Deeper Insights into Hematological Oncology Disorders via Single-Cell Phospho-Signaling Analysis

Abstract

An understanding of hematological cancer cell signaling processes poses one of the most complex and intractable problems in modern biomedical inquiry. While we understand some of the fundamental players that contribute to oncogenic processes, significant effort is focused upon determining how these individual players relay information to each other to create the composite functions of a cancer cell. Efforts designed to understand these processes at the single cell level will undoubtedly allow for understanding of the heterogeneity of hematological tumors as well as, simultaneously, the function of the 'responding' immune system. I will relate some of the insights our laboratory has developed over the last several years applying single-cell phospho-flow cytometry to the study of signaling in primary patient material and murine models. While it is clear that this analysis now allows us to accomplish phosphosignaling biochemistry at the single cell level with primary cell material, we are only beginning to develop some of the bioinformatics tools to appropriately display the vast amount of information collected by such approaches. These approaches, however, have already allowed us to develop approaches that prognosticate patient outcomes based on signaling status, prior to any treatment, as well as subgroup patient subtypes according to signaling states. The modest efforts to date presage a time where it should be possible to provide far more tailored therapies specific to the varied diseases represented by the hematological malignancies.

The Stat3/5 Signaling Biosignature in Hematopoietic Stem/Progenitor Cells Predicts Response and Outcome in Myelodysplastic Syndrome Patients Treated with Azacitidine

“…In both adult and pediatric AML, functional phenotyping of Stat3/5 signaling networks by phospho-protein flow cytometry provides important prognostic information and pathobiologic insights (12,27,37). Yet, despite the reciprocal interactions of DNA methylation with Stat3/5 signaling (19,29,38), the high rate of mutations (39) and aberrant methylation of genes involved in cell signaling (18) in high risk MDS, no current study addresses the Stat3/5 signaling alterations at the single HSPC level in such patients.…”

Diverse phosphorylation patterns of B cell receptor-associated signaling in naïve and memory human B cells revealed by phosphoflow, a powerful technique to study signaling at the single cell level

“…The use of fluorescently labeled monoclonal antibodies for specific phosphorylated epitopes and the development of advanced multichromatic FC techniques have allowed the development of a new technology (phosphoflow) for the study of signaling pathways in primary human cells (Krutzik and Nolan, 2003; Irish et al, 2006; Schulz et al, 2007; Galligan et al, 2009; Krutzik et al, 2011b). This novel technology, in spite of still being in development, has already been used in clinical studies, particularly in blood cancer research (e.g., lymphomas), to identify basic aspects of the cell biology of cancerous cells and susceptibility to chemotherapeutic agents (Irish et al, 2004; Nolan, 2006; Chen et al, 2008; Galligan et al, 2009). However, the potential use of this technology to understand basic aspects of B cell biology in normal and pathologic conditions is enormous and has yet to be realized.…”

Single cell network profiling assay in bladder cancer

“…Although the utility of SCNP assay has been well demonstrated in hematologic malignancies (5,8,9), to date, its application to solid tumors has not been widely studied, with the exception of a limited number of studies focused on lung cancer (10). Although the utility of SCNP assay has been well demonstrated in hematologic malignancies (5,8,9), to date, its application to solid tumors has not been widely studied, with the exception of a limited number of studies focused on lung cancer (10).…”