HER-2/neu Testing and Therapy in Gastroesophageal Adenocarcinoma

Despite ongoing advances in the treatment of gastroesophageal cancer, prognosis remains poor. The best promise to improve this poor survival is provided by new targeted agents. Of these, human epidermal growth factor receptor 2 (HER2) is currently in the spotlight. In this review, we provide an overview of recent developments in HER2 testing and results of clinical trials targeting HER2 in gastroesophageal adenocarcinoma. Based on the encouraging ToGA trial findings it is now expected that routine HER2 testing will be included in the diagnostic work-up of patients with advanced gastric cancer. With regard to this testing, overexpression of the HER2 protein seems to possess the best predictive properties. However, HER2 immunohistochemistry (IHC) is subject to assay and interobserver variability, so standardization and internal and external proficiency testing is an absolute prerequisite, especially as the IHC scoring system in gastric cancer is different from that of breast cancer. Further study is needed to investigate the clinical meaning of the significant heterogeneity observed in both gene amplification and protein overexpression in gastroesophageal cancer. Highly effective therapies for gastroesophageal cancer can only be accomplished by a multi-targeted approach, considering crosstalk between pathways and continuing to optimize chemotherapy

Low frequency of HER2 amplification and overexpression in early onset gastric cancer

BACKGROUND: The recent ToGA trial results indicated that trastuzumab is a new, effective, and well-tolerated treatment for HER2-positive gastric cancer (GC). Although GC mainly affects older patients, fewer than 10% of GC patients are considered early-onset (EOGC) (presenting at the age of 45 years or younger). These EOGC show different clinicopathological and molecular profiles compared to late onset GC suggesting that they represent a separate entity within gastric carcinogenesis. In light of potential trastuzumab benefit, subpopulations of GC such as EOGC (versus late onset) should be evaluated for their frequency of amplification and overexpression using currently available techniques. METHODS: Tissue microarray (TMA) blocks of 108 early onset GC and 91 late onset GC were stained by immunohistochemistry (IHC, Hercep test, DAKO) and chromogenic in situ hybridization (CISH, SPoT-Light, Invitrogen). RESULTS: Overall, we found only 5% HER2 high level amplification and 3% HER2 3+ overexpression (6/199). In addition, 8 patients (4%) showed a low level CISH amplification and 9 patients (4.5%) showed a 2+ IHC score. IHC and CISH showed 92% concordance and CISH showed less heterogeneity than IHC. In 2/199 cases (1%), IHC showed clinically relevant heterogeneity between TMA cores, but all cases with focal IHC 3+ expression were uniformly CISH high level amplified. Early onset GCs showed a significantly lower frequency of HER2 amplification (2%) and overexpression (0%) than late onset GCs (8% and 7% respectively) (p = 0.085 and p = 0.008 respectively). Proximal GC had more HER2 amplification (9% versus 3%) and overexpression (7% versus 2%) than distal tumours although this difference was not significant (p = 0.181 and p = 0.182 respectively). HER2 CISH showed more high level amplification in the intestinal type (7%, 16% if low-level included) compared to the mixed (5%, 5% if low-level included) and diffuse type (3%, 4% if low-level included) GCs (p = 0.029). A similar association was seen for HER2 IHC and histologic type (p = 0.008). Logistic regression indicated a significant association between HER2 expression and age, which remained significant when adjusted for both location and histological type. CONCLUSIONS: Even focal HER2 overexpression in GC points to uniform HER2 amplification by CISH. We show for the first time that early onset GC has a lower frequency of HER2 amplification and overexpression than late onset GC, and confirm that intestinal type GC shows the highest rate of HER2 amplification and overexpression

High-throughput sequencing of gastric cancer patients : unravelling genetic predispositions towards an early-onset subtype

Background: Gastric cancer is the fourth most common cause of cancer-related death. Currently, it is broadly accepted that the molecular complexity and heterogeneity of gastric cancer, both inter- and intra-tumor, display important barriers for finding specific biomarkers for the early detection and diagnosis of this malignancy. Early-onset gastric cancer is not as prevalent as conventional gastric carcinoma, but it is a preferable model for studying the genetic background, as young patients are less exposed to environmental factors, which influence cancer development. Aim: The main objective of this study was to reveal age-dependent genotypic characteristics of gastric cancer subtypes, as well as conduct mutation profiling for the most frequent alterations in gastric cancer development, using targeted next-generation sequencing technology. Patients and methods: The study group included 53 patients, consisting of 18 patients with conventional gastric cancer and 35 with an early-onset subtype. The DNA of all index cases was used for next-generation sequencing, employing a panel of 94 genes and 284 single nucleotide polymorphisms (SNPs) (TruSight Cancer Panel, Illumina), which is characteristic for common and rare types of cancer. Results: From among the 53 samples processed for sequencing, we were able to identify seven candidate genes (STK11, RET, FANCM, SLX4, WRN, MEN1, and KIT) and nine variants among them: one splice_acceptor, four synonymous, and four missense variants. These were selected for the age-dependent differentiation of gastric cancer subtypes. We found four variants with C-Score ≥ 10, as 10% of the most deleterious substitutions: rs1800862 (RET), rs10138997 (FANCM), rs2230009 (WRN), and rs2959656 (MEN1). We identified 36 different variants, among 24 different genes, which were the most frequent genetic alterations among study subjects. We found 16 different variants among the genes that were present in 100% of the total cohort: SDHB (rs2746462), ALK (rs1670283), XPC (rs2958057), RECQL4 (rs4925828; rs11342077, rs398010167; rs2721190), DDB2 (rs326212), MEN1 (rs540012), AIP (rs4930199), ATM (rs659243), HNF1A (rs1169305), BRCA2 (rs206075; rs169547), ERCC5 (rs9514066; rs9514067), and FANCI (rs7183618). Conclusions: The technology of next-generation sequencing is a useful tool for studying the development and progression of gastric carcinoma in a high-throughput way. Our study revealed that early-onset gastric cancer has a different mutation frequency profile in certain genes compared to conventional subtype

Multiplex spatial omics reveals changes in immune-epithelial crosstalk during inflammation and dysplasia development in chronic IBD patients

Patients with long-standing inflammatory bowel disease (IBD) face an increased risk of developing colitis-associated cancer (CAC). Although IBD-induced prolonged inflammation seems to be involved in CAC pathogenesis, the specific molecular changes that contribute remain unknown. Here, we applied digital spatial RNA profiling, RNAscope, and imaging mass cytometry to examine paired uninflamed, inflamed, and early dysplastic mucosa of patients with IBD. We observed robust type 3 (IL-17) responses during inflammation, accompanied by elevated JAK-STAT signaling and phosphorylated STAT3 (P-STAT3) levels, with both inflamed and dysplastic mucosa displaying immune cell activation. Higher stromal P-STAT3 was detected in uninflamed and inflamed mucosa of patients who eventually developed dysplasia. CD8a+ T cells did not infiltrate inflamed or dysplastic epithelial regions in these patients, while control patients showed elevated CD8a in inflamed mucosa. Our study reveals distinct inflammatory patterns throughout CAC development, marked by an activated IL-17 pathway, engaged STAT3, and diminished cytotoxic T cell infiltration

Performance of gastrointestinal pathologists within a national digital review panel for Barrett's oesophagus in the Netherlands: Results of 80 prospective biopsy reviews

Aims: The histopathological diagnosis of low-grade dysplasia (LGD) in Barrett's oesophagus (BO) is associated with poor interobserver agreement and guidelines dictate expert review. To facilitate nationwide expert review in the Netherlands, a web-based digital review panel has been set up, which currently consists of eight 'core' pathologists. The aim of this study was to evaluate if other pathologists from the Dutch BO expert centres qualify for the expert panel by assessing their performance in 80 consecutive LGD reviews submitted to the panel. Methods: Pathologists independently assessed digital slides in two phases. Both phases consisted of 40 cases, with a group discussion after phase I. For all cases, a previous consensus diagnosis made by five core pathologists was available, which was used as reference. The following criteria were used: (1) percentage of 'indefinite for dysplasia' diagnoses, (2) percentage agreement with consensus diagnosis and (3) proportion of cases with a consensus diagnosis of dysplasia underdiagnosed as non-dysplastic. Benchmarks were based on scores of the core pathologists. Results: After phase I, 1/7 pathologists met the benchmark scor

LKB1 as the ghostwriter of crypt history

Familial cancer syndromes present rare insights into malignant tumor development. The molecular background of polyp formation and the cancer prone state in Peutz-Jeghers syndrome remain enigmatic to this day. Previously, we proposed that Peutz-Jeghers polyps are not pre-malignant lesions, but an epiphenomenon to the malignant condition. However, Peutz-Jeghers polyp formation and the cancer-prone state must both be accounted for by the same molecular mechanism. Our contribution focuses on the histopathology of the characteristic Peutz-Jeghers polyp and recent research on stem cell dynamics and how these concepts relate to Peutz-Jeghers polyposis. We discuss a protracted clonal evolution scenario in Peutz-Jeghers syndrome due to a germline LKB1 mutation. Peutz-Jeghers polyp formation and malignant transformation are separately mediated through the same molecular mechanism played out on different timescales. Thus, a single mechanism accounts for the development of benign Peutz-Jeghers polyps and for malignant transformation in Peutz-Jeghers syndrome

A novel pathogenic MLH1 missense mutation, c.112A > C, p.Asn38His, in six families with Lynch syndrome

<p>Abstract</p> <p>Background</p> <p>An unclassified variant (UV) in exon 1 of the <it>MLH1 </it>gene, c.112A > C, p.Asn38His, was found in six families who meet diagnostic criteria for Lynch syndrome. The pathogenicity of this variant was unknown. We aim to elucidate the pathogenicity of this <it>MLH1 </it>variant in order to counsel these families adequately and to enable predictive testing in healthy at-risk relatives.</p> <p>Methods</p> <p>We studied clinical data, microsatellite instability and immunohistochemical staining of MMR proteins, and performed genealogy, haplotype analysis and DNA testing of control samples.</p> <p>Results</p> <p>The UV showed co-segregation with the disease in all families. All investigated tumors showed a microsatellite instable pattern. Immunohistochemical data were variable among tested tumors. Three families had a common ancestor and all families originated from the same geographical area in The Netherlands. Haplotype analysis showed a common haplotype in all six families.</p> <p>Conclusions</p> <p>We conclude that the <it>MLH1 </it>variant is a pathogenic mutation and genealogy and haplotype analysis results strongly suggest that it is a Dutch founder mutation. Our findings imply that predictive testing can be offered to healthy family members. The immunohistochemical data of MMR protein expression show that interpreting these results in case of a missense mutation should be done with caution.</p

Genomic characterization of malignant progression in neoplastic pancreatic cysts

Intraductal papillary mucinous neoplasms (IPMNs) and mucinous cystic neoplasms (MCNs) are non-invasive neoplasms that are often observed in association with invasive pancreatic cancers, but their origins and evolutionary relationships are poorly understood. In this study, we analyze 148 samples from IPMNs, MCNs, and small associated invasive carcinomas from 18 patients using whole exome or targeted sequencing. Using evolutionary analyses, we establish that both IPMNs and MCNs are direct precursors to pancreatic cancer. Mutations in SMAD4 and TGFBR2 are frequently restricted to invasive carcinoma, while RNF43 alterations are largely in non-invasive lesions. Genomic analyses suggest an average window of over three years between the development of high-grade dysplasia and pancreatic cancer. Taken together, these data establish non-invasive IPMNs and MCNs as origins of invasive pancreatic cancer, identifying potential drivers of invasion, highlighting the complex clonal dynamics prior to malignant transformation, and providing opportunities for early detection and intervention

Validation of tissue microarray technology in squamous cell carcinoma of the esophagus

Tissue microarray (TMA) technology has been developed to facilitate high-throughput immunohistochemical and in situ hybridization analysis of tissues by inserting small tissue biopsy cores into a single paraffin block. Several studies have revealed novel prognostic biomarkers in esophageal squamous cell carcinoma (ESCC) by means of TMA technology, although this technique has not yet been validated for these tumors. Because representativeness of the donor tissue cores may be a disadvantage compared to full sections, the aim of this study was to assess if TMA technology provides representative immunohistochemical results in ESCC. A TMA was constructed containing triplicate cores of 108 formalin-fixed, paraffin-embedded squamous cell carcinomas of the esophagus. The agreement in the differentiation grade and immunohistochemical staining scores of CK5/6, CK14, E-cadherin, Ki-67, and p53 between TMA cores and a subset of 64 randomly selected donor paraffin blocks was determined using kappa statistics. The concurrence between TMA cores and donor blocks was moderate for Ki-67 (κ = 0.42) and E-cadherin (κ = 0.47), substantial for differentiation grade (κ = 0.65) and CK14 (κ = 0.71), and almost perfect for p53 (κ = 0.86) and CK5/6 (κ = 0.93). TMA technology appears to be a valid method for immunohistochemical analysis of molecular markers in ESCC provided that the staining pattern in the tumor is homogeneous