Whole-exome and targeted gene sequencing of large-cell lung carcinoma reveals recurrent mutations in the PI3K pathway.

BACKGROUND: Large cell lung carcinoma (LCLC) is an exceptionally aggressive disease with a poor prognosis. At present, little is known about the molecular pathology of LCLC. METHODS: Ultra-deep sequencing of cancer-related genes and exome sequencing were used to detect the LCLC mutational in 118 tumor-normal pairs. The cell function test was employed to confirm the potential carcinogenic mutation of PI3K pathway. RESULTS: The mutation pattern is determined by the predominance of A > C mutations. Genes with a significant non-silent mutation frequency (FDR) < 0.05) include TP53 (47.5%), EGFR (13.6%) and PTEN (12.1%). Moreover, PI3K signaling (including EGFR, FGRG4, ITGA1, ITGA5, and ITGA2B) is the most mutated pathway, influencing 61.9% (73/118) of the LCLC samples. The cell function test confirmed that the potential carcinogenic mutation of PI3K pathway had a more malignant cell function phenotype. Multivariate analysis further revealed that patients with the PI3K signaling pathway mutations have a poor prognosis (P = 0.007). CONCLUSIONS: These results initially identified frequent mutation of PI3K signaling pathways in LCLC and indicate potential targets for the treatment of this fatal type of LCLC.

Liquid biopsy in lung cancer: significance in diagnostics, prediction, and treatment monitoring.

Primary lung cancer is one of the most common malignant tumors in China. Approximately 60% of lung cancer patients have distant metastasis at the initial diagnosis, so it is necessary to find new tumor markers for early diagnosis and individualized treatment. Tumor markers contribute to the early diagnosis of lung cancer and play important roles in early detection and treatment, as well as in precision medicine, efficacy monitoring, and prognosis prediction. The pathological diagnosis of lung cancer in small biopsy specimens determines whether there are tumor cells in the biopsy and tumor type. Because biopsy is traumatic and the compliance of patients with multiple biopsies is poor, liquid biopsy has become a hot research direction. Liquid biopsies are advantageous because they are nontraumatic, easy to obtain, reflect the overall state of the tumor, and allow for real-time monitoring. At present, liquid biopsies mainly include circulating tumor cells, circulating tumor DNA, exosomes, microRNA, circulating RNA, tumor platelets, and tumor endothelial cells. This review introduces the research progress and clinical application prospect of liquid biopsy technology for lung cancer.

The power and the promise of circRNAs for cancer precision medicine with functional diagnostics and prognostic prediction.

Circular RNA (circRNA) is a large class of covalently closed circRNA. As a member of competitive endogenous RNA, it participates in the regulation of circRNA-miRNA-mRNA network and plays an important role in the regulation of physiology and pathology. CircRNA is produced by the reverse splicing of exon, intron or both, forming exon or intron circRNA. Studies have shown that circRNA is a ubiquitous molecule, which exceeds the linear mRNA distributed in human cells. Because of its covalent closed-loop structure, circRNA is resistant to RNase R, which is more stable than linear mRNA; circRNA is highly conserved in different species. It was found that circRNA competitively adsorbs miRNA, as a miRNA sponge, to involve in the expression regulation of a variety of genes and plays an important role in tumor development, invasion, metastasis and other processes. These molecules offer new potential opportunities for therapeutic intervention and serve as biomarkers for diagnosis. In this paper, the origin, characteristics and functions of circRNA and its role in tumor development, invasion and metastasis, diagnosis and prognosis are reviewed.

microRNA-320b suppresses HNF4G and IGF2BP2 expression to inhibit angiogenesis and tumor growth of lung cancer.

We examined the effect of microRNA-320b (miR-320b) on tumor growth and angiogenesis in lung cancer and also determined its downstream molecular mechanisms. Lung cancer tissues and adjacent non-cancerous tissues were collected from 66 patients with lung cancer. miR-320b expression was experimentally determined to be expressed at low level in cancer tissues. The results of gain-of-function experiments suggested that miR-320b overexpression suppressed cancer cell invasion, tube formation, tumor volume and angiogenesis in xenografted nude mice. Hepatocyte nuclear factor 4 gamma (HNF4G) was identified as a target of miR-320b based on in silico analysis. Dual-luciferase reporter gene assays further identified the binding relationship between HNF4G and miR-320b. Lung cancer tissues exhibited increased expression of HNF4G and insulin-like growth factor 2 mRNA-binding protein 2 (IGF2BP2). Meanwhile, HNF4G knockdown suppressed IGF2BP2 expression, thereby repressing cancer cell invasion and tube formation. Furthermore, IGF2BP2 modified m6A to increase the expression of thymidine kinase 1 (TK1), thus promoting angiogenesis. In nude mice, restoration of TK1 reversed the suppressive effect of miR-320b overexpression on tumor growth rate and CD31 expression. In conclusion, miR-320b suppresses lung cancer growth and angiogenesis by inhibiting HNF4G, IGF2BP2 and TK1.

Cancer-associated fibroblast-derived exosomal microRNA-24-3p enhances colon cancer cell resistance to MTX by down-regulating CDX2/HEPH axis.

MicroRNA-24-3p (miR-24-3p) has been implicated as a key promoter of chemotherapy resistance in numerous cancers. Meanwhile, cancer-associated fibroblasts (CAFs) can secret exosomes to transfer miRNAs, which mediate tumour development. However, little is known regarding the molecular mechanism of CAF-derived exosomal miR-24-3p in colon cancer (CC). Hence, this study intended to characterize the functional relevance of CAF-derived exosomal miR-24-3p in CC cell resistance to methotrexate (MTX). We identified differentially expressed HEPH, CDX2 and miR-24-3p in CC through bioinformatics analyses, and validated their expression in CC tissues and cells. The relationship among HEPH, CDX2 and miR-24-3p was verified using ChIP and dual-luciferase reporter gene assays. Exosomes were isolated from miR-24-3p inhibitor-treated CAFs (CAFs-exo/miR-24-3p inhibitor), which were used in combination with gain-of-function and loss-of-function experiments and MTX treatment. CCK-8, flow cytometry and colony formation assays were conducted to determine cell viability, apoptosis and colony formation, respectively. Based on the findings, CC tissues and cells presented with high expression of miR-24-3p and low expression of HEPH and CDX2. CDX2 was a target gene of miR-24-3p and could up-regulate HEPH. Under MTX treatment, overexpressed CDX2 or HEPH and down-regulated miR-24-3p reduced cell viability and colony formation and elevated cell apoptosis. Furthermore, miR-24-3p was transferred into CC cells via CAF-derived exosomes. CAF-derived exosomal miR-24-3p inhibitor diminished cell viability and colony formation and increased cell apoptosis in vitro and inhibited tumour growth in vivo under MTX treatment. Altogether, CAF-derived exosomal miR-24-3p accelerated resistance of CC cells to MTX by down-regulating CDX2/HEPH axis.

KDM5A silencing transcriptionally suppresses the FXYD3-PI3K/AKT axis to inhibit angiogenesis in hepatocellular cancer via miR-433 up-regulation.

Hepatocellular cancer (HCC) has been reported to belong to one of the highly vascularized solid tumours accompanied with angiogenesis of human umbilical vein endothelial cells (HUVECs). KDM5A, an attractive drug target, plays a critical role in diverse physiological processes. Thus, this study aims to investigate its role in angiogenesis and underlying mechanisms in HCC. ChIP-qPCR was utilized to validate enrichment of H3K4me3 and KDM5A on the promotor region of miR-433, while dual luciferase assay was carried out to confirm the targeting relationship between miR-433 and FXYD3. Scratch assay, transwell assay, Edu assay, pseudo-tube formation assay and mice with xenografted tumours were conducted to investigate the physiological function of KDM5A-miR-433-FXYD3-PI3K-AKT axis in the progression of HCC after loss- and gain-function assays. KDM5A p-p85 and p-AKT were highly expressed but miR-433 was down-regulated in HCC tissues and cell lines. Depletion of KDM5A led to reduced migrative, invasive and proliferative capacities in HCC cells, including growth and a lowered HUVEC angiogenic capacity in vitro. Furthermore, KDM5A suppressed the expression of miR-433 by demethylating H3K4me3 on its promoterregion. miR-433 negatively targeted FXYD3. Depleting miR-433 or re-expressing FXYD3 restores the reduced migrative, invasive and proliferative capacities, and lowers the HUVEC angiogenic capacity caused by silencing KDM5A. Therefore, KDM5A silencing significantly suppresses HCC tumorigenesis in vivo, accompanied with down-regulated miR-433 and up-regulated FXYD3-PI3K-AKT axis in tumour tissues. Lastly, KDM5A activates the FXYD3-PI3K-AKT axis to enhance angiogenesis in HCC by suppressing miR-433.

Long non-coding RNA NORAD promotes pancreatic cancer stem cell proliferation and self-renewal by blocking microRNA-202-5p-mediated ANP32E inhibition.

BACKGROUND: Cancer stem cells (CSCs) are key regulators in the processes of tumor initiation, progression, and recurrence. The mechanism that maintains their stemness remains enigmatic, although the role of several long noncoding RNAs (lncRNAs) has been highlighted in the pancreatic cancer stem cells (PCSCs). In this study, we first established that PCSCs overexpressing lncRNA NORAD, and then investigated the effects of NORAD on the maintenance of PCSC stemness. METHODS: Expression of lncRNA NORAD, miR-202-5p and ANP32E in PC tissues and cell lines was quantified after RNA isolation. Dual-luciferase reporter assay, RNA pull-down and RIP assays were performed to verify the interactions among NORAD, miR-202-5p and ANP32E. We then carried out gain- and loss-of function of miR-202-5p, ANP32E and NORAD in PANC-1 cell line, followed by measurement of the aldehyde dehydrogenase activity, cell viability, apoptosis, cell cycle distribution, colony formation, self-renewal ability and tumorigenicity of PC cells. RESULTS: LncRNA NORAD and ANP32E were upregulated in PC tissues and cells, whereas the miR-202-5p level was down-regulated. LncRNA NORAD competitively bound to miR-202-5p, and promoted the expression of the miR-202-5p target gene ANP32E thereby promoting PC cell viability, proliferation, and self-renewal ability in vitro, as well as facilitating tumorigenesis of PCSCs in vivo. CONCLUSION: Overall, lncRNA NORAD upregulates ANP32E expression by competitively binding to miR-202-5, which accelerates the proliferation and self-renewal of PCSCs.

The role of ceRNA-mediated diagnosis and therapy in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related death worldwide due to its high degree of malignancy, high incidence, and low survival rate. However, the underlying mechanisms of hepatocarcinogenesis remain unclear. Long non coding RNA (lncRNA) has been shown as a novel type of RNA. lncRNA by acting as ceRNA can participate in various biological processes of HCC cells, such as tumor cell proliferation, migration, invasion, apoptosis and drug resistance by regulating downstream target gene expression and cancer-related signaling pathways. Meanwhile, lncRNA can predict the efficacy of treatment strategies for HCC and serve as a potential target for the diagnosis and treatment of HCC. Therefore, lncRNA serving as ceRNA may become a vital candidate biomarker for clinical diagnosis and treatment. In this review, the epidemiology of HCC, including morbidity, mortality, regional distribution, risk factors, and current treatment advances, was briefly discussed, and some biological functions of lncRNA in HCC were summarized with emphasis on the molecular mechanism and clinical application of lncRNA-mediated ceRNA regulatory network in HCC. This paper can contribute to the better understanding of the mechanism of the influence of lncRNA-mediated ceRNA networks (ceRNETs) on HCC and provide directions and strategies for future studies.

New Therapeutic Options for Advanced Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC), one of the most common lethal diseases in the world, has a 5-year survival rate of only 7%. Hepatocellular carcinoma has no symptoms in the early stage but obvious symptoms in the late stage, leading to delayed diagnosis and reduced treatment efficacy. In recent years, as the scope of HCC research has increased in depth, the clinical development and application of molecular targeted drugs and immunotherapy drugs have brought new breakthroughs in HCC treatment. Targeted therapy drugs for HCC have high specificity, allowing them to selectively kill tumor cells and minimize damage to normal tissues. At present, these targeted drugs are mainly classified into 3 categories: small molecule targeted drugs, HCC antigen-specific targeted drugs, and immune checkpoint targeted drugs. This article reviews the latest research progress on the targeted drugs for HCC.

Power and Promise of Next-Generation Sequencing in Liquid Biopsies and Cancer Control.

Traditional methods of cancer treatment are usually based on the morphological and histological diagnosis of tumors, and they are not optimized according to the specific situation. Precision medicine adjusts the existing treatment regimen based on the patient's genomic information to make it most suitable for patients. Detection of genetic mutations in tumors is the basis of precise cancer medicine. Through the analysis of genetic mutations in patients with cancer, we can tailor the treatment plan for each patient with cancer to maximize the curative effect, minimize damage to healthy tissues, and optimize resources. In recent years, next-generation sequencing technology has developed rapidly and has become the core technology of precise targeted therapy and immunotherapy for cancer. From early cancer screening to treatment guidance for patients with advanced cancer, liquid biopsy is increasingly used in cancer management. This is as a result of the development of better noninvasive, repeatable, sensitive, and accurate tools used in early screening, diagnosis, evaluation, and monitoring of patients. Cell-free DNA, which is a new noninvasive molecular pathological detection method, often carries tumor-specific gene changes. It plays an important role in optimizing treatment and evaluating the efficacy of different treatment options in clinical trials, and it has broad clinical applications.

Correction to: Proteogenomic characterization and comprehensive integrative genomic analysis of human colorectal cancer liver metastasis.

Following publication of the original article [1], the authors reported an error in affiliation 5.

miR-30 Family Reduction Maintains Self-Renewal and Promotes Tumorigenesis in NSCLC-Initiating Cells by Targeting Oncogene TM4SF1.

Increasing evidence indicates that tumor-initiating cells (TICs) are responsible for the occurrence, development, recurrence, and development of the drug resistance of cancer. MicroRNA (miRNA) plays a significant functional role by directly regulating targets of TIC-triggered non-small-cell lung cancer (NSCLC), but little is known about the function of the miR-30 family in TICs. In this study, we found the miR-30 family to be downregulated during the spheroid formation of NSCLC cells, and patients with lower miR-30a/c expression had shorter overall survival (OS) and progression-free survival (PFS). Moreover, transmembrane 4 super family member 1 (TM4SF1) was confirmed to be a direct target of miR-30a/c. Concomitant low expression of miR-30a/c and high expression of TM4SF1 correlated with a shorter median OS and PFS in NSCLC patients. miR-30a/c significantly inhibited stem-like characteristics in vitro and in vivo via suppression of its target gene TM4SF1, and then it inhibited the activity of the mTOR/AKT-signaling pathway. Thus, our data provide the first evidence that TM4SF1 is a direct target of miR-30a/c and miR-30a/c inhibits the stemness and proliferation of NSCLC cells by targeting TM4SF1, suggesting that miR-30a/c and TM4SF1 may be useful as tumor biomarkers for the diagnosis and treatment of NSCLC patients.

Proteogenomic characterization and comprehensive integrative genomic analysis of human colorectal cancer liver metastasis.

BACKGROUND: Proteogenomic characterization and integrative and comparative genomic analysis provide a functional context to annotate genomic abnormalities with prognostic value. METHODS: Here, we analyzed the proteomes and performed whole exome and transcriptome sequencing and single nucleotide polymorphism array profiling for 2 sets of triplet samples comprised of normal colorectal tissue, primary CRC tissue, and synchronous matched liver metastatic tissue. RESULTS: We identified 112 CNV-mRNA-protein correlated molecules, including up-regulated COL1A2 and BGN associated with prognosis, and four strongest hot spots (chromosomes X, 7, 16 and 1) driving global mRNA abundance variation in CRC liver metastasis. Two sites (DMRTB1R202H and PARP4V458I) were revealed to frequent mutate only in the liver metastatic cohort and displayed dysregulated protein abundance. Moreover, we confirmed that the mutated peptide number has potential prognosis value and somatic variants displayed increased protein abundance, including high MYH9 and CCT6A expression, with clinical significance. CONCLUSIONS: Our proteogenomic characterization and integrative and comparative genomic analysis provides a new paradigm for understanding human colon and rectal cancer liver metastasis. TRIAL REGISTRATION: ClinicalTrials, NCT02917707. Registered 28 September 2016, https://clinicaltrials.gov/ct2/show/NCT02917707 .

Downregulation of miR-223 and miR-19a induces differentiation and promotes recruitment of osteoclast cells in giant-cell tumor of the bone via the Runx2/TWIST-RANK/RANKL pathway.

Giant-cell tumor (GCT) of the bone is an invasiveness and high recurrent bone tumor that is considered borderline or potentially malignant. To explore the molecular mechanism leading to bone destruction and identify novel targets for treatment, we conducted silencing of miR-223 and miR-19a in stromal giant cells and identified TWIST and Runx2 as their target genes. We investigated the impact of these microRNAs and their target genes on stromal giant cells that promote the differentiation of monocyte/macrophages into osteoclast cells and recruitment to the bone microenvironment, which in turn enhances the bone destruction capacity of GCT. MiR-223 and miR-19a were found to regulate the expression of TWIST and Runx2, influence the RANKL-RANK pathway and the expression of MCP-1, and finally regulate the pathophysiological process of osteolytic bone destruction. Our results indicate that re-expression of miR-223 and miR-19a induces an inhibitory effect on the bone destruction capacity of GCT, suggesting that re-expression of miR-223 and miR-19a can be a novel strategy for the treatment of GCT.

Reduced hsa-miR-124-3p levels are associated with the poor survival of patients with hepatocellular carcinoma.

Hsa-MicroRNA-124a-3p (hsa-miR-124-3p) is involved in tumor progression in certain malignant tumors. However, its function and clinical implication in hepatocellular carcinoma (HCC) have not yet been illustrated. In this study, we explored the expression and prognostic value of hsa-miR-124-3p in patients with HCC. Hsa-miR-124-3p expression in HCC was analyzed in silico, which was subsequently confirmed by quantitative PCR in 155 HCC biopsy samples. Overall survival (OS) and disease-free survival in HCC patients was evaluated by Kaplan-Meier survival analysis, and univariate and multivariate Cox proportional hazard models were used. The in silico results demonstrated that hsa-miR-124-3p was reduced in cell lines and tissues of HCC, and hsa-miR-124-3p expression was lower in HCC tumor samples than in normal liver tissues. Moreover, a decrease in hsa-miR-124-3p expression was closely correlated with tumor diameter (≥ 5 cm) and number of lesions (multiple). Lower hsa-miR-124-3p expression was shown to be correlated with a shorter OS and poor prognosis in HCC. Our findings demonstrate that hsa-miR-124-3p might be a potential target for the diagnosis and prognosis of HCC.

Upregulation of miR-181a suppresses the formation of glioblastoma stem cells by targeting the Notch2 oncogene and correlates with good prognosis in patients with glioblastoma multiforme.

Glioblastoma stem-like cells (GSCs) are responsible for the initiation and progression of glioblastoma multiforme (GBM), and microRNAs (miRNAs) play an important role in this disease. However, the mechanisms underlying the role of miRNAs in the stemness of GSCs have not been completely elucidated. We previously showed that miR-181a is downregulated in GBM and may predict prognosis in patients with this disease. Here, we demonstrate that the upregulation of miR-181a suppressed GSC formation and inhibited GBM tumorigenesis by targeting the Notch2 oncogene. We found that miR-181a was downregulated in GSCs derived from human glioblastoma U87MG and U373MG cells. The high expression of miR-181a inhibited the levels of stemness-related markers CD133 and BMI1, attenuated sphere proliferation, promoted cell apoptosis, and reduced the tumorigenicity of GSCs. MiR-181a decreased the expression of Notch2 by targeting the 3'-untranslated region of its mRNA. Notch2 overexpression inhibited the effects of miR-181a downregulation on GSCs, and was negatively correlated with miR-181a expression. Moreover, high Notch2 expression together with low miR-181a expression was correlated with a shorter median overall survival for GBM patients. Together, these data show that miR-181a may play an essential role in GSC formation and GBM progression by targeting Notch2, suggesting that Notch2 and miR-181a have potential prognostic value as tumor biomarkers in GBM patients.

Up-regulated microRNA-299 corrected with poor prognosis of glioblastoma multiforme patients by targeting ELL2.

BACKGROUND: An increasing understanding of the genes and molecular pathways of glioblastoma multiforme (GBM) can provide us a useful insight for the development of more effective targeted therapeutic. METHODS: To investigate the expression and clinical significance of miR-299 and its target genes in GBM, the expression levels of miR-299 and its target gene in human normal brain tissues and GBM were analyzed in silico using genes microarray and hierarchical clustering analysis followed by validation with quantitative RT-PCR. RESULTS: Our results show that miR-299 is up-regulated in GBM patients. Moreover, patients with low miR-299 expression had longer overall survival (OS) compared with those with high miR-299 expression. RNA polymerase II elongation factor, ELL2, was identified as a miR-299 direct target. High expression of ELL2 together with miR-299 down-regulation correlated with a shorter median OS. CONCLUSIONS: Our results provide the first evidence that ELL2 is a direct target of miR-299 and increased ELL2 expression and down-regulation of miR-299 are associated with GBM progression and poor prognosis in patients, suggesting that ELL2 and miR-299 might have potential prognostic value and be used as tumor biomarkers for the diagnosis of patients with GBM.

Association of glutathione S-transferase M1, T1, and P1 polymorphisms with renal cell carcinoma: evidence from 11 studies.

The glutathione S-transferases (GSTs) are a gene superfamily of phase II metabolic enzymes that has attracted a considerable attention as a candidate gene for renal cell carcinoma (RCC) based on its enzyme function as a key factor in biotransformation pathways. In the past decade, a number of case-control studies were conducted to investigate the association of GST genetic polymorphisms and RCC risk. However, studies on the association between GST (GSTM1, GSTT1, and GSTP1) polymorphisms and RCC remain to be conflicting. To derive a more precise estimation of the relationship, a meta-analysis of 2,189 cases and 3,817 controls from 11 case-control studies was performed. Overall, the summarized odds ratio for RCC of the GSTM1 null and GSTT1 null polymorphisms was 1.02 (95% confidence interval (CI) 0.91-1.15, P = 0.70) and 1.28 (95% CI 0.96-1.72, P = 0.09), respectively. No significant results were observed in heterozygous and homozygous genotypes when compared with wild-type genotype for GSTP1 I105V polymorphism. However, the GSTM1-GSTT1 interaction analysis showed that the dual null genotype of GSTM1/GSTT1 was significantly associated with an increased RCC risk (odds ratio (OR) = 1.42, 95% CI 1.14-1.76, P = 0.001). In the stratified analyses by ethnicity, significant gene-disease association was obtained among Asians for GSTT1 and GSTP1 polymorphisms. In our meta-analysis, the associations between variations of GSTs and RCC may vary in different ethnic populations, and the interaction between unfavorable GST genotypes may exist.

Differential gene expression of the key signalling pathway in para-carcinoma, carcinoma and relapse human pancreatic cancer.

Pancreatic cancer (PC) has a high rate of mortality and a poorly understood mechanism of progression. Investigation of the molecular mechanism of PC and exploration of the specific markers for early diagnosis and specific targets of therapy are key points to prevent and treat PC effectively and to improve their prognosis. In our study, expression profiles experiment of para-carcinoma, carcinoma and relapse human PC was performed using Agilent human whole genomic oligonucleotide microarrays with 45 000 probes. Differentially expressed genes related with PC were screened and analysed further by Gene Ontology term analysis and Kyoto encyclopaedia of genes and genomes pathway analysis. Our results showed that there were 3853 differentially expressed genes associated with pancreatic carcinogenesis and relapse. In addition, our study found that PC was related to the Jak-STAT signalling pathway, PPAR signalling pathway and Calcium signalling pathway, indicating their potential roles in pancreatic carcinogenesis and progress.

KLK10/LIPH/PARD6B/SLC52A3 are promising molecular biomarkers for the prognosis of pancreatic cancer through a ceRNA network.

Pancreatic adenocarcinoma (PAAD) remains challenging to diagnose and treat clinically due to its difficult early diagnosis, low surgical resection rate, and high risk of postoperative recurrence and metastasis. SMAD4 is a classical mutated gene in pancreatic cancer and is lost in up to 60%-90 % of PAAD patients, and its mutation often predicts a poor prognosis and treatment resistance. In this study, based on the expression profile data in The Cancer Genome Atlas database, we identified a ceRNA network composed of 2 lncRNAs, 1 miRNA, and 4 mRNAs through differential expression analysis and survival prognosis analysis. Among them, high expression of KLK10/LIPH/PARD6B/SLC52A3 influenced the prognosis and overall survival of PAAD patients. We confirmed the high expression of these target genes in pancreatic tissue of pancreatic-specific SMAD4-deficient mice. In addition, immune infiltration analysis showed that the high expression of these target genes affects the tumor immune environment and contributes to the progression of PAAD. Abnormal overexpression of these target genes may be caused by hypermethylation. In conclusion, we found that KLK10/LIPH/PARD6B/SLC52A3 is a potential prognostic marker for PAAD based on a competing endogenous RNA-mediated mechanism and revealed the potential pathogenic mechanism by which deficient expression of SMAD4 promotes pancreatic cancer progression, which provides a new pathway and theoretical basis for targeted therapy or improved prognosis of pancreatic cancer. These data will help reveal potential therapeutic targets for pancreatic cancer and improve the prognosis of pancreatic cancer patients.