Identification of biomarker(s) of monocyte precursors in human bone marrow

Clinical Center

Investigators

Abstract

Monocytes originate in the bone marrow, and travel through the blood to tissues where they become macrophages or dendritic cells. An increased number of monocytes in the blood (monocytosis) occurs in response to chronic infections, autoimmune disorders, and in certain acute and chronic leukemias. Among the leukemias, various types have been recognized, including acute myelomonocytic, acute monoblastic, acute monocytic, and chronic myelomonocytic leukemia. Other terms, such as acute myeloid leukemia with monocytic differentiation, are also used, adding to the already confusing nomenclature. At present, the main techniques utilized in the identification of monocytic lineage are cytomorphology, cytochemistry and flow cytometry, but the exact delineation of the various types of acute and chronic leukemias and their differentiation from benign monocytosis has remained a serious challenge. This is largely because the identification of blastic, dysplastic or reactive monocytes is far from being accurate or precise. Genetic mutations are often observed in monocytic leukemias but again, they are not specific since they are observed in other neoplastic processes. This uncertainty translates into serious diagnostic or even therapeutic decisions, which are often based on determining the nature of a monocytic proliferation, or an accurate blast count. Most non-monocytic acute leukemias (myeloid or lymphoid) recapitulate the phenotype of early normal precursors and thus express highly informative antigens like CD34, CD117, or TdT. These leukemic-associated antigens are not specific for the neoplastic cells. They are simply antigens that are expressed in early stages of normal cell development in the marrow. It is very likely that markers expressed in early stages of monocyte development could as well serve as proper identifiers for blasts in monocytic leukemias. However, in contrast to granulocytic or lymphoid leukemias, there are no single markers that allow the unequivocal recognition of leukemic monocytes. This is largely because human monocyte precursors have not been adequately characterized. Based on preliminary observations, we propose to identify and purify putative monocytic precursor from normal human bone marrow using flow cytometry/sorting and: 1. Demonstrate that they are indeed monocytic precursors and, 2. Compare their transcriptome with that of all other cells in the marrow using the RNAseq technique in order to identify specific RNA markers (and eventually products) that would allow their identification. Appropriate recognition of monocyte precursors (and likely monocytic blasts) should enable us to establish better diagnosis and classification of monocytic leukemias and help designing proper therapeutic stratification. We have already completed 2 sorts of various cell types from marrows from normal volunteers. After RNA extraction and RNAseq analysis, we were able to compare the expression of multiple genes between the presumptive monocyte precursors and other cells types, including mature monocytes, CD34+ cells, granulocytes, nucleated red blood cells and lymphocytes. Although we found a few good gene candidates with very high differential expression, the results between the 2 normal marrows were not reproducible. We will perform additional experiments to determine which gene targets, if any, may be informative and useful for the original application planned.