Matrix metalloproteinase-9 (MMP-9) is related to tumour development and progression in colorectal cancer (CRC) and its utility as biomarker has been suggested. The aim of our study was to measure serum MMP-9 in asymptomatic first-degree relatives of CRC patients, and to analyse its diagnostic accuracy for the detection of advanced neoplasia (AN: advanced adenomas and CRC). Additionally, we compared its diagnostic capability with the most used non-invasive faecal immunochemical test (FIT). Serum MMP-9 was quantified by ELISA in 516 asymptomatic individuals that underwent a colonoscopy and a FIT. MMP-9 levels were significantly related to age and gender and therefore the concentration was corrected by these confounders. Corrected MMP-9 (cMMP-9) levels were higher in individuals with advanced adenomas (AA; p-value = 0.029) and AN (p-value = 0.056) compared to individuals with no neoplasia. Moreover, elevated cMMP-9 concentration was associated with more severe characteristics of adenomas (number of lesions, size and histology). Nevertheless, the diagnostic accuracy of cMMP-9 was considerably lower than that of FIT for identifying AA (22.64% vs. 47.17% sensitivity, 90% specificity) or AN (19.30% vs. 52.63% sensitivity, 90% specificity). According to our results, serum MMP-9 cannot be considered of utility for the diagnosis of AN in CRC family-risk population screening.