Clinical Performance of Friend Leukemia Virus Integration 1 Methylation as a Biomarker for Colorectal Carcinoma

Objectives Accumulating evidence supports the reliability of molecular biomarkers and cancer, the current situation is lacking. This present study was aimed to investigate clinical performance of friend leukemia virus integration 1 (FLI1) methylation as a biomarker for colorectal cancer (CRC). Methods The datasets of UALCAN, GEPIA, cBioPortal, STRING, TIMER2.0, GEO, and KOBAS were utilized in this present study. In order to testify the methylation function of FLI1 in CRC, we studied 6 CRC cell lines and 20 pairs of tumor tissues. The transcriptional levels of FLI1 were tested by reverse transcription PCR quantitatively and western blot. Cell viability, transwell assays and plate clone assay were utilized to assess the cell function. Results FLI1 was up-regulated in the GBM, KIRC, LAML, etc. Meanwhile, it was down-regulated in BLCA, BRCA, CESC, etc (p<0.05). The CPTAC dataset showed higher total protein in the primary tissues of KIRC, lower protein in the BRCA, OV, LUAD, UCEC than normal tissues(p<0.05). Highly expressed FLI1 was linked to poor prognosis of overall survival (OS) in LGG, UVM (p<0.05). Low expression of FLI1 was associated with short OS in KIRC, LUAD, SKCM (p<0.05). Compared with normal tissues, the methylation level of FLI1 was increased in BRCA, CESC, CHOL, COAD, etc (p<0.05). In the contrary, it was decreased in KIRP, PCPG obviously (p<0.05). Moreover, it showed a reduced phosphorylation for selected probes (BRCA: NP_002008.2:S39, NP_002008.2:S3241; OV: NP_002008.2:S39; LUAD: NP_002008.2:S79, NP_002008.2:S3241; all p<0.05). Meanwhile, it showed an enhanced phosphorylation level of FLI1 in KIRC for selected probes (NP_002008.2:S241, p<0.05). Besides, a statistical negative correlation was found between FLI1 and Treg cells, neutrophils, monocytes, macrophages, NK cells, cancer-associated fibroblasts, and common immune checkpoint gene levels (p<0.05). Besides, in vivo experience showed that 5-Aza-2’-deoxycytidine diminished FLI1 methylation level and restored transcriptional levels (p>0.05) in CRC. In vitro experiments demonstrated that the proliferation, colony formation, invasion and migration of CRC cells were inhibited by FLI1 overexpression through FMNL1 (p<0.05). Conclusions This present study offers a comprehensive understanding of FLI1 in carcinomas with oncogenesis and immunotherapeutic implications. Moreover, FLI1 reduced the proliferation, colony formation, invasion and migration of CRC by FMNL1.

Diverse tumorigenic consequences of human papillomavirus integration in primary oropharyngeal cancers

Human papillomavirus (HPV) causes 5% of all cancers and frequently integrates into host chromosomes. The HPV oncoproteins E6 and E7 are necessary but insufficient for cancer formation, indicating that additional secondary genetic events are required. Here, we investigate potential oncogenic impacts of virus integration. Analysis of 105 HPV-positive oropharyngeal cancers by whole-genome sequencing detects virus integration in 77%, revealing five statistically significant sites of recurrent integration near genes that regulate epithelial stem cell maintenance (i.e., SOX2, TP63, FGFR, MYC) and immune evasion (i.e., CD274). Genomic copy number hyperamplification is enriched 16-fold near HPV integrants, and the extent of focal host genomic instability increases with their local density. The frequency of genes expressed at extreme outlier levels is increased 86-fold within ±150 kb of integrants. Across 95% of tumors with integration, host gene transcription is disrupted via intragenic integrants, chimeric transcription, outlier expression, gene breaking, and/or de novo expression of noncoding or imprinted genes. We conclude that virus integration can contribute to carcinogenesis in a large majority of HPV-positive oropharyngeal cancers by inducing extensive disruption of host genome structure and gene expression.

Long read sequencing reveals sequential complex rearrangements driven by Hepatitis B virus integration

Integration of Hepatitis B (HBV) virus into human genome disrupts genetic structures and cellular functions. Here, we conducted multiplatform long read sequencing on two cell lines and five clinical samples of HBV-induced hepatocellular carcinomas (HCC). We resolved two types of complex viral integration induced genome rearrangements and established a Time-phased Integration and Rearrangement Model (TIRM) to depict their formation progress by differentiating inserted HBV copies with HiFi long reads. We showed that the two complex types were initialized from focal replacements and the fragile virus-human junctions triggered subsequent rearrangements. We further revealed that these rearrangements promoted a prevalent loss-of-heterozygosity at chr4q, accounting for 19.5% of HCC samples in ICGC cohort and contributing to immune and metabolic dysfunction. Overall, our long read based analysis reveals a novel sequential rearrangement progress driven by HBV integration, hinting the structural and functional implications on human genomes.

Wingless-type mouse mammary tumor virus integration site (WNT) regulation of bovine theca cells

Ovarian paracrine mediation by components of the wingless-type mouse mammary tumor virus integration site ligands (WNT1 to 11) and their receptors, Frizzled family members (FZD1 to 10), has been proposed. Secreted truncated forms of FZD proteins (e.g., SFRP4) block the action of WNT ligands. Dickkopf-1 (DKK1) is another WNT antagonist, and R-spondin-1 (RSPO1) is one of a group of four secreted proteins that enhance WNT/β-catenin signaling. Our hypothesis was that granulosa cells signal theca cells (TC) via SFRP4, DKK1, RSPO1 and WNT secretion to regulate TC differentiation and proliferation. Therefore, in vitro experiments were conducted to study the effects of WNT family member 3A (WNT3A), WNT5A, RSPO1, DKK1, insulin-like growth factor 1 (IGF1), bone morphogenetic protein 7 (BMP7), Indian hedgehog (IHH), and fibroblast growth factor-9 (FGF9) on bovine TC proliferation and steroidogenesis. Theca cells of large (8 to 20 mm) and small (3 to 6 mm) follicles were collected from bovine ovaries, TC monolayers were established in vitro, and treated with various doses of recombinant human WNT3A, WNT5A, RSPO1, DKK1, IGF1, FGF9, BMP7, IHH and/or ovine LH in serum-free medium for 48 h. In experiment 1 using LH-treated TC, IGF1, IHH and WNT3A increased (P &lt; 0.05) cell numbers and androstenedione production, whereas WNT3A and BMP7 inhibited (P &lt; 0.05) progesterone production. In experiment 2, FGF9 blocked (P &lt; 0.05) the WNT3A-induced increase in androstenedione production in LH plus IGF1-treated TC. In experiment 3, RSPO1 further increased (P &lt; 0.05) LH plus IGF1-induced progesterone and androstenedione production. In experiment 4, SFRP4 and DKK1 alone had no significant effect on TC proliferation or progesterone production of large-follicle TC, but both blocked the inhibitory effect of WNT5A on androstenedione production. In contrast, DKK1 alone inhibited (P &lt; 0.05) small-follicle TC androstenedione production whereas SFRP4 was without effect. We conclude that the ovarian TC WNT system is functional in cattle, with WNT3A increasing proliferation and androstenedione production of TC.

SARS-CoV-2-Host Chimeric RNA-Sequencing Reads Do Not Necessarily Arise From Virus Integration Into the Host DNA

The human genome bears evidence of extensive invasion by retroviruses and other retroelements, as well as by diverse RNA and DNA viruses. High frequency of somatic integration of the RNA virus severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) into the DNA of infected cells was recently suggested, based on a number of observations. One key observation was the presence of chimeric RNA-sequencing (RNA-seq) reads between SARS-CoV-2 RNA and RNA transcribed from human host DNA. Here, we examined the possible origin specifically of human-SARS-CoV-2 chimeric reads in RNA-seq libraries and provide alternative explanations for their origin. Chimeric reads were frequently detected also between SARS-CoV-2 RNA and RNA transcribed from mitochondrial DNA or episomal adenoviral DNA present in transfected cell lines, which was unlikely the result of SARS-CoV-2 integration. Furthermore, chimeric reads between SARS-CoV-2 RNA and RNA transcribed from nuclear DNA were highly enriched for host exonic, rather than intronic or intergenic sequences and often involved the same, highly expressed host genes. Although these findings do not rule out SARS-CoV-2 somatic integration, they nevertheless suggest that human-SARS-CoV-2 chimeric reads found in RNA-seq data may arise during library preparation and do not necessarily signify SARS-CoV-2 reverse transcription, integration in to host DNA and further transcription.

Successful Radical Pneumonectomy for a Primitive Neuroendodermal Tumor in the Lung: A Case Report and Review of the Literature

Peripheral primitive neuroendodermal tumors (PNETs) and Ewing's sarcoma belong to the Ewing family of tumors and are small round-cell malignancies originating from spinal cord cells. These tumors account for 5% of all small round-cell malignant neoplasms. PNETs that arise from the lung parenchyma without pleural or chest wall involvement are very rare. We report a case of an adult female with a large pulmonary PNET who had given birth just 1 month prior to the diagnosis. She had cough and expectoration for 6 months, and the preoperative examination showed no metastases. Thus, we performed radical pneumonectomy and lymph node dissection. The patient recovered well without surgical complications and was discharged 7 days after the surgery. Postoperative pathology confirmed that the tumor was a small round-cell malignancy, and the tumor cells were positive for CD99, Friend leukemia virus integration 1 (FLI-1), and neuron-specific enolase (NSE), which was consistent with the diagnosis of a PNET. For primary large pulmonary PNETs, radical pneumonectomy may be a safe surgical method, worthy of further application in clinical practice.

SARS-CoV-2-host chimeric RNA-sequencing reads do not necessarily signify virus integration into the host DNA

ABSTRACTThe human genome bears evidence of extensive invasion by retroviruses and other retroelements, as well as by diverse RNA and DNA viruses. High frequency of somatic integration of the RNA virus severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) into the DNA of infected cells was recently suggested, partly based on the detection of chimeric RNA-sequencing (RNA-seq) reads between SARS-CoV-2 RNA and RNA transcribed from human host DNA. Here, we examined the possible origin of human-SARS-CoV-2 chimeric reads in RNA-seq libraries and provide alternative explanations for their origin. Chimeric reads were frequently detected also between SARS-CoV-2 RNA and RNA transcribed from mitochondrial DNA or episomal adenoviral DNA present in transfected cell lines, which was unlikely the result of SARS-CoV-2 integration. Furthermore, chimeric reads between SARS-CoV-2 RNA and RNA transcribed from nuclear DNA was highly enriched for host exonic, than intronic or intergenic sequences and often involved the same, highly expressed host genes. These findings suggest that human-SARS-CoV-2 chimeric reads found in RNA-seq data may arise during library preparation and do not necessarily signify SARS-CoV-2 reverse transcription, integration in to host DNA and further transcription.