DNA diagnosis of peritoneal fluid cytology test by CDO1 promoter DNA hypermethylation in gastric cancer.

BACKGROUND: Minimal residual disease of the peritoneum is challenging for early cancer detection in gastric cancer (GC). Utility of PCR amplification of cancer-derived DNA has been considered feasible due to its molecular stability, however such markers have never been available in GC clinics. We recently discovered cancer-specific methylation of CDO1 gene in GC, and investigated the clinical potential to detect the minimal residual disease. METHODS: One hundred and two GC patients were investigated for peritoneal fluid cytology test (CY), and detection level of the promoter DNA methylation of CDO1 gene was assessed by quantitative methylation specific PCR (Q-MSP) in the sediments (DNA CY). RESULTS: (1) CY1 was pathologically confirmed in 8 cases, while DNA CY1 was detected in 18 cases. All 8 CY1 were DNA CY1. (2) DNA CY1 was recognized in 14.3, 25.0, 20.0, and 42.9%, in macroscopic Type II, small type III, large type III, and type IV, respectively, while it was not recognized in Type 0/I/V. (3) DNA CY1 was prognostic relevance in gastric cancer (p = 0.0004), and its significance was robust among Type III/IV gastric cancer (p = 0.006 for overall survival and p = 0.0006 for peritoneal recurrence free survival). (4) The peritoneal recurrence was hardly seen in GC patients with potent perioperative chemotherapy among those with DNA CY1. CONCLUSIONS: DNA CY1 detected by Q-MSP for CDO1 gene promoter DNA methylation has a great potential to detect minimal residual disease of the peritoneum in GC clinics as a novel DNA marker.

Promoter DNA methylation of CDO1 gene and its clinical significance in esophageal squamous cell carcinoma.

We have demonstrated that CDO1 methylation is frequently found in various cancers, including esophageal squamous cell carcinoma (ESCC), but its clinical relevance has remained elusive. CDO1 methylation was investigated in 169 ESCC patients who underwent esophagectomy between 1996 and 2007. CDO1 methylation was assessed by Q-MSP (quantitative methylation specific PCR), and its clinical significance, including its relationship to prognosis, was analyzed. (i) The median TaqMeth value of CDO1 methylation was 9.4, ranging from 0 to 279.5. CDO1 methylation was significantly different between cStage I and cStage II/III (P = 0.02). (ii) On the log-rank plot, the optimal cut-off value was determined to be 8.9; ESCC patients with high CDO1 methylation showed a significantly worse prognosis than those with low CDO1 methylation (P = 0.02). (iii) A multivariate Cox proportional hazards model identified only CDO1 hypermethylation as an independent prognostic factor (HR 2.00, CI 1.09-3.78, P = 0.03). (iv) CDO1 hypermethylation stratified ESCC patients' prognosis in cStage II/III for both neoadjuvant chemo(radio)therapy (NAC)-positive and NAC-negative cases. Moreover, the CDO1 methylation level was significantly lower in cases with Grade 2/3 than in those with Grade 0/1 (P = 0.02) among cStage II/III ESCC patients with NAC. Promoter DNA hypermethylation of CDO1 could be an independent prognostic factor in ESCC; it may also reflect NAC eradication of tumor cells in the primary tumors.

Hypermethylated CDO1 and CELF4 in cytological specimens as triage strategy biomarkers in endometrial malignant lesions.

Objective: Developing a non-invasive and reliable triage test for endometrial malignant lesions is an important goal, as it could help to reduce the number of invasive diagnostic procedures required and improve patient survival. We aimed to estimate the diagnostic value of DNA methylation levels in cervical cytological samples of endometrial cancer (EC) and endometrial atypical hyperplasia (AH). Methods: A total of 607 women who had indications for endometrial biopsy in the Department of Obstetrics and Gynecology of Cangzhou Central Hospital from October 2022 to April 2023 were enrolled in this study. The cervical exfoliated cells were collected for gene methylation before endometrial biopsy. Clinical information, tumor biomarkers, and endometrial thickness (ET) of transvaginal ultrasonography (TVS) were also collected. With endometrial histopathology as the gold standard, multivariate unconditional logistic regression was applied to analyze the risk factors of endometrial malignant lesions. The role of cysteine dioxygenase type 1 (CDO1) and CUGBP Elav-like family member 4 (CELF4) gene methylation as a triage strategy biomarker in endometrial malignant lesions was specifically explored. Results: Multivariate logistic regression analysis showed that premenopausal ET ≥ 11 mm or postmenopausal ET ≥ 5 mm, CDO1 ΔCt ≤ 8.4, or CELF4 ΔCt ≤ 8.8 were the risk factors for AH and EC, with odds ratios (ORs) (95%CI) of 5.03 (1.83-13.82) and 6.92 (1.10-43.44), respectively (p-values < 0.05). The sensitivity and specificity of CDO1/CELF4 dual-gene methylation assay for AH and EC reached 84.9% (95%CI: 75.3%-94.5%) and 86.6% (95%CI: 83.8%-89.5%), respectively. ET combined with DNA methylation detection further improved the specificity to (94.9%, 95%CI: 93.1%-96.8%). Conclusion: The accuracy of cervical cytology DNA methylation is superior to that of other clinical indicators in the non-invasive examination of endometrial malignant lesions. DNA methylation combined with TVS can further improve the specificity and is a promising biomarker triage strategy in women with suspected endometrial lesions.

Hypermethylated CDO1 and ZNF454 in Cytological Specimens as Screening Biomarkers for Endometrial Cancer.

We aimed to estimate the diagnostic value of DNA methylation levels in cytological samples of endometrial cancer (EC) and atypical hyperplasia (AH). Two hypermethylated genes, namely, cysteine dioxygenase type 1 (CDO1) and zinc finger protein 454 (ZNF454), in patients with EC were identified from The Cancer Genome Atlas database. In 103 endometrial histological specimens (the training set), the methylation levels of candidate genes were verified by quantitative methylation-specific polymerase chain reaction (qMSP). The methylation levels of another 120 cytological specimens (the testing set) were evaluated. Sensitivity (Se), specificity (Sp), accuracy, and area under the curve (AUC) were determined, with diagnosis verified by histopathological results. CDO1 and ZNF454 verified hypermethylation in histological specimens of patients with EC and AH compared with those with benign and normal endometrium (P < 0.001). In cytological specimens, hypermethylated CDO1 showed 86.36% Se and 90.79% Sp with the cutoff value of 6.0 to distinguish between malignant and benign groups; ZNF454 showed 79.55% Se and 93.42% Sp with the cutoff value of 7.1. When the two genes were combined, Se increased to 90.91% and Sp was 86.84%. AUC reached 0.931 (95% CI: 0.885-0.976). The diagnostic accuracy with cytology had no significant difference with endometrial tissue (P = 0.847 for CDO1, P = 0.108 for ZNF454, and P = 0.665 for their combination). Hypermethylated CDO1 and ZNF454 in endometrial cytology showed high Se, Sp, and AUC to detect EC and AH. Methylation analysis of endometrial cytology is promising biomarker for the screening of EC and AH.