Inactivation Of Tumor Suppressor Genes Research Articles

Molecular alterations in pancreatic tumors.

Genetic alterations in pancreatic tumors can usually be classified in: (1) Mutational activation of oncogenes; (2) Inactivation of tumor suppressor genes; and (3) Inactivation of genome maintenance genes controlling the repair of DNA damage. Endoscopic ultrasound-guided fine-needle aspiration has improved pre-operative diagnosis, but the management of patients with a pancreatic lesion is still challenging. Molecular testing could help mainly in solving these “inconclusive” specimens. The introduction of multi-gene analysis approaches, such as next-generation sequencing, has provided a lot of useful information on the molecular characterization of pancreatic tumors. Different types of pancreatic tumors (e.g., pancreatic ductal adenocarcinomas, intraductal papillary mucinous neoplasms, solid pseudopapillary tumors) are characterized by specific molecular alterations. The aim of this review is to summarize the main molecular alterations found in pancreatic tumors.

Extracellular Superoxide Dismutase (EC-SOD) Regulates Gene Methylation and Cardiac Fibrosis During Chronic Hypoxic Stress.

Chronic hypoxic stress induces epigenetic modifications mainly DNA methylation in cardiac fibroblasts, inactivating tumor suppressor genes (RASSF1A) and activating kinases (ERK1/2) leading to fibroblast proliferation and cardiac fibrosis. The Ras/ERK signaling pathway is an intracellular signal transduction critically involved in fibroblast proliferation. RASSF1A functions through its effect on downstream ERK1/2. The antioxidant enzyme, extracellular superoxide dismutase (EC-SOD), decreases oxidative stress from chronic hypoxia, but its effects on these epigenetic changes have not been fully explored. To test our hypothesis, we used an in-vitro model: wild-type C57B6 male mice (WT) and transgenic males with an extra copy of human hEC-SOD (TG). The studied animals were housed in hypoxia (10% O2) for 21 days. The right ventricular tissue was studied for cardiac fibrosis markers using RT-PCR and Western blot analyses. Primary C57BL6 mouse cardiac fibroblast tissue culture was used to study the in-vitro model, the downstream effects of RASSF-1 expression and methylation, and its relation to ERK1/2. Our findings showed a significant increase in cardiac fibrosis markers: Collagen 1, alpha smooth muscle actin (ASMA), and SNAIL, in the WT hypoxic animals as compared to the TG hypoxic group (p < 0.05). The expression of DNA methylation enzymes (DNMT 1&3b) was significantly increased in the WT hypoxic mice as compared to the hypoxic TG mice (p < 0.001). RASSF1A expression was significantly lower and ERK1/2 was significantly higher in hypoxia WT compared to the hypoxic TG group (p < 0.05). Use of SiRNA to block RASSF1A gene expression in murine cardiac fibroblast tissue culture led to increased fibroblast proliferation (p < 0.05). Methylation of the RASSF1A promoter region was significantly reduced in the TG hypoxic group compared to the WT hypoxic group (0.59 vs. 0.75, respectively). Based on our findings, we can speculate that EC-SOD significantly attenuates RASSF1A gene methylation and can alleviate cardiac fibrosis induced by hypoxia.

Frontiers in Bladder Cancer Genomic Research.

Most of the etiology studies of bladder cancer focus on genetic changes, mainly including mutation and activation of oncogenes, mutation and inactivation of tumor suppressor genes, and rearrangement or heterozygous deletion of chromosomes. Moreover, bladder cancer is highly heterogeneous mainly due to abnormal changes in the genome and proteome of tumor cells. Surgery is the main treatment for bladder cancer, but because the recurrence rate is high after surgery and most of the muscle-invasive bladder cancer acquires distant metastasis. Therefore, there is a need to combine with chemotherapy to consolidate the treatment effect. However, there are differences in chemosensitivity among patients. In this article, we review the up-to-date genomic researches on bladder cancer occurrence, development, metastasis, and chemosensitivity in patients, in order to provide some theoretical support for the diagnosis and treatment strategy for bladder cancer.

Discovery of Candidate DNA Methylation Cancer Driver Genes.

Epigenetic alterations, such as promoter hypermethylation, may drive cancer through tumor suppressor gene inactivation. However, we have limited ability to differentiate driver DNA methylation (DNAme) changes from passenger events. We developed DNAme driver inference-MethSig-accounting for the varying stochastic hypermethylation rate across the genome and between samples. We applied MethSig to bisulfite sequencing data of chronic lymphocytic leukemia (CLL), multiple myeloma, ductal carcinoma in situ, glioblastoma, and to methylation array data across 18 tumor types in TCGA. MethSig resulted in well-calibrated quantile-quantile plots and reproducible inference of likely DNAme drivers with increased sensitivity/specificity compared with benchmarked methods. CRISPR/Cas9 knockout of selected candidate CLL DNAme drivers provided a fitness advantage with and without therapeutic intervention. Notably, DNAme driver risk score was closely associated with adverse outcome in independent CLL cohorts. Collectively, MethSig represents a novel inference framework for DNAme driver discovery to chart the role of aberrant DNAme in cancer. SIGNIFICANCE: MethSig provides a novel statistical framework for the analysis of DNA methylation changes in cancer, to specifically identify candidate DNA methylation driver genes of cancer progression and relapse, empowering the discovery of epigenetic mechanisms that enhance cancer cell fitness.This article is highlighted in the In This Issue feature, p. 2113.

Proteogenomic landscape of uterine leiomyomas from hereditary leiomyomatosis and renal cell cancer patients

Pathogenic mutations in fumarate hydratase (FH) drive hereditary leiomyomatosis and renal cell cancer (HLRCC) and increase the risk of developing uterine leiomyomas (ULMs). An integrated proteogenomic analysis of ULMs from HLRCC (n = 16; FH-mutation confirmed) and non-syndromic (NS) patients (n = 12) identified a significantly higher protein:transcript correlation in HLRCC (R = 0.35) vs. NS ULMs (R = 0.242, MWU p = 0.0015). Co-altered proteins and transcripts (228) included antioxidant response element (ARE) target genes, such as thioredoxin reductase 1 (TXNRD1), and correlated with activation of NRF2-mediated oxidative stress response signaling in HLRCC ULMs. We confirm 185 transcripts previously described as altered between HLRCC and NS ULMs, 51 co-altered at the protein level and several elevated in HLRCC ULMs are involved in regulating cellular metabolism and glycolysis signaling. Furthermore, 367 S-(2-succino)cysteine peptides were identified in HLRCC ULMs, of which sixty were significantly elevated in HLRCC vs. NS ULMs (LogFC = 1.86, MWU p < 0.0001). These results confirm and define novel proteogenomic alterations in uterine leiomyoma tissues collected from HLRCC patients and underscore conserved molecular alterations correlating with inactivation of the FH tumor suppressor gene.

Mechanism, Clinical Significance, and Treatment Strategy of Warburg Effect in Hepatocellular Carcinoma

Hepatocellular carcinoma (HCC) is a primary malignancy of the liver and the third leading cause of cancer death worldwide. The incidence of HCC accounts for more than 90% of primary HCC. Like most solid malignancies, the occurrence and development of HCC are closely related to the Warburg effect. The Warburg effect of HCC is mainly manifested as increased glucose uptake by HCC cells, increased glycolysis, restricted mitochondrial oxidative phosphorylation, increased pentose phosphate pathway in HCC cells, and increased glutamine decomposition. As the contribution of glycolysis to the total ATP of tumor cells generally does not exceed 50% to 60%, oxidative phosphorylation (OXPHOS) still makes a considerable contribution to the ATP of tumor cells. In some cases, there will be an anti-Warburg effect. HCC Warburg effect is closely related to HCC cell proliferation, apoptosis, immune escape, migration and invasion, chemotherapy resistance, and treatment failure. The mechanism of the Warburg effect in HCC is complex, involving the expression of stimulating the key glycolysis enzymes by hypoxia-inducible factor-1(HIF-1), the activation of oncogenes and the inactivation of tumor suppressor genes, the continuous activation of related signaling pathways, the participation of noncoding RNA, and the rate of metabolism gene mutation of enzyme. This article synthetically discusses the characteristics of glucose metabolism in HCC cells, the mechanism of Warburg effect, clinical significance, and corresponding treatment strategies and provides new perspectives for the prevention and treatment of HCC.

Genomic Instability in Cancer: Molecular Mechanisms and Therapeutic Potentials.

DNA damage usually happens in all cell types, which may originate from endogenous sources (i.e., DNA replication errors) or be emanated from radiations or chemicals. These damages range from changes in few nucleotides to significant structural abnormalities on chromosomes and, if not repaired, could disturb the cellular homeostasis or cause cell death. As the most significant response to DNA damage, DNA repair provides biological pathways by which DNA damages are corrected and returned into their natural circumstance. However, an aberration in the DNA repair mechanisms may result in genomic and chromosomal instability and the accumulation of mutations. The activation of oncogenes and/or inactivation of tumor suppressor genes is a serious consequence of genomic and chromosomal instability and may bring the cells into a cancerous phenotype. Therefore, genomic and chromosomal instability is usually considered a crucial factor in carcinogenesis and an important hallmark of various human malignancies. In the present study, we review our current understanding of the most updated mechanisms underlying genomic instability in cancer and discuss the potential promises of these mechanisms in finding new targets for the treatment of cancer.

Examination of ATM, BRCA1, and BRCA2 promoter methylation in patients with pancreatic cancer

BackgroundPancreatic cancer is a lethal disease with a poor 5-year survival rate. Pathogenic germline variants in the coding regions of ATM, BRCA1, and BRCA2 are found in up to 4.8% of pancreatic cancer patients. Germline promoter methylation and gene silencing arising from a germline variant or through other mechanisms have been described as a cause of tumor suppressor gene inactivation. MethodsWe measured the level of promoter methylation of the ATM, BRCA1, and BRCA2 genes in peripheral blood lymphocytes from 655 patients with pancreatic cancer using real-time PCR. ResultsNo evidence of germline promoter methylation of any of these genes was found. Promoter methylation levels were minimal with no patient having promoter methylation greater than 3.4%, 3.3%, and 7.6% for ATM, BRCA1 and BRCA2, respectively, well below levels found in patients who have inherited promoter methylation (∼50%). ConclusionsWe found no evidence of germline promoter methylation for the pancreatic susceptibility genes ATM, BRCA1 and BRCA2 in patients with pancreatic cancer. This study reveals that constitutive germline methylation of promoter CpG islands is rare in pancreatic cancer.

Chronic Lung Allograft Dysfunction: Role for Tumor Suppressor Gene Liver Kinase B1

Purpose Epithelial mesenchymal transformation (EMT) has been proposed to play a role in chronic lung allograft dysfunction (CLAD). Liver kinase B1 (LKB1), a tumor suppressor gene, can regulate EMT. However, its role in CLAD development following lung transplantation remains unknown. Goal of this study is to determine the role of LKB1 on Rab27 mediated exosome release and immune-pathogenesis of CLAD. Methods We first analyzed EMT on airway epithelial cell line (BEAS-2B) with exosomes from bronchiolitis obliterans syndrome (BOS) and stable. Role for LKB1 using SiRNA mediated transfection of BEAS-2B and in a hypoxia/reperfusion model were determined. Using qRT-PCR, we analyzed LKB1 and miR-155 levels in biopsies from lung transplant recipients with BOS and stable and fibrosis in a murine lung transplant model of chronic rejection. Results In BEAS-2B, knockdown of LKB1 increased exosome release (p=0.0001). Co-culture of BEAS-2B with exosomes from BOS increased vimentin (p=0.001) and α-SMA (p=0.001). LKB1 knockdown increased Rab27A (p=0.01), α-SMA (p=0.01), and vimentin (p=0.01) following hypoxia/reperfusion. qRT-PCR of biopsies from lung transplant recipients with BOS demonstrated downregulation of LKB1 (p=0.002), upregulation of Rab27A (p=0.006) and miR-155 (p=0.03) compared to stable biopsies. Using a murine lung transplant model of chronic rejection, we demonstrated correlation between development of lung fibrosis to downregulation of LKB1 (p=0.01), upregulation of vimentin (p=0.01) and α-SMA (p=0.01. Conclusion We identified a novel molecular mechanism in which exosomes released from transplanted lungs undergoing chronic rejection with inactivated tumor suppressor gene LKB1 results in EMT leading to the pathogenesis of CLAD following human lung transplantation.

Improving high-resolution copy number variation analysis from next generation sequencing using unique molecular identifiers

BackgroundRecently, copy number variations (CNV) impacting genes involved in oncogenic pathways have attracted an increasing attention to manage disease susceptibility. CNV is one of the most important somatic aberrations in the genome of tumor cells. Oncogene activation and tumor suppressor gene inactivation are often attributed to copy number gain/amplification or deletion, respectively, in many cancer types and stages. Recent advances in next generation sequencing protocols allow for the addition of unique molecular identifiers (UMI) to each read. Each targeted DNA fragment is labeled with a unique random nucleotide sequence added to sequencing primers. UMI are especially useful for CNV detection by making each DNA molecule in a population of reads distinct.ResultsHere, we present molecular Copy Number Alteration (mCNA), a new methodology allowing the detection of copy number changes using UMI. The algorithm is composed of four main steps: the construction of UMI count matrices, the use of control samples to construct a pseudo-reference, the computation of log-ratios, the segmentation and finally the statistical inference of abnormal segmented breaks. We demonstrate the success of mCNA on a dataset of patients suffering from Diffuse Large B-cell Lymphoma and we highlight that mCNA results have a strong correlation with comparative genomic hybridization.ConclusionWe provide mCNA, a new approach for CNV detection, freely available at https://gitlab.com/pierrejulien.viailly/mcna/ under MIT license. mCNA can significantly improve detection accuracy of CNV changes by using UMI.

Epigenetic Mechanisms beyond Tumour-Stroma Crosstalk.

Simple SummaryIn this review article, we will deepen the topic of cancer cell communication with the surrounding non-cancerous cells. In particular, the non-mutational events that modified gene expression, namely “epigenetics”, involved in cell–cell interaction will be the center of this work. Many studies have described the mechanism of back-and-forth communication between cancer and stromal cells, and very recent studies suggested that epigenetics may play an important role in this setting. This work will describe recent advances in the field of epigenetic mechanisms involved in tumour–stroma crosstalk. Understanding these processes can be useful in future cancer therapy design.Despite cancer having been usually considered the result of genetic mutations, it is now well established that epigenetic dysregulations play pivotal roles in cancer onset and progression. Hence, inactivation of tumour suppressor genes can be gained not only by genetic mutations, but also by epigenetic mechanisms such as DNA methylation and histone modifications. To occur, epigenetic events need to be triggered by genetic alterations of the epigenetic regulators, or they can be mediated by intracellular and extracellular stimuli. In this last setting, the tumour microenvironment (TME) plays a fundamental role. Therefore, to decipher how epigenetic changes are associated with TME is a challenge still open. The complex signalling between tumour cells and stroma is currently under intensive investigation, and most of the molecules and pathways involved still need to be identified. Neoplastic initiation and development are likely to involve a back-and-forth crosstalk among cancer and stroma cells. An increasing number of studies have highlighted that the cancer epigenome can be influenced by tumour microenvironment and vice versa. Here, we discuss about the recent literature on tumour–stroma interactions that focus on epigenetic mechanisms and the reciprocal regulation between cancer and TME cells.

5-hydroxymethylcytosine as a liquid biopsy biomarker in mCRPC.

148 Background: 5-hydroxymethylcytosine (5hmC) is an epigenetic modification which regulates gene expression and is associated with active transcription. The optimization of 5hmC sequencing in cell-free DNA (cfDNA) could therefore enable assessment of gene activity through a liquid biopsy. We aimed to investigate the 5hmC landscape of metastatic castration-resistant prostate cancer (mCRPC) and to evaluate the potential of 5hmC modifications in cfDNA as biomarkers of outcome in mCRPC patients. Methods: Genome-wide 5hmC modifications were analyzed with a low-input whole-genome 5hmC sequencing method based on selective chemical labeling in DNA from 93 mCRPC tissue biopsies previously profiled with whole-genome sequencing (WGS), RNA-sequencing and whole-genome bisulfite sequencing (WGBS). In addition, we analyzed 64 cell-free DNA (cfDNA) samples, from men with mCRPC before first-line abiraterone or enzalutamide, with both 5hmC sequencing and a conventional targeted ctDNA panel assessing common genomic alterations. Results: In mCRPC tissue samples, 5hmC enrichment was more strongly associated with gene expression than promoter methylation or copy number. Among cancer hallmark pathways, the androgen response genes had the strongest association between 5hmC and gene expression, suggesting a disease specific marking of gene activation. 5hmC patterns in cfDNA could be used to estimate the circulating tumor DNA fraction (ct-fraction), which was prognostic for overall survival (tertiles of ct-fraction, HR = 1.6 95%CI 1.1-2.3, p = 0.007). Further, 5hmC levels were indicative of gain of oncogene activity (such as AR, MYC, and PIK3CA) and loss of tumor suppressor gene activity (such as RB1, TP53 and BRCA2). The number of alterations, by 5hmC levels, of common drivers of mCRPC was prognostic for overall survival, also after adjusting for ct-fraction (adjusted p = 0.00001), and the prognostic value of common alterations detected by 5hmC sequencing versus conventional targeted ctDNA sequencing was similar. Finally, 5hmC levels in cfDNA of genes not significantly altered by copy number gain or loss (and thus not routinely included in targeted ctDNA sequencing assays), such as TOP2A and EZH2, identified a high-risk subgroup of mCRPC, which was highly prognostic for overall survival independent of ct-fraction (adjusted HR = 1.8 95%CI 1.2-2.8, p = 0.007). Conclusions: 5hmC in mCRPC tissue demonstrated an association with gene expression that was highest for prostate cancer driver genes, highlighting the ability to track disease-specific biology. 5hmC in cfDNA from men with mCRPC can be used to estimate the ct-fraction of the sample, infer activity gain and loss of common drivers of mCRPC, and identify high-risk groups of mCRPC based on alterations not commonly detected with conventional ctDNA sequencing, showing its potential as a liquid biomarker. Further studies are aimed at optimizing and validating 5hmC-based biomarkers in larger cohorts.

Signs of carcinogenicity induced by parathion, malathion, and estrogen in human breast epithelial cells (Review).

Cancer development is a multistep process that may be induced by a variety of compounds. Environmental substances, such as pesticides, have been associated with different human diseases. Organophosphorus pesticides (OPs) are among the most commonly used insecticides. Despite the fact that organophosphorus has been associated with an increased risk of cancer, particularly hormone-mediated cancer, few prospective studies have examined the use of individual insecticides. Reported results have demonstrated that OPs and estrogen induce a cascade of events indicative of the transformation of human breast epithelial cells. In vitro studies analyzing an immortalized non-tumorigenic human breast epithelial cell line may provide us with an approach to analyzing cell transformation under the effects of OPs in the presence of estrogen. The results suggested hormone-mediated effects of these insecticides on the risk of cancer among women. It can be concluded that, through experimental models, the initiation of cancer can be studied by analyzing the steps that transform normal breast cells to malignant ones through certain substances, such as pesticides and estrogen. Such substances cause genomic instability, and therefore tumor formation in the animal, and signs of carcinogenesis in vitro. Cancer initiation has been associated with an increase in genomic instability, indicated by the inactivation of tumor-suppressor genes and activation of oncogenes in the presence of malathion, parathion, and estrogen. In the present study, a comprehensive summary of the impact of OPs in human and rat breast cancer, specifically their effects on the cell cycle, signaling pathways linked to epidermal growth factor, drug metabolism, and genomic instability in an MCF-10F estrogen receptor-negative breast cell line is provided.

IMMUNOHISTOCHEMICAL EXPRESSION OF P53 AND CYCLIN D1 IN SQUAMOUS CELL CARCINOMAAND EPITHELIAL HYPERPLASIA OF ORAL MUCOSA.

Oral cancer is among the top three types of cancers in India. Most invasive oral carcinomas are preceded by a pre-invasive stage, that may last for many years. . Understanding the molecular mechanisms involved in the initiation and progression to malignancy of oral cancer will help to improve its prognosis and in the elaboration of new forms of treatment. The most common genetic alterations, include gene amplication and overexpression of oncogenes such as myc, erbB-2, Epidermal Growth Factor Receptor (EGFR), Cyclin D1 and mutations, deletions and hypermethylation leading to p16 and p53 tumor suppressor gene inactivation. There still remains the problem of differentiating epitheliomatous hyperplasia from squamous cell carcinoma (SCC) especially on biopsies. AIMS: 1. To study and evaluate the expression of p53 and Cyclin D1 in oral SCC and epithelial hyperplasia. 2. To correlate the expression of p53 and Cyclin D1 in different grades of oral SCC. SETTINGS AND DESIGN:Cross-sectional study conducted over a period of 18 months. METHODS AND MATERIAL: 30 cases (n=30) of oral squamous cell carcinoma &amp; 30 cases (n=30) of epithelial hyperplasia diagnosed on H &amp; E were included in this study. All these cases of oral SCC were further classied into different histological grades according to BRODERS' HISTOLOGICALGRADE. STATISTICAL ANALYSIS: The data was collected and the association between different groups was analysed by using Chi square test and quantitative variables were compared using Independent Ttest/Mann-Whitney Test (when the data sets were not normally distributed) between the two groups and ANOVA/Kruskal Wallis test between three groups. Ap value of ≤0.05 was considered statistically signicant. RESULTS: There was a signicant increase in the expression of p53 in oral SCC compared to epithelial hyperplasia. The results were statistically signicant (p value &lt;0.0001). Similarly , Cyclin D1 also showed an increased expression in cases of SCC in contrast to epithelial hyperplasia (p value &lt;0.0001). The expression of p53 signicantly increased with increase in grade of the tumor. The results being statistically signicant, p value=0.004. Similarly, poorly and moderately differentiated SCC also showed an increased expression of Cyclin D1 compared to well differentiated SCC (p value=0.03, statistically signicant). CONCLUSION: Thus the author concludes that the expression of both p53 and Cyclin D1 is increased in oral SCC compared to epithelial hyperplasia of oral mucosa and these markers can be used to differentiate between these two lesion especially on biopsy cases.

In search of novel synthetic lethality anti-cancer drug targets in intrahepatic cholangiocarcinoma: MTAP genomic loss.

337 Background: The association of BRCA tumor suppressor gene (TSG) inactivation and PARP inhibitor efficacy through a mechanism of synthetic lethality (SL) is now well-established. Recently, the PRMT5 arginine methyltransferase dependence in cells with MTAP (S-methyl-5'-thioadenosine phosphorylase) genomic alterations (GA) has been proposed as a new SL based anti-tumor strategy and under consideration for development for intrahepatic cholangiocarcinoma (IC). Methods: 2,170 cases of clinically advanced IC were submitted for comprehensive genomic profiling (CGP) to detect genomic alterations (GA), tumor mutational burden (TMB) and microsatellite instability (MSI) using a hybrid capture based assay (F1CDx). PD-L1 expression in tumor cells (Dako 22C3) was measured by IHC and scored using the tumor proportion score (TPS) method. Results: 328 (15%) ICs featured MTAP inactivation (MTAP-IC). 48% of MTAP-IC cases were female and 52% were male with a median age of 66 years (range 26-93 years). MTAP-IC featured 6.3 GA per sample. The median TMB was 2.5 mut/Mb with only 2% of cases with TMB ≥10 mut/Mb. No (0%) MTAP-IC cases were MSI-high. Of 112 MTAP-IC IHC stained for PD-L1, 4% were PD-L1 high (&gt; 50%), 14% were PD-L1 low (1-49%) and 82% were PD-L1 negative. 99% of MTAP inactivation was by copy number (CN) loss and 1% was due to non-CN mutation. In MTAP-IC, 99% featured CDKN2A loss and 95% featured CDKN2B loss (9p21 co-deletion). TP53 was mutated in 31% and KRAS in 24% (3% G12C). Other currently non-targetable GA included ARID1A (19%), BAP1 (16%) and SMAD4 (13%). Among potentially targetable GA, the MTAP-IC featured 12% FGFR2 GA (8% rearrangements), 10% BRAF (8% V600E), 7% IDH1, 7% PIK3CA, 4% BRCA2, 4% ERBB2 (2% amplification) and 3% MET. Conclusions: Comprehensive genomic profiling elucidated that MTAP GA are frequent in IC and features relatively high GA/tumor, including 9p21 co-deletion, but relatively low TMB and PD-L1 expression. Isolated MTAP deletion without CDKN2A/B co-deletion was extremely rare. In addition to the potential targeting of PRMT5 in MTAP-IC tumors, co-altered genes known to be targets in IC such as FGFR2, IDH1 and BRAF are still prevalent in MTAP-IC, indicating potential for combining targeted therapies in some MTAP-IC patients.

Promoterless Transposon Mutagenesis Drives Solid Cancers via Tumor Suppressor Inactivation.

Simple SummaryOver the past two decades, there have been many published studies reporting high-copy SB transgenic lines in which the transposon allele includes both gene-trap and internal promoter elements to drive tumorigenesis in vivo. Cancer gene discovery from end-stage solid tumors in these studies performed by our labs and others has found few proto-oncogenic insertions. However, the question remains whether these tumors are initiated by SB insertions in proto-oncogenes to promote permissible phenotypes for tumor initiation, which become dispensable for tumor maintenance. Here, our study expands upon these indirect observations to demonstrate that high-copy SB transposon alleles designed with only gene-trap elements that inactivate genes (Onc2.3) can drive tumor initiation, progression, and maintenance to end-stage tumors in the absence of sensitizing mutations.A central challenge in cancer genomics is the systematic identification of single and cooperating tumor suppressor gene mutations driving cellular transformation and tumor progression in the absence of oncogenic driver mutation(s). Multiple in vitro and in vivo gene inactivation screens have enhanced our understanding of the tumor suppressor gene landscape in various cancers. However, these studies are limited to single or combination gene effects, specific organs, or require sensitizing mutations. In this study, we developed and utilized a Sleeping Beauty transposon mutagenesis system that functions only as a gene trap to exclusively inactivate tumor suppressor genes. Using whole body transposon mobilization in wild type mice, we observed that cumulative gene inactivation can drive tumorigenesis of solid cancers. We provide a quantitative landscape of the tumor suppressor genes inactivated in these cancers and show that, despite the absence of oncogenic drivers, these genes converge on key biological pathways and processes associated with cancer hallmarks.

In This Issue

See article, p. 32.Inflammatory signaling perturbs hematopoietic stem cell (HSC) differentiation through a process termed emergency hematopoiesis. Whether an equivalent process has a role in regulating leukemia stem cells (LSC) is unclear. Here Xie and colleagues demonstrate that sphingosine-1-phosphate receptor 3 (S1RP3), a receptor for proinflammatory and bioactive lipid sphingosine-1-phosphate (S1P), drives myeloid differentiation in both HSC and LSC. S1RP3 expression is high in CD33+ myeloid cells and low in primitive CD34+ cells. When S1RP3 is upregulated, it promotes myeloid differentiation in synergy with proinflammatory signaling pathways in HSC and LSC. In addition, S1RP3 expression level could be utilized to stratify acute myeloid leukemia (AML) patients into more differentiated and more primitive cases. Moreover, pharmacologic disruption of S1P signaling by the S1P prodrug FTY720 reduces LSC frequency and decreases leukemia burden in vivo. The results suggest that S1RP3 could serve as both a biomarker and therapeutic target in human AML.See article, p. 54.Specific alterations in DNA methylation patterns are common in cancer cells across tissue origins and genetic drivers. In this study, Kretzmer and colleagues asked how early cancer-associated methylation patterns emerge during chronic lymphocytic leukemia (CLL) oncogenesis. They have profiled DNA methylomes of single cells and bulk samples across cell types representing key stages of oncogenesis, from normal germinal center B-cell subsets to preneoplastic monoclonal B-cell lymphocytosis (MBL) to serial CLL samples at diagnosis and throughout disease progression and therapy. The authors identify cancer-specific methylation patterns distinguishing normal B-cell subtypes from MBL and CLL. These methylation patterns are shared among all CLL clones in each patient case and are already present in nonmalignant MBL cell state. Persistence of the methylation pattern is particularly remarkable when considered on the backdrop of genetic and transcriptional heterogeneity. These findings shed light on possible CLL cell of origin and provide support to the concept that precancerous methylome may be a prerequisite for oncogenic transformation and maintenance of neoplastic state.See article, p. 70.Activated B-cell–like diffuse large B-cell lymphoma (ABC-DLBCL) is associated with poor prognosis when compared with germinal center B-cell–like (GCB)-DLBCL, and there is an urgent need to develop a preclinical animal model that closely models human ABC-DLBCL for the development of effective targeting therapies. MYD88 mutation and gain of BCL2 copy numbers are two of the most frequent genomic aberrations in ABC-DLBCL. Here Flümann and colleagues characterize the phenotypes of a Myd88/BCL2-driven (MBC) mouse lymphoma model and find that this model closely resembles the morphologic and gene expression signatures of human ABC-DLBCL. Moreover, MBC-derived lymphomas undergo apoptosis in response to venetoclax, revealing their druggable vulnerabilities against BCL2. Furthermore, combined treatment of venetoclax plus anti–PD-1 antibody RMP1-14 extends survival of MBC mice after they develop tumors far longer than either drug alone. These findings validate MBC mice as a tool to study ABC-DLBCL and demonstrate preclinical evidence that targeting both BCL2 and PD-1 could serve as a novel therapy for the treatment of ABC-DLBCL.See article, p. 92.PTEN is the second most frequently mutated tumor suppressor gene in cancer. In this study, Tottone and colleagues identified a distal regulatory element enhancing PTEN expression (PE) as a recurrent deletion in human T-ALL. PE displays enhancer-associated chromatin marks in hematopoietic lineage, but not solid tissues. In several human T-ALL cell lines, the PE region is occupied by multiple transcription factors including Notch, is bidirectionally transcribed, and engages in long-range interactions with the PTEN promoter. PE deletion in these cells results in decreased PTEN mRNA and decreased proliferation. PE is conserved across species, and its germline as well as conditional deletion in mice results in dose-dependent diminished PTEN expression in T cells, but not other lineages. Notch1-driven leukemia development is accelerated on PE-null mouse background, providing functional evidence of PE tumor-suppressive role consistent with its deletion profile in human T-ALL. Cancer-associated genetic inactivation of tumor suppressor genes is mostly documented within their promoter or protein-coding regions. This study reveals that distal enhancer deletion is another mechanism by which a classic and ubiquitous tumor suppressor gene can be inactivated in cancer. This work identifies previously uncharacterized PTEN enhancer activity in normal T cells and in T-ALL. It invites exploration of lineage-specific enhancer regulation of PTEN and other classic tumor suppressors in physiology and oncogenesis.In This Issue is written by Blood Cancer Discovery editorial staff. Readers are encouraged to consult the original articles for full details.

The involvement of mitochondrial proteins in tumorigenesis

Mitochondria are the key organelles for life. Mitochondria are important mediators of tumorigenesis. Mitochondria are key participants in tumorigenesis and development. In-depth study on the role of mitochondria in the survival and growth of tumor cells is not only conducive to our understanding of tumor genesis and development, including oncogene activation, tumor suppressor gene inactivation and other aspects, but also of far-reaching significance in the diagnosis and treatment of tumors.

Copy neutral loss of heterozygosity (cnLOH) patterns in synchronous colorectal cancer.

Copy neutral loss of heterozygosity (cnLOH) is a common event in several human malignancies-positing this as a mechanism of carcinogenesis. However, the role of cnLOH in synchronous colorectal cancer (SCRC), a unique CRC subtype, is not well understood. The aim of this study was to establish a cnLOH profile of SCRC using a single-nucleotide polymorphism array (SNP-A), and to explore associations between cnLOH and the genomic landscape of frequently mutated genes in SCRC. Among 74 paired SCRC cases, the most frequently altered regions were 16p11.2-p11.1 (59.5%) and 11p11.2-p11.12 (28.4%). Notably, the 6q11.21-q11.22 region altered by cnLOH was uniquely associated with polyclonal SCRCs (p = 0.038). Together, our analysis suggests that inactivation of tumor suppressor genes and cnLOH are rare events among SCRC cases. This study defines distinct patterns of cnLOH in SCRC, and provides initial evidence of a role for cnLOH in SCRC etiology.

Prohibitin regulates mTOR pathway via interaction with FKBP8.

The ability of tumor cells to sustain continuous proliferation is one of the major characteristics of cancer. The activation of oncogenes and the mutation or inactivation of tumor suppressor genes ensure the rapid proliferation of tumor cells. The PI3K-Akt-mTOR axis is one of the most frequently modified signaling pathways whose activation sustains cancer growth. Unsurprisingly, it is also one of the most commonly attempted targets for cancer therapy. FK506 binding protein 8 (FKBP8) is an intrinsic inhibitor of mTOR kinase that also exerts an anti-apoptotic function. We aimed to explain these contradictory aspects of FKBP8 in cancer by identifying a "switch" type regulator. We identified through immunoprecipitation-mass spectrometry-based proteomic analysis that the mitochondrial protein prohibitin 1 (PHB1) specifically interacts with FKBP8. Furthermore, the downregulation of PHB1 inhibited the proliferation of ovarian cancer cells and the mTOR signaling pathway, whereas the FKBP8 level in the mitochondria was substantially reduced. Moreover, concomitant with these changes, the interaction between FKBP8 and mTOR substantially increased in the absence of PHB1. Collectively, our finding highlights PHB1 as a potential regulator of FKBP8 because of its subcellular localization and mTOR regulating role.