A mouse model featuring tissue-specific deletion of p53 and Brca1 gives rise to mammary tumors with genomic and transcriptomic similarities to human basal-like breast cancer.

PURPOSE AND METHODS: In human basal-like breast cancer, mutations and deletions in TP53 and BRCA1 are frequent oncogenic events. Thus, we interbred mice expressing the CRE-recombinase with mice harboring loxP sites at TP53 and BRCA1 (K14-Cre; p53f/f Brca1f/f) to test the hypothesis that tissue-specific deletion of TP53 and BRCA1 would give rise to tumors reflective of human basal-like breast cancer. RESULTS: In support of our hypothesis, these transgenic mice developed tumors that express basal-like cytokeratins and demonstrated intrinsic gene expression features similar to human basal-like tumors. Array comparative genomic hybridization revealed a striking conservation of copy number alterations between the K14-Cre; p53f/f Brca1f/f mouse model and human basal-like breast cancer. Conserved events included MYC amplification, KRAS amplification, and RB1 loss. Microarray analysis demonstrated that these DNA copy number events also led to corresponding changes in signatures of pathway activation including high proliferation due to RB1 loss. K14-Cre; p53f/f Brca1f/f also matched human basal-like breast cancer for a propensity to have immune cell infiltrates. Given the long latency of K14-Cre; p53f/f Brca1f/f tumors (~ 250 days), we created tumor syngeneic transplant lines, as well as in vitro cell lines, which were tested for sensitivity to carboplatin and paclitaxel. These therapies invoked acute regression, extended overall survival, and resulted in gene expression signatures of an anti-tumor immune response. CONCLUSION: These findings demonstrate that this model is a valuable preclinical resource for the study of human basal-like breast cancer.

A complete durable response of vaginal clear cell carcinoma with pembrolizumab: A case report.

Primary clear cell adenocarcinoma of the vagina represents a rare form of cancer historically correlated with in-utero diethylstibestrol (DES) exposure. Mainstay treatment modalities include surgery, radiation, and chemotherapy. There has been a growing interest in immunotherapy in the field of oncology. KEYNOTE 826 demonstrated that patients with persistent, recurrent, or metastatic cervical cancer including patients who had adenocarcinoma showed improved progression and overall survival by the addition Pembrolizumab to chemotherapy plus or minus bevacizumab. To date, there are no documented cases using pembrolizumab as adjuvant treatment for active or recurrent vaginal clear cell adenocarcinoma. We present a case of a young patient with recurrent vaginal clear cell carcinoma who showed a complete and durable response to Pembrolizumab.

Sample transport and electrokinetic injection in a microchip device for chemical cytometry.

Sample transport and electrokinetic injection bias are well characterized in capillary electrophoresis and simple microchips, but a thorough understanding of sample transport on devices combining electroosmosis, electrophoresis, and pressure-driven flow is lacking. In this work, we evaluate the effects of electric fields from 0 to 300 V/cm, electrophoretic mobilities from 10(-4) to 10(-6) cm(2)/Vs, and pressure-driven fluid velocities from 50 to 250 µm/s on sample injection in a microfluidic chemical cytometry device. By studying a continuous sample stream, we find that increasing electric field strength and electrophoretic mobility result in improved injection and that COMSOL simulations accurately predict sample transport. The effects of pressure-driven fluid velocity on injection are complex, and relative concentration values lie on a surface defined by pressure-driven flow rates. For high-mobility analytes, this surface is flat, and sample injection is robust despite fluctuations in flow rate. For lower mobility analytes, the surface becomes steeper, and injection depends strongly on pressure-driven flow. These results indicate generally that device design must account for analyte characteristics and specifically that this device is suited to high-mobility analytes. We demonstrate that for a suitable pair of peptides fluctuations in injection volume are correlated; electrokinetic injection bias is minimized; and electrophoretic separation is achieved.

Constitutive Ras signaling and Ink4a/Arf inactivation cooperate during the development of B-ALL in mice.

Despite recent advances in treatment, human precursor B-cell acute lymphoblastic leukemia (B-ALL) remains a challenging clinical entity. Recent genome-wide studies have uncovered frequent genetic alterations involving RAS pathway mutations and loss of the INK4A/ARF locus, suggesting their important role in the pathogenesis, relapse, and chemotherapy resistance of B-ALL. To better understand the oncogenic mechanisms by which these alterations might promote B-ALL and to develop an in vivo preclinical model of relapsed B-ALL, we engineered mouse strains with induced somatic KrasG12D pathway activation and/or loss of Ink4a/Arf during early stages of B-cell development. Although constitutive activation of KrasG12D in B cells induced prominent transcriptional changes that resulted in enhanced proliferation, it was not sufficient by itself to induce development of a high-grade leukemia/lymphoma. Instead, in 40% of mice, these engineered mutations promoted development of a clonal low-grade lymphoproliferative disorder resembling human extranodal marginal-zone lymphoma of mucosa-associated lymphoid tissue or lymphoplasmacytic lymphoma. Interestingly, loss of the Ink4a/Arf locus, apart from reducing the number of apoptotic B cells broadly attenuated KrasG12D-induced transcriptional signatures. However, combined Kras activation and Ink4a/Arf inactivation cooperated functionally to induce a fully penetrant, highly aggressive B-ALL phenotype resembling high-risk subtypes of human B-ALL such as BCR-ABL and CRFL2-rearranged. Ninety percent of examined murine B-ALL tumors showed loss of the wild-type Ink4a/Arf locus without acquisition of highly recurrent cooperating events, underscoring the role of Ink4a/Arf in restraining Kras-driven oncogenesis in the lymphoid compartment. These data highlight the importance of functional cooperation between mutated Kras and Ink4a/Arf loss on B-ALL.

An oncogenic Ezh2 mutation induces tumors through global redistribution of histone 3 lysine 27 trimethylation.

B cell lymphoma and melanoma harbor recurrent mutations in the gene encoding the EZH2 histone methyltransferase (EZH2), but the carcinogenic role of these mutations is unclear. Here we describe a mouse model in which the most common somatic Ezh2 gain-of-function mutation (EZH2(Y646F) in human; Ezh2(Y641F) in mouse) is conditionally expressed. Expression of Ezh2(Y641F) in mouse B cells or melanocytes caused high-penetrance lymphoma or melanoma, respectively. Overexpression of the anti-apoptotic protein Bcl2, but not the oncoprotein Myc, or loss of the tumor suppressor protein p53 (encoded by Trp53 in mice) further accelerated lymphoma progression. Expression of the mutant Braf but not the mutant Nras oncoprotein further accelerated melanoma progression. Although expression of Ezh2(Y641F) globally increased the abundance of trimethylated Lys27 of histone H3 (H3K27me3), it also caused a widespread redistribution of this repressive mark, including a loss of H3K27me3 that was associated with increased transcription at many loci. These results suggest that Ezh2(Y641F) induces lymphoma and melanoma through a vast reorganization of chromatin structure, inducing both repression and activation of polycomb-regulated loci.

Fasnall, a Selective FASN Inhibitor, Shows Potent Anti-tumor Activity in the MMTV-Neu Model of HER2(+) Breast Cancer.

Many tumors are dependent on de novo fatty acid synthesis to maintain cell growth. Fatty acid synthase (FASN) catalyzes the final synthetic step of this pathway, and its upregulation is correlated with tumor aggressiveness. The consequences and adaptive responses of acute or chronic inhibition of essential enzymes such as FASN are not fully understood. Herein we identify Fasnall, a thiophenopyrimidine selectively targeting FASN through its co-factor binding sites. Global lipidomics studies with Fasnall showed profound changes in cellular lipid profiles, sharply increasing ceramides, diacylglycerols, and unsaturated fatty acids as well as increasing exogenous palmitate uptake that is deviated more into neutral lipid formation rather than phospholipids. We also showed that the increase in ceramide levels contributes to some extent in the mediation of apoptosis. Consistent with this mechanism of action, Fasnall showed potent anti-tumor activity in the MMTV-Neu model of HER2(+) breast cancer, particularly when combined with carboplatin.

Coexistent ARID1A-PIK3CA mutations promote ovarian clear-cell tumorigenesis through pro-tumorigenic inflammatory cytokine signalling.

Ovarian clear-cell carcinoma (OCCC) is an aggressive form of ovarian cancer with high ARID1A mutation rates. Here we present a mutant mouse model of OCCC. We find that ARID1A inactivation is not sufficient for tumour formation, but requires concurrent activation of the phosphoinositide 3-kinase catalytic subunit, PIK3CA. Remarkably, the mice develop highly penetrant tumours with OCCC-like histopathology, culminating in haemorrhagic ascites and a median survival period of 7.5 weeks. Therapeutic treatment with the pan-PI3K inhibitor, BKM120, prolongs mouse survival by inhibiting the tumour cell growth. Cross-species gene expression comparisons support a role for IL-6 inflammatory cytokine signalling in OCCC pathogenesis. We further show that ARID1A and PIK3CA mutations cooperate to promote tumour growth through sustained IL-6 overproduction. Our findings establish an epistatic relationship between SWI/SNF chromatin remodelling and PI3K pathway mutations in OCCC and demonstrate that these pathways converge on pro-tumorigenic cytokine signalling. We propose that ARID1A protects against inflammation-driven tumorigenesis.

Genetic changes in neoplasms arising in congenital melanocytic nevi: differences between nodular proliferations and melanomas.

Large congenital melanocytic nevi (CMN) are at an increased risk of developing melanoma. Several forms of secondary proliferations can arise in congenital nevi on rare occasions. Although some of these closely resemble melanoma both clinically and histologically, metastasis is rare. We used comparative genomic hybridization to analyze chromosomal aberrations in different types of proliferations arising in CMN and compared them to typical congenital nevi, clear-cut melanomas arising in congenital nevi, as well as primary cutaneous melanomas that were not associated with a CMN. Cases of CMN and CMN with secondary proliferations were assigned to six groups according to the predominant histological pattern: group I, bland congenital nevi (n = 6); group II, congenital nevi with foci of increased cellularity (n = 4); group III, CMN with a proliferation simulating superficial spreading melanoma in situ (n = 3); group IV, CMN with a proliferation simulating nodular melanoma (n = 9); group V, proliferating neurocristic hamartoma (n = 1); and group VI, melanoma arising in congenital nevus (n = 6). No aberrations were found in groups I to III, whereas seven of nine cases of group IV, and one of one case of group V, showed aberrations. In group IV six of seven cases with aberrations (86%) showed numerical aberrations of whole chromosomes exclusively. This pattern differed significantly from the findings in melanoma that arose within CMN (n = 6), group VI, or independent of CMN (n = 122) in which only 5% showed numerical changes only. The single case in group V showed aberrations similar to melanoma. The finding of frequent numerical chromosomal aberrations in atypical nodular proliferations arising in CMN identifies these as clonal neoplasms with a genomic instability consistent with a mitotic spindle checkpoint defect. This difference compared to the aberration pattern found in melanoma might explain their more benign clinical behavior and may be of diagnostic value in ambiguous cases.