Pathological complete response (pCR) after neoadjuvant systemic treatment (NST) is an important prognostic factor in HER2-positive breast cancer. The majority of HER2-positive breast cancers are amplified at the HER2 gene locus, several genes are co-amplified with HER2, and a subset of them are co-expressed. The STARD3 gene belongs to the HER2 amplicon, and its role as a predictive marker was never addressed. The objective of this study was to investigate the predictive value of STARD3 protein expression on NST pathological response in HER2-positive breast cancer. In addition, we studied the prognostic value of this marker. Methods. We conducted a retrospective study between 2007 and 2020 on 112 patients with non-metastatic HER2-positive breast cancer treated by NST and then by surgery. We developed an immunohistochemistry assay for STARD3 expression and subcellular localization and determined a score for STARD3-positivity. As STARD3 is an endosomal protein, its expression was considered positive if the intracellular signal pattern was granular. Results: In this series, pCR was achieved in half of the patients. STARD3 was positive in 86.6% of cases and was significantly associated with pCR in univariate analysis (p = 0.013) and after adjustment on other known pathological parameters (p = 0.044). Performances on pCR prediction showed high sensitivity (96%) and negative predictive value (87%), while specificity was 23% and positive predictive value was 56%. Overall, specific, relapse-free, and distant metastasis-free survivals were similar among STARD3 positive and negative groups, independently of other prognosis factors. Conclusion: NST is an opportunity for HER2-positive cancers. In this series of over a hundred HER2-positive and non-metastatic patients, a STARD3-negative score was associated with the absence of pathological complete response. This study suggests that determining STARD3 overexpression status on initial biopsies of HER2-positive tumors is an added value for the management of a subset of patients with high probability of no pathological response.