Mutation analysis of the HIF-1α oxygen-dependent degradation domain in invasive breast cancer

Abstract

Hypoxia inducible factor-1 (HIF-1) is an important transcription factor that stimulates tumor growth and metastases via several pathways. Activation of HIF-1 depends on the presence of its a-subunit. Hypoxia increases HIF-1α levels by inhibiting prolyl-hydroxylasedmediated hydroxylation and thereby preventing proteosome degradation. Various other mechanisms might also contribute to HIF-1α expression, such as mutation of the oxygen dependent degradation domain (ODD), which prevents binding of prolyl-hydroxylases. Therefore, the presence of ODD mutations was evaluated as a possible explanation for diffuse HIF-1α protein expression often seen in invasive breast cancer. From a group of 200 primary breast cancers, 24 strong diffusely HIF-1α-positive tumor samples were identified with HIF-1α immunohistochemistry. DNA from these tumors was extracted from microdissected paraffin material and, after nested polymerase chain reaction, sequence analysis was performed to detect hif-1α ODD mutations. Additionally, five perinecrotically HIF-1α-positive breast cancers were analyzed as controls. All 24 diffuse and perinecrotic HIF-1α-positive breast cancers showed wild-type DNA sequences in the ODD domain. No mutations seem to occur in the ODD of hif-1α in HIF-1α overexpressing invasive breast cancer, which rules ODD mutations out as a possible explanation for the diffuse HIF-1α expression pattern often seen in this cancer.