Cuproptosis-related gene CDKN2A as a molecular target for IPF diagnosis and therapeutics.

BACKGROUND: Idiopathic pulmonary fibrosis (IPF) is a progressive chronic interstitial lung disease with limited therapeutic options. Cuproptosis is a recently proposed novel form of programmed cell death, which has been strongly implicated in the development of various human diseases. However, the prognostic and therapeutic value of cuproptosis-related genes (CRGs) in IPF remains to be elucidated. METHODS: In the present study, weighted gene co-expression network analysis (WGCNA) was employed to identify the key CRGs associated with the development of IPF. The subsequent GSEA, immune cell correlation analysis, and single-cell RNA-Seq analysis were conducted to explore the potential role of the identified CRGs in IPF. In addition, ROC curves and survival analysis were used to assess the prognostic value of the key CRGs in IPF. Moreover, we explored the molecular mechanisms of participation of identified key CRGs in the development of pulmonary fibrogenesis through in vivo and in vitro experiments. RESULTS: The expression level of cyclin-dependent kinase inhibitor 2A (CDKN2A) is upregulated in the lung tissues of IPF patients and associated with disease severity. Notably, CDKN2A was constitutively expressed by fibrosis-promoting M2 macrophages. Decreased CDKN2A expression sensitizes M2 macrophages to elesclomol-induced cuproptosis in vitro. Inhibition of CDKN2A decreases the number of viable macrophages and attenuates bleomycin-induced pulmonary fibrosis in mice. CONCLUSION: These findings indicate that CDKN2A mediates the resistance of fibrosis-promoting M2 macrophages to cuproptosis and promotes pulmonary fibrosis in mice. Our work provides fresh insights into CRGs in IPF with potential value for research in the pathogenesis, diagnosis, and a new therapy strategy for IPF.

CDKN2A deregulation in fatty liver disease and its accelerative role in the process of lipogenesis.

Previous literature has indicated that cyclin-dependent kinase inhibitor 2 A (CDKN2A) is upregulated, while the Protein Inhibitor of Activated STAT1 (PIAS1) is downregulated in the liver tissues of obese mice. The current study aimed to investigate the relationship between CDKN2A and PIAS1 in the lipogenesis of fatty liver disease. In the C57BL/6J db/db mouse model and hepatocyte model of fatty liver, the expression pattern of CDKN2A, PIAS1, Protein arginine methyltransferase 1 (PRMT1) and CASP8 and FADD-like apoptosis regulator (CFLAR) was characterized by RNA quantitative and Western blot analysis. The lipogenesis-related genes (Srebp1c and Fas) in the liver tissues and cells were employed in the assessment of lipogenesis in response to gain- or loss-of-function of CDKN2A, PIAS1, PRMT1, and CFLAR, while triglyceride and fat content were evaluated in relation to fat accumulation. Western blot analysis was conducted to determine c-Jun amino-terminal kinase (JNK) phosphorylation, while the ubiquitination of CFLAR and SUMOylation of PIAS1 was examined by immunoprecipitation. PIAS1 and CFLAR were downregulated, while CDKN2A, PRMT1, and phosphorylation of JNK was elevated in the tissues and cells of the fatty liver models. Our results suggested that CDKN2A enhanced the SUMOylation of PIAS1 to reduce the expression of PIAS1. PRMT1 downregulated CFLAR by triggering its ubiquitination, while CFLAR repressed phosphorylation of JNK. The in vitro and in vivo results indicated that CDKN2A silencing prevented lipogenesis and fat accumulation by impairing the PRMT1-dependent ubiquitination of CFLAR and blocking the phosphorylation of JNK. Taken together, the central observations of our study demonstrate that targeting CDKN2A contributes to the suppression of lipogenesis and fat accumulation in fatty liver disease. The findings of our study highlight the potential of CDKN2A as a promising target against fatty liver.

Knockout of the caspase 8-associated protein 2 gene improves recombinant protein expression in HEK293 cells through up-regulation of the cyclin-dependent kinase inhibitor 2A gene.

Cell lines used in bioproduction are routinely engineered to improve their production efficiency. Numerous strategies, such as random mutagenesis, RNA interference screens, and transcriptome analyses have been employed to identify effective engineering targets. A genome-wide small interfering RNA screen previously identified the CASP8AP2 gene as a potential engineering target for improved expression of recombinant protein in the HEK293 cell line. Here, we validate the CASP8AP2 gene as an engineering target in HEK293 cells by knocking it out using CRISPR/Cas9 genome editing and assessing the effect of its knockout on recombinant protein expression, cell growth, cell viability, and overall gene expression. HEK293 cells lacking CASP8AP2 showed a seven-fold increase in specific expression of recombinant luciferase and a 2.5-fold increase in specific expression of recombinant SEAP, without significantly affecting cell growth and viability. Transcriptome analysis revealed that the deregulation of the cell cycle, specifically the upregulation of the cyclin-dependent kinase inhibitor 2A (CDKN2A) gene, contributed to the improvement in recombinant protein expression in CASP8AP2 deficient cells. The results validate the CASP8AP2 gene is a viable engineering target for improved recombinant protein expression in the HEK293 cell line.

Association between rs3088440 (G > A) polymorphism at 9p21.3 locus with the occurrence and severity of coronary artery disease in an Iranian population.

BACKGROUND: Several genome-wide association studies showed that a series of genetic variants located at the chromosome 9p21 locus are strongly associated with coronary artery disease (CAD). RATIONALE AND PURPOSE OF THE STUDY: In the present study, the relationship of rs3088440 (G > A) in cyclin-dependent kinase inhibitor 2A (CDKN2A) gene site with the presence of coronary artery disease (CAD) and its severity was evaluated in an Iranian population. METHODS AND RESULTS: The presence of rs3088440 (G > A) genotypes was assessed by polymerase chain reaction-based restriction fragment length polymorphism (PCR-RFLP) technique in 324 CAD patients and 148 normal controls. rs3088440 (G > A) polymorphism was associated with increased risk of CAD in the total population (adjusted OR = 1.76, 95% CI = 1.10-2.82; p-value = 0.017) or in women (adjusted OR = 2.96, 95% CI = 1.34-6.55; p-value = 0.007), but not in the men (adjusted OR = 1.35, 95% CI = 0.70-2.6; p-value = 0.368). CONCLUSIONS: Our findings suggest that the presence of rs3088440 (G > A) is potentially linked with the risk of CAD and its severity in whole study subjects or in women only, independent of CAD risk factors.

CDKN2A, CDK1, and CCNE1 overexpression in sebaceous gland carcinoma of eyelid.

PURPOSE: To investigate the overexpression of genes in sebaceous gland carcinoma (SGC) of the eyelid compared to sebaceous adenoma of the eyelid in order to elucidate the molecular mechanism underlying pathogenesis. METHODS: We performed histopathological examination of eyelid tissues surgically removed from four patients diagnosed with SGC (cases 1-3) and sebaceous adenoma (case 4) of the eyelid. Next, we performed global gene expression analysis of surgical tissue samples using a GeneChip® system and the Ingenuity Pathways Knowledge Base. The results of the GeneChip® analysis were explored with quantitative real-time polymerase chain reaction (qRT-PCR) analysis. RESULTS: In the SGC samples, we found that 211, 199, and 199 genes, respectively, showed ≥ 2.0-fold higher expression than those in the sebaceous adenoma sample (case 4); 194 genes were common to all three SGC samples. For the 194 genes with upregulated expression, functional category analysis showed that SGC of the eyelid employed a unique gene network, including cyclin-dependent kinase inhibitor 2A (CDKN2A), cyclin-dependent kinase 1 (CDK1), and cyclin E1 (CCNE1), which are related to cell cycle progression, incidence of tumor, and cell viability. Furthermore, qRT-PCR analysis showed that the expression levels of CDKN2A, CDK1, and CCNE1 were significantly upregulated in all SGC cases compared to those in the sebaceous adenoma case. These data were similar to the results of microarray analysis. CONCLUSION: Overexpression of cell cycle-related genes CDKN2A, CDK1, CCNE1, and their gene network may help elucidate the pathogenic pathway of SGC of the eyelid at the molecular level.

Comparative susceptibilities and immune-related gene expressions of brown trout strains and their hybrids infected with Lactococcus garvieae and Yersinia ruckeri.

Brown trout are polymorphic salmonid species, and it is of importance to investigate whether hybridization affects disease resistance. In this study, susceptibility of brown trout (Salmo trutta Abant, Anatolian, Black Sea, and Caspius) strains and their hybrids to Lactococcus garvieae and Yersinia ruckeri as well as their immune-related gene expression profiles were studied. Results indicated that reciprocal hybridization did not affect disease resistance in brown trout strains. Purebred Black Sea strain of brown trout was the most resistant group against Y. ruckeri, followed by other Black Sea strain hybrids. On the other hand, purebred Anatolian strain was the most resistant group to L. garvieae, followed by other Anatolian strain hybrids. Expression pattern of target genes differed in families, but the overall gene expression was comparatively high in Y. ruckeri infected families. Upregulations were mainly significant at 7 and 28 d post infection while marginal regulations were observed 8 h after infection. Disease resistance status of strains was supported by high expression of immune-related genes such as major histocompatibility complex class I (MHC-I), immunoglobulin light chain (IgL), and antioxidant- and hemoglobin-related gene expression. Therefore, our findings suggest that Black Sea and Anatolian strains could be used to develop fish stock that are resistant for yersiniosis and lactocaccosis, respectively.

Pediatric melanoma in melanoma-prone families.

BACKGROUND: In the United States, only approximately 0.4% of all melanomas are diagnosed in patients aged <20 years. To the authors' knowledge, melanoma in pediatric members of melanoma-prone families has not been fully investigated to date. The objective of the current study was to evaluate pediatric patients with melanoma with extensive follow-up in melanoma-prone families with and without cyclin-dependent kinase inhibitor 2A (CDKN2A) mutations. METHODS: For this non-population-based study, families were followed prospectively for up to 40 years. A total of 60 families with ≥ 3 patients with melanoma were included for analysis: 30 CDKN2A mutation-positive (CDKN2A+) and 30 CDKN2A mutation-negative (CDKN2A-) families. Age at the time of first melanoma and number of melanomas were obtained for each patient and summarized by family or sets (CDKN2A + vs CDKN2A-). For set comparisons and categorical variables (occurrence of melanoma in pediatric patients, number of melanomas, number of patients with single or multiple melanomas), the Pearson chi-square or Fisher exact test was used. RESULTS: Regardless of CDKN2A status, melanoma-prone families were found to have 6-fold to 28-fold higher percentages of patients with pediatric melanoma compared with the general population of patients with melanoma in the United States. Within CDKN2A + families, pediatric patients with melanoma were significantly more likely to have multiple melanomas compared with their relatives who were diagnosed at age >20 years (71% vs 38%, respectively; P = .004). CDKN2A + families had significantly higher percentages of pediatric patients with melanoma compared with CDKN2A- families (11.1% vs 2.5%; P = .004). CONCLUSIONS: These observations have implications for the prevention of melanoma as well as clinical care for its early detection. Children in melanoma-prone families should have careful sun protection from an early age and skin surveillance to reduce their risk of melanoma.

Melanoma: What do all the mutations mean?

Melanoma is one of the most highly mutated malignancies, largely as a function of its generation through ultraviolet light and other mutational processes. The wide array of mutations in both "driver" and "passenger" genes can present a confusing array of data for practitioners, particularly within the context of the recent revolutions in targeted and immune therapy. Although mutations in BRAF V600 clearly confer sensitivity to BRAF and mitogen-activated protein kinase kinase (MEK) inhibitors, the clinical implications of most other mutations are less often discussed and understood. In this review, we provide an overview of the high-frequency genomic alterations and their prognostic and therapeutic relevance in melanoma.

Expression of p16INK4A in cervical precancerous lesions that is unlikely to be preventable by human papillomavirus vaccines.

BACKGROUND: Whether higher grade cervical intraepithelial neoplasia (CIN grade 2 or greater [CIN ≥ 2]) that develops because of human papillomavirus (HPV) genotypes not included in vaccines may progress to cervical cancer is largely unknown. The objectives of this study were to document expression of the cyclin-dependent kinase inhibitor 2A (p16) tumor-suppressor protein p16INK4A as a biomarker of cervical carcinogenesis or of malignant potential and to evaluate whether its expression differs between lesions associated with vaccine and nonvaccine high-risk (HR) human papillomavirus (HPV) genotypes. METHODS: The study population consisted of 371 women who had not received HPV vaccines. Women were categorized into vaccine and nonvaccine HR-HPV genotypes and lesions associated with those types. Logistic regression analyses were used to determine the association between positive expression p16INK4A and the risk of being diagnosed with CIN 2 or CIN 3. Differences in the proportion of CIN ≥2 lesions that were positive for p16INK4A expression by vaccine-related or nonvaccine-related HR-HPV genotype were determined using the Pearson chi-square test. RESULTS: Specimens that were positive for p16INK4A expression were 5.3 and 16.6 times more likely to be diagnosed as CIN 2 and CIN 3 lesions, respectively, compared to CIN 1 lesions. CIN ≥ 2 lesions that were negative for the bivalent and 9-valent HR-HPV genotypes had similar rates of positive p16INK4A expression compared with lesions that were positive for those HR-HPV genotypes. CONCLUSIONS: Lesions that may develop because of HR-HPV genotypes not included in HPV vaccines are likely to have similar malignant potential, suggesting that well developed screening programs combined with nonvaccine-based approaches may be needed to manage the residual risk of developing cervical cancer in the post-HPV vaccination era. Cancer 2016;122:3615-23. © 2016 American Cancer Society.

Comprehensive genomic profiling of 295 cases of clinically advanced urothelial carcinoma of the urinary bladder reveals a high frequency of clinically relevant genomic alterations.

BACKGROUND: In the current study, the authors present a comprehensive genomic profile (CGP)-based study of advanced urothelial carcinoma (UC) designed to detect clinically relevant genomic alterations (CRGAs). METHODS: DNA was extracted from 40 µm of formalin-fixed, paraffin-embedded sections from 295 consecutive cases of recurrent/metastatic UC. CGP was performed on hybridization-captured, adaptor ligation-based libraries to a mean coverage depth of 688X for all coding exons of 236 cancer-related genes plus 47 introns from 19 genes frequently rearranged in cancer, using process-matched normal control samples as a reference. CRGAs were defined as GAs linked to drugs on the market or currently under evaluation in mechanism-driven clinical trials. RESULTS: All 295 patients assessed were classified with high-grade (International Society of Urological Pathology classification) and advanced stage (stage III/IV American Joint Committee on Cancer) disease, and 294 of 295 patients (99.7%) had at least 1 GA on CGP with a mean of 6.4 GAs per UC (61% substitutions/insertions/deletions, 37% copy number alterations, and 2% fusions). Furthermore, 275 patients (93%) had at least 1 CRGA involving 75 individual genes with a mean of 2.6 CRGAs per UC. The most common CRGAs involved cyclin-dependent kinase inhibitor 2A (CDKN2A) (34%), fibroblast growth factor receptor 3 (FGFR3) (21%), phosphatidylinositol 3-kinase catalytic subunit alpha (PIK3CA) (20%), and ERBB2 (17%). FGFR3 GAs were diverse types and included 10% fusions. ERBB2 GAs were equally divided between amplifications and substitutions. ERBB2 substitutions were predominantly within the extracellular domain and were highly enriched in patients with micropapillary UC (38% of 32 cases vs 5% of 263 nonmicropapillary UC cases; P<.0001). CONCLUSIONS: Using a CGP assay capable of detecting all classes of GA simultaneously, an extraordinarily high frequency of CRGA was identified in a large series of patients with advanced UC. Cancer 2016;122:702-711. © 2015 American Cancer Society.

Multiple primary melanomas (MPMs) and criteria for genetic assessment: MultiMEL, a multicenter study of the Italian Melanoma Intergroup.

BACKGROUND: Multiple primary melanoma (MPM), in concert with a positive family history, is a predictor of cyclin-dependent kinase (CDK) inhibitor 2A (CDKN2A) germline mutations. A rule regarding the presence of either 2 or 3 or more cancer events (melanoma and pancreatic cancer) in low or high melanoma incidence populations, respectively, has been established to select patients for genetic referral. OBJECTIVE: We sought to determine the CDKN2A/CDK4/microphthalmia-associated transcription factor mutation rate among Italian patients with MPM to appropriately direct genetic counseling regardless of family history. METHODS: In all, 587 patients with MPM and an equal number with single primary melanomas and control subjects were consecutively enrolled at the participating centers and tested for CDKN2A, CDK4, and microphthalmia-associated transcription factor. RESULTS: CDKN2A germline mutations were found in 19% of patients with MPM versus 4.4% of patients with single primary melanoma. In familial MPM cases the mutation rate varied from 36.6% to 58.8%, whereas in sporadic MPM cases it varied from 8.2% to 17.6% in patients with 2 and 3 or more melanomas, respectively. The microphthalmia-associated transcription factor E318K mutation accounted for 3% of MPM cases altogether. LIMITATIONS: The study was hospital based, not population based. Rare novel susceptibility genes were not tested. CONCLUSION: Italian patients who developed 2 melanomas, even in situ, should be referred for genetic counseling even in the absence of family history.

Dysplastic nevus: Fact and fiction.

The term "dysplastic nevus" (DN) implies that this nevus exists as a distinct and defined entity of potential detriment to its host. We examine the current data, which suggest that this entity exists as histologically and possibly genetically different from common nevus, with some overlapping features. Studies show that a melanoma associated with a nevus is just as likely to arise in a common nevus as in DN. Furthermore, there is no evidence that a histologically defined DN evolves into a melanoma or that the presence of 1 or more DN on an individual patient confers any increased melanoma risk. We suggest that the term "dysplastic nevus" be abandoned so that the focus can shift to confirmed and relevant indicators of melanoma risk, including high nevus counts and large nevus size.

Eradication of gastric cancer is now both possible and practical.

In 1994, Helicobacter pylori was declared a human carcinogen. Evidence has now accumulated to show that at least 95% of gastric cancers are etiologically related to H. pylori. An extensive literature regarding atrophic gastritis and its effects on acid secretion, gastric microflora, and its tight association with gastric cancer has been rediscovered, confirmed, and expanded. Methods to stratify cancer risk based on endoscopic and histologic findings or serologic testing of pepsinogen levels and H. pylori testing have been developed producing practical primary and secondary gastric cancer prevention strategies. H. pylori eradication halts progressive mucosal damage. Cure of the infection in those with non-atrophic gastritis will essentially prevent subsequent development of gastric cancer. For all, the age-related progression in cancer risk is halted and likely reduced as eradication reduces or eliminates mucosal inflammation and reverses or reduces H. pylori-associated molecular events such aberrant activation-induced cytidine deaminase expression, double strand DNA breaks, impaired DNA mismatch repair and aberrant DNA methylation. Those who have developed atrophic gastritis/gastric atrophy however retain some residual risk for gastric cancer which is proportional to the extent and severity of atrophic gastritis. Primary and secondary cancer prevention starts with H. pylori eradication and cancer risk stratification to identify those at higher risk who should also be considered for secondary cancer prevention programs. Japan has embarked on population-wide H. pylori eradication coupled with surveillance targeted to those with significant remaining risk. We anticipate that countries with high gastric cancer burdens will follow their lead. We provide specific recommendations on instituting practical primary and secondary gastric cancer prevention programs as well identifying research needed to make elimination of gastric cancer both efficient and cost effective.

Poor Prognosis in Low-Grade Endometrial Stromal Sarcoma With Cyclin-Dependent Kinase Inhibitor 2A Homozygous Deletion: A Case Study.

Low-grade endometrial stromal sarcoma (LGESS) typically has a favorable prognosis. Hormone therapy is considered the first choice of treatment for recurrent LGESS. In this report, we describe a case of recurrent LGESS where hormone therapy was ineffective, chemotherapy showed a partial response (PR), and pazopanib resulted in stable disease (SD). A 50-year-old patient with LGESS underwent a simple total hysterectomy and bilateral adnexectomy (pT1aN0M0, stage IA). Five years later, pelvic tumors and ascites were observed. Exploratory laparoscopy revealed bloody ascites, an 8 cm pelvic tumor, and extensive peritoneal dissemination. Nuclear atypia of the tumor cells was mild, pleomorphism and mitotic figures could not be confirmed, and necrosis was not observed. Immunostaining was positive for CD10 and estrogen receptor, negative for the BCL6 corepressor (BCOR), and showed a low Ki-67 index. Fluorescence in situ hybridization (FISH) examination of the tissue showed rearrangement of the JAZF zinc finger 1 (JAZF1) gene. Multigene panel testing revealed a homozygous deletion of cyclin-dependent kinase inhibitor 2A (CDKN2A). Accordingly, the patient was diagnosed with recurrent LGESS and was treated with an aromatase inhibitor, followed by medroxyprogesterone acetate; both were ineffective. The patient had a PR to chemotherapy (doxorubicin/ifosfamide) and SD to pazopanib. The patient died 1.5 years after recurrence. In conclusion, we present a case of LGESS with a poor prognosis where hormone therapy was ineffective, and chemotherapy and pazopanib were both partially effective. The poor prognosis may have been associated with the CDKN2A homozygous deletion.

Clinical significance and prospective mechanism of increased CDKN2A expression in small cell lung cancer.

BACKGROUND: Although it has been shown that cyclin dependent kinase inhibitor 2A (CDKN2A) plays a significant role in a number of malignancies, its clinicopathological value and function in small cell lung cancer (SCLC) is unclear and warrants additional research. METHODS: The clinical significance of CDKN2A expression in SCLC was examined by multiple methods, including comprehensive integration of mRNA level by high throughput data, Kaplan-Meier survival analysis for prognostic value, and validation of its protein expression using in-house immunohistochemistry. RESULTS: The expression of CDKN2A mRNA in 357 cases of SCLC was evidently higher than that in the control group (n = 525) combing the data from 20 research centers worldwide. The standardized mean difference (SMD) was 3.07, and the area under the curve (AUC) of summary receiver operating characteristic curve (sROC) was 0.97 for the overexpression of CDKN2A. ACC, COAD, KICH, KIRC, PCPG, PRAD, UCEC, UVM patients with higher CDKN2A expression had considerably worse overall survival rates than those with lower CDKN2A expression with the hazard ratio (HR) > 1. CONCLUSION: CDKN2A upregulation extensively enhances the carcinogenesis and progression of SCLC.

A Comprehensive Pan-Cancer Analysis Reveals Cyclin-Dependent Kinase Inhibitor 2A Gene as a Potential Diagnostic and Prognostic Biomarker in Colon Adenocarcinoma.

Introduction Cyclin-dependent kinase inhibitor 2A (CDKN2A) is a suppressor carcinogenic gene that is upregulated across various types of cancer including breast, liver, thyroid, and bile duct cancer due to its crucial role in cell cycle regulation and cell division. Nevertheless, it is mostly investigated at the genetic level, but it is still poorly studied on pan-cancer analysis as a biomarker and this study shows its significant potential diagnostic and prognostic characteristics. However, this study aims to investigate the role of CDKN2A as a diagnostic and prognostic biomarker across various types of cancer focusing primarily on colon adenocarcinoma (COAD). Methods We investigated CDKN2A gene expression in a pan-cancer analysis across different types of cancer to show its diagnostic potential characteristics by using various bioinformatic tools, including Tumor Immune Estimation Resource (TIMER) 2.0, Gene Expression Profiling Interactive Analysis (GEPIA), and University of Alabama at Birmingham Cancer Data Analysis Portal (UALCAN) database. TIMER was used to profile gene expression across 32 types of cancer composed of 10,000 RNA-seq samples obtained from the Cancer Genome Atlas (TCGA) and to analyze the tumor-infiltrating immune cells. In addition, GEPIA and UALCAN were further used to analyze gene expression, in terms of gene regulation, pathological stages, and clinical parameters, including gender, age, and race. Therefore, we used GEPIA, UALCAN, and Kaplan-Meier plotter particularly across adenocarcinoma to investigate CDKN2A prognosis by studying its high expression association with the patient's overall survival rate to show the tumor progression. Then, we looked into the genetic alteration of CDKN2A by using the cBio Cancer Genomics Portal (cBioPortal), including 10 pan-cancer studies. We concluded the analysis with gene validation by using a public cohort in Gene Expression Omnibus (GEO). Results CDKN2A showed a trend of upregulation in most cancers and it was significantly upregulated in five cancers, which were commonly identifiable in three databases, including breast invasive carcinoma (p < 0.001), kidney chromophobe (p < 0.001), kidney renal clear cell carcinoma (p < 0.001), kidney renal papillary cell carcinoma (p < 0.001), and COAD (p < 0.001). The upregulation was significantly different in association with pathogenic stages II and III (pr(>F) = 0.00234) which was identifiable significantly in COAD more than in other cancers. The gene showed a high upregulation in association with poor prognosis of patient survival in three cancers, including COAD (log-rank p = 0.011), mesothelioma (log-rank p = 5.9e-07), and liver hepatocellular carcinoma (log-rank p = 0.0045). Therefore, COAD was the only comprehensively analyzed tumor to show a diagnostic and prognostic potential characteristic during high upregulation of CDKN2A. Furthermore, CDKN2A displayed a rare mutation in the form of deep deletion (9%) and revealed an upregulation associated with CD4+ T cells (p = 0.0108), macrophage (p = 0.0073), and neutrophils (p = 0.0272) as immune cells infiltrating COAD.  Conclusion Our study demonstrates the pan-cancer relevance of CDKN2A and revealed a novelty in showing CDKN2A underscores its potential as a diagnostic prognostic biomarker in COAD since CDKN2A is mostly studied at a genetic level across COAD.

Cellular senescence molecules expression in type 2 diabetes mellitus: CDKN2A, CDKN2B, and lncRNA ANRIL.

AIMS: Cellular senescence in type 2 diabetes mellitus (T2DM) has received widespread attention. However, the cellular senescence molecules involved in T2DM are unclear. Furthermore, there are no consistent biomarkers for cellular senescence in T2DM. Therefore, this study aimed to identify cellular senescence molecules in T2DM and investigate their expression in peripheral blood mononuclear cells of individuals with T2DM. METHODS: Patients with T2DM (n = 40) and healthy controls (n = 40) were enrolled. We used different databases to identify cellular senescence molecules in T2DM and confirmed the obtained genes and lncRNA using real-time PCR. RESULTS: Bioinformatics analysis indicated that CDKN2A and CDKN2B genes, and long noncoding RNA ANRIL are the most effective cellular senescence molecules in T2DM. Furthermore, CDKN2A and ANRIL expression decreased in individuals with T2DM. CONCLUSIONS: Cellular senescence may have a protective effect against T2DM. In addition, the cellular senescence molecules CDKN2A and ANRIL may be potential biomarkers of cellular senescence in T2DM.

Cyclin-Dependent Kinase Inhibitor 2A is a Key Regulator of Cell Cycle Arrest and Senescence in Endothelial Colony-Forming Cells in Moyamoya Disease.

OBJECTIVE: Endothelial colony-forming cells (ECFCs) have been reported to play an important role in the pathogenesis of moyamoya disease (MMD). We have previously observed stagnant growth in MMD ECFCs with functional impairment of tubule formation. We aimed to verify the key regulators and related signaling pathways involved in the functional defects of MMD ECFCs. METHODS: ECFCs were cultured from peripheral blood mononuclear cells of healthy volunteers (normal) and MMD patients. Low-density lipoproteins uptake, flow cytometry, high content screening, senescence-associated ß-galactosidase, immunofluorescence, cell cycle, tubule formation, microarray, real-time quantitative polymerase chain reaction, small interfering RNA transfection, and western blot analyses were performed. RESULTS: The acquisition of cells that can be cultured for a long time with the characteristics of late ECFCs was significantly lower in the MMD patients than the normal. Importantly, the MMD ECFCs showed decreased cellular proliferation with G1 cell cycle arrest and cellular senescence compared to the normal ECFCs. A pathway enrichment analysis demonstrated that the cell cycle pathway was the major enriched pathway, which is consistent with the results of the functional analysis of ECFCs. Among the genes associated with the cell cycle, cyclin-dependent kinase inhibitor 2A (CDKN2A) showed the highest expression in MMD ECFCs. Knockdown of CDKN2A in MMD ECFCs enhanced proliferation by reducing G1 cell cycle arrest and inhibiting senescence through the regulation of CDK4 and phospho retinoblastoma protein. CONCLUSION: Our study suggests that CDKN2A plays an important role in the growth retardation of MMD ECFCs by inducing cell cycle arrest and senescence.

Association of CDKN2A/B mutations, PD-1, and PD-L1 with the risk of acute lymphoblastic leukemia in children.

PURPOSE: Currently, the significance of CDKN2A/B mutations in the pathogenesis and prognosis of acute lymphoblastic leukemia (ALL) is inconclusive. In this study, we analyzed the genetic and clinical features of children with CDKN2A/B mutations in ALL. In addition, we evaluated the expression and significance of programmed cell death protein 1 (PD-1) and programmed cell death ligand 1 (PD-L1) in serum and explored their role in the susceptibility of childhood ALL. METHODS: We sequenced CDKN2A/B in the peripheral blood of 120 children with ALL and 100 healthy children with physical examination. The levels of CD4+ T, CD8+ T, and NK cells were measured by flow cytometry (FCM). Furthermore, the expression of PD-1 and PD-L1 was detected by ELISA. RESULTS: We found 32 cases of CDKN2A rs3088440 and 11 of CDKN2B rs2069426 in 120 ALL children. Children with ALL in the CDKN2A rs3088440 were more likely to have hepatosplenomegaly (P = 0.019) and high risk (P = 0.014) than the wild group. In contrast, CDKN2B rs2069426 was more likely to develop lymph node metastasis (P = 0.017). The level of PD-L1 in the serum of ALL children was significantly higher than that of the control group, and there was no significant difference in PD-1 (P < 0.001). Additionally, children with CDKN2A rs3088440 had reduced CD8+ T cell counts than the wild group (P = 0.039). CONCLUSION: CDKN2A rs3088440 and CDKN2B rs2069426 may be related to the occurrence and development of ALL in Chinese children. Additionally, PD-1/PD-L1 may be involved in the immune escape process of ALL, which is expected to become a new target for the treatment of the disease.

Next-generation sequencing identifies CDKN2A alterations as prognostic biomarkers in recurrent or metastatic head and neck squamous cell carcinoma predominantly receiving immune checkpoint inhibitors.

Background: This study aimed to identify potential biomarkers in patients with recurrent or metastatic head and neck squamous cell carcinoma (HNSCC) and further probe the prognostic implications of CDKN2A mutations, particularly within a subset receiving immunotherapy. Methods: In this retrospective single-center study, we evaluated the next-generation sequencing (NGS) data from Foundation Medicine (FM) for patients with recurrent or metastatic HNSCC between January 1, 2019, and December 31, 2021. Patients were stratified based on CDKN2A loss-of-function (LOF) versus wild-type (WT) categorizations, with a focused subgroup analysis on those administered immunotherapy. Results: The study encompassed 77 patients, of which 62 had undergone immunotherapy. The median duration of follow-up was 22.6 months. For the CDKN2A LOF group, the median overall survival (OS) was 16.5 months, contrasted with 30.0 months in the CDKN2A WT group (P=0.014). Notably, female gender (hazard ratio [HR]=4.526, 95% confidence interval [CI]: 1.934-10.180, P=0.0003) and CDKN2A LOF (HR=2.311, 95% CI: 1.156-4.748, P=0.019) emerged as independent risk factors for mortality in patients with recurrent or metastatic HNSCC. Within the immunotherapy subset, the median OS was 11.7 months for the CDKN2A LOF group, and 22.5 months for the CDKN2A WT group (P=0.017). Further, the female gender (HR=4.022, 95% CI: 1.417-10.710, P=0.006), CDKN2A LOF (HR=4.389, 95% CI: 1.782-11.460, P=0.002), and a combined positive score below 1 (HR=17.20, 95% CI: 4.134-79.550, P<0.0001) were identified as significant predictors of mortality among patients with recurrent or metastatic HNSCC receiving immunotherapy. Conclusion: Alterations manifesting as LOF in the CDKN2A gene stand as robust indicators of unfavorable survival outcomes in HNSCC patients, including the subset that underwent immunotherapy.