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Aims The goal of this study is to identify the pathological findings and expression of metabolic markers (GLUT-1, PDK-1 and PGC1α) in the tumour microenvironment of both primary and chemoreduced retinoblastoma (Rb) and to correlate with clinicopathological parameters and patient outcome.Methods 81 prospective cases were included, in which 53 cases underwent primary enucleation and 28 cases received chemotherapy before enucleation. Immunohistochemistry, qRT-PCR and western blotting were performed to evaluate the expression pattern of metabolic markers in primary and chemoreduced Rb.Results Tumour microenvironment and histopathological findings were different for both primary and chemoreduced Rb. Increased immunohistochemical expression of GLUT-1, PDK-1 and PGC1α was found in primary Rb as compared with chemoreduced Rb. mRNA expression was also found to be upregulated in primary Rb compared with chemoreduced. On univariate analysis, the presence of more than one histopathological high-risk factor (HRFs>1) and PDK-1 immunoexpression was statistically significant with overall survival. On prognostication in primary and chemoreduced cases with hypoxia, we found increased HR in cases with retrolaminar ON invasion, presence of more than one HRF, and presence of PDK-1 and PGC-1α immunoexpression.Conclusions This is the first of its kind study predicting a relevant role of the metabolic markers in primary and chemoreduced Rb with prognostic significance. Differential expression of these markers in both groups of Rb is a novel finding and might be an interesting and beneficial target for the management of Rb patients.