Publications by authors named "Shannon Carskadon"

42 Publications

Transcriptional network involving ERG and AR orchestrates Distal-less homeobox-1 mediated prostate cancer progression.

Nat Commun 2021 Sep 7;12(1):5325. Epub 2021 Sep 7.

Molecular Oncology Laboratory, Department of Biological Sciences and Bioengineering, Indian Institute of Technology Kanpur, Kanpur, U.P., India.

Distal-less homeobox-1 (DLX1) is a well-established non-invasive biomarker for prostate cancer (PCa) diagnosis, however, its mechanistic underpinnings in disease pathobiology are not known. Here, we reveal the oncogenic role of DLX1 and show that abrogating its function leads to reduced tumorigenesis and metastases. We observed that ~60% of advanced-stage and metastatic patients display higher DLX1 levels. Moreover, ~96% of TMPRSS2-ERG fusion-positive and ~70% of androgen receptor (AR)-positive patients show elevated DLX1, associated with aggressive disease and poor survival. Mechanistically, ERG coordinates with enhancer-bound AR and FOXA1 to drive transcriptional upregulation of DLX1 in ERG-positive background. However, in ERG-negative context, AR/AR-V7 and FOXA1 suffice to upregulate DLX1. Notably, inhibiting ERG/AR-mediated DLX1 transcription using BET inhibitor (BETi) or/and anti-androgen drugs reduce its expression and downstream oncogenic effects. Conclusively, this study establishes DLX1 as a direct-target of ERG/AR with an oncogenic role and demonstrates the clinical significance of BETi and anti-androgens for DLX1-positive patients.
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http://dx.doi.org/10.1038/s41467-021-25623-2DOI Listing
September 2021

Virus-positive Merkel Cell Carcinoma Is an Independent Prognostic Group with Distinct Predictive Biomarkers.

Clin Cancer Res 2021 May 5;27(9):2494-2504. Epub 2021 Feb 5.

Department of Dermatology, University of Michigan, Ann Arbor, Michigan.

Purpose: Merkel cell carcinoma (MCC) is an aggressive cutaneous neuroendocrine carcinoma that can be divided into two classes: virus-positive (VP) MCC, associated with oncogenic Merkel cell polyomavirus (MCPyV); and virus-negative (VN) MCC, associated with photodamage.

Experimental Design: We classified 346 MCC tumors from 300 patients for MCPyV using a combination of IHC, ISH, and qPCR assays. In a subset of tumors, we profiled mutation status and expression of cancer-relevant genes. MCPyV and molecular profiling results were correlated with disease-specific outcomes. Potential prognostic biomarkers were further validated by IHC.

Results: A total of 177 tumors were classified as VP-MCC, 151 tumors were VN-MCC, and 17 tumors were indeterminate. MCPyV positivity in primary tumors was associated with longer disease-specific and recurrence-free survival in univariate analysis, and in multivariate analysis incorporating age, sex, immune status, and stage at presentation. Prioritized oncogene or tumor suppressor mutations were frequent in VN-MCC but rare in VP-MCC. mutation developed with recurrence in one VP-MCC case. Importantly, for the first time we find that VP-MCC and VN-MCC display distinct sets of prognostic molecular biomarkers. For VP-MCC, shorter survival was associated with decreased expression of immune markers including granzyme and IDO1. For VN-MCC, shorter survival correlated with high expression of several genes including .

Conclusions: MCPyV status is an independent prognostic factor for MCC. Features of the tumor genome, transcriptome, and microenvironment may modify prognosis in a manner specific to viral status. MCPyV status has clinicopathologic significance and allows for identification of additional prognostic subgroups.
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http://dx.doi.org/10.1158/1078-0432.CCR-20-0864DOI Listing
May 2021

A Noncoding Variant Near PPP1R3B Promotes Liver Glycogen Storage and MetS, but Protects Against Myocardial Infarction.

J Clin Endocrinol Metab 2021 01;106(2):372-387

Brigham and Women's Hospital, Havard University, Boston, MA, USA.

Context: Glycogen storage diseases are rare. Increased glycogen in the liver results in increased attenuation.

Objective: Investigate the association and function of a noncoding region associated with liver attenuation but not histologic nonalcoholic fatty liver disease.

Design: Genetics of Obesity-associated Liver Disease Consortium.

Setting: Population-based.

Main Outcome: Computed tomography measured liver attenuation.

Results: Carriers of rs4841132-A (frequency 2%-19%) do not show increased hepatic steatosis; they have increased liver attenuation indicative of increased glycogen deposition. rs4841132 falls in a noncoding RNA LOC157273 ~190 kb upstream of PPP1R3B. We demonstrate that rs4841132-A increases PPP1R3B through a cis genetic effect. Using CRISPR/Cas9 we engineered a 105-bp deletion including rs4841132-A in human hepatocarcinoma cells that increases PPP1R3B, decreases LOC157273, and increases glycogen perfectly mirroring the human disease. Overexpression of PPP1R3B or knockdown of LOC157273 increased glycogen but did not result in decreased LOC157273 or increased PPP1R3B, respectively, suggesting that the effects may not all occur via affecting RNA levels. Based on electronic health record (EHR) data, rs4841132-A associates with all components of the metabolic syndrome (MetS). However, rs4841132-A associated with decreased low-density lipoprotein (LDL) cholesterol and risk for myocardial infarction (MI). A metabolic signature for rs4841132-A includes increased glycine, lactate, triglycerides, and decreased acetoacetate and beta-hydroxybutyrate.

Conclusions: These results show that rs4841132-A promotes a hepatic glycogen storage disease by increasing PPP1R3B and decreasing LOC157273. rs4841132-A promotes glycogen accumulation and development of MetS but lowers LDL cholesterol and risk for MI. These results suggest that elevated hepatic glycogen is one cause of MetS that does not invariably promote MI.
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http://dx.doi.org/10.1210/clinem/dgaa855DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7823249PMC
January 2021

Gene fusion characterisation of rare aggressive prostate cancer variants-adenosquamous carcinoma, pleomorphic giant-cell carcinoma, and sarcomatoid carcinoma: an analysis of 19 cases.

Histopathology 2020 Dec 13;77(6):890-899. Epub 2020 Sep 13.

Department of Pathology and Laboratory Medicine and Henry Ford Cancer Institute, Henry Ford Health System, Detroit, MI, USA.

Aims: To evaluate the molecular underpinnings of the rare aggressive prostate cancer variants adenosquamous carcinoma, pleomorphic giant-cell carcinoma, and sarcomatoid carcinoma.

Methods And Results: We retrieved 19 tumours with one or more variant(s), and performed ERG immunohistochemistry, a next-generation sequencing assay targeting recurrent gene fusions, and fluorescence in-situ hybridisation (FISH) for ERG and BRAF. Divergent differentiation included: sarcomatoid carcinoma (n = 10), adenosquamous carcinoma (n = 7), and pleomorphic giant-cell carcinoma (n = 7). Five patients had more than one variant. Four had variants only in metastases. ERG rearrangement was detected in nine (47%, seven via sequencing, showing TMPRSS2-ERG fusions and one GRHL2-ERG fusion, and two via FISH, showing rearrangement via deletion). ERG was immunohistochemically positive in the adenocarcinoma in eight of nine (89%) patients, but was immunohistochemically positive in the variant in only five of nine patients (56%, typically decreased). One patient had a false-positive ERG immunohistochemical result in the sarcomatoid component despite a negative FISH result. Two (11%) harboured BRAF fusions (FAM131A-BRAF and SND1-BRAF).

Conclusions: ERG fusions are present in these rare prostate cancer variants with a frequency close to that in conventional prostate cancer (9/19, 47%). ERG immunohistochemistry usually detects rearrangement in the adenocarcinoma, but is less sensitive for the variant histology, with weak to negative staining. Adenosquamous and sarcomatoid variants can, particularly, occur together. Molecular assessment may be an additional tool in selected cases to confirm the prostatic origin of unusual tumours. The presence of two BRAF rearrangements suggests that this gene fusion may be enriched in this setting, as RAF kinase fusions have been previously reported in 1-2% of prostate cancers.
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http://dx.doi.org/10.1111/his.14205DOI Listing
December 2020

The MD Anderson Prostate Cancer Patient-derived Xenograft Series (MDA PCa PDX) Captures the Molecular Landscape of Prostate Cancer and Facilitates Marker-driven Therapy Development.

Clin Cancer Res 2020 09 23;26(18):4933-4946. Epub 2020 Jun 23.

Department of Genitourinary Medical Oncology and the David H. Koch Center for Applied Research of Genitourinary Cancers, The University of Texas MD Anderson Cancer Center, Houston, Texas.

Purpose: Advances in prostate cancer lag behind other tumor types partly due to the paucity of models reflecting key milestones in prostate cancer progression. Therefore, we develop clinically relevant prostate cancer models.

Experimental Design: Since 1996, we have generated clinically annotated patient-derived xenografts (PDXs; the MDA PCa PDX series) linked to specific phenotypes reflecting all aspects of clinical prostate cancer.

Results: We studied two cell line-derived xenografts and the first 80 PDXs derived from 47 human prostate cancer donors. Of these, 47 PDXs derived from 22 donors are working models and can be expanded either as cell lines (MDA PCa 2a and 2b) or PDXs. The histopathologic, genomic, and molecular characteristics (androgen receptor, ERG, and loss) maintain fidelity with the human tumor and correlate with published findings. PDX growth response to mouse castration and targeted therapy illustrate their clinical utility. Comparative genomic hybridization and sequencing show significant differences in oncogenic pathways in pairs of PDXs derived from different areas of the same tumor. We also identified a recurrent focal deletion in an area that includes the speckle-type POZ protein-like () gene in PDXs derived from seven human donors of 28 studied (25%). is a paralog, and mutations define a molecular subclass of prostate cancer. deletions are found in 7% of The Cancer Genome Atlas prostate cancers, which suggests that our cohort is a reliable platform for targeted drug development.

Conclusions: The MDA PCa PDX series is a dynamic resource that captures the molecular landscape of prostate cancers progressing under novel treatments and enables optimization of prostate cancer-specific, marker-driven therapy.
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http://dx.doi.org/10.1158/1078-0432.CCR-20-0479DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7501166PMC
September 2020

Clonal evaluation of prostate cancer molecular heterogeneity in biopsy samples by dual immunohistochemistry and dual RNA in situ hybridization.

Mod Pathol 2020 09 1;33(9):1791-1801. Epub 2020 Apr 1.

Department of Urology, Vattikuti Urology Institute, Henry Ford Health System, Detroit, MI, USA.

Prostate cancer is frequently multifocal. Although there may be morphological variation, the genetic underpinnings of each tumor are not clearly understood. To assess the inter and intra tumor molecular heterogeneity in prostate biopsy samples, we developed a combined immunohistochemistry and RNA in situ hybridization method for the simultaneous evaluation of ERG, SPINK1, ETV1, and ETV4. Screening of 601 biopsy cores from 120 consecutive patients revealed multiple alterations in a mutually exclusive manner in 37% of patients, suggesting multifocal tumors with considerable genetic differences. Furthermore, the incidence of molecular heterogeneity was higher in African Americans patients compared with Caucasian American patients. About 47% of the biopsy cores with discontinuous tumor foci showed clonal differences with distinct molecular aberrations. ERG positivity occurred in low-grade cancer, whereas ETV4 expression was observed mostly in high-grade cancer. Further studies revealed correlation between the incidence of molecular markers and clinical and pathologic findings, suggesting potential implications for diagnostic pathology practice, such as defining dominant tumor nodules and discriminating juxtaposed but molecularly different tumors of different grade patterns.
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http://dx.doi.org/10.1038/s41379-020-0525-0DOI Listing
September 2020

Therapeutically actionable PAK4 is amplified, overexpressed, and involved in bladder cancer progression.

Oncogene 2020 05 30;39(20):4077-4091. Epub 2020 Mar 30.

Department of Medicine, Dana-Farber Cancer Institute, Boston, MA, USA.

Muscle-invasive bladder carcinomas (MIBCs) are aggressive genitourinary malignancies. Metastatic urothelial carcinoma of the bladder is generally incurable by current chemotherapy and leads to early mortality. Recent studies have identified molecular subtypes of MIBCs with different sensitivities to frontline therapy, suggesting tumor heterogeneity. We have performed multi-omic profiling of the kinome in bladder cancer patients with the goal of identify therapeutic targets. Our analyses revealed amplification, overexpression, and elevated kinase activity of P21 (RAC1) activated kinase 4 (PAK4) in a subset of Bladder cancer (BLCA). Using bladder cancer cells, we confirmed the role of PAK4 in BLCA cell proliferation and invasion. Furthermore, we observed that a PAK4 inhibitor was effective in curtailing growth of BLCA cells. Transcriptomic analyses identified elevated expression of another kinase, protein tyrosine kinase 6 (PTK6), upon treatment with a PAK4 inhibitor and RNA interference of PAK4. Treatment with a combination of kinase inhibitors (vandetanib and dasatinib) showed enhanced sensitivity compared with either drug alone. Thus, PAK4 may be therapeutically actionable for a subset of MIBC patients with amplified and/or overexpressed PAK4 in their tumors. Our results also indicate that combined inhibition of PAK4 and PTK6 may overcome resistance to PAK4. These observations warrant clinical investigations with selected BLCA patients.
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http://dx.doi.org/10.1038/s41388-020-1275-7DOI Listing
May 2020

Androgen deprivation upregulates SPINK1 expression and potentiates cellular plasticity in prostate cancer.

Nat Commun 2020 01 20;11(1):384. Epub 2020 Jan 20.

Molecular Oncology Laboratory, Department of Biological Sciences and Bioengineering, Indian Institute of Technology Kanpur, Kanpur, UP, 208016, India.

Emergence of an aggressive androgen receptor (AR)-independent neuroendocrine prostate cancer (NEPC) after androgen-deprivation therapy (ADT) is well-known. Nevertheless, the majority of advanced-stage prostate cancer patients, including those with SPINK1-positive subtype, are treated with AR-antagonists. Here, we show AR and its corepressor, REST, function as transcriptional-repressors of SPINK1, and AR-antagonists alleviate this repression leading to SPINK1 upregulation. Increased SOX2 expression during NE-transdifferentiation transactivates SPINK1, a critical-player for maintenance of NE-phenotype. SPINK1 elicits epithelial-mesenchymal-transition, stemness and cellular-plasticity. Conversely, pharmacological Casein Kinase-1 inhibition stabilizes REST, which in cooperation with AR causes SPINK1 transcriptional-repression and impedes SPINK1-mediated oncogenesis. Elevated levels of SPINK1 and NEPC markers are observed in the tumors of AR-antagonists treated mice, and in a subset of NEPC patients, implicating a plausible role of SPINK1 in treatment-related NEPC. Collectively, our findings provide an explanation for the paradoxical clinical-outcomes after ADT, possibly due to SPINK1 upregulation, and offers a strategy for adjuvant therapies.
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http://dx.doi.org/10.1038/s41467-019-14184-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6971084PMC
January 2020

Next-generation sequencing implicates oncogenic roles for p53 and JAK/STAT signaling in microcystic adnexal carcinomas.

Mod Pathol 2020 06 19;33(6):1092-1103. Epub 2019 Dec 19.

Department of Pathology, Michigan Medicine, University of Michigan, Ann Arbor, MI, USA.

Microcystic adnexal carcinoma is a locally aggressive sweat gland carcinoma characterized by its infiltrative growth and histopathologic overlap with benign adnexal tumors, often posing challenges to both diagnosis and management. Understanding the molecular underpinnings of microcystic adnexal carcinoma may allow for more accurate diagnosis and identify potential targetable oncogenic drivers. We characterized 18 microcystic adnexal carcinomas by targeted, multiplexed PCR-based DNA next-generation sequencing of the coding sequence of over 400 cancer-relevant genes. The majority of cases had relatively few (<8) prioritized somatic mutations, and lacked an ultraviolet (UV) signature. The most recurrent mutation was TP53 inactivation in four (22%) tumors. Frame-preserving insertions affecting the kinase domain of JAK1 were detected in three (17%) cases, and were nonoverlapping with TP53 mutations. Seven (39%) cases demonstrated copy number gain of at least one oncogene. By immunohistochemistry, p53 expression was significantly higher in microcystic adnexal carcinomas with TP53 mutations compared with those without such mutations and syringomas. Similarly, phospho-STAT3 expression was significantly higher in microcystic adnexal carcinomas harboring JAK1 kinase insertions compared with those with wild-type JAK1 and syringomas. In conclusion, microcystic adnexal carcinomas are molecularly heterogeneous tumors, with inactivated p53 or activated JAK/STAT signaling in a subset. Unlike most other nonmelanoma skin cancers involving sun-exposed areas, most microcystic adnexal carcinomas lack evidence of UV damage, and hence likely originate from a relatively photo-protected progenitor population in the dermis. These findings have implications for the biology, diagnosis, and treatment of microcystic adnexal carcinomas, including potential for therapeutic targeting of p53 or the JAK/STAT pathway in advanced tumors.
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http://dx.doi.org/10.1038/s41379-019-0424-4DOI Listing
June 2020

Clonal evaluation of early onset prostate cancer by expression profiling of ERG, SPINK1, ETV1, and ETV4 on whole-mount radical prostatectomy tissue.

Prostate 2020 01 4;80(1):38-50. Epub 2019 Oct 4.

Department of Urology, Vattikuti Urology Institute, Henry Ford Health System, Detroit, Michigan.

Background: Expression profiles of erythroblast transformation-specific (ETS)-related gene fusions and serine protease inhibitor Kazal-type 1 (SPINK1) in early onset prostate cancer have not been thoroughly explored.

Methods: We retrieved 151 radical prostatectomy specimens from young men with prostate cancer (<55 years) and characterized the expression of ETS-related gene (ERG), SPINK1, ETS Variant 1 (ETV1), and ETV4 by dual immunohistochemistry and dual RNA in situ hybridization. Age, race, family history, preoperative prostate-specific antigen, biochemical recurrence, and pathological variables using whole-mount radical prostatectomy tissue were collected.

Results: A total of 313 tumor nodules from 151 men including 68 (45%) Caucasians and 61 (40%) African Americans were included in the analysis. Positive family history of prostate cancer was seen in 65 (43%) patients. Preoperative prostate-specific antigen ranged from 0.3 to 52.7 ng/mL (mean = 7.04). The follow-up period ranged from 1 to 123.7 months (mean = 30.3). Biochemical recurrence was encountered in 8 of 151 (5%). ERG overexpression was observed in 85 of 151 (56%) cases, followed by SPINK1 in 61 of 151 (40%), ETV1 in 9 of 149 (6%), and ETV4 in 4 of 141 (3%). There were 25 of 151 (17%) cases showing both ERG and SPINK1 overexpression within different regions of either the same tumor focus or different foci. Higher frequency of ERG overexpression was seen in younger patients (≤45 years old; 76% vs 49%, P = .002), Caucasian men (71% vs 41% P = .0007), organ-confined tumors (64% vs 33%, P = .0008), and tumors of Gleason Grade groups 1 and 2 (62% vs 26%, P = .009). SPINK1 overexpression was more in African American men (68% vs 26%, P = .00008), in tumors with high tumor volume (>20%) and with anterior located tumors. ETV1 and ETV4 demonstrated rare overexpression in these tumors, particularly in the higher-grade tumors.

Conclusion: This study expands the knowledge of the clonal evolution of multifocal cancer in young patients and support differences in relation to racial background and genetics of prostate cancer.
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http://dx.doi.org/10.1002/pros.23914DOI Listing
January 2020

CDK7 Inhibition Suppresses Castration-Resistant Prostate Cancer through MED1 Inactivation.

Cancer Discov 2019 11 29;9(11):1538-1555. Epub 2019 Aug 29.

Department of Cancer Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.

Metastatic castration-resistant prostate cancer (CRPC) is a fatal disease, primarily resulting from the transcriptional addiction driven by androgen receptor (AR). First-line CRPC treatments typically target AR signaling, but are rapidly bypassed, resulting in only a modest survival benefit with antiandrogens. Therapeutic approaches that more effectively block the AR-transcriptional axis are urgently needed. Here, we investigated the molecular mechanism underlying the association between the transcriptional coactivator MED1 and AR as a vulnerability in AR-driven CRPC. MED1 undergoes CDK7-dependent phosphorylation at T1457 and physically engages AR at superenhancer sites, and is essential for AR-mediated transcription. In addition, a CDK7-specific inhibitor, THZ1, blunts AR-dependent neoplastic growth by blocking AR/MED1 corecruitment genome-wide, as well as reverses the hyperphosphorylated MED1-associated enzalutamide-resistant phenotype. , THZ1 induces tumor regression of AR-amplified human CRPC in a xenograft mouse model. Together, we demonstrate that CDK7 inhibition selectively targets MED1-mediated, AR-dependent oncogenic transcriptional amplification, thus representing a potential new approach for the treatment of CRPC. SIGNIFICANCE: Potent inhibition of AR signaling is critical to treat CRPC. This study uncovers a driver role for CDK7 in regulating AR-mediated transcription through phosphorylation of MED1, thus revealing a therapeutically targetable potential vulnerability in AR-addicted CRPC...
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http://dx.doi.org/10.1158/2159-8290.CD-19-0189DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7202356PMC
November 2019

Pseudogene Associated Recurrent Gene Fusion in Prostate Cancer.

Neoplasia 2019 10 22;21(10):989-1002. Epub 2019 Aug 22.

Vattikuti Urology Institute, Department of Urology, Henry Ford Health System, Detroit, MI. Electronic address:

We present the functional characterization of a pseudogene associated recurrent gene fusion in prostate cancer. The fusion gene KLK4-KLKP1 is formed by the fusion of the protein coding gene KLK4 with the noncoding pseudogene KLKP1. Screening of a cohort of 659 patients (380 Caucasian American; 250 African American, and 29 patients from other races) revealed that the KLK4-KLKP1 is expressed in about 32% of prostate cancer patients. Correlative analysis with other ETS gene fusions and SPINK1 revealed a concomitant expression pattern of KLK4-KLKP1 with ERG and a mutually exclusive expression pattern with SPINK1, ETV1, ETV4, and ETV5. Development of an antibody specific to KLK4-KLKP1 fusion protein confirmed the expression of the full-length KLK4-KLKP1 protein in prostate tissues. The in vitro and in vivo functional assays to study the oncogenic properties of KLK4-KLKP1 confirmed its role in cell proliferation, cell invasion, intravasation, and tumor formation. Presence of strong ERG and AR binding sites located at the fusion junction in KLK4-KLKP1 suggests that the fusion gene is regulated by ERG and AR. Correlative analysis of clinical data showed an association of KLK4-KLKP1 with lower preoperative PSA values and in young men (<50 years) with prostate cancer. Screening of patient urine samples showed that KLK4-KLKP1 can be detected noninvasively in urine. Taken together, we present KLK4-KLKP1 as a class of pseudogene associated fusion transcript in cancer with potential applications as a biomarker for routine screening of prostate cancer.
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http://dx.doi.org/10.1016/j.neo.2019.07.010DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6713813PMC
October 2019

A pediatric case of pigmented epithelioid melanocytoma with chromosomal copy number alterations in 15q and 17q and a novel NTRK3-SCAPER gene fusion.

J Cutan Pathol 2020 Jan 11;47(1):70-75. Epub 2019 Sep 11.

Department of Urology, Vattikuti Urology Institute, Henry Ford Health System, Detroit, Michigan.

Pigmented epithelioid melanocytoma (PEM) represents a group of rare, heavily pigmented melanocytic tumors encompassing lesions previously designated as "animal-type melanomas" and "epithelioid blue nevi." Despite the association of multiple such tumors in the setting of Carney complex, most cases of PEM occur spontaneously as solitary neoplasms in otherwise healthy patients. PEM may arise in both children and adults, and has a known propensity to spread to the regional lymph nodes. Despite this latter finding, recurrence at the biopsy site or spread beyond the lymph node basin is exceptionally uncommon. Although the molecular basis for PEM continues to be characterized, findings to date suggest that this category of melanocytic neoplasia has genetic alterations distinct from those seen in common nevi, dysplastic nevi, Spitz nevi, and melanoma. Herein, we present an in-depth clinical, histopathologic, and molecular analysis of a case of PEM occurring on the scalp of a young African American girl found to have a novel NTRK3-SCAPER gene fusion.
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http://dx.doi.org/10.1111/cup.13566DOI Listing
January 2020

Prostate cancer with comedonecrosis is frequently, but not exclusively, intraductal carcinoma: a need for reappraisal of grading criteria.

Histopathology 2019 Jun 29;74(7):1081-1087. Epub 2019 Apr 29.

Department of Pathology and Laboratory Medicine, Henry Ford Cancer Institute, Henry Ford Health System, Detroit, MI, USA.

Aims: Comedonecrosis in prostate cancer has always been Gleason pattern 5. However, we aimed to evaluate how intraductal carcinoma (not graded) with comedonecrosis should be considered.

Methods And Results: From 52 radical prostatectomy patients, 40 were informative and evaluated with immunohistochemistry for basal cells. Clinical outcome was assessed for biochemical recurrence, metastatic disease and the need for adjuvant therapy. Comedonecrosis was predominantly located in intraductal carcinoma (24, 60%). However, nine (23%) had comedonecrosis within invasive cancer and seven (18%) within both invasive and intraductal carcinoma. Extraprostatic extension rarely showed comedonecrosis (5, 13%), but rather perineural invasion within cribriform glands. Tumours were largely high-stage (15, 38% pT3a and 19, 48% pT3b), with 15 (37%) having positive lymph nodes and four distant metastases. Most cases (25, 63%) had other patterns of Gleason pattern 5 (single cells, solid), although 10 were reclassified as containing no invasive pattern 5. Of these, most were pT3 (eight of 10), but none had positive lymph nodes. Lymph node metastases were more common in patients with invasive cancer containing comedonecrosis (P = 0.02), and the need for androgen deprivation was near significance (P = 0.07), but biochemical recurrence was not significantly different (P = 0.58).

Conclusions: Prostate cancer with comedonecrosis is often intraductal; however, these tumours are largely high-stage, showing a higher rate of positive lymph nodes with invasive comedonecrosis. Immunohistochemistry may be considered when comedonecrosis may significantly change the tumour grade. However, it is not clear at present that excluding intraductal carcinoma from the grade is superior to including it in grading when it is associated with high-grade invasive cancer.
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http://dx.doi.org/10.1111/his.13833DOI Listing
June 2019

Epigenetic Silencing of miRNA-338-5p and miRNA-421 Drives SPINK1-Positive Prostate Cancer.

Clin Cancer Res 2019 May 26;25(9):2755-2768. Epub 2018 Dec 26.

Molecular Oncology Lab, Department of Biological Sciences and Bioengineering, Indian Institute of Technology Kanpur, Kanpur, Uttar Pradesh, India.

Purpose: Serine peptidase inhibitor, Kazal type-1 (SPINK1) overexpression defines the second most recurrent and aggressive prostate cancer subtype. However, the underlying molecular mechanism and pathobiology of SPINK1 in prostate cancer remains largely unknown.

Experimental Design: miRNA prediction tools were employed to examine the 3'UTR for miRNA binding. Luciferase reporter assays were performed to confirm the 3'UTR binding of shortlisted miR-338-5p/miR-421. Furthermore, miR-338-5p/-421-overexpressing cancer cells (SPINK1-positive) were evaluated for oncogenic properties using cell-based functional assays and a mouse xenograft model. Global gene expression profiling was performed to unravel the biological pathways altered by miR-338-5p/-421. IHC and RNA hybridization were carried out on prostate cancer patients' tissue microarray for SPINK1 and expression, respectively. Chromatin immunoprecipitation assay was performed to examine EZH2 occupancy on the miR-338-5p/-421-regulatory regions. Bisulfite sequencing and methylated DNA immunoprecipitation were performed on prostate cancer cell lines and patients' specimens.

Results: We established a critical role of miRNA-338-5p/-421 in posttranscriptional regulation of . Ectopic expression of miRNA-338-5p/-421 in SPINK1-positive cells abrogates oncogenic properties including cell-cycle progression, stemness, and drug resistance, and shows reduced tumor burden and distant metastases in a mouse model. Importantly, we show that patients with SPINK1-positive prostate cancer exhibit increased EZH2 expression, suggesting its role in epigenetic silencing of miRNA-338-5p/-421. Furthermore, presence of CpG dinucleotide DNA methylation marks on the regulatory regions of miR-338-5p/-421 in SPINK1-positive prostate cancer cells and patients' specimens confirms epigenetic silencing.

Conclusions: Our findings revealed that miRNA-338-5p/-421 are epigenetically silenced in SPINK1-positive prostate cancer, although restoring the expression of these miRNAs using epigenetic drugs or synthetic mimics could abrogate SPINK1-mediated oncogenesis..
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http://dx.doi.org/10.1158/1078-0432.CCR-18-3230DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6517268PMC
May 2019

Neurofilament is superior to cytokeratin 20 in supporting cutaneous origin for neuroendocrine carcinoma.

Histopathology 2019 Feb 5;74(3):504-513. Epub 2018 Dec 5.

Department of Pathology, Michigan Medicine/University of Michigan Health System, Ann Arbor, MI, USA.

Aim: Primary cutaneous neuroendocrine carcinoma, or Merkel cell carcinoma (MCC), cannot be distinguished morphologically from small-cell neuroendocrine carcinomas (SmCC) from other sites. Immunohistochemistry is required to confirm cutaneous origin, and is also used for detection of sentinel lymph node (SLN) metastases of MCC. Cytokeratin 20 (CK20) expression is commonly used for these purposes, but is negative in some MCC cases, and has unclear specificity. We evaluated immunohistochemistry for neurofilament and CK20 in MCC compared with SmCC from other sites.

Methods And Results: We evaluated neurofilament expression in 55 MCC specimens from 39 unique patients, including nine CK20-negative MCC tumours. Neurofilament expression was observed in 42 of 55 (76.4%) MCC cases, including seven of nine (77.8%) CK20-negative MCC cases. Neurofilament was expressed in nine of 12 (75%) Merkel cell polyomavirus-positive tumours and five of 10 (50%) virus-negative tumours. Compared to a standard immunohistochemical panel (cytokeratin cocktail and CK20), neurofilament was 87.5% sensitive for detecting SLN metastases. Neurofilament and CK20 expression was also assessed in 61 extracutaneous SmCC from 60 unique patients, with primary sites including lung (27), bladder (18), cervix (3), gastrointestinal tract (3), sinonasal tract (2) and other sites (7). The specificity of neurofilament and CK20 for MCC versus non-cutaneous SmCC was 96.7% and 59.0%, respectively.

Conclusions: Neurofilament has superior specificity to CK20 in distinguishing MCC from non-cutaneous SmCC. Neurofilament is frequently expressed in CK20- and virus-negative MCC tumours. Limitations of neurofilament immunohistochemistry include lower sensitivity than CK20 and subtle staining in some tumours. However, our findings indicate that neurofilament is useful for excluding non-cutaneous SmCC.
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http://dx.doi.org/10.1111/his.13758DOI Listing
February 2019

Wnt receptor Frizzled 8 is a target of ERG in prostate cancer.

Prostate 2018 12 26;78(16):1311-1320. Epub 2018 Jul 26.

Department of Pathology, University of Alabama at Birmingham, Birmingham, Alabama.

Prostate cancer (PCa) is one of the most frequently diagnosed cancers among men. Many molecular changes have been detailed during PCa progression. The gene encoding the transcription factor ERG shows recurrent rearrangement, resulting in the overexpression of ERG in the majority of prostate cancers. Overexpression of ERG plays a critical role in prostate oncogenesis and development of metastatic disease. Among the downstream effectors of ERG, Frizzled family member FZD4 has been shown to be a target of ERG. Frizzled-8 (FZD8) has been shown to be involved in PCa bone metastasis. In the present study, we show that the expression of FZD8 is directly correlated with ERG expression in PCa. Furthermore, we show that ERG directly targets and activates FZD8 by binding to its promoter. This activation is specific to ETS transcription factor ERG and not ETV1. We propose that ERG overexpression in PCa leads to induction of Frizzled family member FZD8, which is known to activate the Wnt pathway. Taken together, these findings uncover a novel mechanism for PCa metastasis, and indicate that FZD8 may represent a potential therapeutic target for PCa.
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http://dx.doi.org/10.1002/pros.23704DOI Listing
December 2018

Enrichment and mutation detection of circulating tumor cells from blood samples.

Oncol Rep 2018 Jun 30;39(6):2537-2544. Epub 2018 Mar 30.

Celsee Diagnostics, Plymouth, MI 48170, USA.

The potential of circulating tumor cells (CTCs) in the diagnosis and prognosis of cancer patients has become increasingly attractive. However, molecular analysis of CTCs is hindered by low sensitivity and a high level of background leukocytes in CTC enrichment technologies. We have developed a novel protocol using a microfluidic device, which enriches and retrieves CTCs from blood samples. The principle of CTC capturing is that tumor cells are larger and less deformable than normal blood cells. To evaluate the potential of utilizing Celsee PREP100 in CTC molecular analysis, we prepared prostate cancer cell lines PC3 and LNCaP, retrieved the captured cells and analyzed them using PCR amplicon sequencing. We were able to recover an average of 79% of 110‑1,100 PC3 and 60‑1,500 LNCaP cells, and detect the p.K139fs*3 deletion of the p53 gene in PC3 cells and p.T877A mutation of the androgen receptor gene in LNCaP cells. Next, we spiked these two types of cells into normal donor blood samples, captured the cells and analyzed them using PCR amplicon sequencing. The PC3 and LNCaP cells were captured and retrieved with the ratio of captured CTCs to the background leukocytes reaching 1:1.5 for PC3 and 1:2.9 for LNCaP cells. We further revealed that the p.K139fs*3 deletion and p.T877A mutation can be detected in the captured PC3 and LNCaP cells, respectively. We successfully validated this approach using clinical blood samples from patients with metastatic prostate cancer. Our results demonstrated a novel approach for CTC enrichment and illustrated the potential of CTC molecular characterization for diagnosis, prognosis and treatment selection of patients with metastatic malignancy.
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http://dx.doi.org/10.3892/or.2018.6342DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5983925PMC
June 2018

Pseudosarcomatous myofibroblastic proliferations of the genitourinary tract are genetically different from nodular fasciitis and lack USP6, ROS1 and ETV6 gene rearrangements.

Histopathology 2018 Aug 30;73(2):321-326. Epub 2018 May 30.

Department of Pathology and Laboratory Medicine and Henry Ford Cancer Institute, Henry Ford Health System, Detroit, MI, USA.

Aims: Pseudosarcomatous myofibroblastic proliferations of the genitourinary tract have a debatable relationship with inflammatory myofibroblastic tumour (generally lacking ALK rearrangement); however, they share several overlapping features with nodular fasciitis of soft tissue. As rearrangement of the USP6 gene has been recently recognised as a recurrent alteration in soft tissue nodular fasciitis, and several other alternative gene fusions have been recently recognised in inflammatory myofibroblastic tumour, the aim of this study was to investigate whether USP6, ROS1 or ETV6 rearrangements were present in these lesions (12 cases).

Methods And Results: Fluorescence in-situ hybridisation analysis was performed by the use of bacterial artificial chromosome-derived break-apart probes against USP6, ROS1, and ETV6. Two cases with adequate genetic material from recent paraffin tissue blocks were also tested by use of a solid tumour gene fusion detection assay via next-generation sequencing, targeting >50 known genes involved in recurrent fusions. None of the genitourinary pseudosarcomatous myofibroblastic proliferations was found to harbour USP6 (0/12), ROS1 (0/8) or ETV6 (0/7) rearrangements, and no gene fusions were detected in two cases studied by sequencing.

Conclusions: Despite overlap in histological and immunohistochemical features between pseudosarcomatous myofibroblastic proliferation and nodular fasciitis, these tumours lack the recently recognised USP6 rearrangements that occur in nodular fasciitis, as well as alternative fusions found in ALK-negative inflammatory myofibroblastic tumours. At present, this diagnosis remains based primarily on clinical, histological and immunohistochemical features.
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http://dx.doi.org/10.1111/his.13526DOI Listing
August 2018

Increased expression of EZH2 in Merkel cell carcinoma is associated with disease progression and poorer prognosis.

Hum Pathol 2017 09 21;67:78-84. Epub 2017 Jul 21.

Department of Dermatology, University of Michigan Health System, Ann Arbor, MI 48109, USA; Department of Pathology, University of Michigan Health System, Ann Arbor, MI 48109, USA. Electronic address:

Enhancer of zeste homolog 2 (EZH2) is a histone methyltransferase that affects tumorigenesis by epigenetic gene silencing. Merkel cell carcinoma (MCC) is a rare cutaneous neuroendocrine carcinoma that has a high risk of disease progression with nodal and distant metastases. Here, we evaluated EZH2 expression by immunohistochemistry in a cohort of 85 MCC tumors (29 primary tumors, 41 lymph node metastases, 13 in-transit metastases, and 2 distant metastases) with clinical follow-up. We show strong/moderate EZH2 expression in 54% of tumors. Importantly, weak expression of EZH2 in the primary tumor, but not nodal metastases, correlated with improved prognosis compared to moderate/strong EZH2 expression (5-year MCC-specific survival of 68% versus 22%, respectively, P=.024). In addition, EZH2 was expressed at higher levels in nodal metastases compared to primary tumors (P=.005). Our data demonstrate that EZH2 has prognostic value and may play an oncogenic role in MCC.
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http://dx.doi.org/10.1016/j.humpath.2017.07.009DOI Listing
September 2017

Age and Gender Associations of Virus Positivity in Merkel Cell Carcinoma Characterized Using a Novel RNA Hybridization Assay.

Clin Cancer Res 2017 Sep 12;23(18):5622-5630. Epub 2017 Jun 12.

Michigan Center for Translational Pathology, University of Michigan Health System, Ann Arbor, Michigan.

Merkel cell carcinoma (MCC) is a highly aggressive neuroendocrine tumor of the skin. Merkel cell polyomavirus (MCPyV) plays an oncogenic role in the majority of MCCs. Detection of MCPyV in MCCs has diagnostic utility and prognostic potential. We investigated whether RNAscope, an RNA hybridization (ISH) assay for detection of RNA transcripts in tissues, is useful for MCPyV detection. We applied an RNAscope probe targeting MCPyV T antigen transcripts on tissue microarrays (TMA) and whole-tissue sections encompassing 87 MCCs from 75 patients, 14 carcinomas of other types, and benign tissues. For comparison, qPCR was performed on 57 cases of MCC from 52 patients. RNA-ISH demonstrated the presence of MCPyV in 37 of 75 cases (49.3%). Notably, tumors from younger patients (<73 years) had a significantly higher virus positivity than those from elderly patients (≥73 years; 64.9% vs. 34.2%, = 0.011). Female patients had a higher positive rate of MCPyV than male patients (66.7% vs. 39.6%, = 0.032). Data from both RNA-ISH and qPCR were available for 57 samples. Considering MCPyV qPCR as the gold standard for determining MCPyV status, RNAscope had 100% sensitivity and 100% specificity. There was a strong correlation between qPCR copy number and RNA-ISH product score (Spearman correlation coefficient = 0.932, < 0.0001). RNA-ISH is comparably sensitive to qPCR for detection of MCPyV and allows for correlation with tissue morphology. This study also reveals a significant association between age, gender, and MCPyV positivity. .
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http://dx.doi.org/10.1158/1078-0432.CCR-17-0299DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5600832PMC
September 2017

Renal Cell Carcinoma With Chromosome 6p Amplification Including the TFEB Gene: A Novel Mechanism of Tumor Pathogenesis?

Am J Surg Pathol 2017 Mar;41(3):287-298

*Department of Pathology and Laboratory Medicine †Josephine Ford Cancer Institute ∥Department of Urology, Vattikutti Urology Institute, Henry Ford Health System ‡Department of Pathology, Wayne State University School of Medicine, Detroit #Michigan Center for Translational Pathology, University of Michigan, Ann Arbor, MI §Department of Pathology and Laboratory Medicine, Indiana University School of Medicine, Indianapolis, IN ¶Department of Environmental Biotechnology, Bharathidasan University, Thiruchirapalli, India.

Amplification of chromosome 6p has been implicated in aggressive behavior in several cancers, but has not been characterized in renal cell carcinoma (RCC). We identified 9 renal tumors with amplification of chromosome 6p including the TFEB gene, 3 by fluorescence in situ hybridization, and 6 from the Cancer Genome Atlas (TCGA) databases. Patients' ages were 28 to 78 years (median, 61 y). Most tumors were high stage (7/9 pT3a, 2/9 pN1). Using immunohistochemistry, 2/4 were positive for melanocytic markers and cathepsin K. Novel TFEB fusions were reported by TCGA in 2; however, due to a small composition of fusion transcripts compared with full-length transcripts (0.5/174 and 3.3/132 FPKM), we hypothesize that these represent secondary fusions due to amplification. Five specimens (4 TCGA, 1 fluorescence in situ hybridization) had concurrent chromosome 3p copy number loss or VHL deletion. However, these did not resemble clear cell RCC, had negative carbonic anhydrase IX labeling, lacked VHL mutation, and had papillary or unclassified histology (2/4 had gain of chromosome 7 or 17). One tumor each had somatic FH mutation and SMARCB1 mutation. Chromosome 6p amplification including TFEB is a previously unrecognized cytogenetic alteration in RCC, associated with heterogenous tubulopapillary eosinophilic and clear cell histology. The combined constellation of features does not fit cleanly into an existing tumor category (unclassified), most closely resembling papillary or translocation RCC. The tendency for high tumor stage, varied tubulopapillary morphology, and a subset with melanocytic marker positivity suggests the possibility of a unique tumor type, despite some variation in appearance and genetics.
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http://dx.doi.org/10.1097/PAS.0000000000000776DOI Listing
March 2017

Loss of p16 expression and copy number changes of CDKN2A in a spectrum of spitzoid melanocytic lesions.

Hum Pathol 2016 12 26;58:152-160. Epub 2016 Aug 26.

Department of Pathology, University of Michigan Health System, Ann Arbor, MI 48109; Department of Dermatology, University of Michigan Health System, Ann Arbor, MI 48109.

Spitzoid melanocytic lesions, including Spitz nevi (benign), spitzoid melanoma (malignant), and borderline atypical Spitz tumors (ASTs), frequently present challenges for accurate diagnosis and prognosis. Evaluation for loss of the tumor suppressor p16, encoded by CDKN2A gene on chromosome 9p21.3, has been proposed to be useful for evaluation of spitzoid melanocytic lesions. However, reports on the utility of p16 immunohistochemistry for spitzoid lesions have been conflicting, and few studies have directly compared p16 immunohistochemistry with fluorescence in situ hybridization (FISH) for CDKN2A genomic status. We analyzed a spectrum of benign (n=24), borderline (n=27), and malignant (n=19) spitzoid lesions for p16 protein expression by immunohistochemistry and CDKN2A copy number by FISH. Immunohistochemistry was evaluated by 2 scoring methods: H score and 2-tiered score (positive or negative for p16 loss). By immunohistochemistry, loss of p16 expression was not observed in Spitz nevi (0/24) but was seen in ASTs (7/27; 26%) and spitzoid melanomas (3/19; 16%). By H score, p16 expression was significantly higher in Spitz nevi relative to ASTs or spitzoid melanomas. Similarly, copy number aberrations of CDKN2A by FISH were absent in Spitz nevi but were found in 2 (9.5%) of 21 ASTs and 4 (33%) of 12 spitzoid melanomas. Our findings from this large cohort suggest that p16 aberrations are highly specific for borderline and malignant spitzoid neoplasms relative to Spitz nevi. Similar to ASTs, p16 loss in spitzoid melanomas may occur in the presence or absence of genomic CDKN2A loss.
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http://dx.doi.org/10.1016/j.humpath.2016.07.029DOI Listing
December 2016

Morpheaform Basal Cell Carcinomas With Areas of Predominantly Single-Cell Pattern of Infiltration: Diagnostic Utility of p63 and Cytokeratin.

Am J Dermatopathol 2016 Oct;38(10):744-50

Departments of *Pathology and †Dermatology, University of Michigan Health System, Ann Arbor MI; ‡Michigan Center for Translational Pathology, University of Michigan Health System, Ann Arbor MI; and §Department of Urology, Vattikutti Urology Institute, Henry Ford Health System, Detroit, MI.

Background: Morpheaform basal cell carcinoma (BCC) is a variant of BCC characterized by narrow strands and nests of basaloid cells with dense sclerotic stroma. The histologic extent often exceeds the clinical impression, leading to high recurrence rates after standard excision. The authors encountered a case with single-cell invasion distant from the main tumor. To date a systematic review of single-cell infiltration in morpheaform BCC has yet to be performed.

Design: Ten morpheaform BCCs, 10 nonmorpheaform aggressive BCCs, 5 desmoplastic trichoepitheliomas, and 2 microcystic adnexal carcinomas were identified by database search and confirmed on hematoxylin and eosin. Cases were evaluated by hematoxylin and eosin, immunohistochemical staining for p63, and (in a subset) broad-spectrum cytokeratin. Single-cell pattern was defined as individual cells, 2-cell clusters, or single-file invasion.

Results: Three types of single-cell pattern were identified: intratumoral (single cells within the main tumor mass), peripheral, and distant. Single cells were typically a minor component relative to larger tumor nodules and strands. Eight of the 10 cases of morpheaform BCC demonstrated areas of single-cell pattern: 3 intratumoral, 3 peripheral, and 2 with distant spread (0.75 and 1.0 mm from the main tumor). Eight of the 10 aggressive BCC demonstrated a peripheral single-cell pattern. Rare intratumoral single cells were identified in 3/5 desmoplastic trichoepitheliomas and 1/2 microcystic adnexal carcinomas.

Conclusion: Single-cell pattern is frequently a component of morpheaform BCC. Tumor cells at a significant distance from the main component were unique to morpheaform BCC. Thus, when evaluating margins for morpheaform BCC, increased caution is recommended, and immunohistochemical stains for p63 or cytokeratins may be helpful.
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http://dx.doi.org/10.1097/DAD.0000000000000541DOI Listing
October 2016

Fibroblast growth factor family aberrations as a putative driver of head and neck squamous cell carcinoma in an epidemiologically low-risk patient as defined by targeted sequencing.

Head Neck 2016 04 5;38 Suppl 1:E1646-52. Epub 2016 Feb 5.

Department of Otolaryngology-Head and Neck Surgery, University of Michigan Medical School, Ann Arbor, Michigan.

Background: Targeted sequencing of patients with epidemiologically low-risk (ELR) head and neck squamous cell carcinoma (HNSCC) could help identify novel drivers or lost suppressors leading to precision medicine protocols and improved survival rates.

Methods: A patient with ELR-HNSCC was selected for targeted sequencing. We then assessed next generation sequencing cohorts from the Oncomine Powertool Database, which contains pan-cancer data from The Cancer Genome Atlas (TCGA).

Results: Targeted sequencing revealed fibroblast growth factor receptor-1 (FGFR1) amplifications as a putative driver of the patient's tumor. Patients with HNSCC from TCGA data demonstrated fibroblast growth factor (FGF) family mutations, rearrangements, or amplifications in over 35% of HNSCC cases, with a statistically significant higher frequency in African American populations. FGF alterations were unique from activating phosphatidylinositol 3-kinase (PIK3CA) mutations.

Conclusion: Together, these data suggest that FGF signaling may be critical for a subset of patients with HNSCC independent of other known pathways and provides rationale for leveraging patients with ELR-HNSCC to define molecular subsets of high-risk HNSCC. © 2016 Wiley Periodicals, Inc. Head Neck 38: E1646-E1652, 2016.
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http://dx.doi.org/10.1002/hed.24292DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4844767PMC
April 2016

Role and regulation of coordinately expressed de novo purine biosynthetic enzymes PPAT and PAICS in lung cancer.

Oncotarget 2015 Sep;6(27):23445-61

Michigan Center for Translational Pathology, Ann Arbor, MI 48109, USA.

Cancer cells exhibit altered metabolism including aerobic glycolysis that channels several glycolytic intermediates into de novo purine biosynthetic pathway. We discovered increased expression of phosphoribosyl amidotransferase (PPAT) and phosphoribosylaminoimidazole carboxylase, phosphoribosylaminoimidazole succinocarboxamide synthetase (PAICS) enzymes of de novo purine biosynthetic pathway in lung adenocarcinomas. Transcript analyses from next-generation RNA sequencing and gene expression profiling studies suggested that PPAT and PAICS can serve as prognostic biomarkers for aggressive lung adenocarcinoma. Immunohistochemical analysis of PAICS performed on tissue microarrays showed increased expression with disease progression and was significantly associated with poor prognosis. Through gene knockdown and over-expression studies we demonstrate that altering PPAT and PAICS expression modulates pyruvate kinase activity, cell proliferation and invasion. Furthermore we identified genomic amplification and aneuploidy of the divergently transcribed PPAT-PAICS genomic region in a subset of lung cancers. We also present evidence for regulation of both PPAT and PAICS and pyruvate kinase activity by L-glutamine, a co-substrate for PPAT. A glutamine antagonist, 6-Diazo-5-oxo-L-norleucine (DON) blocked glutamine mediated induction of PPAT and PAICS as well as reduced pyruvate kinase activity. In summary, this study reveals the regulatory mechanisms by which purine biosynthetic pathway enzymes PPAT and PAICS, and pyruvate kinase activity is increased and exposes an existing metabolic vulnerability in lung cancer cells that can be explored for pharmacological intervention.
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http://dx.doi.org/10.18632/oncotarget.4352DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4695129PMC
September 2015

Extensive survey of STAT6 expression in a large series of mesenchymal tumors.

Am J Clin Pathol 2015 May;143(5):672-82

Pathology.

Objectives: Expression of strong nuclear STAT6 is thought to be a specific marker for solitary fibrous tumors (SFTs). Little is known about subtle expression patterns in other mesenchymal lesions.

Methods: We performed immunohistochemical studies against the C-terminus of STAT6 in tissue microarrays and whole sections, comprising 2366 mesenchymal lesions.

Results: Strong nuclear STAT6 was expressed in 285 of 2,021 tumors, including 206 of 240 SFTs, 49 of 408 well-differentiated/dedifferentiated liposarcomas, eight of 65 unclassified sarcomas, and 14 of 184 desmoid tumors, among others. Expression in SFTs was predominately limited to the nucleus. Other positive tumors typically expressed both nuclear and cytoplasmic STAT6. Complete absence of STAT6 was most common in pleomorphic liposarcoma and alveolar soft part sarcoma (60% and 72% cases negative, respectively).

Conclusions: Strong nuclear STAT6 is largely specific for SFTs. Physiologic low-level cytoplasmic/nuclear expression is common in mesenchymal neoplasia and is of uncertain significance.
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http://dx.doi.org/10.1309/AJCPN25NJTOUNPNFDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4505928PMC
May 2015

Molecular profiling of ETS and non-ETS aberrations in prostate cancer patients from northern India.

Prostate 2015 Jul 23;75(10):1051-62. Epub 2015 Mar 23.

Michigan Center for Translational Pathology, University of Michigan Medical School, Ann Arbor, Michigan.

Background: Molecular stratification of prostate cancer (PCa) based on genetic aberrations including ETS or RAF gene-rearrangements, PTEN deletion, and SPINK1 over-expression show clear prognostic and diagnostic utility. Gene rearrangements involving ETS transcription factors are frequent pathogenetic somatic events observed in PCa. Incidence of ETS rearrangements in Caucasian PCa patients has been reported, however, occurrence in Indian population is largely unknown. The aim of this study was to determine the prevalence of the ETS and RAF kinase gene rearrangements, SPINK1 over-expression, and PTEN deletion in this cohort.

Methods: In this multi-center study, formalin-fixed paraffin embedded (FFPE) PCa specimens (n = 121) were procured from four major medical institutions in India. The tissues were sectioned and molecular profiling was done using immunohistochemistry (IHC), RNA in situ hybridization (RNA-ISH) and/or fluorescence in situ hybridization (FISH).

Results: ERG over-expression was detected in 48.9% (46/94) PCa specimens by IHC, which was confirmed in a subset of cases by FISH. Among other ETS family members, while ETV1 transcript was detected in one case by RNA-ISH, no alteration in ETV4 was observed. SPINK1 over-expression was observed in 12.5% (12/96) and PTEN deletion in 21.52% (17/79) of the total PCa cases. Interestingly, PTEN deletion was found in 30% of the ERG-positive cases (P = 0.017) but in only one case with SPINK1 over-expression (P = 0.67). BRAF and RAF1 gene rearrangements were detected in ∼1% and ∼4.5% of the PCa cases, respectively.

Conclusions: This is the first report on comprehensive molecular profiling of the major spectrum of the causal aberrations in Indian men with PCa. Our findings suggest that ETS gene rearrangement and SPINK1 over-expression patterns in North Indian population largely resembled those observed in Caucasian population but differed from Japanese and Chinese PCa patients. The molecular profiling data presented in this study could help in clinical decision-making for the pursuit of surgery, diagnosis, and in selection of therapeutic intervention.
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http://dx.doi.org/10.1002/pros.22989DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4832366PMC
July 2015

Cutaneous basal cell carcinosarcomas: evidence of clonality and recurrent chromosomal losses.

Hum Pathol 2015 May 14;46(5):690-7. Epub 2015 Jan 14.

Department of Pathology, University of Michigan Health System, Ann Arbor MI 48109; Department of Dermatology, University of Michigan Health System, Ann Arbor MI 48109.

Cutaneous carcinosarcomas are heterogeneous group of tumors composed of malignant epithelial and mesenchymal components. Although mutation analyses have identified clonal changes between these morphologically disparate components in some subtypes of cutaneous carcinosarcoma, few cases have been analyzed thus far. To our knowledge, copy number variations (CNVs) and copy-neutral loss of heterozygosity (CN-LOH) have not been investigated in cutaneous carcinosarcomas. We analyzed 4 carcinosarcomas with basal cell carcinoma and osteosarcomatous components for CNVs/CN-LOH by comparative genomic hybridization/single-nucleotide polymorphism array, TP53 hot spot mutations by polymerase chain reaction and Sanger sequencing, and TP53 genomic rearrangements by fluorescence in situ hybridization. All tumors displayed multiple CNV/CN-LOH events (median, 7.5 per tumor). Three of 4 tumors displayed similar CNV/CN-LOH patterns between the epithelial and mesenchymal components within each tumor, supporting a common clonal origin. Recurrent changes included allelic loss at 9p21 (CDKN2A), 9q (PTCH1), and 17p (TP53). Allelic losses of chromosome 16 including CDH1 (E-cadherin) were present in 2 tumors and were restricted to the sarcomatous component. TP53 mutation analysis revealed an R248L mutation in both epithelial and mesenchymal components of 1 tumor. No TP53 rearrangements were identified. Our findings indicate that basal cell carcinosarcomas harbor CNV/CN-LOH changes similar to conventional basal cell carcinoma, with additional changes including recurrent 9p21 losses and a relatively high burden of copy number changes. In addition, most cutaneous carcinosarcomas show evidence of clonality between epithelial and mesenchymal components.
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http://dx.doi.org/10.1016/j.humpath.2015.01.006DOI Listing
May 2015

Clear cell melanoma: a cutaneous clear cell malignancy.

Arch Pathol Lab Med 2014 Oct;138(10):1328-36

From the Departments of Pathology (Drs Pletneva, Andea, Palanisamy, Betz, Wang, Patel, Fullen, and Harms, and Ms Carskadon) and Dermatology (Drs Andea Patel, Fullen, and Harms), and the Michigan Center for Translational Pathology (Drs Palanisamy and Harms, and Ms Carskadon), University of Michigan Medical Center, Ann Arbor.

Clear cell melanoma is a rare clear cell malignancy. Accurate diagnosis of clear cell melanoma requires integration of immunohistochemical and morphologic findings, with molecular studies to rule out clear cell sarcoma. The differential diagnosis includes melanoma, carcinoma, perivascular epithelioid cell tumor, and epidermotropic clear cell sarcoma. We use a case of a lesion on the helix of an 86-year-old man as an example. Histologic examination revealed an ulcerated clear cell malignant tumor. Tumor cell cytoplasm contained periodic acid-Schiff-positive, diastase-sensitive glycogen. Tumor cells showed positive labeling for S100, HMB-45, and Melan-A, and negative labeling for cytokeratins, p63, and smooth muscle actin. Molecular studies demonstrated BRAF V600E mutation, copy gains at the 6p25 (RREB1) and 11q13 (CCND1) loci, and absence of EWSR1-ATF1 fusion. These findings supported a diagnosis of clear cell melanoma. The rare pure clear cell morphology occurs due to accumulation of intracytoplasmic glycogen. We review the differential diagnosis of clear cell melanoma and describe the utility of immunohistochemical and molecular studies in confirming this diagnosis.
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http://dx.doi.org/10.5858/arpa.2014-0307-CCDOI Listing
October 2014
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