TWIST 1 and 2 expression restricted to stromal cells. Inverse correlation between TWIST1 protein expression and methylation (Cohort 1) suggests hypermethylation as a mechanism of TWIST1 regulation. TWIST 1 and 2 protein expression significantly correlated with low- and high-grade budding phenotype. LCM of high-grade tumor-budding case with positive TWIST1/2 stroma and no methylation, inverse pattern in low-grade tumor-budding case. TWIST1 (Cohort 2) associated with adverse tumor features and independent prognostic factor. The gene discussed is TWIST1; the disease is neoplasm.