Ashraf Khan

Application of an immunodiagnostic method for improving preoperative diagnosis of nodular thyroid lesions

BACKGROUNDnThyroid cancer is the most common endocrine malignant disease, but preoperative diagnosis remains a challenge. Fine-needle aspiration cytology has greatly improved the clinical management of thyroid nodules, but the preoperative characterisation of follicular lesions is very difficult. Many patients are thus referred to surgery more for diagnosis than for therapeutic necessity. We undertook an international multicentre study to assess the usefulness of immunohistocytochemical staining for two potential markers of malignant thyrocytes.nnnMETHODSnExpression of galectin-3 and CD44v6 was tested on 1009 thyroid lesions (tissue specimens and cytological cell-blocks) and 226 fresh cytological samples obtained preoperatively by ultrasound-guided fine-needle aspiration of thyroid nodules (prospective analysis). The test used monoclonal antibodies specific for CD44v6 and galectin-3, the indirect avidin-biotin complex immunoperoxidase method, and 3-amino-9-ethyl-carbazole as substrate.nnnFINDINGSnThe sensitivity, specificity, positive predictive value, and diagnostic accuracy of this test method (for coexpression of the two markers) in the prospective analysis were 88%, 98%, 91%, and 97%, respectively. The sensitivity and specificity of galectin-3 immunodetection alone in discriminating benign from malignant thyroid lesions were more than 99% and 98% respectively, and the positive predictive value and diagnostic accuracy were 92% and 99%.nnnINTERPRETATIONnThe integration of galectin-3 immunostaining with conventional cytomorphological and clinical diagnostic procedures represents a sensitive and reliable diagnostic approach for preoperative identification of thyroid carcinomas. This test method improves the diagnostic accuracy of conventional cytology and provides the molecular basis for a new nosological assignation of the not yet classified thyroid neoplasms of indeterminate malignant behaviour.

P504S: a new molecular marker for the detection of prostate carcinoma.

* ون ي ،لوئسم هدنس نسح ناديم ،نارهت ،انيس ناتسراميب ،دابآ يژولوتاپ شخب نفلت : 9 66701041 email: ahmadise@tums.ac.ir فده و هنيمز : رب ديكات دوز فشك طرس ماگنه ب تاتسورپ نا ه كمك ارت يفارگونوسارتلوا سن لاـتكر ،ينزوـس يسـپويب و تسيژولوتاپ ار اه ناطرس صيخشت لضعم اب تـسا هدومن هجاوم كچوك ياه . ارـيخا ̋ زا ركراـم P504S صيخشـت تـهج يسپويب رد ناطرس يعطق تسا هدش هدافتسا كچوك ياه . يسررب شور : گنر ركرام يزيمآ P504S يارب 70 سپويب هنومن ي و تاتسورپ ينزوس شش زر هنومن ك هك ارجم قيرط زا نويس يگمه نوناك يواح ياه كوكشم ) پيتآ ـ ي ك ( و دـندوب زـين 40 هنومن تاتسورپ يعطق ناطرس ، لماش تشه لولس ياراد هنومن فك ياه دولآ (foamy) ب ه دمآ لمع . هتفاي اـه : 36 زا هـنومن 40 ،يعطق ناطرس يسپويب زا يتاجرد گنر يارب يريذپ P504S دـنداد ناشن ) تيساسـح 90 (% ؛ ود و كـچوك ناطرـس ود لولس اب ناطرس فك ياه گنر دقاف دولآ دندوب يريذپ . عومجم زا 76 ،كوكشم دروم 18 ـ ب دروم ه لـخاد يزلاپوـئن ناوـنع يپا لااب هجرد يلايلت (HGPIN) دش هتخانش دن هك 16 دروم اهنآ گنر دنداد ناشن رشتنم طسوتم يريذپ . زا 58 دروم يقاب هدـنام ياراد يلورپ نويسارف كچوك ددغ آ يت كيپ 14 ًايوق دروم ب ه ن ف دوب ناطرس ع هك راهچ گـنر دـقاف اـهنآ دروـم يريذـپ P504S دندوب . رد نيب 44 پيتآ دروم ي ك ب ه عفن شوخ ميخ ي ، 14 پيتآ يموندآ يزلاپرپيه دروم ي ك اب ود گنر دروم فيعـض يريذـپ يعضوم و ود ب يپيتآ دروم ه ب يباتوترپ لابند ا كي گنر دروم و دش هدهاشم يعضوم يريذپ 28 رـگيد هـنومن P504S يـفنم دندوب . هجيتن يريگ : تيساسح P504S يم روصت ًلابق هچنآ زا تاتسورپ ناطرس فشك يارب نيياپ دش تـسا رت . يـفنم يـّقلت ندش ناطرس نكمم كچوك ياه ا ب تس ه گـنر ينوگمهاـن ليلد دـشاب يريذـپ . نويـسارفيلورپ رد كـچوك ددـغ تآ پي ـ ي ،ك گنر رشتنم و تبثم يريذپ P504S گنر مدع يلو تسا ناطرس صيخشت يايوگ ًايوق يـمن در ارنآ يريذپ دـنك . تيساسـحThe ability to diagnose prostate carcinoma would be improved by the detection of a tumor-associated antigen. P504S, a cytoplasmic protein, was recently identified by cDNA library subtraction in conjunction with high throughput microarray screening from prostate carcinoma. The aim of this study was to establish the pattern of expression of P504S in prostate carcinoma and benign prostatic tissue. A total of 207 cases, including 137 cases of prostate carcinoma and 70 cases of benign prostate, from prostatectomies (n = 77), prostate needle biopsies (n = 112), and transurethral prostate resections (n = 18) were examined by immunocytochemistry for P504S. P504S showed strong cytoplasmic granular staining in 100% of prostate carcinomas regardless of Gleason scores and diffuse (>75% of tumor) staining in 92% of cases. In contrast, 171 of 194 (88%) of benign prostates, including 56 of 67 (84%) benign prostate cases and 115 of 127 (91%) cases of benign glands adjacent to cancers were negative for P504S. The remainders of benign prostates were focally and weakly positive for P504S. The staining pattern of these normal glands was different and easily distinguishable from that observed in prostate carcinoma. Expression of P504S was not found in basal cell hyperplasia, urothelial cells/metaplasia and small atrophic glands that may mimic prostate carcinoma. Our findings indicate that P504S is a highly sensitive and specific positive marker for prostate carcinoma.

The Role of Cell Cycle Regulatory Protein, Cyclin D1, in the Progression of Thyroid Cancer

Cell cycle progression is facilitated by cyclin-dependent kinases that are activated by cyclins including cyclin D1 and inactivated by cyclin-dependent kinase inhibitors (CDKIs) such as p27. Our previous studies have demonstrated decreased p27 expression in both papillary and more aggressive carcinomas of the thyroid compared to thyroid adenoma and almost similar level of cyclin D1 expression between thyroid adenoma and papillary carcinoma. These results indicate that CDKIs may have an important role in the carcinogenesis of the thyroid and that they probably have a limited role in malignant progression of the thyroid cancer. The role of cyclin D1 in malignant progression of thyroid carcinoma has yet to be established. We studied the expression of cyclin D1 by immunohistochemistry in 34 cases of conventional papillary carcinoma (CPC), 10 cases of minimally invasive follicular carcinoma (MIFC), and 32 cases of more aggressive thyroid carcinoma (ATC), which included 11 tall cell variants, one columnar cell variant of papillary carcinoma, seven insular carcinomas, and 13 anaplastic carcinomas. Cyclin D1 staining was classified by staining score as 0, negative; 1+, less than 25%; 2+, 25 to 50%; and 3+, more than 50% tumor cells staining positive. Kruskal-Wallis one-way ANOVA and Wilcoxon Rank Sum/Mann-Whitney U Test was used to assess the difference in the expression of cyclin D1 between the study groups. Twenty-eight out of the 34 CPCs were cyclin D1 positive, 24 (70%) were 1+, 3 (9%) were 2+, and one (3%) were 3+ positive. Seven of 10 MIFCs were cyclin D1 positive, five (71%) were 1+, and the remaining two (29%) were 2+ positive. On the other hand, 28 of 32 ATCs showed cyclin D1 immunostaining. Of these, three (9%) were 1+, five (13%) were 2+, and 20 (63%) were 3+ positive. This study demonstrates a significant overexpression of cyclin D1 in ATC compared CPC (P < .001) and MIFC (P < .005), suggesting that the cyclin D1 expression may play a role in tumor progression and may have prognostic significance in thyroid cancer.

A functional Notch–survivin gene signature in basal breast cancer

IntroductionBasal-type, or triple-negative, breast cancer (lacking estrogen receptor, progesterone receptor, and human epidermal growth factor receptor-2 expression) is a high-risk disease for which no molecular therapies are currently available. We studied genetic signatures of basal breast cancer potentially suitable for therapeutic intervention.MethodsWe analyzed protein expression of the Notch-1 intracellular domain and survivin by immunohistochemistry in a series of basal breast cancer patients. A hierarchical clustering and overall survival analysis was carried out on a microarray mRNA database of 232 breast cancer patients. Fifteen published mRNA datasets containing estrogen receptor-negative or estrogen receptor-positive samples were subjected to meta-analysis for co-segregated gene expression. Experiments of plasmid transfection and gene silencing were carried out in estrogen receptor-negative MDA-MB-231 breast cancer cells.ResultsThe developmental signaling regulator Notch-1 was highly expressed in breast cancer, compared with normal tissue, and was segregated with basal disease. Higher Notch-1 levels correlated with progressively abbreviated overall survival, and with increased expression of survivin, a tumor-associated cell death and mitotic regulator implicated in stem cell viability. Analysis of Pearsons correlation coefficient indicated that Notch-1 and survivin co-segregated in basal breast cancer. Notch-1 stimulation in MDA-MB-231 cells increased survivin expression, whereas silencing Notch reduced survivin levels.ConclusionsA Notch-1–survivin functional gene signature is a hallmark of basal breast cancer, and may contribute to disease pathogenesis. Antagonists of Notch and survivin currently in the clinic may be tested as novel molecular therapy for these recurrence-prone patients.

The role of cell cycle regulatory proteins, cyclin D1, cyclin E, and p27 in thyroid carcinogenesis.

The cell cycle is controlled in part by cyclin-dependent kinases (CDKs), which are activated by forming complexes with cyclins. CDKs phosphorylate certain substrates to facilitate the proliferating cells through the cell cycle. CDK inhibitors (CDKIs) such as p27 inhibit cyclin-CDK complexes and function as a negative cell cycle regulator. The overexpression of the positive regulators (cyclins) or the underexpression of the negative regulators including p27 has been seen in a variety of neoplasms, but their role and interaction in thyroid carcinogenesis is yet to be established. We studied the expression of cyclins D1 and E, and the CDKI, p27 by immunohistochemistry in 116 cases, including 59 cases of follicular variant of papillary carcinoma (FVPC) and 57 cases of follicular adenoma (FA). The positive staining was divided into four grades: 1+ if less than 10%, 2+ if 11% to 25%, 3+ if 26% to 50%, and 4+ if greater than 50% of the nuclei of tumor cells stained positively. Cyclin D1 expression was seen in 37 (63%) FVPC and 34 (60%) FA. Cyclin E-positive cells were seen in 51 (86%) FVPC and 47 (82%) FA. No significant differences in the grade of cyclins D1 (P = .261) and E (P = .284) staining was seen between FVPC and FA. Of the 59 FVPC, 53 (89%) showed p27-positive cells; of these, 33 were 1+, nine were 2+, seven were 3+ and only four were 4+ positive. Conversely, all 57 FA were p27 positive, 53 were 4+, and four were 3+ positive. This difference in the grade of p27 staining between FVPC and FA was statistically significant (P < .001). This study shows a significant underexpression of p27 in FVPC compared with FA, suggesting that a decrease in p27 expression plays a more important role than overexpression of cyclins D1 and E alone in thyroid carcinogenesis and that p27 immunostaining may be helpful in the diagnosis of FVPC.

Analysis of integrin beta4 expression in human breast cancer: association with basal-like tumors and prognostic significance

Purpose: The β4 integrin has been implicated in functions associated with the genesis and progression of carcinomas based on data obtained from cell lines and mouse models. Data on its expression and relevance to human carcinomas, however, are relatively scant. The aim of this study was to assess its expression and prognostic significance in human breast carcinomas. Experimental Design: We integrated data on β4 expression from multiple gene profiling studies of breast tumors of known clinical outcome with immunohistochemical analysis of 105 breast carcinomas, and we identified genes whose expression correlates with that of β4. Results: The expression of both β4 mRNA and protein is not homogeneous in breast cancer and it associates most significantly with the “basal-like” subtype of breast tumors (P = 0.008). No association between β4 and HER2 expression was evident from either gene profiling or immunohistochemical analysis. To gain insight into the relevance of β4 expression to human breast carcinomas, we generated a 65-gene “β4 signature” based on integration of four published gene profiling studies that included the top 0.1% of genes that correlated with β4, either positively or negatively. This β4 signature predicted decreased time to tumor recurrence and survival of patients when applied to four data sets including two independent ones. Conclusions: These observations indicate that β4 expression in human breast cancer is restricted and associated with basal-like cancers, and they support the hypothesis that β4 may function in concert with a discrete set of proteins to facilitate the aggressive behavior of a subset of tumors.

IMP3 is a novel biomarker for triple negative invasive mammary carcinoma associated with a more aggressive phenotype

IMP3, an oncofetal protein, is a member of the insulin-like growth factor-II (IGF-II) mRNA-binding protein family. Its relevance as a novel biomarker in lung, pancreatic, renal, and cervical adenocarcinoma was recently revealed. However, its role in breast carcinogenesis and tumor progression is not yet established. Basal-like carcinoma was initially identified by gene expression profiling. It accounts for 15% to 30% of all breast cancers. These tumors express basal epithelial markers including cytokeratin 5 but lack expression of the estrogen receptor, progesterone receptor, and human epidermal growth factor receptor 2 (HER2), therefore, are often referred to as triple negative breast cancer. They have been found to be associated with a worse overall and disease-free survival. In this retrospective study, we examined the IMP3 expression in invasive ductal carcinoma of the breast and correlated its expression with morphological and biologic prognostic factors. The study group comprised 138 cases of invasive ductal carcinoma retrieved from the surgical pathologic files for a 10-year period from 1997 to 2006. Survival data and clinical stage were available on all 138 patients. Tumor characteristics including size, grade, lymphovascular invasion, necrosis, lymph node metastasis, estrogen receptor, progesterone receptor, and HER2 status were obtained from pathologic reports. Immunohistochemistry was performed on formalin-fixed paraffin-embedded tissue using mouse monoclonal antibody against IMP3 and CK5/6. Of the 138 breast cancer cases, IMP3 expression was seen in 45 (33%). Twenty-five of the IMP3+ cases were triple negative. We found significant correlation between IMP3 expression and higher grade (P = .001), necrosis (P< .0001) triple negative, and CK5/6 expression (P < .0001 for each). Cox multivariate analysis showed a hazard ratio of IMP3 expression at 3.14 (P = .05). IMP3 is a novel biomarker for triple negative (basal-like) invasive mammary carcinoma, and its expression is associated with a more aggressive phenotype and decreased overall survival.

Pathology of the Thyroid Gland

The thyroid gland develops from the larger median analage and the two lateral analagen. The medial analage, which forms the major portion of the thyroid, is derived from the floor of the foregut and the two lateral analagen are derived from the endoderm of the fourth and the fifth branchial pouches as the ultimobranchial bodies. The medial analage appears by d 24 as median endodermal diverticulum from the base of the tongue in the region of foramen cecum. The diverticulum descends down from the foramen cecum into the neck along the midline attached to the thyroglossal duct. It reaches its final position anterior to the trachea by about 7 wk; it then grows laterally, and becomes bilobed [1]. Early during the fifth week, the thyroglossal duct loses its lumen and shortly afterwards breaks into fragments [2]. However, the caudal end of the thyroglossal duct may persist in some embryos, and this constitutes the pyramidal process, which is present in about 75% of mature human thyroids [3]. The lateral thyroid analage becomes attached to the posterior surface of the thyroid during the fifth week and contributes up to 30% to the thyroid weight [2]. The causes of the fusion of the lateral and medial analage are unknown [2]. It is speculated that migration of the ultimobranchial body controls the growth of the medial analage, or that the growth of the medial analage laterally and caudally inhibits expansion of the ultimobranchial body [2]. The lateral thyroid analage is thought to give rise to the calcitonin producing C cells and the solid cell nests. It is believed that the C cells are derived from the neural crest; they migrate to the ultimobranchial body and are subsequently incorporated into the thyroid [4]. However, the existence of mixed follicular and C-cell tumors raises the possibility of the common stem cell origin for both follicular and C cells as is seen in the gastrointestinal tract [1].

Follicular variant of papillary thyroid carcinoma: a comparative study of histopathologic features and cytology results in 141 patients.

OBJECTIVEnTo characterize the histopathologic features of follicular variant of papillary thyroid carcinoma (FVPC) and its cytology results on fine-needle aspiration (FNA) biopsy and compare them with those of papillary thyroid carcinoma (PC).nnnMETHODSnWe searched the University of Massachusetts Medical Center pathology database for all surgical specimens associated with a diagnosis of FVPC or PC between January 1992 and February 1998 and reviewed the related pathology reports. In addition, the associated preoperative FNA results were analyzed.nnnRESULTSnOn initial assessment, FVPC was associated with a significantly lower incidence of cervical lymph node metastatic involvement in comparison with PC (5.6% versus 35.7%; P<0.001). Even though the mean size of FVPC was larger than that of PC (2.57 cm versus 1.75 cm; P<0.05), FVPC showed a lower incidence of thyroid capsule invasion (5.6% versus 11.4%), infiltrative resection margins (2.8% versus 20.0%; P = 0.01), local soft tissue invasion (7.0% versus 25.7%; P<0.005), and multicentricity (25.4% versus 47.1%; P<0.01). Lymphocytic thyroiditis was a common feature of both FVPC (36.6%) and PC (35.7%). FNA biopsy revealed the presence of malignant cells in 9.8% of patients with FVPC in comparison with 67.5% of patients with PC. Most cytology specimens of FVPC (58.8%) were interpreted as suspicious for a malignant lesion or as a follicular neoplasm.nnnCONCLUSIONnFVPC is associated with a significantly lower incidence of cervical lymph node metastatic lesions and invasive histologic features than is PC. Long-term prospective clinical studies are needed to determine whether these findings translate into a more benign natural history for this variant of PC. Results of FNA biopsy in FVPC are more commonly interpreted as suspicious rather than malignant; this factor has major implications for preoperative planning.

Pseudoangiomatous stromal hyperplasia: an overview.

Pseudoangiomatous stromal hyperplasia (PASH) of the breast is a benign, proliferative mesenchymal lesion with possible hormonal etiology. It typically affects women in the reproductive age group. Pseudoangiomatous stromal hyperplasia is frequently an incidental histologic finding in breast biopsies performed for other benign or malignant lesions. Rarely, it can present as a firm, painless breast mass, which has been referred to as nodular or tumorous PASH. Grossly, tumorous PASH is a well-circumscribed, firm, rubbery mass with solid, homogenous, gray-white cut surface. On histologic examination, it is characterized by the presence of open slitlike spaces in dense collagenous stroma. The spaces are lined by a discontinuous layer of flat, spindle-shaped myofibroblasts with bland nuclei. The spindle cells express progesterone receptors and are positive for vimentin, actin, and CD34. The most important differential diagnosis on histopathology is angiosarcoma. Pseudoangiomatous stromal hyperplasia discovered incidentally does not require any additional specific treatment. Tumorous PASH is treated by local surgical excision with clear margins and the prognosis is excellent, with minimal risk of recurrence after adequate surgical excision.