Exploring Aberrant Expression Of Long Non-coding RNAs And Other Molecular Alterations In Cancer Pathology

Background:Cancer is a serious public health problem globally.Its burden is enormous,and is predicted to intensify by not less than 60%by 2040.They were new recorded cases of about 18.1 million in 2018,and was predicted to increase to 29.4 million in 2040 according to data from The World Health Organization.Most cancer deaths are as a result of poor comprehension of the genetic and molecular bases of the disease which has hindered the development of long-term therapeutic solutions,as well as the lack of reliable diagnostic and prognostic cancer biomarkers which accounts for the poor survival outcome in patients.Detailed comprehension of the processes that fuel carcinogenesis and disease progression,including the molecular and genetic alterations that incite tumor growth are needed for a more successful war against cancer.Hence,the overall aim of this thesis was to detect new molecules dysregulated in cancers,to examine their roles in cancer prognoses,and to uncover their mechanisms,in other to ascertain their clinical relevance as cancer biomarkers or targets for therapy.Specifically,the expression patterns and correlations of six long non-coding RNAs(lncRNAs)(PTPRG-AS1,FOXP4-AS1,BLACAT2,ZXF2,UCC and SAP30L-AS1)in gastric cancer and their clinical significance were investigated.Next,the expressions and prognostic importance of 8 CBX family members in gastric cancer were studied.Then,the expression,prognostic efficacy and molecular mechanism of VCAN in multiple cancers was evaluated.Finally,the involvement of the ribonucleotide reductase regulatory subunit M2(RRM2)in the tumorigenic processes of 30 different human cancers was elucidated.Methods:The expression profiles of six lncRNAs were examined by RT-qPCR analysis of 61 pairs of gastric cancer tissues and adjacent normal tissues(ANTs),and gastric cancer cell lines.Correlation analysis between lncRNA expressions and patients’ clinical features were done using Pearson Chi-squared test.Effects of UCC and SAP30L-AS1 on tumor growth and metastasis were evaluated by functional experiments in-vitro.Clinical values of FOXP4-AS1,BLACAT2,UCC and SAP30L-AS1 were determined using GEPIA database and PTPRG-AS1 in the KMPlot database.cBioPortal for Cancer Genomics was used to examine the frequencies of PTPRG-AS1 and BLACAT2 gene alterations.UALCAN database was used to study expression profile of the 8 CBXs in gastric cancer.This was validated using GEPIA database.Then,the Kaplan-Meier plotter was utilized to study the association of CBXs with prognosis of gastric cancer patients.Subgroup analysis based on different clinicopathological features of patients was also performed.cBioPortal for cancer genomics was used to examine CBX genes mutation in gastric cancer and their relationship with overall survival(OS)and disease-free survival(DFS).GeneMANIA database unveiled the genetic interaction network of CBXs and the functions of the genes in its network.mRNA expression levels of VCAN in many cancers and their subtypes were studied with ONCOMINE,and its protein expression levels were examined in the human protein atlas.The relationship between VCAN expression and clinical characteristics of patients in many cancers and their subtypes was assessed using UALCAN.With GEPIA,UALCAN,OncoLnc and Kaplan-Meier plotter we revealed its clinical importance in cancers.Mutations of VCAN gene and its relationship with prognosis was examined via cBioPortal for cancer genomics.GeneMANIA and STRING were used to construct genetic interaction networks for VCAN.PROMO was used to uncover transcription factors that bind to VCAN,and miRDB,TarBase,and mirDIP,were utilized for prediction of the miRNA targets of VCAN.The role and mechanism of ribonucleotide reductase regulatory subunit M2(RRM2)-a rate-limiting enzyme in the metabolic process required for DNA synthesis and cell replication-in 30 human cancers were investigated using databases and bioinformatics tools such as GEPIA,UALCAN,cBioPortal for cancer genomics,GeneMANIA,STRING,RPISeq,RNAup webserver,and PROMO.Results:PTPRG-AS1,FOXP4-AS1,BLACAT2,ZXF2,UCC and SAPOL-AS1 were overexpressed in gastric cancer tissues compared with ANTs and in gastric cancer cell lines.PTPRG-AS1 and ZXF2 expressions correlated with the cell proliferative marker Ki67 expression status.UCC and SAP30L-AS1 promoted gastric cancer growth in-vitro.UCC correlated with lymph node metastasis and SAP30L-AS1 promoted tumor cell migration in-vitro.Overexpression of FOXP4-AS1 was associated with favorable DFS prognosis,whereas,PTPRG-AS1 overexpression predicted worse prognostic outcomes for patients in different risk groups of gastric cancer.Genetic mutation of BLACAT2 correlated with worse DFS prognosis.All CBXs were found to be significantly deregulated in gastric cancer.CBX1/2/3/4/5/8 were upregulated while CBX6/7 were downregulated.Expressions of CBXs were associated with pathological stages and tumor grades linking CBXs with gastric pathogenesis.CBX2/3/4/5/6/7/8 had significant associations with OS of patients,CBX1/3/4/5/6/7/8 significantly correlated with the time to first progression(FP)for all patients,and all CBXs significantly correlated with post-progression survival(PPS)in all patients.Subgroup analysis also showed significant correlations of CBXs with OS,FP,and PPS based on HER2 status,Lauren classification,tumor stages,tumor grades,and different treatments.Analysis of genetic mutations of CBXs in gastric cancer revealed that CBX genes had 13%alterations.Their interactions with neighbor genes unveiled that all CBXs share the same protein domain,physically interact,and are co-expressed with genes related to histone acetyltransferase activity,methylated histone binding and modification of chromatin structure.Functional enrichment analysis also validated their role in epigenetic regulation necessary for proper differentiation of cells and mammalian development.Both mRNA levels and protein levels of VCAN were overexpressed in most cancers and VCAN expression positively correlated with the clinicopathological characteristics of patients in cancers and their subtypes showing its role in tumor initiation and progression.High VCAN expression correlated with shorter OS times and predicted poor OS prognosis in many tumors.VCAN gene mutation correlated with prognosis in terms of OS and DFS further revealing its association with clinical outcomes.Also,the genetic interactions of VCAN and the biological functions of genes in VCAN network,confirmed VC AN as an oncogene in humans.We identified transcription factors that bind to VCAN,including the famous p53,FOXP3,and others,which may be explorable in unveiling transcriptional regulation of VCAN in cancers.Also,hsa-miR-23b-3p was identified as a miRNA target of VCAN in synovial,brain,kidney,and bone marrow tissues and cell lines based on results of high-throughput experiments.hsa-miR-23b-3p may represent a new insight for the biological regulation of VCAN expression especially since it has also been experimentally proven to influence renal,breast,hepatocellular and gastric cancers.RRM2 was upregulated in all cancer types studied.In most cancers,its expression was associated with the pathological stage and nodal metastasis status of tumor,revealing it proto-oncogenic involvement in disease initiation and progression.Analysis of associated KEGG pathways also reiterated its oncogenicity.Overexpression of RRM2 significantly correlated with worse OS and DFS outcomes in majority of tumors.Genetic mutations of RRM2 predicted worse outcomes in some tumors and favorable outcomes in others,probably due to differences in cell types.RRM2 interacted with major drivers of oncogenesis including the core oncogenic transcription factor E2F1,which may simulate a novel approach towards understanding its mechanism in carcinogenesis,and exploration of the mechanism and associated pathways may be useful in identifying general therapeutic targets for cancers.Conclusions:PTPRG-AS1,FOXP4-AS1,BLACAT2,ZXF2,UCC and SAP30L-AS1 are potential markers for the molecular diagnosis of gastric cancer and their individual roles can be exploited for prognostic predictions and as potential targets for gastric cancer therapy.All CBXs are implicated in gastric tumorigenesis.Individual CBXs have different prognostic relevance in gastric cancer,and the CBXs network may represent a potential target for the treatment of gastric cancer.CBX1/4/5/6/8 expressions,if validated in large-scale studies,may have potentials for application in the prediction of response to chemotherapy and in gastric cancer management.VCAN may be an oncogene in many human cancers whose expression may be useful in prognostic predictions.Its mechanisms and interaction networks represent potential therapeutic strategies for cancers.RRM2 may be an oncogene in human cancers and a potential general tumor marker.Exploration of the mechanisms and pathways associated with RRM2 may be useful in identifying general therapeutic targets for human cancers.