Abstract Malignancy of the prostate remains a leading cause of cancer-related deaths among American men. Chemoprevention using safe and inexpensive phytochemicals from edible or medicinal plants is clinically attractive for reducing both morbidity and mortality from prostate cancer. Sulforaphane (SFN), which is the principal bioactive phytochemical in broccoli sprout extract (BSE), has emerged as a particularly potent chemopreventive agent based on its ability to target multiple mechanisms to control carcinogenesis. The overall goal of the parent NCI-funded R01 award (R01 CA225716-02) was to determine the feasibility of inhibiting fatty acid metabolism for chemoprevention of prostate cancer using SFN and BSE. This revision application will incorporate the IMAT-funded super-resolution microscopy technology that can detect the cumulative changes in disrupted nanoscale chromatin folding at different epigenetic states during carcinogenesis to (a) accurately quantify the reduction of cancer risk from multiple molecular pathways and (b) further understand the alteration of higher-order chromatin folding under the influences of metabolism in the context of chemopreventive intervention for prostate cancer. This revision project will incorporate the IMAT-funded super-resolution microscopy as follows: Specific Aim 1: Determine whether chemoprevention of prostate cancer by SFN and BroccoMax® (which is being used as a source of SFN in the clinical trial in specific aim 3 of the parent R01) in Hi-Myc transgenic mice is associated with alterations in nanoscale chromatin folding. Specific Aim 2: Determine whether higher-order chromatin folding is altered in PTEN-deletion mouse model of prostate cancer, and whether this phenotype is reversed following dietary administration of SFN and BroccoMax®. Specific Aim 3: Validate the human relevance of disrupted chromatin folding and evaluate its feasibility to risk-stratify patients with prostate cancer. Impact: This revision project will open a new avenue for cancer risk stratification and facilitate the evaluation of chemopreventive intervention and establish the relationship between disrupted nanoscale chromatin folding that fuels carcinogenesis and metabolism in the context of chemopreventive intervention.