Andrew T. Turk

Pax8: A marker for carcinoma of Müllerian origin in serous effusions

PAX8 is a nuclear transcription factor with limited expression in normal and neoplastic tissues in a cell lineage‐dependent manner. PAX8 has been detected in embryonic Müllerian ducts, human fallopian tubes, and ovarian carcinomas. However, little is known about its expression in other areas of the female genital tract. In this study, we used immunohistochemistry (IHC) to examine PAX8 expression in the normal uterine corpus and cervix, malignant tumors arising from these sites, and malignant effusions. We reported here that PAX8 was also detected in endometrial epithelial cells and endocervical glands, with a lower expression level in the latter, but not in the stromal cells of these areas. All endometrial carcinomas (N = 52) were positive for PAX8, whereas endocervical adenocarcinomas (N = 5) and uterine leiomyosarcomas (N = 3) were negative for PAX8. PAX8 was detected in 70% (22/31) and 68.8% (11/16) of metastatic carcinomas of the ovary and endometrium in serous effusions, respectively. No PAX8 was detected in carcinomas of nongynecologic origin or noncarcinomas (N = 71) in serous effusions except in one renal‐cell carcinoma and one carcinoma of unknown primary in a woman. In addition, papillary serous carcinomas of the peritoneum (N = 10) were diffusely positive for PAX8, implying a Müllerian origin similar to malignant tumors in the female genital tract. Our findings suggest that PAX8 is an additional IHC marker for carcinomas of Müllerian origin hence we recommend including PAX8 for evaluation of malignant serous effusions in women, especially when tumors of Müllerian origin are in the differential diagnosis. Diagn. Cytopathol. 2011.

Loss of PTEN Expression Is Associated with Poor Prognosis in Patients with Intraductal Papillary Mucinous Neoplasms of the Pancreas

Purpose: Previously, we reported PIK3CA gene mutations in high-grade intraductal papillary mucinous neoplasms (IPMN). However, the contribution of phosphatidylinositol-3 kinase pathway (PI3K) dysregulation to pancreatic carcinogenesis is not fully understood and its prognostic value unknown. We investigated the dysregulation of the PI3K signaling pathway in IPMN and its clinical implication. Experimental Design: Thirty-six IPMN specimens were examined by novel mutant-enriched sequencing methods for hot-spot mutations in the PIK3CA and AKT1 genes. PIK3CA and AKT1 gene amplifications and loss of heterozygosity at the PTEN locus were also evaluated. In addition, the expression levels of PDPK1/PDK1, PTEN, and Ki67 were analyzed by immunohistochemistry. Results: Three cases carrying the E17K mutation in the AKT1 gene and one case harboring the H1047R mutation in the PIK3CA gene were detected among the 36 cases. PDK1 was significantly overexpressed in the high-grade IPMN versus low-grade IPMN (P = 0.034) and in pancreatic and intestinal-type of IPMN versus gastric-type of IPMN (P = 0.020). Loss of PTEN expression was strongly associated with presence of invasive carcinoma and poor survival in these IPMN patients (P = 0.014). Conclusion: This is the first report of AKT1 mutations in IPMN. Our data indicate that oncogenic activation of the PI3K pathway can contribute to the progression of IPMN, in particular loss of PTEN expression. This finding suggests the potential employment of PI3K pathway-targeted therapies for IPMN patients. The incorporation of PTEN expression status in making surgical decisions may also benefit IPMN patients and should warrant further investigation. Clin Cancer Res; 19(24); 6830–41. ©2013 AACR.

Implementation of next generation sequencing into pediatric hematology-oncology practice: moving beyond actionable alterations

BackgroundMolecular characterization has the potential to advance the management of pediatric cancer and high-risk hematologic disease. The clinical integration of genome sequencing into standard clinical practice has been limited and the potential utility of genome sequencing to identify clinically impactful information beyond targetable alterations has been underestimated.MethodsThe Precision in Pediatric Sequencing (PIPseq) Program at Columbia University Medical Center instituted prospective clinical next generation sequencing (NGS) for pediatric cancer and hematologic disorders at risk for treatment failure. We performed cancer whole exome sequencing (WES) of patient-matched tumor-normal samples and RNA sequencing (RNA-seq) of tumor to identify sequence variants, fusion transcripts, relative gene expression, and copy number variation (CNV). A directed cancer gene panel assay was used when sample adequacy was a concern. Constitutional WES of patients and parents was performed when a constitutionally encoded disease was suspected. Results were initially reviewed by a molecular pathologist and subsequently by a multi-disciplinary molecular tumor board. Clinical reports were issued to the ordering physician and posted to the patient’s electronic medical record.ResultsNGS was performed on tumor and/or normal tissue from 101 high-risk pediatric patients. Potentially actionable alterations were identified in 38% of patients, of which only 16% subsequently received matched therapy. In an additional 38% of patients, the genomic data provided clinically relevant information of diagnostic, prognostic, or pharmacogenomic significance. RNA-seq was clinically impactful in 37/65 patients (57%) providing diagnostic and/or prognostic information for 17 patients (26%) and identified therapeutic targets in 15 patients (23%). Known or likely pathogenic germline alterations were discovered in 18/90 patients (20%) with 14% having germline alternations in cancer predisposition genes. American College of Medical Genetics (ACMG) secondary findings were identified in six patients.ConclusionsOur results demonstrate the feasibility of incorporating clinical NGS into pediatric hematology-oncology practice. Beyond the identification of actionable alterations, the ability to avoid ineffective/inappropriate therapies, make a definitive diagnosis, and identify pharmacogenomic modifiers is clinically impactful. Taking a more inclusive view of potential clinical utility, 66% of cases tested through our program had clinically impactful findings and samples interrogated with both WES and RNA-seq resulted in data that impacted clinical decisions in 75% of cases.

Gleason score concordance on biopsy-confirmed prostate cancer: is pathological re-evaluation necessary prior to radical prostatectomy?

Study Type – Diagnosis (case series)

Oncogenic PIK3CA Mutation and Dysregulation in Human Salivary Duct Carcinoma

Salivary duct carcinoma (SDC) is an aggressive malignant tumor with a high mortality, which resembles high-grade breast ductal carcinoma in morphology. The parotid gland is the most common location. Its molecular genetic characteristics remain largely unknown. We have previously reported high incidence of PIK3CA somatic mutations in head and neck squamous cell carcinoma, particularly in pharyngeal cancers. Here we examined the PIK3CA gene expression status and hotspot mutations in six cases of SDC by immunohistochemistry and genomic DNA sequencing. Immunohistochemistry showed that PIK3CA expression was elevated in all six patients with SDC. By DNA sequencing, two hotspot mutations of the PIK3CA gene, E545K (exon 9) and H1047R (exon 20), were identified in two of the six cases. Our results support that oncogenic PIK3CA is upregulated and frequently mutated in human SDC, adding evidence that PIK3CA oncogenic pathway is critical in the tumorigenesis of SDC, and may be a plausible drug target for this rare disease.

A case study of an integrative genomic and experimental therapeutic approach for rare tumors: identification of vulnerabilities in a pediatric poorly differentiated carcinoma.

BackgroundPrecision medicine approaches are ideally suited for rare tumors where comprehensive characterization may have diagnostic, prognostic, and therapeutic value. We describe the clinical case and molecular characterization of an adolescent with metastatic poorly differentiated carcinoma (PDC). Given the rarity and poor prognosis associated with PDC in children, we utilized genomic analysis and preclinical models to validate oncogenic drivers and identify molecular vulnerabilities.MethodsWe utilized whole exome sequencing (WES) and transcriptome analysis to identify germline and somatic alterations in the patient’s tumor. In silico and in vitro studies were used to determine the functional consequences of genomic alterations. Primary tumor was used to generate a patient-derived xenograft (PDX) model, which was used for in vivo assessment of predicted therapeutic options.ResultsWES revealed a novel germline frameshift variant (p.E1554fs) in APC, establishing a diagnosis of Gardner syndrome, along with a somatic nonsense (p.R790*) APC mutation in the tumor. Somatic mutations in TP53, MAX, BRAF, ROS1, and RPTOR were also identified and transcriptome and immunohistochemical analyses suggested hyperactivation of the Wnt/ß-catenin and AKT/mTOR pathways. In silico and biochemical assays demonstrated that the MAX p.R60Q and BRAF p.K483E mutations were activating mutations, whereas the ROS1 and RPTOR mutations were of lower utility for therapeutic targeting. Utilizing a patient-specific PDX model, we demonstrated in vivo activity of mTOR inhibition with temsirolimus and partial response to inhibition of MEK.ConclusionsThis clinical case illustrates the depth of investigation necessary to fully characterize the functional significance of the breadth of alterations identified through genomic analysis.

Micropapillary urothelial carcinoma: Cytologic features in a retrospective series of urine specimens

Background: The micropapillary variant of urothelial carcinoma (uPC) is a rare variant of urothelial carcinoma that carries a poor prognosis. Definitive surgery may represent optimal management of low stage tumors. Urine cytology is indispensable in the screening and follow-up of urinary tract cancer. However, cytopathological criteria for diagnosis of uPC and its differentiation from conventional urothelial carcinoma (CUC) are not well-defined. Materials and Methods: Twenty-five cases of histologically confirmed micropapillary uPC from 21 patients were compared to 25 cases of histologically confirmed high-grade CUC. Results: In uPC cases, cell clusters were identified in 13 of 25 specimens from 10 patients. Six of the 13 specimens containing cell clusters corresponded to surgical pathology specimens in which micropapillary carcinoma accounted for at least 50% of total carcinoma. In contrast, only 1 of the 12 urine specimens devoid of cell clusters corresponded to surgical specimens in which micropapillary carcinoma accounted for at least 50% of total carcinoma. Cytomorphologic features of urinary specimens from patients with histologically confirmed micropapillary carcinoma were generally similar to those from patients with high-grade CUC, making it difficult to distinguish these entities in exfoliative urine specimens. Conclusions and Summary: Further investigation of the core cytopathological characteristics of uPC is warranted to refine its diagnostic criteria by exfoliative urine cytology.

Pathological confirmation of nerve-sparing types performed during robot-assisted radical prostatectomy (RARP): Pathological confirmation of nerve-sparing during RARP

Incremental nerve‐sparing techniques (NSTs) improve postoperative erectile function after robot‐assisted radical prostatectomy (RARP). However, there are no studies to date that histologically confirm the surgeon intended NST. Thus, in the present study, we histologically confirmed that the surgeon performed the nerve preservation as his intended NSTs during RARP. Also, we found that there was more variability in fascia width outcome on the left side compared with the right. Therefore, when performing nerve preservation on the surgeons non‐dominant side, we need to pay more close attention.

Clinical Genomic Profiling of a Diverse Array of Oncology Specimens at a Large Academic Cancer Center: Identification of Targetable Variants and Experience with Reimbursement

Large cancer panels are being increasingly used in the practice of precision medicine to generate genomic profiles of tumors with the goal of identifying targetable variants and guiding eligibility for clinical trials. To facilitate identification of mutations in a broad range of solid and hematological malignancies, a 467-gene oncology panel (Columbia Combined Cancer Panel) was developed in collaboration with pathologists and oncologists and is currently available and in use for clinical diagnostics. Herein, we share our experience with this testing in an academic medical center. Of 255 submitted specimens, which encompassed a diverse range of tumor types, we were able to successfully sequence 92%. The Columbia Combined Cancer Panel assay led to the detection of a targetable variant in 48.7% of cases. However, although we show good clinical performance and diagnostic yield, third-party reimbursement has been poor. Reimbursement from government and third-party payers using the 81455 Current Procedural Terminology code was at 19.4% of billed costs, and 55% of cases were rejected on first submission. Likely contributing factors to this low level of reimbursement are the delays in valuation of the 81455 Current Procedural Terminology code and in establishing national or local coverage determinations. In the absence of additional demonstrations of clinical utility and improved patient outcomes, we expect the reimbursement environment will continue to limit the availability of this testing more broadly.

Precision Medicine in Children and Young Adults with Hematologic Malignancies and Blood Disorders: The Columbia University Experience

Background The advent of comprehensive genomic profiling has markedly advanced the understanding of the biology of pediatric hematological malignancies, however, its application to clinical care is still unclear. We present our experience integrating genomic data into the clinical management of children with high-risk hematologic malignancies and blood disorders and describe the broad impact that genomic profiling has in multiple aspects of patient care. Methods The Precision in Pediatric Sequencing Program at Columbia University Medical Center instituted prospective clinical next-generation sequencing (NGS) for high-risk malignancies and blood disorders. Testing included cancer whole exome sequencing (WES) of matched tumor-normal samples or targeted sequencing of 467 cancer-associated genes, when sample adequacy was a concern, and tumor transcriptome (RNA-seq). A multidisciplinary molecular tumor board conducted interpretation of results and final tiered reports were transmitted to the electronic medical record according to patient preferences. Results Sixty-nine samples from 56 patients with high-risk hematologic malignancies and blood disorders were sequenced. Patients carried diagnoses of myeloid malignancy (n = 25), lymphoid malignancy (n = 25), or histiocytic disorder (n = 6). Six patients had only constitutional WES, performed for a suspicion of an inherited predisposition for their disease. For the remaining 50 patients, tumor was sequenced with matched normal tissue when available. The mean number of somatic variants per sample was low across the different disease categories (2.85 variants/sample). Interestingly, a gene fusion was identified by RNA-seq in 58% of samples who had adequate RNA available for testing. Molecular profiling of tumor tissue led to clinically impactful findings in 90% of patients. Forty patients (80%) had at least one targetable gene variant or fusion identified in their tumor tissue; however, only seven received targeted therapy. Importantly, NGS findings contributed to the refinement of diagnosis and prognosis for 34% of patients. Known or likely pathogenic germline alterations were discovered in 24% of patients involving cancer predisposition genes in 12% of cases. Conclusion Incorporating whole exome and transcriptome profiling of tumor and normal tissue into clinical practice is feasible, and the value that comprehensive testing provides extends beyond the ability to target-specific mutations.