DNA hypomethylation restrains early antitumor immunity in prostate cancer [Nanopore]

Cancer is characterized by hypomethylation-associated silencing of large chromatin domains, whose contribution to tumorigenesis is uncertain. Through high-resolution single cell DNA methylation sequencing in prostate cancer, we identify 40 core hypomethylation domains, consistently hypomethylated across tumor cells and arising at early stages of malignancy. Transcriptionally silenced genes within these domains are enriched for immune-related genes; nested among repressive domains are small loci with preserved methylation, encoding cell proliferation genes that escape silencing. Prominent among hypomethylation-silenced genes is a gene cluster harboring all five CD1 genes that present lipid antigens to NKT cells, and four IFI16-related interferon-inducible genes implicated in innate immunity. Re-expression of CD1 or IFI16 murine orthologs in immunocompetent mice abrogates prostate tumorigenesis, accompanied by activation of anti-tumor immunity. Thus, early epigenetic changes in cancer may shape tumorigenesis, targeting co-located genes within defined chromosomal loci. Hypomethylation domains are detectable in blood specimens enriched for circulating tumor cells. CTCs are shed into the blood by invasive localized prostate cancers long before they establish metastases, raising the possibility that they may provide an orthogonal assay for early cancer detection. Given the specificity of DNA hypomethylation domains in prostate cancer cells and their large genomic size, we reasoned that they may provide high sensitivity and quantitative signal for cancer detection, following CTC enrichment in blood specimens. Oxford Nanopore long-read native sequencing typically produces sequencing reads up to 100 kb, and directly identifies methylated CpG residues, without requiring bisulfite conversion in library preparation. In contrast to the short Illumina sequencing reads (usually harboring <5 CpG sites per read), mathematical modeling indicates that the long reads generated by Nanopore sequencing would empower detection with significantly higher precision for rare signal. We therefore processed 10 ml blood specimens from patients with either localized or metastatic prostate cancer, using microfluidic enrichment to deplete leukocytes (10^4-fold depletion), but without further CTC purification or individual CTC micromanipulation.