hematopoietic stem/progenitor cells are also found in circulation. These cells are shed from bone marrow (BM) and thus may serve as the 'liquid biopsy' of BM tissue. However, circulating hematopoietic cells in blood are not only rare but also as heterogeneous as the hematopoietic cell hierarchy in bone marrow, requiring single-cell resolution analysis. Recent studies show that both mutant and non-mutant bone marrow cells are producing a wide range of cytokines and the abnormal cytokine secreting cells are mechanistically linked to systemic cytokine elevation and pathogenesis. Therefore, it is highly promising to detect cellular mechanism of myeloproliferative disease in patients and potentially for therapeutic stratification by measuring the cytokine function profile f individual circulating hematopoietic cells. In the past years, we have devised an innovative microchip platform for highly multiplex detection of cytokine secretion at the single-cell level. This technology represents the highest multiplexing recorded to date in terms of single-cell protein secretion assay and requires very small quantities of cell samples, which uniquely address the aforementioned challenges. This application aims to conduct validation and advanced development of this emerging single-cell technology for analyzing cytokine functions of individual circulating hematopoietic cells in patients with the potential to identify malignant cells and for early diagnosis and therapeutic stratification. Specifically, we will validate this technology for detecting abnormal cytokine-secreting hematopoietic cells in blood from patients and conduct further development to create an integrated microsystem for frequent monitoring of progression or therapeutic response. While this project is designed to address a specific cancer research need in myeloproliferative neoplasm, ultimately it may offer a versatile technology platform for minimally invasive diagnosis and stratification of a range of hematologic malignancies and other inflammatory malignancies.