Divergent clonal selection dominates medulloblastoma at recurrence.

The development of targeted anti-cancer therapies through the study of cancer genomes is intended to increase survival rates and decrease treatment-related toxicity. We treated a transposon-driven, functional genomic mouse model of medulloblastoma with 'humanized' in vivo therapy (microneurosurgical tumour resection followed by multi-fractionated, image-guided radiotherapy). Genetic events in recurrent murine medulloblastoma exhibit a very poor overlap with those in matched murine diagnostic samples (<5%). Whole-genome sequencing of 33 pairs of human diagnostic and post-therapy medulloblastomas demonstrated substantial genetic divergence of the dominant clone after therapy (<12% diagnostic events were retained at recurrence). In both mice and humans, the dominant clone at recurrence arose through clonal selection of a pre-existing minor clone present at diagnosis. Targeted therapy is unlikely to be effective in the absence of the target, therefore our results offer a simple, proximal, and remediable explanation for the failure of prior clinical trials of targeted therapy.

Functional characterization of rare variants in human dopamine receptor D4 gene by genotype-phenotype correlations.

Next generation sequencing technologies have facilitated a notable shift from common disease common variant hypothesis to common disease rare variant, as also witnessed in recent literature on schizophrenia. Dopamine receptor D4 (DRD4), a G-protein-coupled receptor is associated with psychiatric disorders and has high affinity for atypical antipsychotic clozapine. We investigated the functional role of rare genetic variants in DRD4 which may have implications for translational medicine. CHO-K1 cells independently expressing four rare non-synonymous variants of DRD4 namely R237L, A281P, S284G located in the third cytosolic loop and V194G, located in the fifth transmembrane domain were generated. Their genotype-phenotype correlations were evaluated using [³H]spiperone binding, G-protein activation and molecular dynamics-simulation studies. A281P and S284G were functionally similar to wildtype (WT). With R237L, potency of dopamine and quinpirole reduced ∼sixfold and threefold respectively compared to WT; [³H]spiperone binding studies showed a reduction in total number of binding sites (∼40%) but not binding affinity, in silico docking studies revealed that binding of both dopamine and spiperone to R237L was structurally similar to WT. Of note, V194G variant failed to inhibit forskolin-stimulated adenylate cyclase activity and phosphorylate extracellular signal-regulated kinase; showed significant reduction in binding affinity (K(d)=2.16 nM) and total number of binding sites (∼66%) compared to WT in [³H]spiperone binding studies; and ligand docking studies showed that binding of dopamine and spiperone is superficial due to probable structural alteration. Transmembrane variant V194G in DRD4.4 results in functional alteration warranting continuing functional analysis of rare variants.