PIK3CA alterations in Middle Eastern ovarian cancers

<p>Abstract</p> <p>Background</p> <p>PI3K/AKTsignaling pathway plays an important role in cell growth, proliferation, and tumorgenesis of various malignancies. This signaling pathway has been shown to be frequently altered in several human cancers including ovarian cancers. However the role of this oncogenic signaling pathway has not been explored in the Middle Eastern epithelial ovarian cancer (EOC). Therefore, we investigated PI3K/AKT genetic alterations such as PIK3CA amplification, PIK3CA mutation, PTEN protein loss and their relationships with various clinicopathological characteristics in 156 EOCs.</p> <p>Results</p> <p>Fluorescence <it>in situ </it>hybridization (FISH) technique and DNA sequencing were used to analyze PIK3CA amplification and mutation respectively. Expression of PIK3CA protein expression (p110 α), PTEN, p-AKT and Ki-67 was analyzed by immunohistochemistry. <it>PIK3CA </it>amplification was seen in 54 of 152 (35.5%) EOC cases analyzed; PIK3CA gene mutations in 6/153 EOC (3.9%); <it>KRAS </it>mutations in 3/154 EOC (1.9%), BRAF mutations in 3/156 EOC (1.9%), p53 mutation in 50/154 EOC (32.5%), and loss of PTEN protein expression in 33/144 EOC (22.9%). p110 α overexpression was associated with increased phosphorylation of AKT-Ser 473 and with the proliferation marker Ki-67.</p> <p>Conclusion</p> <p>Our data showed mutual exclusivity between the molecular event of PIK3CA amplification and mutations in <it>PIK3CA</it>, <it>KRAS</it>, <it>BRAF </it>genes, which suggests that each of these alterations may individually be sufficient to drive ovarian tumor pathogenesis independently. High prevalence of genetic alterations in PI3K/AKT pathway in a Middle Eastern ovarian carcinoma provides genetic evidence supporting the notion that dysregulated PI3K/AKT pathways play an important role in the pathogenesis of ovarian cancers.</p

ALK alteration is a frequent event in aggressive breast cancers

Introduction: Breast cancer is the most common female malignancy worldwide and, despite improvements in treatment modalities, there are increased chances of recurrence and metastasis in a substantial number of cases and it remains one of the major causes of mortality among female cancer patients. Anaplastic lymphoma kinase (ALK) gene has been found to be altered in several solid and hematologic tumors. We aimed to comprehensively study the prevalence of ALK expression, and changes in copy number and translocation in a large cohort of breast cancer cases in a Middle Eastern population. Methods: ALK protein expression was investigated by immunohistochemistry and numerical and structural variations of the ALK gene were analyzed by fluorescence in situ hybridization (FISH) in a tissue microarray format in a cohort of more than 1000 Middle Eastern breast cancers. The data were correlated with clinicopathologic parameters and other important molecular biomarkers.Results: Immunohistochemical analysis showed ALK overexpression in 36.0 % of the breast cancer patients and gene amplification was present in 13.3 % of cases, seen by FISH analyses. ALK overexpression was significantly associated with ALK gene amplification (p = 0.0031). ALK-overexpressing tumors showed significant association with high-grade tumors (p = 0.0039), ductal histologic subtype (p = 0.0076), triple-negative phenotype (p = 0.0034), and high Ki-67 (p = 0.0001) and p-AKT (p \u3c0.0001). Conclusions: Immunohistochemical analysis showed ALK is overexpressed in a substantial proportion of breast cancers and possibly plays a significant role in the aggressive behavior of this cancer. Gene amplification is hypothesized to be a possible cause for a significant proportion of this overexpression. Based on these findings, a potential role for an ALK inhibitor, as a therapeutic agent targeting aggressive subtypes of breast cancer, merits further investigation

Bortezomib-mediated expression of p27Kip1 through S-phase kinase protein 2 degradation in epithelial ovarian cancer

S-phase kinase protein 2 (SKP2), an F-box protein, targets cell-cycle regulators including cyclin-dependent kinase inhibitor p27Kip1 through ubiquitin-mediated degradation. SKP2 is frequently overexpressed in variety of cancers. We investigated the function of SKP2 and its ubiquitin–proteasome pathway in a large series (156) of epithelial ovarian cancer (EOC) patient samples, using a panel of cell lines, and nude mouse model. Using immunohistochemistry, we detected SKP2 in 13.2% tumor samples and found that it was inversely associated with p27Kip1. EOC subset with high level of SKP2 and low level of p27Kip1 showed a strong association with proliferative marker Ki167 (P\u3c0.0014). Treatment of EOC cell lines with bortezomib or expression of siRNA of SKP2 causes downregulation of SKP2 and accumulation of p27Kip1. In addition, co-treatment of EOC with bortezomib and cisplatin causes more pronounced effect on cell proliferation, apoptosis and downregulation of SKP2 leading to accumulation of p27kip1. Bortezomib treatment of EOC cells causes apoptosis by involving mitochondrial pathway, activation of caspases and downregulation of XIAP, and survivin. Finally, treatment of EOC cell line xenografts with bortezomib resulted in growth inhibition of tumors in nude mice through downregulation of SKP2 and accumulation of p27Kip1. Altogether, our results suggest that SKP2 and ubiquitin–proteasome pathway may be a potential target for therapeutic intervention for treatment of EOC

Novel noncoding RNA from human Y distal heterochromatic block (Yq12) generates testis-specific chimeric CDC2L2

The human Y chromosome, because it is enriched in repetitive DNA, has been very intractable to genetic and molecular analyses. There is no previous evidence for developmental stage- and testis-specific transcription from the male-specific region of the Y (MSY). Here, we present evidence for the first time for a developmental stage- and testis-specific transcription from MSY distal heterochromatic block. We isolated two novel RNAs, which localize to Yq12 in multiple copies, show testis-specific expression, and lack active X-homologs. Experimental evidence shows that one of the above Yq12 noncoding RNAs (ncRNAs) trans-splices with CDC2L2 mRNA from chromosome 1p36.3 locus to generate a testis-specific chimeric β sv13 isoform. This 67-nt 5′UTR provided by the Yq12 transcript contains within it a Y box protein-binding CCAAT motif, indicating translational regulation of the β sv13 isoform in testis. This is also the first report of trans-splicing between a Y chromosomal and an autosomal transcript

Molecular signatures in childhood acute lymphoblastic leukemia in the kingdom of Saudi Arabia

Acute Lymphoblastic Leukemia (ALL) is the most common malignancy in childhood population. ALL is a heterogeneous group of disease that consists of various subtypes that allows to risk stratify the patients into favorable and unfavorable groups. Risk stratification of patients has definitely played an important role in the improvement of overall survival in childhood ALL. With the introduction of microarray technology, it has become increasingly clear that leukemia with different risk factors display distinct gene-expression profiles. The aim of this study was to investigate the gene expression profile on pediatric patients diagnosed with ALL using microarray analysis and study the common genetic subtypes in ALL samples using multiplex real-time RT-PCR. We studied 77 patients of newly diagnosed ALL at the King Faisal Specialist Hospital and Research center, Riyadh, Saudi Arabia. Using multiple parameters including age, sex and WBC count; we analyzed samples for differential expression of genes using the GeneChip® human genome U133 plus 2.0 microarray from Affymetrix. Based on the age of the patient, we compared samples between 1-9 years (favorable prognosis) to more than 10 years (unfavorable prognosis) of age. Using Avadis software from Strand genomics, we were able to identify 156 genes that were differentially expressed using a cutoff p value of 0.05. Out of this list, three genes were of interest to us as they were involved in either, cell cycle and/or apoptosis. They include the pro-apoptotic genes; TGF-beta induced apoptosis protein 2 (TAIP2) and Caspase Recruitment Domain member 14 (CARD14) that were 10.16 and 4.12 folds over-expressed in the favorable prognosis group respectively. In addition, another gene, cyclin A1 (CCNA1) that is involved in cell cycle was found to be down-regulated 4.84 folds in the favorable group. These genes have important clinical implications and can be used as prognostic markers in this age group. Interestingly, CyclinA1 is being currently targeted using siRNA to inhibit tumor growth and induce apoptosis in certain cancers. In addition to gene expression profiling, we also studied the four common genetic translocations detected in ALL by multiplex real-time RT-PCR. They include the t(12;21) TEL/AML1 fusion gene, t(9;22) BCR/ABL fusion gene, t(4;11) MLL1/AF4 fusion gene and t(1;19) E2A/PBX1 fusion gene. We found that the genetic translocation detected in our samples varied from the published data on western population. We detected 15% of TEL/AML1 translocation which was less than that found in the western population (22%), 4% of BCR/ABL that was higher than the western population (2.2%) and 6.5% of E2A/PBX1, that was higher than the western population (3.8%). These data have important clinical significance as they allow us to better understand the underlying biology of ALL as well as risk stratifies ALL patients according to their gene expression profiles and their genetic subtypes

Role of x-linked inhibitor of apoptosis as a prognostic marker and therapeutic target in papillary thyroid carcinoma

Context: Papillary thyroid cancer (PTC) is the second most common cancer in females in Saudi Arabia. However, the pathogenesis of PTC is still not fully elucidated. Objective: To identify potential genes that play important role in progression of PTC, we studied the role of X-linked inhibitor of apoptosis protein (XIAP) as a potential prognostic marker and therapeutic target in a large cohort of PTC samples and cell lines.Design: A DNA microarray chip was used to screen for gene copy number. XIAP expression was assessed by immunohistochemistry in a tissue microarray format on a cohort of 1022 clinical samples. In vitro and in vivo studies were performed using Embelin and/or LY294002 on PTC cell lines. Results: XIAP was found to be amplified in 14 of 29 and overexpressed in 48.8% of PTC cases. XIAP overexpression was significantly associated with old age, extrathyroidal extension, tumor size, nodal involvement, tall-cell variant, advanced stage disease, and significantly poor disease-free survival (P = .0341). XIAP was also significantly associated with phosphorylated AKT (P \u3c .0001), Bcl-Xl (P \u3c .0001), and Ki67 (P = .0006) proteins. Embelin treatment caused growth inhibition and apoptosis in PTC cell lines and induced tumor regression in PTC xenograft in nude mice. Finally, the combination of suboptimal doses of Embelin and LY294002 induced a synergistic apoptotic response in PTC cellsConclusion: XIAP dysregulation in PTC confers an aggressive phenotype with poor outcome. In vitro and in vivo studies using an XIAP inhibitor suggest that this subgroup of PTC with overexpression of XIAP can be therapeutically targeted, either alone or in combination, to induce efficient apoptosis in these cancers

FoxM1 and its association with matrix metalloproteinases (MMPs) signaling pathway in papillary thyroid carcinoma

Context: Forkhead boxM1 (FoxM1) transcription factor has been shown to promote pathogenesis of several malignancies. FoxM1 has also been shown to be associated with matrix metalloproteinases (MMP) in various cancers. However, little is known about its function in papillary thyroid carcinoma (PTC).Objective: In this study, we investigated the role of FoxM1 in pathogenesis in a large series of PTC in a tissue microarray format followed by in vitro and in vivo studies using PTC cell lines and nude mice. Design: Expression of FoxM1 and its associated proteins were investigated in Middle Eastern PTC samples by immunohistochemistry. Apoptosis was measured by flow cytometry and immunoblotting. Invasion and migration studies were performed using 8-μm Transwell plates. Results: FoxM1 was overexpressed in 28.4% of PTC and significantly associated with activated matrix metalloproteinase-9 (MMP-9) (P = 0.0004), X-linked inhibitor of apoptosis protein (XIAP) (P = 0.0024), and B-cell lymphoma-extra large (Bcl-XL) (P = 0.0014) expression. Treatment of PTC cell lines with thiostrepton, an inhibitor of FoxM1, resulted in inhibition of cell viability via induction of apoptosis. In addition, thiostrepton treatment of PTC cells or expression of FoxM1-specific small interfering RNA down-regulated expression of FoxM1 accompanied with decreased MMP-2 and MMP-9 expression. Furthermore, inhibition of FoxM1 attenuated migration and invasion of PTC cells. Interestingly, overexpression of FoxM1 rescued the effects of thiostrepton in PTC cell lines. Finally, treatment of PTC cell line xenografts with thiostrepton resulted in growth inhibition of tumors in nude mice via down-regulation of FoxM1 and MMP-9 and MMP-2. Conclusion: Altogether, this is the first study showing that FoxM1 and its associated signaling pathway play a critical role in the pathogenesis of PTC and may be a potential target for therapeutic intervention for treatment of these cancers

Frequent PIK3CA gene amplification and its clinical significance in colorectal cancer

Using a DNA microarray approach to screen for gene copy number changes in 20 colorectal (CR) carcinoma samples and filtering for high-level DNA copy number changes, we detected an amplicon at 3q26 containing the PIK3CA gene. Fluorescence in situ hybridization was employed for evaluation of PIK3CA amplification on a progression CR tissue microarray containing 448 CR carcinomas, normal mucosa, and adenomas with follow-up information. PIK3CA amplification (ratio PIK3CA/centromere 3≥2.0) was found in 38% of cancers, while another 19% of tumours had PIK3CA gains (ratio \u3e1.0 but \u3c2.0). Both PIK3CA amplification and gains were associated with high levels of PIK3CA protein expression and no association was seen between PIK3CA amplification and PIK3CA mutation. In a subset of 220 patients who received adjuvant chemotherapy and/or radiotherapy, survival in patients with PIK3CA-amplified cancers was significantly longer compared with patients with cancers without amplification. This association was independent of stage, grade, histology subtype, gender, and age categories. Interestingly, PIK3CA amplification was also seen in CR adenomas, indicating an early genetic alteration, and was also a frequent event in colorectal carcinogenesis. Furthermore, PIK3CA amplification is an independent prognostic marker for better survival and may be one of the promising markers to define CRC subsets that may maximally benefit from adjuvant therapy. Copyright © 2009 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd

Additional file 2: Figure S1. of ALK alteration is a frequent event in aggressive breast cancers

Biochemical fractionation of ALK expression in breast cancer cell lines. (A) Nuclear-free cytosolic and nuclear extracts were isolated from MDA-MB231 and MCF12A cells and immunoblotted with antibodies against ALK and tubulin. (B) Relative expression of ALK was calculated using spot densitometry on cytosolic and nuclear immunoblots of MDA-MB231 and MCF12A cells. Expression of ALK was normalized with tubulin