Joshua Wisell

Epithelial stem cell mutations that promote squamous cell carcinoma metastasis.

Squamous cell carcinomas (SCCs) originate in stratified epithelia, with a small subset becoming metastatic. Epithelial stem cells are targets for driver mutations that give rise to SCCs, but it is unknown whether they contribute to oncogenic multipotency and metastasis. We developed a mouse model of SCC by targeting two frequent genetic mutations in human SCCs, oncogene Kras(G12D) activation and Smad4 deletion, to mouse keratin 15-expressing (K15+) stem cells. We show that transgenic mice developed multilineage tumors, including metastatic SCCs. Among cancer stem cell-enriched (CSC-enriched) populations, those with increased side population (SP) cells correlated with epithelial-mesenchymal transition (EMT) and lung metastasis. We show that microRNA-9 (miR-9) contributed to SP expansion and metastasis, and miR-9 inhibition reduced the number of SP cells and metastasis. Increased miR-9 was detected in metastatic human primary SCCs and SCC metastases, and miR-9-transduced human SCC cells exhibited increased invasion. We identified α-catenin as a predominant miR-9 target. Increased miR-9 in human SCC metastases correlated with α-catenin loss but not E-cadherin loss. Our results demonstrate that stem cells with Kras(G12D) activation and Smad4 depletion can produce tumors that are multipotent and susceptible to EMT and metastasis. Additionally, tumor initiation and metastatic properties of CSCs can be uncoupled, with miR-9 regulating the expansion of metastatic CSCs.

Grape seed extract and resveratrol prevent 4‐nitroquinoline 1‐oxide induced oral tumorigenesis in mice by modulating AMPK activation and associated biological responses

Preventive measures against oral carcinogenesis are urgently warranted to lower the high morbidity and mortality associated with this malignancy worldwide. Here, we investigated the chemopreventive efficacy of grape seed extract (GSE) and resveratrol (Res) in 4‐nitroquinoline‐1‐oxide (4NQO)‐induced tongue tumorigenesis in C57BL/6 mice. Following 8 weeks of 4NQO exposure (100 µg/ml in drinking water), mice were fed with either control AIN‐76A diet or diet containing 0.2% GSE (w/w) or 0.25% Res (w/w) for 8 subsequent weeks, while continued on 4NQO. Upon termination of the study at 16 weeks, tongue tissues were histologically evaluated for hyperplasia, dysplasia, and papillary lesions, and then analyzed for molecular targets by immunohistochemistry. GSE and Res feeding for 8 weeks, moderately decreased the incidence, but significantly prevented the multiplicity and severity of 4NQO‐induced preneoplastic and neoplastic lesions, without any apparent toxicity. In tongue tissues, both 4NQO + GSE and 4NQO + Res treatment correlated with a decreased proliferation (BrdU labeling index) but increased apoptotic death (TUNEL‐positive cells) as compared to the 4NQO group. Furthermore, tongue tissues from both the 4NQO + GSE and 4NQO + Res groups showed an increase in activated metabolic regulator phospho‐AMPK (Thr172) and decreased autophagy flux marker p62. Together, these findings suggest that GSE and Res could effectively prevent 4NQO‐induced oral tumorigenesis through modulating AMPK activation, and thereby, inhibiting proliferation and inducing apoptosis and autophagy, as mechanisms of their efficacy.

Kinase Gene Fusions in Defined Subsets of Melanoma.

Genomic rearrangements resulting in activating kinase fusions have been increasingly described in a number of cancers including malignant melanoma, but their frequency in specific melanoma subtypes has not been reported. We used break‐apart fluorescence in situ hybridization (FISH) to identify genomic rearrangements in tissues from 59 patients with various types of malignant melanoma including acral lentiginous, mucosal, superficial spreading, and nodular. We identified four genomic rearrangements involving the genes BRAF, RET, and ROS1. Of these, three were confirmed by Immunohistochemistry (IHC) or sequencing and one was found to be an ARMC10‐BRAF fusion that has not been previously reported in melanoma. These fusions occurred in different subtypes of melanoma but all in tumors lacking known driver mutations. Our data suggest gene fusions are more common than previously thought and should be further explored particularly in melanomas lacking known driver mutations.

Whole-exome sequencing identifies recurrent SF3B1 R625 mutation and comutation of NF1 and KIT in mucosal melanoma

Mucosal melanomas are a rare subtype of melanoma, arising in mucosal tissues, which have a very poor prognosis due to the lack of effective targeted therapies. This study aimed to better understand the molecular landscape of these cancers and find potential new therapeutic targets. Whole-exome sequencing was performed on mucosal melanomas from 19 patients and 135 sun-exposed cutaneous melanomas, with matched peripheral blood samples when available. Mutational profiles were compared between mucosal subgroups and sun-exposed cutaneous melanomas. Comparisons of molecular profiles identified 161 genes enriched in mucosal melanoma (P<0.05). KIT and NF1 were frequently comutated (32%) in the mucosal subgroup, with a significantly higher incidence than that in cutaneous melanoma (4%). Recurrent SF3B1 R625H/S/C mutations were identified and validated in 7 of 19 (37%) mucosal melanoma patients. Mutations in the spliceosome pathway were found to be enriched in mucosal melanomas when compared with cutaneous melanomas. Alternative splicing in four genes were observed in SF3B1-mutant samples compared with the wild-type samples. This study identified potential new therapeutic targets for mucosal melanoma, including comutation of NF1 and KIT, and recurrent R625 mutations in SF3B1. This is the first report of SF3B1 R625 mutations in vulvovaginal mucosal melanoma, with the largest whole-exome sequencing project of mucosal melanomas to date. The results here also indicated that the mutations in SF3B1 lead to alternative splicing in multiple genes. These findings expand our knowledge of this rare disease.

Cutaneous spindle cell carcinoma misdiagnosed as atypical fibroxanthoma based on immunohistochemical stains

Poorly differentiated spindle cell neoplasms pose a diagnostic challenge for dermatopathologists because of a lack of specific morphologic features on standard histopathology. Further characterization with immunohistochemical (IHC) studies is frequently required. We present a fatal case of spindle cell squamous cell carcinoma (SCC) misdiagnosed as atypical fibroxanthoma (AFX) based on a limited panel of IHC markers.

Discordance Between Intraoperative Consultation by Frozen Section and Final Diagnosis: A Classification Model to Guide Quality Improvement

Discrepancies between intraoperative consultations with frozen section diagnosis and the final pathology report have the potential to alter treatment decisions and affect patient care. Monitoring these correlations is a key component of laboratory quality assurance, however identifying specific areas for improvement can be difficult to attain. Our goal is to develop a standardized method utilizing root cause analysis and a modified Eindhoven classification schematic to identify the source of discrepancies and deferrals and subsequently to guide performance improvement initiatives. A retrospective review of intraoperative consultations performed at a tertiary level hospital and cancer center over a 6-month period identified deferrals and discrepancies between the intraoperative consult report and the final pathology report. We developed and applied a classification tool to identify the process errors and cognitive errors leading to discrepant results. A total of 48 (4.6%) discrepancies and 24 (2.3%) deferrals were identified from the 1042 frozen sections. Within the entire data set of frozen sections, the process errors (n = 26, 54.2%) were due to gross sampling (n = 16, 33.3%), histologic sampling (n = 8, 16.7%), and surgical sampling (n = 2, 4.2%). Interpretation errors (n = 22, 45.8%) included undercalls/false negatives (n=8, 16.7%), overcalls/false positives (n = 10, 20.8%), and misclassification errors (n = 4, 8.3%). Application of our classification tool demonstrated that the root cause of discrepancies and deferrals varied both between organ systems and by specific organs and that classification models may be utilized as a standardized method to identify focused areas for improvement.

Sox10 nuclear immunostaining lacks diagnostic utility for CNS granular cell tumors.

Sox10 immunohistochemistry has recently been shown to have diagnostic utility in differentiating typical granular cell tumors (GCTs) in skin and soft tissue from histological mimics (1–3). The overwhelming majority of cases studied have been of cutaneous origin (1–3), although Karamchandani et al included an unspecified number of soft tissue, laryngeal, and esophageal examples (2), and Heerema et al added 2 esophageal GCTs and 2 GCTs of neurohypophysis (3). All tumors from cutaneous and deep sites demonstrated strong diffuse nuclear immunoreactivity for Sox10, while the 2 neurohypophyseal examples were negative (3). To our knowledge, CNS GCTs, which are usually of high-grade astrocytic origin (4), have not yet been explored Given the important role of this transcription factor in neural crest, peripheral nervous system, and CNS development (albeit particularly for oligodendrocyte, not astrocytic, maturation in the CNS) (5), we hypothesized that CNS granular cell tumors might show Sox10 nuclear expression. Therefore, we immunostained CNS GCTs utilizing a polyclonal commercial antibody (Cell Marque Corp., Rocklin, CA). We first paralleled and extended the previous work with a study of 9 cutaneous (Fig. 1A) and 7 deep tissue GCTs from esophageal, colon, breast (n = 2), tongue, lung carina, and tracheal sites. These had been previously characterized at the time of initial diagnosis with hematoxylin and eosin (H&E) stain alone, S100 immunostaining (Fig. 1B), or additional antibodies, but not Sox10. All granular …

Advanced acral melanoma.

In the United States, the incidence of melanoma is increasing faster than any other preventable cancer. Acral melanoma is the fourth most common type of cutaneous melanoma and is frequently diagnosed later in its course compared with melanoma in other anatomic locations. Here we describe a case of highly advanced acral melanoma.

Abstract P6-05-10: Progestins exert divergent growth effects and regulate tumor-unique gene cohorts in patient derived breast cancer xenografts

Background: Progesterone is an important hormone for development and normal function of the breast; however, its role in established breast cancers is less clear. Progestins have been implicated in regulating tumor cell growth, signaling, differentiation state, and stem/progenitor properties in breast cancer cells. High dose progestin treatment can be an effective therapy for some advanced breast tumors, through an unknown mechanism. Progesterone receptors (PRs) are considered positive prognostic indicators, although their levels vary considerably among luminal subtype breast tumors. Progestin/PR regulated genes in tumor cells have only been determined in cultured breast cancer cells and mouse mammary tumor models. Here we define tumor unique progestin-dependent growth effects and gene regulation profiles in patient-derived luminal breast tumor xenografts. Methods: For these studies, three luminal estrogen receptor (ER)+PR+ transplantable xenografts were used that were derived from one pleural effusion and two previously untreated primary tumors. All three tumors were estrogen dependent to various degrees and contained variable levels of ER (5–90%) and PR (5–90%). Tumors were grown in vivo under continuous placebo, estrogen, or estrogen plus progestin conditions for 8–10 weeks and growth parameters monitored. Gene expression profiles were determined from dissected tumors using Affymetrix® GeneChip human gene 1.1 ST microarrays and results analyzed using Partek Genomics Suite software. Specific gene regulation was confirmed by q-RT-PCR and immunohistochemistry. Results: Progestins had an inhibitory, neutral, and stimulatory effect on estrogen dependent growth in each of the three tumor lines. Accordingly, there was relatively little overlap in PR regulated genes between the three tumors. In the tumor in which progestin treatment had a potent anti-tumor effect, progestins were strong repressors of estrogen/ER-regulated genes. Gene expression patterns between the natural hormone progesterone and the synthetic drug MPA were compared in one tumor line and found to be mostly similar. The percent of PR+ tumor cells may have influenced the potency of gene regulation. Unique progestin regulated genes were discovered in this human tumor model system that were involved in immune response, cytokine signaling, and stem/progenitor cell features. Conclusions: Women with breast cancer are exposed to progestins naturally or through hormonal therapies and these may have a profound effect on tumor biology. In some tumors, progestins are potently anti-proliferative, while in others they elicit the opposite effect and potentiate estrogen induced tumor growth. The diversity of progestin-regulated genes in each tumor underscores the hormone9s context dependent effects. Determining progestin dependent gene signatures in patient tumors may pinpoint appropriate candidates for progestin therapy in advanced tumors and/or provide a prognostic tool for predicting tumor progression. Funded by: NIH R01 CA140985 (C.A.S.), the Grohne Fund (P.K.), and the University of Colorado Cancer Center (C.A.S., P.K.) Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr P6-05-10.