ABSTRACT
The biological heterogeneity of neuroblastoma underscores the need for an in vitro model of each molecularly defined subgroup to investigate tumorigenesis and develop targeted therapies. We have established a permanently growing cell line from a 12-year-old girl who developed a late recurrent stage MS, MDM2-amplified neuroblastoma arising in the liver and performed histological, molecular, cytogenetic, exome, and telomere analyses of the recurrent tumor and the cell line. On histology, the recurrent tumor was immunoreactive for TP53, CDKN1A, and MDM2. A molecular cytogenetic study of the recurrent tumor revealed the amplification of MDM2 but no amplification of MYCN. The established cell line, NBM-SHIM, showed amplification of both MDM2 and MYCN on double-minute chromosomes. A copy number evaluation based on exome data confirmed the finding for MYCN and MDM2 and further identified high ploidy on CDK4 and GLI2 loci in the recurrent tumor and the cell line. The telomere maintenance mechanism on the cell line is unusual in terms of the low expression of TERT despite MYCN amplification and alternative lengthening of telomeres suggested by positive value for C-circle assay and telomere contents quantitative assay. The cell line is unique because it was established from a MYCN-nonamplified, MDM2-amplified, late-relapsed stage MS neuroblastoma, and MYCN amplification was acquired during cell culture. Therefore, the cell line is a valuable tool for investigating neuroblastoma tumorigenesis and new molecular targeted therapies for disrupted ARF-TP53-MDM2 pathway and amplification of MDM2 and CDK4.
Subject(s)
Gene Amplification , N-Myc Proto-Oncogene Protein , Neuroblastoma , Proto-Oncogene Proteins c-mdm2 , Humans , Neuroblastoma/genetics , Neuroblastoma/pathology , Proto-Oncogene Proteins c-mdm2/genetics , Proto-Oncogene Proteins c-mdm2/metabolism , N-Myc Proto-Oncogene Protein/genetics , Female , Child , Gene Amplification/genetics , Cell Line, Tumor , Neoplasm Recurrence, Local/genetics , Neoplasm Recurrence, Local/pathology , Liver Neoplasms/genetics , Liver Neoplasms/pathology , Telomere/genetics , Cyclin-Dependent Kinase 4/genetics , Cyclin-Dependent Kinase 4/metabolism , Telomere Homeostasis/geneticsABSTRACT
Increasing the proportion of locally produced plant protein in currently meat-rich diets could substantially reduce greenhouse gas emissions and loss of biodiversity1. However, plant protein production is hampered by the lack of a cool-season legume equivalent to soybean in agronomic value2. Faba bean (Vicia faba L.) has a high yield potential and is well suited for cultivation in temperate regions, but genomic resources are scarce. Here, we report a high-quality chromosome-scale assembly of the faba bean genome and show that it has expanded to a massive 13 Gb in size through an imbalance between the rates of amplification and elimination of retrotransposons and satellite repeats. Genes and recombination events are evenly dispersed across chromosomes and the gene space is remarkably compact considering the genome size, although with substantial copy number variation driven by tandem duplication. Demonstrating practical application of the genome sequence, we develop a targeted genotyping assay and use high-resolution genome-wide association analysis to dissect the genetic basis of seed size and hilum colour. The resources presented constitute a genomics-based breeding platform for faba bean, enabling breeders and geneticists to accelerate the improvement of sustainable protein production across the Mediterranean, subtropical and northern temperate agroecological zones.
Subject(s)
Crops, Agricultural , Diploidy , Genetic Variation , Genome, Plant , Genomics , Plant Breeding , Plant Proteins , Vicia faba , Chromosomes, Plant/genetics , Crops, Agricultural/genetics , Crops, Agricultural/metabolism , DNA Copy Number Variations/genetics , DNA, Satellite/genetics , Gene Amplification/genetics , Genes, Plant/genetics , Genetic Variation/genetics , Genome, Plant/genetics , Genome-Wide Association Study , Geography , Plant Breeding/methods , Plant Proteins/genetics , Plant Proteins/metabolism , Recombination, Genetic , Retroelements/genetics , Seeds/anatomy & histology , Seeds/genetics , Vicia faba/anatomy & histology , Vicia faba/genetics , Vicia faba/metabolismABSTRACT
BACKGROUND: Human epidermal growth factor receptor 2 (HER2)-positive tumors are defined by protein overexpression (3+) or gene amplification using immunohistochemistry (IHC) or fluorescence in situ hybridization (FISH), respectively. HER2-positive tumors have historically included both IHC(3+) and IHC(2+, equivocal)/FISH(+) tumors and received the same treatment. Differences in biology between these two tumor types, however, are poorly understood. Considering anti-HER2 drugs bind directly to HER2 protein on the cell surface, we hypothesized anti-HER2 therapies would be less effective in IHC(2+)/FISH(+) tumors than in IHC(3+) tumors, leading to differences in patient outcomes. METHODS: A total of 447 patients with HER2-positive invasive carcinoma who underwent curative surgery were retrospectively investigated. HER2 status was assessed in surgical specimens, except in patients who received neo-adjuvant chemotherapy, where biopsy specimens were employed. RESULTS: Age, tumor size, lymph node status and ER status were independent factors relating to disease-free-survival, but no difference was observed between IHC(3+) and IHC(2+)/FISH(+) tumors. Kaplan-Meier analysis found patient outcomes did not differ, even after stratifying into those that did (n = 314), or did not (n = 129), receive chemotherapy with anti-HER2 drugs. In 134 patients who received NAC, pathological complete response rates in IHC(3+) and IHC(2+)/FISH(+) tumors were 45% and 21%, respectively. Survival after developing metastasis was significantly shorter in the IHC(2+)/FISH(+) group. CONCLUSIONS: The prognosis of patients with IHC(2+)/FISH(+) tumors did not differ from IHC(3+) tumors. However, the significance of HER2 protein overexpression in relation to treatment response remains unclear and warrants further investigations.
Subject(s)
Breast Neoplasms/genetics , Carcinoma/genetics , Gene Amplification/genetics , Gene Expression/genetics , Receptor, ErbB-2/metabolism , Biomarkers, Tumor/genetics , Breast Neoplasms/mortality , Breast Neoplasms/therapy , Carcinoma/mortality , Carcinoma/therapy , Chemotherapy, Adjuvant , Female , Humans , Immunohistochemistry , In Situ Hybridization, Fluorescence , Middle Aged , Prognosis , Retrospective Studies , Treatment OutcomeABSTRACT
Mycobacterium tuberculosis (Mtb) possesses five type VII secretion systems (T7SS), virulence determinants that include the secretion apparatus and associated secretion substrates. Mtb strains deleted for the genes encoding substrates of the ESX-3 T7SS, esxG or esxH, require iron supplementation for in vitro growth and are highly attenuated in vivo. In a subset of infected mice, suppressor mutants of esxG or esxH deletions were isolated, which enabled growth to high titers or restored virulence. Suppression was conferred by mechanisms that cause overexpression of an ESX-3 paralogous region that lacks genes for the secretion apparatus but encodes EsxR and EsxS, apparent ESX-3 orphan substrates that functionally compensate for the lack of EsxG or EsxH. The mechanisms include the disruption of a transcriptional repressor and a massive 38- to 60-fold gene amplification. These data identify an iron acquisition regulon, provide insight into T7SS, and reveal a mechanism of Mtb chromosome evolution involving "accordion-type" amplification.
Subject(s)
Mycobacterium tuberculosis/genetics , Type VII Secretion Systems/genetics , Animals , Bacterial Secretion Systems/genetics , Biological Evolution , Evolution, Molecular , Gene Amplification/genetics , Mice , Mycobacterium tuberculosis/metabolism , Type VII Secretion Systems/physiology , Virulence , Virulence Factors/geneticsABSTRACT
High-level amplification of MDM2 and other genes in the 12q13-15 locus is a hallmark genetic feature of well-differentiated and dedifferentiated liposarcomas (WDLPS and DDLPS, respectively). Detection of this genomic aberration in plasma cell-free DNA may be a clinically useful assay for non-invasive distinction between these liposarcomas and other retroperitoneal tumors in differential diagnosis, and might be useful for the early detection of disease recurrence. In this study, we performed shallow whole genome sequencing of cell-free DNA extracted from 10 plasma samples from 3 patients with DDLPS and 1 patient with WDLPS. In addition, we studied 31 plasma samples from 11 patients with other types of soft tissue tumors. We detected MDM2 amplification in cell-free DNA of 2 of 3 patients with DDLPS. By applying a genome-wide approach to the analysis of cell-free DNA, we also detected amplification of other genes that are known to be recurrently affected in DDLPS. Based on the analysis of one patient with DDLPS with longitudinal plasma samples available, we show that tracking MDM2 amplification in cell-free DNA may be potentially useful for evaluation of response to treatment. The patient with WDLPS and patients with other soft tissue tumors in differential diagnosis were negative for the MDM2 amplification in cell-free DNA. In summary, we demonstrate the feasibility of detecting amplification of MDM2 and other DDLPS-associated genes in plasma cell-free DNA using technology that is already routinely applied for other clinical indications. Our results may have clinical implications for improved diagnosis and surveillance of patients with retroperitoneal tumors.
Subject(s)
Cell Dedifferentiation/genetics , Cell-Free Nucleic Acids/genetics , Gene Amplification/genetics , Liposarcoma/genetics , Proto-Oncogene Proteins c-mdm2/genetics , Aged , Cell Differentiation/genetics , Diagnosis, Differential , Female , Humans , Male , Middle Aged , Soft Tissue Neoplasms/genetics , Whole Genome Sequencing/methodsABSTRACT
PURPOSE: In this study, we aimed to evaluate the efficacy and safety of pyrotinib, a pan-HER inhibitor, in patients with HER2-amplified non-small cell lung cancer (NSCLC). PATIENTS AND METHODS: In this prospective, multicenter, single-arm trial (ChiCTR1800020262), patients with advanced NSCLC with HER2 amplification, as determined by next-generation sequencing, were enrolled and administered pyrotinib orally at 400 mg per day. The primary endpoint was 6-month progression-free survival (PFS) rate. Other endpoints included objective response rate (ORR), disease control rate (DCR), PFS, overall survival (OS), and safety. RESULTS: The enrolled cohort included 27 patients with HER2 amplification. The 6-month PFS rate was 51.9% [95% confidence interval (CI), 34.0-69.3]. The median PFS (mPFS) was 6.3 months (95% CI, 3.0-9.6 months), and median OS was 12.5 months (95% CI, 8.2-16.8 months). Pyrotinib elicited a confirmed ORR of 22.2% (95% CI, 10.6%-40.8%). Patients administered pyrotinib as first-line treatment achieved an mPFS of 12.4 months. Moreover, 30.8% of the patients who had progressed on EGFR tyrosine kinase inhibitor (TKI) responded to pyrotinib. Patients with brain metastases had an ORR of 40%. Treatment-related adverse events (TRAE) occurred in all patients (grade 3, 22.2%), but no grade 4 or higher TRAEs were documented. Diarrhea was the most frequent TRAE (all, 92.6%; grade 3, 7.4%). Loss of HER2 amplification was detected upon disease progression. CONCLUSIONS: Pyrotinib provided antitumor efficacy with a manageable safety profile in HER2-amplified patients with NSCLC.
Subject(s)
Acrylamides/administration & dosage , Aminoquinolines/administration & dosage , Carcinoma, Non-Small-Cell Lung/drug therapy , Carcinoma, Non-Small-Cell Lung/genetics , Gene Amplification/genetics , Lung Neoplasms/drug therapy , Lung Neoplasms/genetics , Receptor, ErbB-2/genetics , Receptor, ErbB-2/metabolism , Acrylamides/adverse effects , Administration, Oral , Adult , Aged , Aminoquinolines/adverse effects , Carcinoma, Non-Small-Cell Lung/mortality , Disease-Free Survival , Female , High-Throughput Nucleotide Sequencing , Humans , Lung Neoplasms/mortality , Male , Middle Aged , Prospective Studies , Time Factors , Treatment OutcomeABSTRACT
Superenhancer usages in single cancer form such as colorectal cancer (CRC) may provide novel efficient targeting candidates. It is unclear whether CRC contains recurrent superenhancers that confer a predisposition to malignancy. We investigated the superenhancer profile of CRC cell line HCT116 and compared it to that of a healthy sigmoid colon. We found that HCT116 had lost most of the normal colon superenhancer activities but gained a new set of tumor-favoring superenhancers that facilitate tumor proliferation, growth signalling, and hypoxia resistance. Inhibiting the superenhancers by JQ-1 treatment had significantly decreased the colony formation capability of HCT116. Then, by comparing the superenhancer genes and robust CRC upregulated genes, we identified a superenhancer associated with a common CRC upregulated oncogene, POU5f1B. POU5f1B overexpression is related to the worse outcome in CRCs. Via performing ChIP-PCR in 35 clinical samples and investigating CRC anti-H3K27ac ChiP-seq public dataset consisting of 36 samples, we further identified that the superenhancer of oncogene POU5F1B is recurrently activated in CRCs, taking 62 and 72 per cent, respectively. Moreover, JQ-1 treatment successfully inhibited the POU5F1B expression in 5 out of 6 POU5F1B superenhancer-positive samples. Therefore, we concluded that the superenhancer activation of POU5F1B contributes partially to its high expression in CRCs, in addition to the well-known gene amplification aetiology.
Subject(s)
Colorectal Neoplasms/genetics , Homeodomain Proteins/genetics , Oncogenes/genetics , Cell Line, Tumor , Cell Movement/genetics , Cell Proliferation/genetics , Gene Amplification/genetics , Gene Expression Regulation, Neoplastic/genetics , HCT116 Cells , Humans , Signal Transduction/genetics , Up-Regulation/geneticsABSTRACT
Cutinases are enzymes produced by phytopathogenic fungi like Moniliophthora roreri. The three genome-located cutinase genes of M. roreri were amplified from cDNA of fungi growing in different induction culture media for cutinase production. The mrcut1 gene was expressed in the presence of a cacao cuticle, while the mrcut2 and mrcut3 genes were expressed when an apple cuticle was used as the inducer. The sequences of all genes were obtained and analyzed by bioinformatics tools to determine the presence of signal peptides, introns, glycosylation, and regulatory sequences. Also, the theoretical molecular weight and pI were obtained and experimentally confirmed. Finally, cutinase 1 from M. roreri (MRCUT1) was selected for heterologous expression in Escherichia coli. Successful overexpression of MRCUT1 was observed with the highest enzyme activity of 34,036 U/mg under the assay conditions at 40°C and pH 8. Furthermore, the degradation of different synthetic polyesters was evaluated; after 21 days, 59% of polyethylene succinate (PES), 43% of polycaprolactone (PCL), and 31% of polyethylene terephthalate (PET) from plastic residues were degraded. IMPORTANCE Plastic pollution is exponentially increasing; even the G20 has recognized an urgent need to implement actions to reduce it. In recent years, searching for enzymes that can degrade plastics, especially those based on polyesters such as PET, has been increasing as they can be a green alternative to the actual plastic degradation process. A promising option in recent years refers to biological tools such as enzymes involved in stages of partial and even total degradation of some plastics. In this context, the MRCUT1 enzyme can degrade polyesters contained in plastic residues in a short time. Besides, there is limited knowledge about the biochemical properties of cutinases from M. roreri. Commonly, fungal enzymes are expressed as inclusion bodies in E. coli with reduced activity. Interestingly, the successful expression of one cutinase of M. roreri in E. coli with enhanced activity is described.
Subject(s)
Agaricales/metabolism , Biodegradation, Environmental , Carboxylic Ester Hydrolases/metabolism , Polyesters/metabolism , Polyethylene Terephthalates/metabolism , Polyethylenes/metabolism , Succinates/metabolism , Agaricales/enzymology , Agaricales/genetics , Amino Acid Sequence , Base Sequence , Cacao/genetics , Carboxylic Ester Hydrolases/genetics , Environmental Pollutants/metabolism , Environmental Pollution/analysis , Escherichia coli/genetics , Escherichia coli/metabolism , Gene Amplification/genetics , Gene Expression/genetics , Nucleic Acid Amplification Techniques , Plastics/metabolismABSTRACT
BACKGROUND: Hypoxia is a striking feature of most solid tumors and could be used to discriminate tumors from normoxic tissues. Therefore, the design of hypoxia-conditioned Chimeric Antigen Receptor (CAR) T cells is a promising strategy to reduce on-target off-tumor toxicity in adoptive cell therapy. However, existing hypoxia-conditioned CAR-T designs have been only partially successful in enhancing safety profile but accompanied with reduced cytotoxic efficacy. Our goal is to further improve safety profile with retained excellent antitumor efficacy. METHODS: In this study, we designed and constructed a hypoxia-inducible transcription amplification system (HiTA-system) to control the expression of CAR in T (HiTA-CAR-T) cells. CAR expression was determined by Flow cytometry, and the activation and cytotoxicity of HiTA-CAR-T cells in vitro were evaluated in response to antigenic stimulations under hypoxic or normoxic conditions. The safety of HiTA-CAR-T cells was profiled in a mouse model for its on-target toxicity to normal liver and other tissues, and antitumor efficacy in vivo was monitored in murine xenograft models. RESULTS: Our results showed that HiTA-CAR-T cells are highly restricted to hypoxia for their CAR expression, activation and cytotoxicity to tumor cells in vitro. In a mouse model in vivo, HiTA-CAR-T cells targeting Her2 antigen showed undetectable CAR expression in all different normoxic tissues including human Her2-expresing liver, accordingly, no liver and systemic toxicity were observed; In contrast, regular CAR-T cells targeting Her2 displayed significant toxicity on human Her2-expression liver. Importantly, HiTA-CAR-T cells were able to achieve signiï¬cant tumor suppression in murine xenograft models. CONCLUSION: Our HiTA system showed a remarkable improvement in hypoxia-restricted transgene expression in comparison with currently available systems. HiTA-CAR-T cells presented significant antitumor activities in absence of any significant liver or systemic toxicity in vivo. This approach could be also applied to design CAR-T cell targeting other tumor antigens.
Subject(s)
Cell Hypoxia/genetics , Gene Amplification/genetics , Immunotherapy/methods , Neoplasms/genetics , Receptors, Chimeric Antigen/metabolism , Animals , Cell Line, Tumor , Disease Models, Animal , Humans , MiceABSTRACT
The diagnostic accuracy of the multigene panel test (MPT) and OncoScan™ in the determination of HER2 amplification in breast tumors remains controversial. In the present study, HER2 copy number was analyzed using both MPT and OncoScan™ in 45 breast tumors and was compared with that in fluorescent in situ hybridization (FISH) analysis. Tumors with low cellularity were examined using tumor cell enrichment and fluorescenceactivated cell sorting. Both MPT and OncoScan™ exhibited significant correlations with FISH with respect to the determination of HER2 amplification in breast tumors. However, the correlation coefficient was significantly higher for the comparison of MPT and FISH (r=0.770) compared with that between OncoScan™ and FISH (r=0.564). The accuracy of MPT (93.3%) was slightly higher compared with that in OncoScan™ (84.4%) in determining the HER2 status, which was mostly explained by the higher sensitivity of MPT in tumors with low cellularity (83.3 vs. 33.3%), but not in those with high cellularity (81.8 vs. 72.7%). The specificity was 100% for both tests. The MPT exhibited higher sensitivity in the determination of the amplification of other genes, including MYC, fibroblast growth factor receptor 1 and GATA binding protein 3 in tumors with low cellularity compared with that in tumors with high cellularity. OncoScan™ exhibited low sensitivity without tumor cell enrichment. The results suggested that MPT could be a promising method to determine HER2 status in breast tumors and that it could exhibit improved accuracy compared with that in OncoScan™ in tumors with low cellularity.
Subject(s)
Breast Neoplasms/genetics , DNA Mutational Analysis/methods , Gene Amplification/genetics , Genes, erbB-2/genetics , In Situ Hybridization, Fluorescence/methods , Receptor, ErbB-2/genetics , Adult , Aged , Aged, 80 and over , Biomarkers, Tumor/genetics , Breast Neoplasms/metabolism , Female , Humans , Immunohistochemistry , Middle Aged , Receptor, ErbB-2/metabolismABSTRACT
BACKGROUND: CCND1 copy number increase is characteristic of acral melanoma and is useful in distinguishing benign and malignant acral melanocytic lesions. Increase of the gene copy number may result in protein overexpression. This raises the possibility that detection of high expression of cyclin D1 by immunohistochemistry (IHC) may be used as a surrogate for direct evaluation of increase in the CCND1 gene copy number. METHODS: We examined increases in CCND1 copy number with fluorescence in situ hybridization (FISH), and examined cyclin D1 protein expression with IHC in 61 acral melanomas. RESULTS: Using FISH, 29 acral melanomas (29/61, 47.5%) showed increase in the CCND1 copy number, including 8 (8/61, 13.1%) which showed low-level increase in the CCND1 copy number and 21 (21/61, 34.4%) with high-level increase in the CCND1 copy number. By analysis of IHC, the median IHC score was 15% (range: 1-80%) in acral melanomas with no CCND1 copy number alteration. In acral melanomas with low-level CCND1 copy number increase, the median IHC score was 25% (range: 3-90%). In acral melanomas with high-level CCND1 copy number increase, the median IHC score was 60% (range: 1-95%). Comparing FISH and IHC, cyclin D1 protein expression level has no corelation with the CCND1 copy number in acral melanomas which have no CCND1 copy number alteration and low-level CCND1 copy number increase (P = 0.108). Cyclin D1 protein expression level correlated positively with CCND1 copy number in acral melanomas with high-level CCND1 copy number increase (P = 0.038). The sensitivity, specificity and positive predictive value of using cyclin D1 IHC to predict CCND1 FISH result was 72.4, 62.5 and 63.6%. Increase in CCND1 copy number was associated with Breslow thickness in invasive acral melanoma. CONCLUSION: High-level increase in the CCND1 copy number can induce high cyclin D1 protein expression in acral melanomas. However low-level increase and normal CCND1 copy number have no obvious correlation with protein expression. Cyclin D1 IHC cannot serve as a surrogate for CCND1 FISH in acral melanomas.
Subject(s)
Cyclin D1/metabolism , DNA Copy Number Variations/genetics , Immunohistochemistry , In Situ Hybridization, Fluorescence , Melanoma/genetics , Skin Neoplasms/genetics , Adult , Aged , Aged, 80 and over , China , Cohort Studies , Cyclin D1/genetics , Female , Gene Amplification/genetics , Gene Dosage , Humans , Immunohistochemistry/methods , In Situ Hybridization, Fluorescence/methods , Male , Middle Aged , Melanoma, Cutaneous MalignantABSTRACT
Systematic perturbation screens provide comprehensive resources for the elucidation of cancer driver genes. The perturbation of many genes in relatively few cell lines in such functional screens necessitates the development of specialized computational tools with sufficient statistical power. Here we developed APSiC (Analysis of Perturbation Screens for identifying novel Cancer genes) to identify genetic drivers and effectors in perturbation screens even with few samples. Applying APSiC to the shRNA screen Project DRIVE, APSiC identified well-known and novel putative mutational and amplified cancer genes across all cancer types and in specific cancer types. Additionally, APSiC discovered tumor-promoting and tumor-suppressive effectors, respectively, for individual cancer types, including genes involved in cell cycle control, Wnt/ß-catenin and hippo signalling pathways. We functionally demonstrated that LRRC4B, a putative novel tumor-suppressive effector, suppresses proliferation by delaying cell cycle and modulates apoptosis in breast cancer. We demonstrate APSiC is a robust statistical framework for discovery of novel cancer genes through analysis of large-scale perturbation screens. The analysis of DRIVE using APSiC is provided as a web portal and represents a valuable resource for the discovery of novel cancer genes.
Subject(s)
Cell Transformation, Neoplastic/genetics , Genes, Neoplasm/genetics , Genomics , Neoplasms/genetics , Apoptosis/genetics , Cell Line, Tumor , Gene Amplification/genetics , Humans , Neoplasms/pathology , RNA, Small Interfering/genetics , Signal Transduction/geneticsABSTRACT
Background: Retinoblastoma (Rb) is a childhood tumor of the developing retina where predisposition is caused by RB1 pathogenic variants. MYCN amplification (MYCNA) has been implicated in around 2% of sporadic unilateral Rb tumors with no detectable RB1 variants. We audited data from tumors collected between 1993 and 2019 to determine if this is the case for patients treated at Barts Health NHS Trust, and how often it occurred alongside RB1 variants. Materials and methods: Screening for MYCNA was carried out by Multiple Ligation Probe Analysis of tumor and blood samples collected for RB1 genetic screening. The cohort consisted of 149 tumors, of which 114 had matched blood samples. Results: 10/149 (6.7%) tumors were positive for MYCNA in a population containing a disproportionate number of cases negative for RB1 pathogenic variants. Of 65 unbiased tumors collected from 2014 to 2019, 2 (3.1%) had MYCNA. All MYCNA samples were from sporadic, unilateral patients and 3/10 (30%) had RB1 pathogenic variants. MYCNA was not detected in any blood sample. No MYCNA tumor had 6p gain which is usually a common alteration in Rbs. Conclusions:MYCNA occurs in a small fraction of Rbs and can occur in the presence of pathogenic RB1 variants. However, where it occurs alongside RB1 alterations, the age of onset appears to be later. MYCNA has yet to be seen as a heritable change. In sporadic cases with early diagnosis, Rbs with no RB1 pathogenic variant identified should be tested for MYCNA. Conversely, tumors with MYCNA should still be screened for RB1 pathogenic variants.
Subject(s)
Gene Amplification/genetics , N-Myc Proto-Oncogene Protein/genetics , Retinal Neoplasms/genetics , Retinoblastoma/genetics , Child , Child, Preschool , Exons , Female , Genetic Testing , Humans , Infant , Male , Nucleic Acid Amplification Techniques/methods , Retinal Neoplasms/pathology , Retinoblastoma/pathology , Retinoblastoma Binding Proteins/genetics , Ubiquitin-Protein Ligases/geneticsABSTRACT
Fibroblast growth factor receptor 1 (FGFR1) has recently been identified as a promising novel therapeutic target and prognostic marker in different types of cancer. In the present study, a meta-analysis was performed to clarify the correlation between FGFR1 and the survival outcomes of head and neck squamous cell carcinoma (HNSCC) patients. PubMed, Embase, and Web of Science were systematically searched for relevant studies in order to explore the prognostic significance of FGFR1 in HNSCC. Hazards ratios (HR) and 95% confidence intervals (CI) were collected to estimate the correlation between overexpression and amplification of FGFR1 and survival outcomes of HNSCC patients. Nine studies including 2708 patients with HNSCC were finally selected for the meta-analysis. The results indicated that FGFR1 predicted poor overall survival (OS) (HR, 1.97; 95% CI, 1.49-2.61, P<0.001) in HNSCC patients. Futhermore, FGFR1 was related to poor OS in human papillomavirus (HPV) negative HNSCC not in HPV positive HNSCC patients. Subgroup analysis stratified by molecular abnormalities, such as overexpression or amplification showed the similar results. The present study demonstrated that HNSCC patients with FGFR1 overexpression and amplification were more likely to exhibit poorer survival.
Subject(s)
Head and Neck Neoplasms/genetics , Head and Neck Neoplasms/pathology , Receptor, Fibroblast Growth Factor, Type 1/metabolism , Squamous Cell Carcinoma of Head and Neck/genetics , Squamous Cell Carcinoma of Head and Neck/pathology , Adult , Aged , Aged, 80 and over , Biomarkers, Tumor/genetics , Biomarkers, Tumor/metabolism , Gene Amplification/genetics , Head and Neck Neoplasms/mortality , Humans , Middle Aged , Prognosis , Receptor, Fibroblast Growth Factor, Type 1/genetics , Squamous Cell Carcinoma of Head and Neck/mortality , Young AdultABSTRACT
There are contradictory data regarding the correlation between HER2 amplification level determined by in situ hybridization and evolution after treatment with anti-HER2 therapies. The aim of this study was to correlate quantitative results of FISH (ratio HER2/CEP17 and number of HER2 signals/nucleus) with pathological response achieved after neoadjuvant treatment with trastuzumab and chemotherapy. For this purpose, we analysed 100 consecutive HER2-positive cases of breast carcinoma treated with neoadjuvant therapy. HER2 amplification determined by FISH was found in 92 of the 100 cases studied. pCR was obtained in 58% of the patients whose tumours presented amplification. In contrast, no pCR was obtained in the 8 patients with non-amplified tumours. A significant direct correlation between HER2 high amplification (HER2/CEP17 ratio > 5 or HER2 signals/nucleus > 10) and pCR was found. In conclusion, HER2 amplification levels are clinically relevant because they provide oncologists with valuable information on the possibilities of achieving pCR after neoadjuvant treatment.
Subject(s)
Breast Neoplasms/genetics , Receptor, ErbB-2/genetics , Adult , Aged , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Biomarkers, Pharmacological , Breast Neoplasms/drug therapy , Breast Neoplasms/pathology , Female , Gene Amplification/genetics , Humans , In Situ Hybridization, Fluorescence/methods , Middle Aged , Neoadjuvant Therapy/methods , Receptor, ErbB-2/drug effects , Trastuzumab/therapeutic use , Treatment OutcomeABSTRACT
There were rare clinical reports on clear cell tumor of the lung (CCTL). The clinical characteristics and underlying genetic mutation status of CCTL are poorly understood. From 2012 to 2017, patients pathologically diagnosed with CCTL in our hospital were investigated and analyzed based on clinical manifestations, pathological characteristics, prognosis and full gene mutation status through next generation sequencing (NGS) technology. During a 6-year period, four eligible patients were diagnosed with CCTL through surgical resection and were included in this study. All patients showed solitary nodules or lumps located in the left lung. The average maximum diameter of lesions was 2.5 ± 1.1 cm. Computed tomography (CT) imaging characteristics of these nodules/lumps demonstrated the features of benign tumors. The hematoxylin-eosin (HE) morphology and immunohistochemistry were consistent with the histopathological features of benign CCTL. Subsequent NGS analysis showed frame shift mutations of F2421/E2419, K1466E mutation, and p. 1450_1456 deletion mutation in mTOR gene in two of four patient samples and amplifications of MCL1 were observed in three of four samples. CCTL is a rare type of primary pulmonary mesenchymal tumor with good prognosis. Preliminary diagnosis on CT is usually sclerosing pneumocytoma. It is still unclear whether the occurrence and development of the disease are related to specific gene mutation. In this study, the genomic findings of frame shift mutation of mTOR genes and amplification of MCL1 gene in CCTL suggest that these mutations might play a role in proliferation of CCTL.
Subject(s)
Adenocarcinoma, Clear Cell/genetics , Adenocarcinoma, Clear Cell/pathology , Lung Neoplasms/genetics , Lung Neoplasms/pathology , Myeloid Cell Leukemia Sequence 1 Protein/genetics , TOR Serine-Threonine Kinases/genetics , Adenocarcinoma, Clear Cell/diagnosis , Female , Frameshift Mutation/genetics , Gene Amplification/genetics , Glycogen/metabolism , High-Throughput Nucleotide Sequencing , Humans , Lung Neoplasms/diagnosis , Male , Middle Aged , Prognosis , Tomography, X-Ray ComputedABSTRACT
Non-small cell lung cancer (NSCLC) targeted therapies are mostly based on activating mutations and rearrangements which are rare events in Lung Squamous Cell Carcinomas (LUSC). Recently advances in immunotherapy have improved the therapeutic repository for LUSC, but there is still an urgent need for novel targets and biomarkers. We examined 73 cases of LUSC for relative copy number amplification of DCUN1D1, BCL9, FGFR1 and ERBB2 genes and searched for correlations with molecular alterations and clinicopathological characteristics. In our cohort BCL9 gene was amplified in 57.5 % of the cases, followed by DCUN1D1 in 37 %, FGFR1 in 19 % whereas none of the cases were amplified in ERBB2 gene. The majority of the samples exhibited amplification in at least one gene while half of them displayed concurrent amplification of two/three genes. Interestingly, 93 % of the FGFR1 amplified cases were also found co amplified with DCUN1D1 and/or BCL9 genes. Linear correlations were found between BCL9 and DCUN1D1 as well as BCL9 and FGFR1 gene amplification. BCL9 and DCUN1D1 genes' amplification was correlated with poorly differentiated tumors (p = 0.035 and p = 0.056 respectively), implying their possible role in tumor aggressiveness. This is the first study, to the best of our knowledge that examines the correlation of DCUN1D1 and BCL9 genes relative copy number amplification with molecular alterations and clinicopathologic characteristics of squamous cell lung cancer tissue samples. Our findings show concurrent amplification of genes in different chromosomes, with possible involvement in tumor aggressiveness. These results support the complexity of LUSC tumorigenesis and imply the necessity of multiple biomarkers / targets for a more effective therapeutic result in LUSC.
Subject(s)
Biomarkers, Tumor/genetics , Carcinoma, Squamous Cell/pathology , Intracellular Signaling Peptides and Proteins/genetics , Lung Neoplasms/pathology , Receptor, Fibroblast Growth Factor, Type 1/genetics , Transcription Factors/genetics , Aged , Aged, 80 and over , Carcinoma, Squamous Cell/genetics , Female , Gene Amplification/genetics , Gene Dosage/genetics , Humans , Lung Neoplasms/genetics , Male , Middle Aged , Retrospective StudiesABSTRACT
Assessing Erb-b2 receptor tyrosine kinase 2 (ERBB2) amplification status in breast and gastric cancer is necessary for deciding the best therapeutic strategy. Immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) are currently used for assessing protein levels and gene copy number (CN), respectively. The use of next-generation sequencing (NGS) to measure ERBB2 CN in breast cancer is approved by the United States Federal Drug Administration as a companion diagnostic. However, a CN of less than 8 is evaluated as "equivocal", which means that some ERBB2 amplification cases diagnosed as "HER2 negative" might be false-negative cases. We reviewed the results of gene profiling targeting 160 cancer-related genes in breast (N = 90) and non-breast (N = 19) cancer tissue, and compared the ERBB2 CN results with the IHC/FISH scores. We obtained an estimated CN from the measured CN by factoring in the histological proportion of tumor cells and found that an ERBB2-estimated CN of 3.2 or higher was concordant with the combined IHC/FISH outcome in 98.4% (88/90) of breast cancer cases, while this was not always evident among non-breast cancer cases. Therefore, NGS-estimated ERBB2 CN could be considered a diagnostic test for anti-HER2 therapy after the completion of adequate prospective clinical trials.
Subject(s)
Breast Neoplasms/genetics , Gene Amplification/genetics , High-Throughput Nucleotide Sequencing , Receptor, ErbB-2/genetics , Adult , Aged , Aged, 80 and over , Biomarkers, Tumor/genetics , Biomarkers, Tumor/metabolism , Breast Neoplasms/diagnosis , Breast Neoplasms/metabolism , Female , Gene Dosage , Genetic Testing/standards , Humans , Immunohistochemistry , In Situ Hybridization, Fluorescence , Middle Aged , Neoplasms/diagnosis , Neoplasms/genetics , Neoplasms/metabolism , Receptor, ErbB-2/metabolismABSTRACT
PURPOSE: The identification of HER2 overexpression in a subset of gastric adenocarcinoma (GA) patients represents a significant step forward in unveiling the molecular complexity of this disease. The predictive and prognostic value of HER2 amplification in advanced HER2 inhibitor-treated GA patients has been investigated. However, its predictive value in resectable patients remains elusive. METHODS: We enrolled 98 treatment-naïve resectable Chinese GA patients with HER2 overexpression assessed using IHC. Capture-based targeted sequencing using a panel consisting of 41 gastrointestinal cancer-related genes was performed on tumor tissues. Furthermore, we also investigated the correlation between HER2 copy number (CN) and survival outcomes. RESULTS: Of the 98 HER2-overexpressed patients, 90 had HER2 CN amplification assessed using next-generation sequencing, achieving 92% concordance. The most commonly seen concurrent mutations were occurring in TP53, EGFR and PIK3CA. We found HER2 CN as a continuous variable was an independent predictor associated with DFS (p = 0.029). Our study revealed HER2 CN-high patients showed a trend of intestinal-type GA predominant (p = 0.075) and older age (p = 0.07). The median HER2 CN was 15.34, which was used to divide the cohort into CN-high and CN-low groups. Patients with high HER2 CN had a significantly shorter DFS than patients with low HER2 CN (p = 0.002). Furthermore, HER2 CN as a categorical variable was also an independent predictor associated with DFS in patients. CONCLUSION: We elucidated the mutation spectrum of HER2-positive resectable Chinese GA patients and the association between HER2 CN and DFS. Our work revealed HER2 CN as an independent risk factor predicted unfavorable prognosis in HER2-positive GA patients and allowed us to further stratify HER2-positive resectable GA patients for disease management.
Subject(s)
Adenocarcinoma/genetics , Adenocarcinoma/mortality , DNA Copy Number Variations/genetics , Receptor, ErbB-2/genetics , Stomach Neoplasms/genetics , Stomach Neoplasms/mortality , Adult , Aged , Aged, 80 and over , Cohort Studies , Disease-Free Survival , Female , Gene Amplification/genetics , Humans , Male , Middle Aged , Mutation/genetics , PrognosisABSTRACT
Uterine carcinosarcoma (UCS) is an uncommon and highly aggressive tumor. There is no HER2 testing protocol for UCS despite the development of HER2 antibody conjugates. We aimed to elucidate histopathological HER2 expression details in UCS, to compare HER2 scores between ASCO/CAP criteria for gastric and breast cancer, and to propose requirements for HER2 testing for UCS. Eighty-nine specimens from 84 patients with metastatic/recurrent UCS were prospectively collected from May 2018 to July 2020. We performed HER2 immunohistochemistry (IHC) for 89 specimens and FISH for 44 specimens. HER2 expression details and HER2 score were evaluated according to the latest ASCO/CAP criteria for gastric (2016) and breast cancer (2018). HER2 IHC scores according to the gastric cancer criteria were 0 in 31 cases (35%), 1+ in 26 (29%), 2+ in 22 (25%), and 3+ in 10 cases (11%) of the 89 specimens. A lateral/basolateral membranous staining pattern was observed in 28/32 (88%) specimens with HER2 scores of 2+/3+. HER2 intratumoral heterogeneity was identified in 28/32 (88%) of the specimens with HER2 scores of 2+/3+. The overall concordance rate of HER2 score was 70% between the gastric and breast criteria. FISH revealed HER2 gene amplification in 10/44 (23%) specimens containing only lateral/basolateral membranous staining pattern. Based on the histopathological features of HER2 expression in UCS, a scoring system that accepts lateral/basolateral staining patterns should be applied. Furthermore, we proposed specific requirements for UCS testing, including specimen selection, scoring system, and calculating the proportion of HER2-positive cells.