DNA methylation data for 489 high stage, high grade serous ovarian tumors and eight normal fallopian tube samples was obtained from http://tcga.cancer.gov/dataportal/ . In addition, buffy coat samples from two female individuals were obtained. All data were generated with Illumina Infinium HumanMethylation 27 arrays, which interrogate 27,578 CpG sites located in proximity to the transcription start sites of 14,475 consensus coding sequencing in the NCBI Database (Genome Build 36). The level of DNA methylation at each probe was summarized with beta values ranging from 0 (unmethylated) to 1 (methylated) 58 (link).
The leukocyte methylation signature was derived as follows. Each probe was ranked by the difference in mean beta value in buffy coat and fallopian tube samples. We retained the 100 probes with the largest positive difference and the 100 with the largest negative difference between mean DNA methylation in normal fallopian tube tissues and peripheral blood leukocytes, designated BC and FT, (buffy coat and fallopian tube enriched, respectively). Let Tik denote the beta value for probe k in tumor sample i. Let Bk denote the average beta value of buffy coat samples for each probe. Let Tk denote the minimum observed beta value across all tumor samples for the BC probes and the maximum for the FT probes. Denote by fB the fraction of buffy coat components in the sample, then we have the following equation for each probe: Tik = BkfB + Tk(lāfB). Solving this equation for fB gives: fB = (Tik ā Tk)/(Bk ā Tk). The values of fB for each of the 200 probes in the signature were calculated and a kernel density estimate was obtained. The leukocyte signature was then calculated as the mode of this density estimate.
The leukocyte methylation signature was derived as follows. Each probe was ranked by the difference in mean beta value in buffy coat and fallopian tube samples. We retained the 100 probes with the largest positive difference and the 100 with the largest negative difference between mean DNA methylation in normal fallopian tube tissues and peripheral blood leukocytes, designated BC and FT, (buffy coat and fallopian tube enriched, respectively). Let Tik denote the beta value for probe k in tumor sample i. Let Bk denote the average beta value of buffy coat samples for each probe. Let Tk denote the minimum observed beta value across all tumor samples for the BC probes and the maximum for the FT probes. Denote by fB the fraction of buffy coat components in the sample, then we have the following equation for each probe: Tik = BkfB + Tk(lāfB). Solving this equation for fB gives: fB = (Tik ā Tk)/(Bk ā Tk). The values of fB for each of the 200 probes in the signature were calculated and a kernel density estimate was obtained. The leukocyte signature was then calculated as the mode of this density estimate.
