Experimental and spontaneous metastasis assays can result in divergence in clonal architecture.

Intratumoural heterogeneity is associated with poor outcomes in breast cancer. To understand how malignant clones survive and grow in metastatic niches, in vivo models using cell lines and patient-derived xenografts (PDX) have become the gold standard. Injections of cancer cells in orthotopic sites (spontaneous metastasis assays) or into the vasculature (experimental metastasis assays) have been used interchangeably to study the metastatic cascade from early events or post-intravasation, respectively. However, less is known about how these different routes of injection impact heterogeneity. Herein we directly compared the clonality of spontaneous and experimental metastatic assays using the human cell line MDA-MB-231 and a PDX model. Genetic barcoding was used to study the fitness of the subclones in primary and metastatic sites. Using spontaneous assays, we found that intraductal injections resulted in less diverse tumours compared to other routes of injections. Using experimental metastasis assays via tail vein injection of barcoded MDA-MB-231 cells, we also observed an asymmetry in metastatic heterogeneity between lung and liver that was not observed using spontaneous metastasis assays. These results demonstrate that these assays can result in divergent clonal outputs in terms of metastatic heterogeneity and provide a better understanding of the biases inherent to each technique.

Computational Screening of Anti-Cancer Drugs Identifies a New BRCA Independent Gene Expression Signature to Predict Breast Cancer Sensitivity to Cisplatin.

The development of therapies that target specific disease subtypes has dramatically improved outcomes for patients with breast cancer. However, survival gains have not been uniform across patients, even within a given molecular subtype. Large collections of publicly available drug screening data matched with transcriptomic measurements have facilitated the development of computational models that predict response to therapy. Here, we generated a series of predictive gene signatures to estimate the sensitivity of breast cancer samples to 90 drugs, comprising FDA-approved drugs or compounds in early development. To achieve this, we used a cell line-based drug screen with matched transcriptomic data to derive in silico models that we validated in large independent datasets obtained from cell lines and patient-derived xenograft (PDX) models. Robust computational signatures were obtained for 28 drugs and used to predict drug efficacy in a set of PDX models. We found that our signature for cisplatin can be used to identify tumors that are likely to respond to this drug, even in absence of the BRCA-1 mutation routinely used to select patients for platinum-based therapies. This clinically relevant observation was confirmed in multiple PDXs. Our study foreshadows an effective delivery approach for precision medicine.

Loss of TAF8 causes TFIID dysfunction and p53-mediated apoptotic neuronal cell death.

Mutations in genes encoding general transcription factors cause neurological disorders. Despite clinical prominence, the consequences of defects in the basal transcription machinery during brain development are unclear. We found that loss of the TATA-box binding protein-associated factor TAF8, a component of the general transcription factor TFIID, in the developing central nervous system affected the expression of many, but notably not all genes. Taf8 deletion caused apoptosis, unexpectedly restricted to forebrain regions. Nuclear levels of the transcription factor p53 were elevated in the absence of TAF8, as were the mRNAs of the pro-apoptotic p53 target genes Noxa, Puma and Bax. The cell death in Taf8 forebrain regions was completely rescued by additional loss of p53, but Taf8 and p53 brains failed to initiate a neuronal expression program. Taf8 deletion caused aberrant transcription of promoter regions and splicing anomalies. We propose that TAF8 supports the directionality of transcription and co-transcriptional splicing, and that failure of these processes causes p53-induced apoptosis of neuronal cells in the developing mouse embryo.

The site of breast cancer metastases dictates their clonal composition and reversible transcriptomic profile.

Intratumoral heterogeneity is a driver of breast cancer progression, but the nature of the clonal interactive network involved in this process remains unclear. Here, we optimized the use of optical barcoding to visualize and characterize 31 cancer subclones in vivo. By mapping the clonal composition of thousands of metastases in two clinically relevant sites, the lungs and liver, we found that metastases were highly polyclonal in lungs but not in the liver. Furthermore, the transcriptome of the subclones varied according to their metastatic niche. We also identified a reversible niche-driven signature that was conserved in lung and liver metastases collected during patient autopsies. Among this signature, we found that the tumor necrosis factor-α pathway was up-regulated in lung compared to liver metastases, and inhibition of this pathway affected metastasis diversity. These results highlight that the cellular and molecular heterogeneity observed in metastases is largely dictated by the tumor microenvironment.

Loss of p53 Causes Stochastic Aberrant X-Chromosome Inactivation and Female-Specific Neural Tube Defects.

Neural tube defects (NTDs) are common birth defects in humans and show an unexplained female bias. Female mice lacking the tumor suppressor p53 display NTDs with incomplete penetrance. We found that the combined loss of pro-apoptotic BIM and p53 caused 100% penetrant, female-exclusive NTDs, which allowed us to investigate the female-specific functions of p53. We report that female p53-/- embryonic neural tube samples show fewer cells with inactive X chromosome markers Xist and H3K27me3 and a concomitant increase in biallelic expression of the X-linked genes, Huwe1 and Usp9x. Decreased Xist and increased X-linked gene expression was confirmed by RNA sequencing. Moreover, we found that p53 directly bound response elements in the X chromosome inactivation center (XIC). Together, these findings suggest p53 directly activates XIC genes, without which there is stochastic failure in X chromosome inactivation, and that X chromosome inactivation failure may underlie the female bias in neural tube closure defects.

MOZ regulates B-cell progenitors and, consequently, Moz haploinsufficiency dramatically retards MYC-induced lymphoma development.

The histone acetyltransferase MOZ (MYST3, KAT6A) is the target of recurrent chromosomal translocations fusing the MOZ gene to CBP, p300, NCOA3, or TIF2 in particularly aggressive cases of acute myeloid leukemia. In this study, we report the role of wild-type MOZ in regulating B-cell progenitor proliferation and hematopoietic malignancy. In the Eµ-Myc model of aggressive pre-B/B-cell lymphoma, the loss of just one allele of Moz increased the median survival of mice by 3.9-fold. MOZ was required to maintain the proliferative capacity of B-cell progenitors, even in the presence of c-MYC overexpression, by directly maintaining the transcriptional activity of genes required for normal B-cell development. Hence, B-cell progenitor numbers were significantly reduced in Moz haploinsufficient animals. Interestingly, we find a significant overlap in genes regulated by MOZ, mixed lineage leukemia 1, and mixed lineage leukemia 1 cofactor menin. This includes Meis1, a TALE class homeobox transcription factor required for B-cell development, characteristically upregulated as a result of MLL1 translocations in leukemia. We demonstrate that MOZ localizes to the Meis1 locus in pre-B-cells and maintains Meis1 expression. Our results suggest that even partial inhibition of MOZ may reduce the proliferative capacity of MEIS1, and HOX-driven lymphoma and leukemia cells.