Evolution and Impact of Subclonal Mutations in Papillary Thyroid Cancer.

Unlike many cancers, the pattern of tumor evolution in papillary thyroid cancer (PTC) and its potential role in relapse have not been elucidated. In this study, multi-region whole-exome sequencing (WES) was performed on early-stage PTC tumors (n = 257 tumor regions) from 79 individuals, including 17 who had developed relapse, to understand the temporal and spatial framework within which subclonal mutations catalyze tumor evolution and its potential clinical relevance. Paired primary-relapse tumor tissues were also available for a subset of individuals. The resulting catalog of variants was analyzed to explore evolutionary histories, define clonal and subclonal events, and assess the relationship between intra-tumor heterogeneity and relapse-free survival. The multi-region WES approach was key in correctly classifying subclonal mutations, 40% of which would have otherwise been erroneously considered clonal. We observed both linear and branching evolution patterns in our PTC cohort. A higher burden of subclonal mutations was significantly associated with increased risk of relapse. We conclude that relapse in PTC, while generally rare, does not follow a predictable evolutionary path and that subclonal mutation burden may serve as a prognostic factor. Larger studies utilizing multi-region sequencing in relapsed PTC case subjects with matching primary tissues are needed to confirm these observations.

Lynch Syndrome Identification in Saudi Cohort of Endometrial Cancer Patients Screened by Universal Approach.

Lynch syndrome (LS) is the most common cause of inherited endometrial cancer (EC). The prevalence and molecular characteristic of LS in Middle Eastern women with EC have been underexplored. To evaluate the frequency of LS in a cohort of EC patients from Saudi Arabia, a total of 436 EC cases were screened utilizing immunohistochemistry (IHC), MLH1 promoter methylation analysis and next-generation sequencing technology. A total of 53 of 436 (12.2%) ECs were classified as DNA mismatch repair-deficient (dMMR). MLH1 promoter hypermethylation was detected in 30 ECs (6.9%). Three ECs (0.7%) were found to be LS harboring germline pathogenic variants (PVs)/likely pathogenic variants (LPVs): two in the MSH2 gene and one in the MSH6 gene. Three ECs (0.7%) were Lynch-like syndrome (LLS) carrying double somatic MSH2 PVs/LPVs. Seven cases were found to have variants of uncertain significance in cancer-related genes other than MMR genes. Our results indicate that LS prevalence is low among Saudi EC patients and LLS is as common as LS in this ethnicity. Our findings could help in better understanding of the prevalence and mutational spectrum of this syndrome in Saudi Arabia, which may help in defining best strategies for LS identification, prevention and genetic counseling for EC patients.

Genetic heterogeneity and evolutionary history of high-grade ovarian carcinoma and matched distant metastases.

BACKGROUND: High-grade serous ovarian carcinoma (HGSOC) is the most frequent type of ovarian carcinoma, associated with poor clinical outcome and metastatic disease. Although metastatic processes are becoming more understandable, the genomic landscape and metastatic progression in HGSOC has not been elucidated. METHODS: Multi-region whole-exome sequencing was performed on HGSOC primary tumours and their metastases (n = 33 tumour regions) from six patients. The resulting somatic variants were analysed to delineate tumour evolution and metastatic dissemination, and to compare the repertoire of events between primary HGSOC and metastasis. RESULTS: All cases presented branching evolution patterns in primary HGSOC, with three cases further showing parallel evolution in which different mutations on separate branches of a phylogenetic tree converge on the same gene. Furthermore, linear metastatic progression was observed in 67% of cases with late dissemination, in which the metastatic tumour mostly acquires the same mutational process active in primary tumour, and parallel metastatic progression, with early dissemination in the remaining 33.3% of cases. Metastatic-specific SNVs were further confirmed as late dissemination events. We also found the involvement of metastatic-specific driver events in the Wnt/ß-catenin pathway, and identified potential clinically actionable events in individual patients of the metastatic HGSOC cohort. CONCLUSIONS: This study provides deeper insights into clonal evolution and mutational processes that can pave the way to new therapeutic targets.

XIAP over-expression is an independent poor prognostic marker in Middle Eastern breast cancer and can be targeted to induce efficient apoptosis.

BACKGROUND: Breast cancer is the most common cancer in females and is ranked second in cancer-related deaths all over the world in women. Despite improvement in diagnosis, the survival rate of this disease has still not improved. X-linked Inhibitor of Apoptosis (XIAP) has been shown to be over-expressed in various cancers leading to poor overall survival. However, the role of XIAP in breast cancer from Middle Eastern region has not been fully explored. METHODS: We examined the expression of XIAP in more than 1000 Middle Eastern breast cancer cases by immunohistochemistry. Apoptosis was measured by flow cytometry. Protein expression was determined by western blotting. Finally, in vivo studies were performed on nude mice following xenografting and treatment with inhibitors. RESULTS: XIAP was found to be over-expressed in 29.5% of cases and directly associated with clinical parameters such as tumor size, extra nodal extension, triple negative breast cancer and poorly differentiated breast cancer subtype. In addition, XIAP over-expression was also significantly associated with PI3-kinase pathway protein; p-AKT, proliferative marker; Ki-67 and anti-apoptotic marker; PARP. XIAP over-expression in our cohort of breast cancer was an independent poor prognostic marker in multivariate analysis. Next, we investigated inhibition of XIAP using a specific inhibitor; embelin and found that embelin treatment led to inhibition of cell viability and induction of apoptosis in breast cancer cells. Finally, breast cancer cells treated with combination of embelin and PI3-kinase inhibitor; LY294002 synergistically induced apoptosis and caused tumor growth regression in vivo. CONCLUSION: These data suggest that XIAP may be playing an important role in the pathogenesis of breast cancer and can be therapeutically targeted either alone or in combination with PI3-kinase inhibition to induce efficient apoptosis in breast cancer cells.

Molecular markers and pathway analysis of colorectal carcinoma in the Middle East.

BACKGROUND: Colorectal cancer (CRC) is one of the most common cancers in the world. A newly proposed integrated pathway comprising traditional, alternate, and serrated pathways by genetic and epigenetic factors was defined recently and hypothesized to play a role in the pathogenesis of CRC; however, to the authors' knowledge, there is a paucity of information regarding these proposed molecular pathways in different ethnic groups. METHODS: Molecular characterization of 770 CRC specimens was performed for microsatellite instability, BRAF, and KRAS by polymerase chain reaction and 500 cases for CpG island methylator phenotype (CIMP) high phenotype by MethyLight technology. Tumors were assigned to different molecular pathways and examined for clinicopathological correlation and survival analysis. RESULTS: The traditional pathway constituted 33.4% of CRC cases, the alternate pathway comprised 11.6%, and the serrated molecular pathway accounted for only 0.8% of Middle Eastern CRC cases. Approximately 54.2% of CRC cases did not qualify to fit into any pathway and thus were designated as an unassigned group. Molecular pathways were found to be significantly associated with tumor site and grade. A subset of cases with an uncategorized pathway demonstrated a significant survival difference (P = .0079). CONCLUSIONS: The serrated pathway was found to account for a very low percentage of the CRC patient cohort in the current study. The unassigned group accounted for the majority of Middle Eastern CRC cases, and therefore methods of CRC pathway analysis might not be applicable to this ethnic group. The current study demonstrates the need to unravel the molecular genetic basis of this disease to further subcategorize these CRC cases. It also identifies a need for further studies on different populations for a better understanding of their exact role and incidence.

Predicting factors and clinical outcome of biochemical incomplete response in middle eastern differentiated thyroid carcinoma.

PURPOSE: The aim of this study was evaluate biochemical incomplete response (BIR) in Middle Eastern differentiated thyroid cancer (DTC), identify factors that could predict BIR before radioactive iodine (RAI) ablation and to investigate the long-term clinical outcome of DTC patient exhibiting BIR to initial therapy. METHODS: We retrospectively evaluated 1286 DTCs from Middle Eastern ethnicity who underwent total thyroidectomy and RAI therapy. Demograpic and clinico-pathological factors predicting BIR were evaluated. The outcome of these patients was analyzed using primary outcome of structural disease and disease-free survival (DFS). RESULTS: With a median follow-up of 10 years, 266 (20.7%) patients had BIR. High pre-ablation stimulated thyroglobulin (presTg), presence of lymph node metastasis, male gender and delayed initial RAI therapy (≥3 months) after thyroidectomy were significant independent predictors of BIR. Upon evaluating long-term clinical outcomes in 266 patients with BIR, we found 36.8% of patients developed structural disease. Male sex (OR = 1.56; 95% CI = 1.05-2.30; p = 0.0272) and increasing Tg after initial therapy (OR = 4.25; 95% CI = 1.93-10.82; p = 0.0001) were independent risk factors for structural disease in patients with BIR. DFS was significantly worse if both these risk factors existed concomitantly (p < 0.0001). CONCLUSION: To achieve the fair efficacy of RAI therapy, early prediction of BIR before RAI ablation is desirable. Our finding of the clinico-pathological factors (high presTg level, LNM, delayed RAI therapy and male gender) could serve as easy and robust early predictors of BIR. In addition, DTC patients exhibiting BIR had a high risk of structural disease and hence personalized management approach would be preferable for BIR patients to ensure best clinical outcome.

Radioactive iodine refractoriness in Middle Eastern differentiated thyroid cancer: clinical outcome and risk factor analysis.

Background: Radioactive iodine refractory differentiated thyroid cancer (RAIR-DTC) has received increasing attention due to its poor prognosis. However, outcomes may vary among patients with RAIR-DTC. The role of clinico-pathological and molecular prognostic factors in survival remains controversial, resulting in difficulty in selecting patients for new targeted therapies. We assessed mortality rate and DTC-specific survival in Middle Eastern RAIR-DTC to identify prognostic factors associated with survival. Methods: This single center, retrospective study enrolled 268 patients with RAIR-DTC. Mortality rate and DTC-specific survival were analyzed to identify prognostic factors related to survival. Univariate and multivariate analysis were performed using Cox proportional hazards model. Results: Of the 268 cases of RAIR-DTC, 40.3% (108/268) had absent 131I uptake (either on diagnostic or post-therapy whole body scan), 15.3% (41/268) had progressive disease (PD) despite 131I, 7.5% (20/268) had persistent disease despite cumulative activity of I131 of >600 mCi and 36.9% (n=99/268) developed distant metastasis. On multivariate analysis, age (more than 45 years), presence of metastatic disease and tumors harboring telomerase reverse transcriptase (TERT) promoter mutations were independent prognostic factors for poor DTC-specific survival. Subjects were divided into 3 groups according to the number of risk factors; low risk (no risk factors); intermediate (≤ 2 risk factors); and high risk (all the 3 risk factors). Ten-year DTC-specific survival rates in low, intermediate and high-risk groups were 100.0%, 92.9% and 53.6%, respectively. Conclusions: The contribution of age greater than 45 years to RAIR-DTC mortality is impactful. Older age, presence of distant metastasis and TERT mutations could be used as early predictors of RAIR-DTC cases. The identification of prognostic factors for poor survival in RAIR-DTC may improve the selection of patients for more personalized surveillance and therapeutic modalities.

The Prognostic Impact of Tumor Size and BRAF Mutational Status in Middle Eastern Differentiated Thyroid Cancer.

PURPOSE: Tumor size at diagnosis has been widely used as a major mortality risk factor in risk stratification of DTC. The current study was designed to analyze whether tumor size at diagnosis is a major prognostic factor in Middle Eastern DTC. METHODS: We conducted a comparative study of the relationship between tumor size at diagnosis and event free survival (EFS) with respect to BRAF status in 1709 consecutive patients treated surgically for DTC. Patients were divided into four groups according to the size of tumor and BRAF mutation status: Group 1 (≤4 cm without BRAF mutation), Group 2 (≤4 cm with BRAF mutation), Group 3 (>4 cm without BRAF mutation) and Group 4 (>4 cm with BRAF mutation). Predictors of EFS were compared using the Log-rank test and Cox proportional hazards models. RESULTS: Tumor size >4cm was associated with adverse clinico-pathologic characteristics, such as older age, male gender, bilateral tumors, extrathyroidal extension, lymphovascular invasion, advanced tumor stage and persistent/recurrent disease. Tumor size was also inversely associated with BRAF mutation. Both tumor size (> 4cm) and BRAF mutation were associated with EFS on univariate analysis. On subgroup analysis, larger tumor size was an independent predictor of EFS (Group 3 vs. Group 1), irrespective of BRAF mutation status. Also, within the BRAF mutant tumors, larger tumor size was still an independent predictor of EFS (Group 4 vs. Group 2). CONCLUSION: Tumor size is an independent predictor of EFS in Middle Eastern DTC patients, regardless of BRAF mutational status.

Whole Exome-Wide Association Identifies Rare Variants in GALNT9 Associated with Middle Eastern Papillary Thyroid Carcinoma Risk.

Papillary thyroid carcinoma (PTC) is the commonest thyroid cancer. The majority of inherited causes of PTC remain elusive. However, understanding the genetic underpinnings and origins remains a challenging endeavor. An exome-wide association study was performed to identify rare germline variants in coding regions associated with PTC risk in the Middle Eastern population. By analyzing exome-sequencing data from 249 PTC patients (cases) and 1395 individuals without any known cancer (controls), GALNT9 emerged as being strongly associated with rare inactivating variants (RIVs) (4/249 cases vs. 1/1395 controls, OR = 22.75, p = 5.09 × 10-5). Furthermore, three genes, TRIM40, ARHGAP23, and SOX4, were enriched for rare damaging variants (RDVs) at the exome-wide threshold (p < 2.5 × 10-6). An additional seven genes (VARS1, ZBED9, PRRC2A, VWA7, TRIM31, TRIM40, and COL8A2) were associated with a Middle Eastern PTC risk based on the sequence kernel association test (SKAT). This study underscores the potential of GALNT9 and other implicated genes in PTC predisposition, illuminating the need for large collaborations and innovative approaches to understand the genetic heterogeneity of PTC predisposition.

Predictive risk factors for distant metastasis in pediatric differentiated thyroid cancer from Saudi Arabia.

Background: Despite their excellent prognosis, children and young adults (CAYA) with differentiated thyroid cancer (DTC) tend to have more frequent occurrence of distant metastasis (DM) compared to adult DTC. Data about DM in CAYA from Middle Eastern ethnicity is limited. Methods: Medical records of 170 patients with DTC ≤18 years were retrospectively reviewed. Clinico-pathological factors associated with lung metastasis in CAYA, their clinical presentation and outcome were analyzed. Rick factors related to distant metastasis-free survival (DMFS) for the whole cohort were evaluated. Results: DM was observed in 27 patients and all were lung metastasis. Lung metastasis was significantly associated with younger age (≤15 years), extrathyroidal extension (ETE), multifocal tumors, bilaterality, presence of lymph node (LN) disease and high post-operative stimulated thyroglobulin (sTg). Highest negative predictive values were seen with low post-operative sTg (97.9%), absence of LN disease (93.8%), absence of ETE (92.2%) and age older than 15 years (92.9%). Post-therapy whole body scan (WBS) identified most of the lung metastasis (21 of 27; 77.8%). Upon evaluating patients response according to ATA guidelines, excellent response was seen in only one patient, while biochemical persistence and structural persistence were seen in 11.1% (3/27) and 77.8% (21/27), respectively. Elevated post-operative sTg (>10ng/ml) was the only risk factor found to be significantly associated with both biochemical persistence (with or without structural persistence (p = 0.0143)) and structural persistence (p = 0.0433). Cox regression analysis identified age and post-operative sTg as independent risk factors related to DMFS. Based on these two risk factors for DMFS, patients were divided into 3 groups: low risk (no risk factors), intermediate risk (1 risk factor) and high risk (both risk factors). 20-year DMFS rates in the low-, intermediate- and high-risk groups were 100.0%, 81.3% and 23.7% respectively (p < 0.0001). Conclusion: Higher suspicion for metastatic pediatric DTC should be considered in patients who are young, have LN disease, extrathyroidal extension and elevated post-operative sTg. Persistent disease, despite therapy, is very common and it appears to be related to post-operative sTg level. Hence, risk adaptive management is desirable in CAYA with DTC.

Identification and characterization of ATM founder mutation in BRCA-negative breast cancer patients of Arab ethnicity.

Breast cancer (BC) is the most prevalent malignancy among women worldwide with germline pathogenic variants/likely pathogenic variants (PVs/LPVs) in BRCA1/2 accounting for a large portion of hereditary cases. Recently, heterozygous PVs/LPVs in the ATM serine/threonine kinase or Ataxia-telangiectasia mutated gene (ATM) has been identified as a moderate susceptibility factor for BC in diverse ethnicities. However, the prevalence of ATM PVs/LPVs in BC susceptibility in Arab populations remains largely unexplored. This study investigated the prevalence of ATM PVs/LPVs among BC patients from Saudi Arabia, employing capture-sequencing technology for ATM PVs/LPVs screening in a cohort of 715 unselected BC patients without BRCA1/2 PVs/LPVs. In addition, founder mutation analysis was conducted using the PHASE program. In our entire cohort, four unique PVs/LPVs in the ATM gene were identified in six cases (0.8%). Notably, one recurrent LPV, c.6115G > A:p.Glu2039Lys was identified in three cases, for which haplotype analysis confirmed as a novel putative founder mutation traced back to 13 generations on average. This founder mutation accounted for half of all identified mutant cases and 0.4% of total screened cases. This study further reveals a significant correlation between the presence of ATM mutation and family history of BC (p = 0.0127). These findings underscore an approximate 0.8% prevalence of ATM germline PVs/LPVs in Arab BC patients without BRCA1/2 PVs/LPVs and suggest a founder effect of specific recurrent ATM mutation. These insights can help in the design of a genetic testing strategy tailored to the local population in Saudi Arabia, thereby, enabling more accurate clinical management and risk prediction.

PALB2 germline mutations in a large cohort of Middle Eastern breast-ovarian cancer patients.

The PALB2 gene is a breast cancer (BC) and ovarian cancer (OC) predisposition gene involved in the homologous recombination repair pathway. However, the prevalence and clinicopathological association of PALB2 pathogenic/likely pathogenic (PV/LPV) variants in Middle East is still not fully explored. Total 918 BC/OC patients from Saudi Arabia were selected for PALB2 mutations screening using capture sequencing technology. Five heterozygous PVs or LPVs were identified in six cases, accounting for 0.65% (6/918) of entire cohort. Two cases (33.3%) harbored PVs and four cases (66.7%) carried LPVs. Four PVs/LPVs (80%) were frameshift along with one novel splicing LPV (c.2835-2_2835-1delinsTT). One recurrent LPV (c.3425delT: p.L1142fs) was identified in two cases. All six affected carriers have breast cancer diagnosis with median age of 39.5 years (range 34-49 years). Only two cases (33%) have documented family history of cancer. Breast cancer phenotype was invasive ductal unilateral cancer in all cases with 66.7% of hormone receptor positive and 16% of triple negative tumors. Germline PVs/LPVs in the PALB2 gene were observed in low frequency of 0.65% in Saudi BC and/or OC. Our study confirms one recurrent LPV and one novel LPV in Saudi breast cancer patients.

The biological and clinical impact of inhibition of NF-κB-initiated apoptosis in diffuse large B cell lymphoma (DLBCL).

NF-κB is frequently over-expressed in a variety of non-Hodgkin's lymphomas (NHLs) and has been implicated in lymphomagenesis; however, its role in diffuse large B cell lymphoma (DLBCL) as a prognostic biomarker has not been fully elucidated. Therefore, we investigated the role of NF-κB and its association with clinicopathological features in a tissue microarray cohort of 230 DLBCL patient samples. We then elucidated the role of NF-κB inhibition on cell viability and apoptosis in vitro, using DLBCL cell lines. Using immunohistochemistry, NF-κB was detected in 25.6% (52/203) DLBCL tumours, was associated with activated B cell (ABC) phenotype (p = 0.0054), Epstein-Barr virus (EBV; p = 0.0080) and over-expression of the anti-apoptotic marker XIAP (p = 0.0013). DLBCL cases with nuclear expression of NF-κB showed a significantly poorer overall survival as compared to those without NF-κB expression (p = 0.0236). In a multivariate analysis using a Cox proportional hazard model for IPI and NF-κB expression, the relative risk was 2.97 for high NF-κB expression (95% CI 1.27-6.94; p = 0.0113) and 7.55 for the high-IPI group (95% CI 3.34-18.35; p < 0.0001). In vitro, Bay 11-7085 inhibited constitutively active NF-κB expression in a dose-dependent manner and inhibition of NF-κB also down-regulated expression of the downstream target gene products Bcl-2, Bcl-XL (BCL2L1), XIAP and Survivin, leading to apoptosis via activation of the mitochondrial apoptotic pathway. NF-κB over-expression was found to be an independent prognostic marker for poor survival in DLBCL. Altogether, these results suggest that NF-κB may be a useful prognostic biomarker and a potential target for therapeutic intervention in DLBCL.

Lymph node ratio is superior to AJCC N stage for predicting recurrence in papillary thyroid carcinoma.

OBJECTIVE: Recently, lymph node ratio (LNR) has emerged as an alternative to American Joint Committee on Cancer (AJCC) N stage, with superior prognostic value. The utility of LNR in Middle Eastern papillary thyroid carcinoma (PTC) remains unknown. Therefore, we retrospectively analyzed a large cohort of 1407 PTC patients for clinicopathological associations of LNR. METHODS: Receiver operating characteristics (ROC) curve was used to determine the cut-off for LNR. We also performed multivariate logistic regression analysis to determine whether LNR or AJCC N stage was superior in predicting recurrence in PTC. RESULTS: Based on ROC curve analysis, a cut-off of 0.15 was chosen for LNR. High LNR was significantly associated with adverse clinicopathological characteristics such as male sex, extrathyroidal extension, lymphovascular invasion, multifocality, bilateral tumors, T4 tumors, lateral lymph node (N1b) involvement, distant metastasis, advanced tumor stage, American Thyroid Association (ATA) high-risk category and tumor recurrence. On multivariate analysis, we found that LNR was a better predictor of tumor recurrence than AJCC N stage (odds ratio: 1.96 vs 1.30; P value: 0.0184 vs 0.3831). We also found that LNR combined with TNM stage and ATA risk category improved the prediction of recurrence-free survival, compared to TNM stage or ATA risk category alone. CONCLUSIONS: The present study suggests LNR is an independent predictor of recurrence in Middle Eastern PTC. Integration of LNR with 8th edition AJCC TNM staging system and ATA risk stratification will improve the accuracy to predict recurrence in Middle Eastern PTC and help in tailoring treatment and surveillance strategies in these patients.

Exome sequencing revealed comparable frequencies of RNF43 and BRAF mutations in Middle Eastern colorectal cancer.

Mutation-induced activation of Wnt-ß Catenin signaling pathway is frequent in CRC. The E3 ubiquitin ligase, RNF43, has been reported to negatively regulate the Wnt signaling pathway and RNF43 mutations are frequently seen in CRC. However, its role in Middle Eastern CRC remains unclear. Therefore, we employed Exome and Sanger sequencing technology to assess the frequency of RNF43 mutations and its association with other clinico-pathological features in Middle Eastern CRC. RNF43 mutations were found in 5.9% (13/220) of CRC cases and was inversely correlated to APC and TP53 mutations. A strong association of RNF43 mutations with right sided and sporadic microsatellite instable (MSI) CRC was observed. No association was identified between RNF43 mutation and other clinico-pathological features including BRAF mutation, age, tumor histological subtype, tumor grade or patients' prognosis. Multivariate logistic regression analysis revealed that MSI status and wild type APC were independent predictor of RNF43 mutation. We conclude that RNF43 mutations occur in Middle Eastern CRC at comparable frequencies with BRAF mutations and represent a distinct molecular subtype which further enhances our understanding of how different mutational subsets of Wnt tumor suppressor genes link to distinct tumor characteristics, which might be considered for treatment strategies for CRC patients.

TERT Promoter Mutations Are an Independent Predictor of Distant Metastasis in Middle Eastern Papillary Thyroid Microcarcinoma.

Background: Papillary thyroid microcarcinomas (PTMCs) have been attributed to the recent increased incidence of thyroid cancer. Although indolent, a subset of PTMC could potentially develop distant metastasis (DM). This study aimed to evaluate the clinico-pathological features and molecular characteristics of PTMC and identify the risk factors for DM in PTMC patients from Middle Eastern ethnicity. Methods: We retrospectively analyzed 210 patients with histologically confirmed PTMC. Clinico-pathological associations for DM, BRAF mutation and TERT mutation were analyzed successfully in 184 patients. Multivariate analysis was performed using Cox proportional hazards model and logistic regression analysis. Results: Among the PTMC patients included in this cohort, DM was noted in 6.0% (11/184), whereas tumor relapse occurred in 29/184 (15.8%). Of the 11 cases with DM, lung metastasis occurred in 8 cases, bone metastasis in 2 cases and brain metastasis in 1 case. Presence of extrathyroidal extension and male sex were significantly associated with DM. Molecular analysis showed BRAF V600E mutations to be the most frequent, being detected in 45.7% (84/184). TERT promoter mutations were detected in 16 (8.7%) cases and were significantly associated with DM and shorter metastasis-free survival in multivariate analysis. Conclusions: Our study indicates a surprisingly high frequency of TERT promoter mutation in Saudi patients with PTMC. Identifying TERT promoter mutations as an independent predictor of DM in patients with microcarcinoma could explain the inherent aggressive nature of PTMC from Middle Eastern ethnicity and magnify its role in patient risk stratification, which might help in improving therapeutic strategy for these patients.

APOBEC SBS13 Mutational Signature-A Novel Predictor of Radioactive Iodine Refractory Papillary Thyroid Carcinoma.

Standard surgery followed by radioactive iodine (131I, RAI) therapy are not curative for 5−20% of papillary thyroid carcinoma (PTC) patients with RAI refractory disease. Early predictors indicating therapeutic response to RAI therapy in PTC are yet to be elucidated. Whole-exome sequencing was performed (at median depth 198x) on 66 RAI-refractory and 92 RAI-avid PTCs with patient-matched germline. RAI-refractory tumors were significantly associated with distinct aggressive clinicopathological features, including positive surgical margins (p = 0.016) and the presence of lymph node metastases at primary diagnosis (p = 0.012); higher nonsilent tumor mutation burden (p = 0.011); TERT promoter (TERTp) mutation (p < 0.0001); and the enrichment of the APOBEC-related single-base substitution (SBS) COSMIC mutational signatures 2 (p = 0.030) and 13 (p < 0.001). Notably, SBS13 (odds ratio [OR] 30.4, 95% confidence intervals [CI] 1.43−647.22) and TERTp mutation (OR 41.3, 95% CI 4.35−391.60) were revealed to be independent predictors of RAI refractoriness in PTC (p = 0.029 and 0.001, respectively). Although SBS13 and TERTp mutations alone highly predicted RAI refractoriness, when combined, they significantly increased the likelihood of predicting RAI refractoriness in PTC. This study highlights the APOBEC SBS13 mutational signature as a novel independent predictor of RAI refractoriness in a distinct subgroup of PTC.

Microscopic Extrathyroidal Extension Results in Increased Rate of Tumor Recurrence and Is an Independent Predictor of Patient's Outcome in Middle Eastern Papillary Thyroid Carcinoma.

BACKGROUND: Papillary Thyroid Cancer (PTC) is the most common endocrine malignancy, with recurrence rate as high as 30%. A great deal of controversy surrounds the significance of microscopic extrathyroidal extension (m-ETE) as a prognostic factor. The most recent edition (8th) of American Joint Committee on Cancer (AJCC) staging system has removed m-ETE from the definition of pT3, which suggests that m-ETE may lack prognostic impact in PTC patients. Moreover, data about m-ETE prevalence and clinical impact on Middle Eastern PTC remains unknown. We therefore investigate the prevalence of m-ETE and its clinico-pathological correlation and prognostic impact in Middle Eastern PTC. We also compared the AJCC 7th and 8th staging systems and their prognostic performance. METHODS: PTCs from 1430 consecutive adult (> 18 years) patients from single tertiary care hospital were included in this study. A retrospective analysis of PTC patients' survival and recurrence were compared between AJCC 8th and AJCC 7th staging systems using Proportion of Variation Explained (PVE) and Harrell's C-index. RESULTS: Median follow up of the study cohort was 9.3 years. 31.2% (446/1430) of patients had m-ETE. In the overall cohort, m-ETE was associated with multiple adverse features such as older age (p < 0.0001), male sex (p = 0.0245), tall cell variant (p < 0.0001), bilateral tumors (p < 0.0001), multifocality (p < 0.0001), lymphovascular invasion (p < 0.0001), lymph node metastasis (p < 0.0001), distant metastasis (p = 0.0166), tumor recurrence (p < 0.0001), radioactive iodine refractoriness (p < 0.0001), BRAF mutation (p < 0.0001) and reduced recurrence-free survival (RFS; HR = 1.75; 95% CI = 1.30 - 2.35; p < 0.0001) irrespective of tumor size. Of the 611 patients with T3 disease based on AJCC 7th edition, 359 (58.8%) were down-staged in AJCC 8th edition classification. Overall, the prognostic performance of AJCC 8th edition was inferior to AJCC 7th on the basis of lower PVE (3.04% vs. 3.73%) and lower C-index (0.40 vs. 0.48). CONCLUSIONS: In Middle Eastern PTC, m-ETE is significantly associated with compromised survival and acts as an independent predictor of RFS. Given these findings, m-ETE should be included in the thyroid cancer treatment guidelines.

Prognostic significance of TRAIL death receptors in Middle Eastern colorectal carcinomas and their correlation to oncogenic KRAS alterations.

BACKGROUND: Tumour necrosis factor-related apoptosis-inducing ligand (TRAIL) is a member of the tumour necrosis factor cytokine family that induces apoptosis upon binding to its death domain containing receptors, TRAIL receptor 1 (DR4) and TRAIL receptor 2 (DR5). Expression of TRAIL receptors is higher in colorectal carcinoma (CRC) as compared to normal colorectal mucosa and targeted therapy with TRAIL leads to preferential killing of tumor cells sparing normal cells. METHODS: We investigated the expression of TRAIL and its receptors in a tissue microarray cohort of 448 Middle Eastern CRC. We also studied the correlation between TRAIL receptors and various clinico-pathological features including key molecular alterations and overall survival. RESULTS: CRC subset with TRAIL-R1 expression was associated with a less aggressive phenotype characterized by early stage (p = 0.0251) and a histology subtype of adenocarcinomas (p = 0.0355). Similarly CRC subset with TRAIL-R2 expression was associated with a well-differentiated tumors (p < 0.0001), histology subtype of adenocarcinomas (p = 0.0010) and tumors in left colon (p = 0.0009). Over expression of pro apoptotic markers: p27KIP1 and KRAS4A isoforms was significantly higher in CRC subset with TRAIL-R1 and TRAIL-R2 expression; TRAIL-R1 expression was also associated with cleaved caspase-3(p = 0.0011). Interestingly, TRAIL-R2 expression was associated with a microsatellite stable (MS--S/L) phenotype (p = 0.0003) and with absence of KRAS mutations (p = 0.0481). CONCLUSION: TRAIL-R1 expression was an independent prognostic marker for better survival in all CRC samples and even in the CRC group that received adjuvant therapy. The biological effects of TRAIL in CRC models, its enhancement of chemosensitivity towards standard chemotherapeutic agents and the effect of endogenous TRAIL receptor levels on survival make TRAIL an extremely attractive therapeutic target.

A multicentre matched case control study of risk factors for preeclampsia in healthy women in Pakistan.

BACKGROUND: Preeclampsia is one of the leading causes of maternal and perinatal morbidity and mortality world-wide. The risk for developing preeclampsia varies depending on the underlying mechanism. Because the disorder is heterogeneous, the pathogenesis can differ in women with various risk factors. Understanding these mechanisms of disease responsible for preeclampsia as well as risk assessment is still a major challenge. The aim of this study was to determine the risk factors associated with preeclampsia, in healthy women in maternity hospitals of Karachi and Rawalpindi. METHODS: We conducted a hospital based matched case-control study to assess the factors associated with preeclampsia in Karachi and Rawalpindi, from January 2006 to December 2007. 131 hospital-reported cases of PE and 262 controls without history of preeclampsia were enrolled within 3 days of delivery. Cases and controls were matched on the hospital, day of delivery and parity. Potential risk factors for preeclampsia were ascertained during in-person postpartum interviews using a structured questionnaire and by medical record abstraction. Conditional logistic regression was used to estimate matched odds ratios (ORs) and 95% confidence intervals (95% CIs). RESULTS: In multivariate analysis, women having a family history of hypertension (adjusted OR 2.06, 95% CI; 1.27-3.35), gestational diabetes (adjusted OR 6.57, 95% CI; 1.94 -22.25), pre-gestational diabetes (adjusted OR 7.36, 95% CI; 1.37-33.66) and mental stress during pregnancy (adjusted OR 1.32; 95% CI; 1.19-1.46, for each 5 unit increase in Perceived stress scale score) were at increased risk of preeclampsia. However, high body mass index, maternal age, urinary tract infection, use of condoms prior to index pregnancy and sociodemographic factors were not associated with higher risk of having preeclampsia. CONCLUSIONS: Development of preeclampsia was associated with gestational diabetes, pregestational diabetes, family history of hypertension and mental stress during pregnancy. These factors can be used as a screening tool for preeclampsia prediction. Identification of the above mentioned predictors would enhance the ability to diagnose and monitor women likely to develop preeclampsia before the onset of disease for timely interventions and better maternal and fetal outcomes.