Multiplatform genomic analyses have identified 93 frequently altered genes in breast cancer. Of these, as many as 49 genes are directly or indirectly involved in transcription. These include constitutive and inducible DNA-binding transcription factors (DB-TFs, 13 genes), corepressors/coactivators (14 genes), epigenetic (10), and mediator/splicing/rRNA (3) factors. At least nine additional genes are immediate upstream regulators of transcriptional cofactors. G:profiler analysis reveals that these alterations affect cell cycle, development/differentiation, steroid hormone, and chromatin modification pathways. A notable observation is that DB-TFs that mediate major oncogenic signaling (e.g., WNT, receptor tyrosine kinase (RTK), NOTCH, and HIPPO), which switch from default repression (signal OFF) to transcriptional activation (signal ON), are not altered in breast cancer. Instead, corepressors (e.g., pRb for E2F1 downstream of various proliferation signals) or upstream factors (e.g., APC and AXIN for TCF, downstream of canonical WNT signaling) are lost, or coactivators (e.g., NOTCH1/2 for CSL/RBPJk) are induced. In contrast, constitutive (MYC, TBX3) and signal-induced (TP53, FOXA1) DB-TFs that do not mediate default repression are directly altered in breast cancer. Some of these TFs have been implicated in the establishment of super-enhancers and positive transcriptional elongation. In addition, oncogenic transcription is induced by mutations affecting regulatory elements or chromatin conformation that create new TF-binding sites in promoters and enhancers of oncogenic genes to promote tumorigenesis. Here we review these diverse oncogenic alterations in TFs in BC and discuss implications for therapy.

We present a comprehensive comparison of PAX5,IKZF1, and CDKN2A/B abnormalities in 21 B-cell precursor acute lymphoblastic leukemia (B-ALL) patients studied by aCGH and gene-specific FISH assays. In our cohort of B-ALL patients, alterations of IKZF1, PAX5, and CDKN2A/B were detected by aCGH analysis in 43, 52, and 57% of samples, respectively. Deletions of IKZF1 were present in 9 samples, including 5 cases positive for both PAX5 and IKZF1 deletions, implying digenic impairment. Furthermore, all cases with IKZF1 deletions also had additional genomic alterations, including BCR-ABL1 gene fusions, PAX5 deletions, CDKN2A/B deletions, and FLT3 amplification. Deletions of CDKN2A/B represented the most frequent abnormalities in our group of patients. Our study demonstrates the high incidence of PAX5, IKZF1, and CDKN2A/B alterations in B-ALL detected by aCGH analysis. Due to the small size and variability in the deletion breakpoints, FISH studies showed false-negative results in 10, 40, and 28% of the samples tested for the IKZF1,PAX5, and CDKN2A/B gene deletions, respectively. The PAX5 and IKZF1 abnormalities are highly specific to B-ALL and can be used as diagnostic markers. Moreover, IKZF1 alterations frequently coexist with a BCR-ABL gene fusion. Our study revealed multiple additional B-ALL-specific genomic alterations and showed that aCGH is a more sensitive method than FISH, allowing whole genome profiling and identification of aberrations of diagnostic and prognostic significance in patients with B-ALL.

MicroRNA (miR)-19a, a member of the miR-17-92 cluster, functions as an oncomiRNA in multiple kinds of cancers. However, its involvement in human osteosarcomas remains unclear. In this study, to analyze the expression pattern of miR-19a and to investigate its clinical implication in human osteosarcomas, quantitative reverse-transcription polymerase chain reaction was performed to detect expression levels of miR-19a in 166 self-pairs of osteosarcoma and noncancerous bone tissues. Associations between miR-19a expression and various clinicopathological parameters and patients' prognosis of osteosarcomas were further evaluated. As a results, miR-19a expression in osteosarcoma tissues was significantly higher than that in corresponding noncancerous bone tissues (P<0.001). Osteosarcoma patients with high miR-19a expression more frequently had large tumor size (P=0.03), advanced clinical stage (P=0.01), positive distant metastasis (P=0.008) and poor response to chemotherapy (P=0.01) than those with low miR-19a expression. Additionally, kaplan-Meier analysis showed that both overall and disease-free survivals of osteosarcoma patients with high miR-19a expression were shorter than those with low miR-19a expression (both P<0.001). Further multivariate analysis identified miR-19a expression as an independent prognostic factor for both overall (P=0.001) and disease-free (P=0.006) survivals. In conclusion, the aberrant expression of miR-19a may play a crucial role in development and progression of human osteosarcomas. MiR-19a may act as a novel prognostic marker for patients with this malignancy.

The epithelial to mesenchymal transition (EMT) is associated with acquired resistance to epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) in certain non-small cell lung cancers that harbor EGFR mutations. Because no currently available drugs specifically kill cancer cells via EMT, novel treatment strategies that overcome or prevent EMT are needed. A recent report suggested that dasatinib (an ABL/Src kinase inhibitor) inhibits EMT induced by transforming growth factor (TGF)-beta in lung cancer cells (Wilson et al., 2014). In this study, we analyzed effects of dasatinib on the resistance mechanism in HCC4006 cells, which tend to acquire resistance to EGFR-TKIs via EMT.

The Phase III, randomized, open-label IPASS study (NCT00322452) of first-line epidermal growth factor receptor tyrosine kinase inhibitor (EGFR TKI) gefitinib versus carboplatin/paclitaxel for Asian patients with advanced non-small-cell lung cancer (NSCLC) showed that investigator-assessed progression-free survival (PFS) and objective response rate (ORR) were significantly prolonged in patients with EGFR mutation-positive NSCLC who received gefitinib versus patients with EGFR mutation-negative NSCLC. We report post-hoc analyses of IPASS data by blind independent central review (BICR), performed at the request of the US FDA, in a subset of patients with EGFR mutation-positive NSCLC.

Programmed cell death-ligand 1 (PD-L1) expressed in tumor tissues is a key molecule for immune suppression, given its role in immune checkpoints. The significance and implication of soluble PD-L1 (sPD-L1) in the blood of lung cancer patients remain unknown.

Chromosomal instability (CIN) is classically defined as an increase in the rate at which numerical or structural chromosomal aberrations are acquired in a cancer cell. The number of somatic copy number abnormalities (CNAs) revealed by high resolution genomic array can be considered as a surrogate marker for CIN, but several points, related to sample processing and data analysis, need to be standardized. In this work we analyzed 51 CRC samples and matched normal mucosae by whole genome SNP arrays and compared different bioinformatics tools in order to identify broad (>25% of a chromosomal arm) and focal somatic copy number abnormalities (BCNAs and FCNAs respectively). In 15 tumors, two samples, separated by at least 1 cm, were taken from the same tumor mass (double-sampling pairs) in order to evaluate differences in detection of chromosomal abnormalities between distant regions of the same tumor and their influence on CIN quantitative and qualitative analysis. Our data show a high degree of correlation of the quantitative CIN index (somatic BCNA number) between distant tumor regions. On the contrary, a lower correlation is observed in terms of chromosomal distribution of BCNAs, as summarized by a simplified cytogenetic table. Quantitative or qualitative analysis of FCNAs, including homozygous deletions and high level amplifications, did not add further information on the CIN status. The use of the index "somatic BCNA number" can be proposed for a robust classification of tumors as CIN positive or negative even in the presence of a significant tumor regional heterogeneity.

A next-generation FISH probe mapping to the MDM2 locus-specific region has recently been designed. The level of MDM2 gene amplification (high versus low) may allow selection of patients for cancer treatment with MDM2 inhibitors and may predict their responsiveness. We investigated the spectrum of MDM2 gene alterations using the new probes in vivo after visualizing single neoplastic cells in situ from a series of glioblastomas. Signals from next-generation repeat-free FISH interphase probes were identified in tissue microarrays that included 3 spots for each of the 48 cases. The murine double minutes (MDM2)-specific DNA probe and the satellite enumeration probe for chromosome 12 were used. Three cases (6%) showed more than 25 signals (high gene amplification), and 7 (15%) showed 3-10 signals (gains); among these, 4 cases (8%) had an equal number of MDM2 and centromeric signals on chromosome 12 (polyploidy). Genomic heterogeneity was observed only in 3 cases with low gene amplification. In our series, 6% of glioblastomas exhibited high MDM2 amplification (in vivo) with a pattern related to the known double minutes/chromothripsis phenomenon (in situ), and only cases with low amplification showed genomic heterogeneity. We concluded that the rate of MDM2 gene amplification can be a useful predictive biomarker to improve patient selection.

Germline mutations in the BRCA tumor suppressor genes account for a substantial proportion of hereditary breast/ovarian cancer. However, this contribution is lower than expected. This underestimation can partly be explained by the BRCA alterations missed by using Sanger sequencing methods. Thus, large genomic rearrangements (LGRs) in BRCA1 and BRCA2 are responsible for 4-28% of all inherited BRCA mutations. In this study, Multiplex ligation-dependent probe amplification (MLPA) assay was used for detection of large rearrangements of BRCA1 and BRCA2 genes in 36 unrelated high-risk breast/ovarian cancer patients negative for BRCA1/2 point mutations. MLPA assay for all exons of both genes and for 1100delC variant of CHEK2 gene were performed. Positive MLPA results were confirmed by real-time quantitative PCR (qPCR). Two different rearrangements in the BRCA1 gene were identified consisting of exon 5 deletion and exon 20 duplication. MLPA analysis did not reveal any large genomic rearrangements in BRCA2 gene. Overall BRCA1/2 LGRs prevalence among high-risk Tunisian patients was 5.5%. Quantitative real-time PCR confirmed MPLA findings. Our results suggest the usefulness of screening for LGRs in BRCA genes in the Tunisian population. To avoid false-negative results, we suggest that MLPA should be used in genetic testing programs. These results are important for guidance counseling and clinical management of Tunisian high-risk patients.

This study evaluates prognostic markers as predictors of clonal evolution (CE) and assesses the impact of CE on overall survival (OS) in a population-based cohort of 159 consecutive eligible patients with chronic lymphocytic leukemia (CLL) obtained from the British Columbia Provincial CLL Database. CE was detected by interphase fluorescence in situ hybridization (FISH) in 34/159 patients (21%) with 65% of CE patients acquiring deletion 17p or 11q. CD38 positive status (≥30%) on flow cytometry predicted 2.7 times increased risk of high-risk CE (acquisition of deletion 17p or 11q) on multivariate analysis. Prior CLL therapy was not a significant predictor of CE. CE was associated with 4.1 times greater risk of death when analyzed as a time-dependent variable for OS after adjusting for age, lymphocyte count, and FISH timing. High-risk CE was associated with worse OS while acquisition of low/intermediate-risk abnormalities (trisomy 12, deletion 13q, and IGH translocation) had no difference in OS. Our study demonstrates the negative impact of CE detected by FISH on OS in this population-based cohort. These data provide support for repeating FISH testing during CLL follow-up as patients with high-risk CE have reduced survival and may require closer observation.

Gene fusions and their encoded products (fusion RNAs and proteins) are viewed as one of the hallmarks of cancer. Traditionally, they were thought to be generated solely by chromosomal rearrangements. However, recent discoveries of trans-splicing and cis-splicing events between neighboring genes, suggest that there are other mechanisms to generate chimeric fusion RNAs without corresponding changes in DNA. In addition, chimeric RNAs have been detected in normal physiology, complicating the use of fusions in cancer detection and therapy. On the other hand, "intergenically spliced" fusion RNAs represent a new repertoire of biomarkers and therapeutic targets. Here, we review current knowledge on chimeric RNAs and implications for cancer detection and treatment, and discuss outstanding questions for the advancement of the field.

Cancer of the gastroesophageal junction (GEJ), although rare, is now considered a separate entity with a distinct pathophysiological and molecular profile. Although much progress has been made over the past decades in delineating the multiple environmental and genetic pathways involved GEJ carcinoma, the exact molecular mechanisms underlying disease initiation and progression are still poorly understood. This is of paramount importance for the treating physician as the disease bears a poor therapeutic response. This review defines the GEJ and types of GEJ carcinoma, and provides useful insight in its pathophysiology. Future aspects include better understanding of GEJ oncogenesis, early detection of precursor lesions, the use of biomarkers and targeted therapy (through molecular profiling) so as to increase overall survival. (Acta gastroenterol. belg., 2016, 79, 471-479).

We present the case of a 76-year-old male found to have a large tumor involving the left lateral lobe of the liver, presumed to be hepatocellular carcinoma (HCC). After resection, pathologic features demonstrated both high-grade HCC and high-grade neuroendocrine carcinoma (NEC). Areas of NEC stained strongly for synaptophysin, which was not present in HCC component. The HCC component stained strongly for Hep-Par 1, which was not present in the NEC component. The patient underwent genetic analysis for biomarkers common to both tumor cell types. Both tumor components contained gene mutations in CTNNB1 gene (S33F located in exon 3). They also shared mutations in PD-1, PGP, and SMO. Mixed HCC/NEC tumors have been rarely reported in the literature with generally poor outcomes. This patient has been referred for adjuvant platinum-based chemotherapy; genetic biomarker analysis may provide some insight to guide targeted chemotherapy.

FHIT, located at FRA3B, is one of the most commonly deleted genes in human cancers, and loss of FHIT protein is one of the earliest events in cancer initiation. However, location of FHIT at a chromosomal fragile site, a locus prone to breakage and gap formation under even mild replication stress, has encouraged claims that FHIT loss is a passenger event in cancers. We summarize accumulated evidence that FHIT protein functions as a genome "caretaker" required to protect the stability of genomes of normal cells of most tissues from agents causing intrinsic and extrinsic DNA damage. FHIT loss leads to intracellular replication stress and subsequent genome instability, which provides an opportunistic mutational landscape in preneoplasias for selection of a variety of other cancer-driving mutations. We also review evidence showing that FHIT loss leads to enhanced activation of other common fragile sites, including the FRA16D/WWOX locus, and creates optimal single-stranded DNA substrates for the hypermutator enzyme, APOBEC3B.

Microsatellite instability (MSI) is a hypermutable phenotype that usually arises from either a germline mutation in components of the mismatch repair (MMR) machinery (i.e. hMLH1, MSH2, MSH6 and PMS2) in patients with Lynch syndrome (LS) or somatic hypermethylation of the hMLH1 promoter in sporadic carcinomas. In all colorectal cancers (CRC) is possible to identify the MMR deficiency through protein expression by immunoistochemistry (IHC). Recently, the predictive role of MMR deficiency in reduced chemotherapy benefit and the introduction of universal screening for Lynch syndrome suggest to include MMR testing into routine clinical practice. In this scenario is mandatory to update the minimal requirements for MMR IHC standardization and evaluation. According to international guidelines, these are the GIPAD and AIFEG suggestions for MMR IHC testing.

Gastrointestinal stromal tumors (GISTs) are the most common mesenchymal tumors of the gastrointestinal tract. The discovery of the occurrence of activating KIT mutations and KIT expression in GISTs opened the way to the unequivocal diagnosis of these tumors and to their successful treatment with imatinib, a tyrosin kinase inhibitor. Since then, research progress revealed molecular GIST triggers alternative to KIT, implying heterogeneous analytic approaches and prognostic expectations. Several targeted therapies, variably specific for each GIST trigger, have been developed or are being investigated. Thus, GISTs eventually revealed a family of diseases rather than a single tumor type. All these events had an unprecedented impact on pathology practice, constituting at the same time a heavy burden and an exciting challenge, ultimately putting pathologists in the spotlight as never before. This review will discuss the most recent advances concerning GISTs, highlighting the tasks of pathologists facing these tumors, with an emphasis on traps potentially compromising a correct diagnosis.