Low level of microsatellite instability correlates with short disease-free survival of gastric cancer patients undergoing neoadjuvant chemotherapy.

Limited studies have been reported about the function of low level of microsatellite instability (MSI-L) in cancer. The aim of our study is to unveil the prognostic role of MSI-L in gastric cancer (GC). One hundred nine patients with locally advanced GC (T3-4a, N+, M0) who underwent neoadjuvant chemotherapy plus gastrectomy with extended (D2) lymph node dissection were collected. Clinicopathological characteristics, tumour regression score, disease-free survival (DFS), and overall survival (OS) were analysed and correlated with the MSI status. The MSI status of 96 patients was identified (7 (7.3%) with MSI-H, 12 (12.5%) with MSI-L, and 77 (80.2%) with MSS). MSI-L was significantly correlated with perineural invasion (P = 0.009) and decreased MUC5AC expression (P = 0.042). Poor response to neoadjuvant chemotherapy in MSI-L patients (83.3% assessed as poor response) was observed (P = 0.501). Compared with patients with MSS tumours, patients with MSI-L tumours showed poor DFS (P = 0.018) with a hazard ratio (HR) of 2.839 (95% CI 1.131-7.124, P = 0.026) from multivariable cox regression analysis. However, this was not associated with OS (P = 0.063). MSI-L is an independent poor prognostic biomarker for the locally advanced gastric cancer treated with neoadjuvant chemotherapy. Further studies with larger sample sizes are needed for validation.

Early-onset colorectal cancer: A distinct entity with unique genetic features.

The aim of the present study was to elucidate the genetic features of early-onset colorectal cancer (CRC), particularly the genetic mutations that may be regarded as prognostic and/or predictive markers in CRC and other malignancies. In total, 40 patients with non-polyposis CRC aged 35 or younger were selected. The formalin-fixed, paraffin-embedded tumors acquired were subjected to mismatch repair (MMR) protein immunochemical staining and gene analysis with next-generation sequencing (44 exons, 17 genes; Ion Torrent Sequencing Platform). A total of 11 (27.5%) tumors presented with MMR protein deficiency (dMMR) and 26 (65%) tumors harbored one or more genetic mutations, including K-RAS proto-oncogene (35%), phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA; 20%), B-Raf proto-oncogene (5%), erb-b2 receptor tyrosine kinase 2 (5%), discoidin domain receptor tyrosine kinase 2 (5%), N-RAS proto-oncogene (2.5%), KIT proto-oncogene (2.5%), TSC complex subunit 1 (2.5%), DNA methyltransferase 3 alpha (2.5%) and ABL proto-oncogene 1 (2.5%). Of the dMMR tumors, 81.8% (9/11) of cases presented with mutations in the tested genes, while only 58.6% (17/29) of the MMR-proficient (pMMR) tumors presented with these (P=0.158). PI3KCA was frequently mutated in dMMR tumors compared to pMMR tumors (P=0.025). In a subgroup with a family history of CRC, the dMMR status (P<0.001) and PIK3CA genetic mutation status (P=0.01) were more frequently observed compared to the other two groups (with a family history of other cancer types or no malignancy). Almost all patients who had relatives with CRC presented with both dMMR and other genetic mutations, while this was not observed in the patients who had relatives with other types of carcinoma. Certain genetic mutations that are rarely reported in CRC were only identified in those patients with a family history of carcinoma. In conclusion, non-polyposis CRC in young adults presents as a distinct entity with a unique set of genetic features. However, investigation of more cases in further studies is required to verify the present results.

Prediction model of the response to neoadjuvant chemotherapy in breast cancers by a Naive Bayes algorithm.

BACKGROUND AND OBJECTIVE: Chemotherapy is useful to many breast cancer patients, however, it is not therapeutic for some patients. Pathologic complete response (pCR) is an indicator to good response in Neoadjuvant chemotherapy (NAC). In this study, we aimed to develop a way to predict pCR before NAC. METHODS: We retrospectively collected 287 stage II-III breast cancer cases either to a training set (N = 197) or to a test set (N = 90). Fourteen candidate genes were selected from four public microarray data sets. A prediction model was built, by using these fourteen candidate genes and three reference genes expression which were tested by TaqMan probe-based quantitative polymerase chain reaction, after selecting a better algorithm. RESULTS: The Naive Bayes algorithm had a relatively higher predictive value, compared with random forest, support vector machine (SVM), and k-nearest neighbor (knn) algorithms (P < 0.05). This 17-gene prediction model showed a high positive correlation with pCR (odds ratio, 8.914, 95% confidence interval, 4.430-17.934, P < 0.001). By using this model, the enrolled patients were classified into sensitive (SE) and insensitive (INS) groups. The pCR rates between the SE and INS groups were highly different (42.3% vs.7.6%, P < 0.001). The sensitivity and specificity of this prediction model were 84.5% and 62.0%. CONCLUSIONS: Instead of whole transcriptome-based technologies, panel gene expression with tens of essential genes implemented in a machine learning model has predictive potential for chemosensitivity in breast cancers.

[BRAF V600E Mutation and TERT Promoter Mutation in Papillary Thyroid Carcinomas and Their Association with Clinicopathological Characteristics].

OBJECTIVE: To explore the relationships of BRAF V600E and TERT promoter mutations with the clinicopathological features in papillary thyroid carcinoma (PTC). METHODS: The mutations of BRAF V600E and TERT promoters were examined by PCR-direct sequencing in tumor tissues from 326 PTC patients, while the relationships between the gene mutations and clinicopathological features were analyzed. RESULTS: BRAF V600E mutation was found in 269/326 (82.52%), and TERT promoter mutation in 11/326 (3.37%) of PTC patients. In site mutations of TERT promoter, 9 cases were C228T and 2 cases were C250T. Single factor analysis showed that BRAF V600E mutations were significantly associated with age and recurrence/distant metastasis of tumor (P < 0.05), while TERT promoter mutations were significantly associated with age, tumor size, extrathyroidal extension, T stage, AJCC stage and recurrence/distant metastasis of tumor (P < 0.05). Coexistence of BRAF V600E and TERT promoter mutations (BRAF+/TERT+) were particularly associated with age, tumor size, extrathyroidal extension, T stage and AJCC stage (P < 0.05). CONCLUSION: Coexistence of BRAF V600E and TERT promoter mutations in PTC shows more aggressive tumor behavior.

Mutation in BRAF and SMAD4 associated with resistance to neoadjuvant chemoradiation therapy in locally advanced rectal cancer.

Our study was done in order to identify novel molecular markers to predict which locally advanced rectal cancers (LARCs) might be resistant to neoadjuvant chemoradiotherapy (nCRT). Seventy-four patients with LARCs treated with nCRT were collected. Pathological evaluation after nCRT was performed according to the tumor regression grading (TRG) system. Next-generation sequencing kit including 279 exons of 59 genes was performed on Illumina Miseq Platform. Sanger sequencing was performed to confirm some mutations. Four of the tumors (4/74, 5.4%) had BRAF mutation, which presented in one TRG 2 tumor and three TRG 3 tumors but was not observed in TRG 0-1 tumors. Higher mutational frequency of BRAF gene in TRG 3 tumors (3/12, 25%) was found in comparison with the TRG 0-2 tumors (1/62, 1.6%; p = 0.012). Eight tumors (8/74, 10.8%) harbored SMAD4 mutations, which was mutated across all TRG groups. However, SMAD4 mutated more in TRG 3 tumors (4/12, 33.3%) compared with that in TRG 0-2 tumors (4/62, 6.5%; p = 0.020). The patients with BRAF-mutated LARCs had shorter progression-free survival (PFS) (p = 0.045) and shorter overall survival (OS) (p = 0.000) than the BRAF wild-type (WT) ones. The patients with SMAD4-mutated tumors had shorter PFS than the WT cases (p = 0.008). BRAF and SMAD4 genetic mutations might be important molecular markers to predict resistance to nCRT and poor prognosis in LARCs. More cases are needed to confirm these findings in the near future.

Whole exome and target sequencing identifies MAP2K5 as novel susceptibility gene for familial non-medullary thyroid carcinoma.

Although the genotype-phenotype for familial medullary thyroid carcinoma (FMTC) is well studied, only few low susceptibility risk loci were identified for familial non-medullary thyroid carcinoma (FNMTC). The aim of this study is to screen and identify high-penetrate genes for FNMTC. A total of 34 families with more than two first-degree relatives diagnosed as papillary thyroid cancer without other familial syndrome were recruited. Whole exome and target gene sequencing were performed for candidate variants. These variants were screened and analyzed with ESP6500, ExAC, 1000 genomes project, and the Cancer Genome Atlas (TCGA) with SIFT score and Polyphen2 prediction. Finally, we identified recurrent genetic mutation of MAP2K5 variants c.G961A and c.T1100C (p. A321T and p.M367 T) as susceptibility loci for FNMTC. The frequencies of MAP2K5 c.G961A and c.T1100C were found, 0.0385 and 0.0259 in FNMTC and 0 and 0.00022523 in healthy Chinese controls (n = 2200, P < 0.001), respectively. Both variants were located in the protein kinase domain. The functional study showed that MAP2K5 A321T or M367 T could consistently phosphorylate downstream protein ERK5 on site Ser731 + Thr733 or Ser496, promoting nuclear translocation and subsequently altering target gene expressions. Our data revealed that MAP2K5 variants A321T or M367 T can activate MAP2K5-ERK5 pathway, alter downstream gene expression, and subsequently induce thyroid epithelial cell malignant transformation. While classic MAP2K1/2(MEK1/2)-ERK1/2 signaling is well known for driving sporadic NMTC, our research indicated that MAP2K5 (MEK5) is a susceptibility gene for FNMTC. These findings highlight the potential application of MAP2K5 for molecular diagnosis as well as early prevention.

[BRAF-V600E mutation and its clinical significance in children with Langerhans cell histiocytosis].

OBJECTIVE: To investigate the clinical significance of BRAF-V600E mutation in children with Langerhans cell histiocytosis (LCH). METHODS: Real-time fluorescence quantitative PCR was used to detect BRAF-V600E mutation in paraffin-embedded tissue samples from 26 children with LCH. A retrospective analysis was performed for the association of BRAF-V600E mutation with clinical features and prognosis of children with LCH. RESULTS: Of the 26 children, 25 received standard chemotherapy, with a 2-year overall survival (OS) rate of 100% and a 2-year event-free survival (EFS) rate of 88%. Of the 26 pathological samples, 18 (70%) came from bone tissue, and the positive rate of BRAF-V600E mutation reached 50% (13/26). The positive rate of BRAF-V600E gene mutation was not associated with age, sex, affected organ, clinical classification, early treatment response, recurrence, and 2-year OS and EFS rates of the children with LCH (P>0.05), but it was associated with clinical grouping of LCH (P<0.05). CONCLUSIONS: Children with LCH tend to have a high OS rate and a high incidence rate of BRAF-V600E mutation. BRAF-V600E mutation is associated with clinical grouping of LCH.

Quantitative real-time polymerase chain reaction is an alternative method for the detection of HER-2 amplification in formalin-fixed paraffin-embedded breast cancer samples.

Fluorescent in situ hybridization (FISH) and immunohistochemistry (IHC) are the most common methods that are used to quantify HER-2 gene and protein levels, respectively, in human breast cancer. However, due to bad sample quality, some samples are unable to be subjected to a FISH assay. We evaluated 71 formalin-fixed paraffin-embedded (FFPE) breast carcinoma specimens by quantitative real-time polymerase chain reaction (qPCR), IHC, and FISH. We also performed qPCR and FISH assays on delayed formalin-fixed (DDF) samples. The qPCR results were in complete concordance with the results of IHC and FISH. In regards to the DDF samples, the HER-2 fluorescent signal seemed decayed compared with that of the DDF samples after 1 h. However, the qPCR method still works well up to 12 hours. Our results indicated that qPCR was obviously superior to FISH in cases that were not fixed in a reasonable amount of time. However, qPCR can be an alternative method by which to perform HER2 amplification assays in breast cancer.

The Presence of EpCAM(-)/CD49f(+) Cells in Breast Cancer Is Associated with a Poor Clinical Outcome.

PURPOSE: It is well established that breast cancer stem cells (BCSCs) play an essential role in tumor invasion for both local and distant metastasis. The aim of this study was to establish whether BCSCs could act as a prognostic and clinical marker. METHODS: We analyzed tumor tissues from 161 breast cancer patients. Dual immunohistochemistry and immunofluorescence were performed on all the slides, and we analyzed the relationship between EpCAM(-)/CD49f(+) tumor cells and key clinical and prognostic factors. RESULTS: Univariate survival analysis using the Kaplan-Meier method showed that the presence of EpCAM(-)/CD49f(+) tumor cells in breast cancer was significantly associated with shorter disease-free survival (DFS) and overall survival (OS). Multivariate analysis showed that the presence of EpCAM(-)/CD49f(+) cells was associated with shorter DFS (p=0.010; hazard ratio [HR], 2.070) and OS (p=0.002; HR, 3.235). Tumors containing EpCAM(-)/CD49f(+) cells were also more likely to metastasize after initial surgery (p=0.048). CONCLUSION: Our study suggests that breast tumors containing EpCAM(-)/CD49f(+) cells are more likely to undergo distant metastasis after initial surgery and are associated with a shorter DFS and OS.

The Clinicopathological Features of BRAF Mutated Papillary Thyroid Cancers in Chinese Patients.

The BRAF(V600E) mutation is commonly found in papillary thyroid cancers (PTCs) at different frequencies in different regions. However, the association between the BRAF(V600E) mutation and clinicopathological features in Chinese PTC patients is unknown. A total of 543 Chinese patients with histologically confirmed PTC were enrolled in this study. For the BRAF mutation assay, the target fragments were amplified and sequenced with an ABI 3500 gene analyzer. In 170 of 543 samples (31.3%), the BRAF(V600E) mutation was detected. In the bivariate analysis, the BRAF(V600E) mutation showed an association with bilaterality, tumor size, extrathyroidal invasion, and lymph node metastases (LNM). However, in the multivariate analysis, the BRAF(V600E) mutation was positively related to only tumor size (>1 cm) and extrathyroidal invasion. In addition, the multivariate analysis also showed that the age at diagnosis (<45 y) and tumor size (>1 cm) were independent predictors for LNM. In this study, the BRAF(V600E) mutation is positively associated with worse prognostic factors, including larger tumor size and the tumor extending to the thyroid capsule or extrathyroidal region; however, it is not an independent predictor for LNM.

The expression of aldehyde dehydrogenase family in breast cancer.

PURPOSE: It is widely accepted that aldehyde dehydrogenase (ALDH) activity is a signature of breast cancer stem cells, and high activity has been reported to be associated with poor clinical outcome. The aim of this study was to assess the expression of members of the ALDH family of isozymes in breast cancer tissues and to evaluate the implications of the results. METHODS: We analyzed paraffin-embedded tumor tissue from 160 patients with breast cancer. Immunohistochemistry (IHC) staining was performed on the slides using antibodies against different ALDH family members. We collated the IHC results with patient clinical characteristics and determined their prognostic value. In addition, we analyzed normal, hyperplastic, and carcinomatous tissues in situ to check their ALDH distributions. RESULTS: All the tested ALDH members were detected in the various tissue types, but at different levels. Only ALDH 1A3 was found to be significantly associated with distant metastasis (p=0.001), disease-free survival (p<0.001), and overall survival (p<0.001). CONCLUSION: The level of ALDH 1A3 in breast cancer tissue is a predictive marker of a poor clinical outcome.