Matrix metalloproteinases (MMPs) play an important role in breast cancer tumor invasion and progression. MMP-9 is a member of the MMP family and is also known as Gelatinase B or type IV collagenases (92 kDa) and possesses proteolytic activity against type IV collagen, a major component of the basement membrane. Our study aims to examine the association of Gelatinase B (-1562C > T) promoter polymorphism with breast cancer invasion and progression. The study involves 200 breast cancer patients and age-matched 191 healthy controls. The SNP-1562C > T (rs3918242) in MMP-9 promoter region was examined by allele-specific polymerase chain reaction and gel electrophoresis. The genotypes were determined and compared between patients and controls, and the influence of the polymorphism on clinicopathological data was analyzed. The T allele of the -1562C > T MMP-9 polymorphism was detected more frequently in breast cancer patients than controls (p < 0.001). Our results suggest the clinical importance of MMP-9 gene polymorphism (-1562C > T) in breast cancer patients. The study may also help in identifying individuals at risk of developing breast cancer.