Alexander D. Boiko

Human melanoma-initiating cells express neural crest nerve growth factor receptor CD271

The question of whether tumorigenic cancer stem cells exist in human melanomas has arisen in the last few years. Here we show that in melanomas, tumour stem cells (MTSCs, for melanoma tumour stem cells) can be isolated prospectively as a highly enriched CD271+ MTSC population using a process that maximizes viable cell transplantation. The tumours sampled in this study were taken from a broad spectrum of sites and stages. High-viability cells isolated by fluorescence-activated cell sorting and re-suspended in a matrigel vehicle were implanted into T-, B- and natural-killer-deficient Rag2−/−γc−/− mice. The CD271+ subset of cells was the tumour-initiating population in 90% (nine out of ten) of melanomas tested. Transplantation of isolated CD271+ melanoma cells into engrafted human skin or bone in Rag2−/−γc−/− mice resulted in melanoma; however, melanoma did not develop after transplantation of isolated CD271− cells. We also show that in mice, tumours derived from transplanted human CD271+ melanoma cells were capable of metastatsis in vivo. CD271+ melanoma cells lacked expression of TYR, MART1 and MAGE in 86%, 69% and 68% of melanoma patients, respectively, which helps to explain why T-cell therapies directed at these antigens usually result in only temporary tumour shrinkage.

VHL loss in renal cell carcinoma leads to up-regulation of CUB domain-containing protein 1 to stimulate PKCδ-driven migration

A common genetic mutation found in clear cell renal cell carcinoma (CC-RCC) is the loss of the von Hippel-Lindau (VHL) gene, which results in stabilization of hypoxia-inducible factors (HIFs), and contributes to cancer progression and metastasis. CUB-domain-containing protein 1 (CDCP1) was shown to promote metastasis in scirrhous and lung adenocarcinomas as well as in prostate cancer. In this study, we established a molecular mechanism linking VHL loss to induction of the CDCP1 gene through the HIF-1/2 pathway in renal cancer. Also, we report that Fyn, which forms a complex with CDCP1 and mediates its signaling to PKCδ, is a HIF-1 target gene. Mechanistically, we found that CDCP1 specifically regulates phosphorylation of PKCδ, but not of focal adhesion kinase or Crk-associated substrate. Signal transduction from CDCP1 to PKCδ leads to its activation, increasing migration of CC-RCC. Furthermore, patient survival can be stratified by CDCP1 expression at the cell surface of the tumor. Taken together, our data indicates that CDCP1 protein might serve as a therapeutic target for CC-RCC.

Therapeutic Implications of Melanoma Heterogeneity

Over the last decade, the treatment of metastatic melanoma has been revolutionized by the translation of molecular insights into therapeutic benefit for patients. These include advances in immunotherapeutic and small‐molecule approaches aimed at destroying cells with immunogenic antigens or gene mutations. Despite these advances, the limited durability of clinical response and eventual disease progression underscores a need for better understanding of mechanisms underlying tumor development. Current targeted therapies are developed partly based on the rationale that tumors are primarily clonal with respect to mutant oncogene or cell surface antigen target. However, with the advancement of cell isolation and transplantation approaches coupled with deep sequencing and mutation detection techniques, it has become increasingly clear that tumors are polyclonal. As a result, sensitive malignant cells are eradicated by treatment while the remaining tumor cell populations are conferred varying degrees of resistance and survival advantages by harbouring or acquiring certain epigenetic and genetic abnormalities. Tumor heterogeneity thus represents a major obstacle to the successful application of current therapies. Gaining insights into the cellular and molecular aspects of tumor diversity will not only facilitate the development and selection of therapeutic targets but also promote the evolution of precision medicine. In this viewpoint, we will discuss the implications of tumor heterogeneity for the treatment of metastatic melanoma and propose approaches to accelerate the translation of scientific discovery into improved clinical outcomes.

Profiling Melanoma Heterogeneity Using Microwell RNA Cytometry

Unraveling heterogeneity of melanoma to discover new subpopulations of cells within the tumor has been fundamental to many advances in cancer biology, including identification of tumor initiating subsets and cells resisting immune-therapeutic approaches (Boiko et al., Nature 466:133-137, 2010; Civenni et al., Cancer Res 71:3098-3109, 2011; Schatton et al., Nature 451:345-349, 2008; Landsberg et al., Nature 490:412-416, 2012; Fang et al., Cancer Res 65:9328-9337, 2005). Traditionally, these discoveries were made possible due to the existence of well-characterized antibodies that enabled identification of cells homogeneous for the expression of specific cell surface antigen. However, further unwinding of heterogeneous cell populations into homogenous subsets in order to more precisely define their functional profile is limited by the availability of highly specific antibodies. Here we describe a technique capable of identifying homogeneous cell populations in heterogeneous sample based on the transcriptome profile. This approach enables semiquantitative measurement of gene expressions in hundreds to thousands of single cells in one step, paving the way to identify homogenous subpopulations of melanoma cells based on gene transcripts, independent of the availability of antibodies.