This paper describes an associated analysis method of DNA methylation for the detection of cancer using an optically amplifying cationic conjugated polymer (CCP, poly{(1,4-phenylene)-2,7-[9,9-bis(6'-N,N,N-trimethyl ammonium)-hexyl fluorene] dibromide)}. Genomic DNA is digested by methylation-sensitive restriction endonuclease, followed by PCR amplification to incorporate fluorescein-labeled dNTP. Only methylated DNA can be amplified by PCR, and the methylation level is detected through fluorescence resonance energy transfer (FRET) between CCP and fluorescein that is incorporated into the PCR product. The methylation levels of RASSF1A, OPCML, and HOXA9 promoters of 35 ovarian cancer samples and 11 normal samples were assayed. In accordance with the degree of methylation levels, they are clustered to three sections and assigned a value. Through an associated analysis, we acquired a threshold for cancer detection with a sensitivity of 85.7%. The assay takes about 20 h to obtain the detection results and shows great potential as a useful tool for diagnostic and screening of cancer.

The therapeutic effect of Fe2O3 nanoparticles combined with magnetic fluid hyperthermia (MFH) on human hepatocarcinoma SMMC-7721 cells in vitro and xenograft liver cancer in nude mice is studied. We examined growth and apoptosis of SMMC-7721 cells treated with MFH containing Fe2O3 nanoparticles at various concentrations (2, 4, 6, and 8 g/liter) for 30-60 min by using MTT, flow cytometry (FCM), and transmission electron microscopy (TEM). We also observed weight and volume inhibitory rates of the tumors of SMMC-7721-bearing nude mice by using animal experiments. The results showed that Fe2O3 nanoparticles combined with MFH could significantly inhibit the proliferation and increase the ratio of apoptosis of SMMC-7721 cells, and the effect was dose-dependent. The inhibitory rate was 26.5%, 33.53%, 54.4%, and 81.2%, respectively, and the apoptosis rate was 30.26%, 38.65%, 50.28%, and 69.33%, respectively. Animal experiments showed that tumors became small. The weight inhibitory ratio was 42.10%, 66.34%, 78.5%, and 91.46%, and the volume inhibitory ratio was 58.77%, 80.44%, 93.40%, and 98.30%, respectively. Compared with the control and experimental groups, each group had statistically significant difference (p < 0.05). So, Fe2O3 nanoparticles combined with MFH could inhibit the proliferation and induce apoptosis of SMMC-7721 cells and also has a significant inhibitory effect on the weight and volume of xenograft liver cancer. However, the mechanism remains to be further investigated.

Medroxyprogesterone (MPA) is used for the conservative treatment of endometrial cancer. Unfortunately, progesterone resistance seriously affects its therapeutic effect. The purpose of the current study was to investigate the influence of deletion of AT-rich interactive domain 1A (ARID1A) in progesterone resistance in Ishikawa cells. Ablation of ARID1A was conducted through the CRISPR/Cas9 technology. Acquired progesterone-resistant Ishikawa (Ishikawa-PR) cells were generated by chronic exposure of Ishikawa cells to MPA. The sensitivity of the parental Ishikawa, Ishikawa-PR, and ARID1A-deficient cells to MPA and/or LY294002 was determined using the Cell Counting Kit-8 (CCK-8) assay and flow cytometry analysis. In addition, Western blot analysis and reverse transcription-polymerase chain reaction was performed to evaluate the mRNA and protein expression levels of ARID1A, progesterone receptor B (PRB), and P-AKT. Both Ishikawa-PR and ARID1A knockout cells showed insensitivity to MPA, downregulation of PRB, and hyperphosphorylation of AKT compared to the parental Ishikawa cells. Pretreatment with LY294002 significantly enhanced the ability of MPA to suppress proliferation and to induce apoptosis in the parental and Ishikawa-PR cells via the inhibition of AKT activation and upregulation of PRB transcriptional activity. However, the PRB transcriptional activity and insensitivity to MPA were irreversible by LY294002 in ARID1A-deficient cells. Ablation of ARID1A is associated with low PRB expression, which serves an important role in primary progesterone resistance. Akt inhibition cannot rescue PRB or sensitize to MPA in ARID1A knockout cells. These findings suggest that ARID1A may act as a reliable biomarker to predict the response for the combination of AKT inhibitor and MPA treatment.

ABSTRACT. DNA methylation plays an important role in carcinogenesis and cancer development. In this study, we examined gene methylation in epithelial ovarian cancer (EOC) using cationic conjugated polymer (CCP)-based fluorescence resonance energy transfer (FRET) to evaluate the application of cumulative methylation alternations of genes associated with cancer antigen 125 for early cancer diagnosis. The methylation status of 3 genes (Ras association domain family 1 isoform A, RASSF1A; opioid-binding protein/cell adhesion molecule, OPCML; homeobox A9, HOXA9) were examined and compared in 35 EOC samples and 11 normal ovarian tissue samples using CCP-based FRET. Gene methylation levels were clustered into 3 sections and assigned a value; values for the 3 genes were summed. Although methylation of the OPCML gene was significantly associated with stage, histological types, grade, and ascites and that of RASSF1A and HOXA9 was not, the sum for the 3 genes was significantly associated with stage and ascites. The sum showed higher sensitivity (85.7%) and specificity (100%) for discriminating EOC from normal ovarian tissues than did the methylation status of RASSF1A, OPCML, and HOXA9

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