Melanoma & Other Cancers

Melanoma originates in the epidermis and becomes metastatic after invasion into the dermis. Prior interactions between melanoma cells and dermis are poorly studied. Here, we show that melanoma cells directly affect the formation of the dermal tumour niche by microRNA trafficking before invasion. 

Melanocytes, cells of melanoma origin, are specialized in releasing pigment vesicles, termed melanosomes. In melanoma in situ, we found melanosome markers in distal fibroblasts before melanoma invasion. The melanosomes carry microRNAs into primary fibroblasts triggering changes, including increased proliferation, migration and pro-inflammatory gene expression, all known features of cancer-associated fibroblasts (CAFs). Specifically, melanosomal microRNA-211 directly targets IGF2R and leads to MAPK signalling activation, which reciprocally encourages melanoma growth. Melanosome release inhibitor prevented CAF formation. Since the first interaction of melanoma cells with blood vessels occurs in the dermis, our data suggest an opportunity to block melanoma invasion by preventing the formation of the dermal tumour niche.

Publications:

Dror et al. (2016) Nature Cell Biol. 18, 1006-1017 [PDF]

RNA interference (RNAi) has become a popular and important tool for the analysis of gene function. Loss-of-function studies, commonly performed by transfection of short interfering RNAs (siRNAs), have greatly facilitated functional analyses of the human transcriptome. However, there are major downsides to siRNA experiments, most importantly the transient inhibition of gene expression as well as their inefficient transfection into non-dividing cells. Overcoming those limitations, short hairpin RNA (shRNA) expression vectors are available, which stably integrate into a target cell's genome via retro- or lentiviral gene transfer. Intracellular processing of shRNAs results in short duplex RNAs with siRNA-like properties. Viral integration ensures a broad range of infectable target cell types and a stable expression of specific shRNAs, resulting in the permanent reduction of the targeted gene product. Complex shRNA expression libraries allow the targeted knockdown of thousands of different genes in a single experiment. 

Using such lentiviral vector shRNA libraries and initially barcode arrays and meanwhile next-generation sequencing analysis for decoding of the pooled RNAi screens, we are able to quantify the abundance of individual shRNAs and thus determine in a complex pool the number of cells infected with an individual shRNA construct. We used the technique to predict anti-proliferative effects of individual shRNAs from pooled negative selection screens, for example, identified synthetic-lethal activities toward combination therapies, defined genes which are required for a stem-cell like phenotype and found tumour suppressor genes by in vivo studies. For more details, see below.

Further studies are under way, both for the elucidation of basic regulative processes associated to cancer and for the identification of pathways that are affected by particular drugs or compounds. In particular, we use the technique for obtaining more detailed information on the functional effects of particularly potentially druggable gene products.

More recently, CRISPR-Cas constructs have been used for similar analyses and other purposes.