Papillary renal cell carcinoma (pRCC) is characterized with underlying genetic disorders and the role enolase 2 (ENO2) in ccRCC is unknown. An in silico exploratory analysis using multiple public genetic datasets was used to establish association between ENO2 expression and clinicopathological parameters. Associations of interest were validated using 49 pRCC samples using immunohistochemistry. In vitro and in vivo assays were carried out to validate findings in tissue. ENO2 was overexpressed and prognostic in pRCC. ENO2 expression was significantly higher in younger patients and in CpG island methylator phenotype subtype. ENO2-overexpressed cases showed significant enrichment in glycolysis. Overexpression of ENO2 significantly increased proliferation and silencing of ENO2 significantly inhibited growth of ACHN cells. Glycolytic genes HK1, HK 2, and lactate dehydrogenase A were decreased when ENO2 was silenced in ACHN. Glycolytic inhibitor TT-232 showed minimal inhibitory effect on ACHN cells yet showed synergistic effect in the presence of ENO2 silencing. ENO2 significantly increased and decreased extracellular glucose, respectively in ACHN cells. Xenograft mouse model showed ENO2 silencing and TT-232 combination treatment showed synergistic effect in ACHN tumors. ENO2 is associated with worsened prognosis in pRCC and is related to glycolysis. ENO2-targeted therapy can be of therapeutic potential.