Meora Feinmesser

MicroRNAs accurately identify cancer tissue origin

MicroRNAs (miRNAs) belong to a class of noncoding, regulatory RNAs that is involved in oncogenesis and shows remarkable tissue specificity. Their potential for tumor classification suggests they may be used in identifying the tissue in which cancers of unknown primary origin arose, a major clinical problem. We measured miRNA expression levels in 400 paraffin-embedded and fresh-frozen samples from 22 different tumor tissues and metastases. We used miRNA microarray data of 253 samples to construct a transparent classifier based on 48 miRNAs. Two-thirds of samples were classified with high confidence, with accuracy >90%. In an independent blinded test-set of 83 samples, overall high-confidence accuracy reached 89%. Classification accuracy reached 100% for most tissue classes, including 131 metastatic samples. We further validated the utility of the miRNA biomarkers by quantitative RT-PCR using 65 additional blinded test samples. Our findings demonstrate the effectiveness of miRNAs as biomarkers for tracing the tissue of origin of cancers of unknown primary origin.

Second neoplasms in patients with Merkel cell carcinoma

Merkel cell carcinoma (MCC) has been associated with a high incidence of other skin tumors and hematological malignancies. The purpose of this study was to analyze data from the Israel Cancer Registry regarding the incidence of second neoplasms in patients with MCC and their impact on survival.

Validation of a microRNA-based qRT-PCR test for accurate identification of tumor tissue origin

Identification of the tissue of origin of a tumor is vital to its management. Previous studies showed tissue-specific expression patterns of microRNA and suggested that microRNA profiling would be useful in addressing this diagnostic challenge. MicroRNAs are well preserved in formalin-fixed, paraffin-embedded (FFPE) samples, further supporting this approach. To develop a standardized assay for identification of the tissue origin of FFPE tumor samples, we used microarray data from 504 tumor samples to select a shortlist of 104 microRNA biomarker candidates. These 104 microRNAs were profiled by proprietary quantitative reverse transcriptase polymerase chain reaction (qRT–PCR) on 356 FFPE tumor samples. A total of 48 microRNAs were chosen from this list of candidates and used to train a classifier. We developed a clinical test for the identification of the tumor tissue of origin based on a standardized protocol and defined the classification criteria. The test measures expression levels of 48 microRNAs by qRT–PCR, and predicts the tissue of origin among 25 possible classes, corresponding to 17 distinct tissues and organs. The biologically motivated classifier combines the predictions generated by a binary decision tree and K-nearest neighbors (KNN). The classifier was validated on an independent, blinded set of 204 FFPE tumor samples, including nearly 100 metastatic tumor samples. The test predictions correctly identified the reference diagnosis in 85% of the cases. In 66% of the cases the two algorithm predictions (tree and KNN) agreed on a single-tissue origin, which was identical to the reference diagnosis in 90% of cases. Thus, a qRT–PCR test based on the expression profile of 48 tissue-specific microRNAs allows accurate identification of the tumor tissue of origin.

Primary cutaneous apocrine carcinoma: A clinico-pathologic analysis of 24 cases

Primary cutaneous apocrine carcinoma is a rare malignancy. This study of 24 examples suggests that the prognosis is not always poor and that grading criteria devised for breast carcinoma may have utility in this group of malignancies. Furthermore, steroid receptor expression should be investigated in these tumors, particularly if a tumor is unlikely to be controlled by surgery alone.

A Second-Generation MicroRNA-Based Assay for Diagnosing Tumor Tissue Origin

BACKGROUND Cancers of unknown primary origin (CUP) constitute 3%-5% (50,000 to 70,000 cases) of all newly diagnosed cancers per year in the United States. Including cancers of uncertain primary origin, the total number increases to 12%-15% (180,000 to 220,000 cases) of all newly diagnosed cancers per year in the United States. Cancers of unknown/uncertain primary origins present major diagnostic and clinical challenges because the tumor tissue of origin is crucial for selecting optimal treatment. MicroRNAs are a family of noncoding, regulatory RNA genes involved in carcinogenesis. MicroRNAs that are highly stable in clinical samples and tissue specific serve as ideal biomarkers for cancer diagnosis. Our first-generation assay identified the tumor of origin based on 48 microRNAs measured on a quantitative real-time polymerase chain reaction platform and differentiated 25 tumor types. METHODS We present here the development and validation of a second-generation assay that identifies 42 tumor types using a custom microarray. A combination of a binary decision-tree and a k-nearest-neighbor classifier was developed to identify the tumor of origin based on the expression of 64 microRNAs. RESULTS Overall assay sensitivity (positive agreement), measured blindly on a validation set of 509 independent samples, was 85%. The sensitivity reached 90% for cases in which the assay reported a single answer (>80% of cases). A clinical validation study on 52 true CUP patients showed 88% concordance with the clinicopathological evaluation of the patients. CONCLUSION The abilities of the assay to identify 42 tumor types with high accuracy and to maintain the same performance in samples from patients clinically diagnosed with CUP promise improved utility in the diagnosis of cancers of unknown/uncertain primary origins.

Expression of the apoptosis-related oncogenes bcl-2, bax, and p53 in Merkel cell carcinoma: Can they predict treatment response and clinical outcome?

Chemotherapy and radiation therapy act predominantly through the induction of apoptosis in malignancies. Merkel cell carcinoma, an aggressive malignancy with prominent apoptosis, has proved to be sensitive to both modes to a certain degree. We used immunohistochemical methods to examine 25 Merkel cell carcinomas and 8 of their lymph node metastases to assess the status of the antiapoptotic gene bcl-2 and 2 proapoptotic genes, wild-type p53 and bax. All tumors showed prominent bax immunopositivity; 76% were positive for bcl-2, and only 28% were positive for p53, the latter presumably reflecting mutated p53. No statistically significant relationship was found between tumor immunopositivity and therapy response or survival. The widespread bax immunopositivity and the apparently low rate of p53 mutations, as suggested by the low rate of p53 immunopositivity, may be related to the presence of prominent apoptosis in Merkel cell carcinoma. The finding of bcl-2 immunopositivity in 76% of the tumors suggests that some of the tumor cells may be resistant to apoptosis-inducing agents.

Applicability of the Sentinel Node Technique to Merkel Cell Carcinoma

Background. Merkel cell carcinoma (MCC) resembles malignant melanoma in several ways. Both are cutaneous lesions of the same embryonic origin. Both have an unpredictable biologic behavior, early regional lymph node involvement, early distant metastases, and high recurrence rate. Objective. To apply the sentinel node technique described for melanoma to MCC in light of the common biologic features of these two tumors. Methods. Preoperative lymphoscintigraphy, intraoperative lymphatic mapping, and sentinel node biopsy and frozen section histology were performed to guide the surgical treatment of three patients with MCC. Results. Application of this approach in patients with MCC is feasible, reproducible, and seems reliable. Conclusion. The use of the sentinel node technique for MCC will reduce the number of unnecessary lymphadenectomies, will enable identification of microscopic metastases to lymph nodes, and will improve the stratification and accrual of patients into adjuvant treatment protocols. It may even lead to a survival benefit.

Occasional involvement of the ovary in Ewing sarcoma

BACKGROUND Ewing sarcoma (EWS) is a highly metastatic malignancy in young patients. Ovarian cryopreservation is often an option for fertility preservation in cancer patients of reproductive age, specifically in minors. Thus, the possibility of ovarian involvement in EWS needs to be elucidated. METHODS Eight patients aged 13-20 years with EWS participated in the study. Ovarian samples were fixed and prepared for light microscopy, and frozen in liquid nitrogen for RNA extraction followed by RT-PCR. Histological studies, including immunostaining for the adhesion receptor CD99, were used to detect histopathological features. Sensitive molecular methods were used to detect translocations causing the formation of tumor-specific EWS-Friend leukemia virus integration site 1 fusion gene (EWS-FLI1). RESULTS In seven patients, there was no evidence of EWS in the ovaries from pathological/molecular studies. However, in one patient, the RT-PCR showed the EWS translocation, although there was no pathological evidence. CONCLUSIONS Ovarian involvement is possible in EWS. Therefore, in patients with EWS ovarian tissue should be examined for traces of malignancy at both the pathological and molecular levels prior to the grafting of cryopreserved tissue in order to minimize the risk of reseeding the cancer.

c-kit expression in primary and metastatic merkel cell carcinoma.

Merkel cell carcinoma (MCC) is an aggressive neuroendocrine tumor of the skin that is associated with a high incidence of recurrence and metastasis. The therapeutic arsenal for this malignancy is limited and once it spreads, there is no effective treatment. c-kit expression has been demonstrated previously in primary MCCs thus raising the possibility of treating MCCs with imatinib mesylate, the tyrosine kinase inhibitor that has shown promise in the management of c-kit expressing tumors. In this study we examine 25 additional primary MCCs and also 6 of their lymph node metastases. Formalin-fixed, paraffin-embedded tissues were stained immunohistochemically with an antibody directed against the KIT receptor. Percentage and intensity of staining were analyzed semiquantitatively using a three-tiered system. Twenty-one of the 25 (84%) primary tumors stained positively for KIT, of which 14 (67%) showed widespread positivity. Five of the 6 lymph nodes (83%) were similarly positive. High mitotic rate and vascular invasion in the primary tumors tended to be associated with prominent staining in the lymph node metastases. No association was found between c-kit expression and outcome. We confirm that the majority of primary MCCs express c-kit and further find that metastases are positive for the KIT receptor as well. Thus, c-kit expression may be an early event in the transformation of MCC, but not a marker for tumor progression.

Telomere length is a prognostic factor in neuroblastoma

Maintenance of telomeres, in most instances by reactivation of telomerase, is obligatory for the indefinite proliferation of tumor cells. The objective of this study was to evaluate telomere length and telomerase activity (TA) as markers for progression and prognosis in neuroblastoma.