Heme Binding Biguanides Target Cytochrome P450-Dependent Cancer Cell Mitochondria

Area of Science:

• Biochemistry
• Cancer Biology
• Pharmacology

Background:

• The role of cancer cell-intrinsic cytochrome P450 (CYP) monooxygenases in tumor progression remains largely unexplored.
• CYP3A4 was identified in breast cancer mitochondria, synthesizing epoxyeicosatrienoic acids (EETs) from arachidonic acid (AA).

Purpose of the Study:

• To elucidate the mechanisms of CYP3A4 in breast cancer.
• To investigate the potential of biguanides, like metformin, as inhibitors of CYP3A4 in cancer therapy.

Main Methods:

• CYP3A4 knockdown experiments.
• Mitochondrial respiration assays.
• AMP-activated protein kinase (AMPKα) activity measurements.
• Autophagy assessment.
• Co-crystal structure determination of metformin bound to CYP3A4.
• Structure-based design of novel biguanide inhibitors.
• In vivo studies using ER+ mammary tumor models.

Main Results:

• CYP3A4 synthesized AA-derived EETs in breast cancer mitochondria, enhancing respiration and inhibiting AMPKα.
• CYP3A4 knockdown activated AMPKα, induced autophagy, and prevented tumor formation.
• Metformin inhibited CYP3A4-mediated EET biosynthesis and depleted intracellular EETs.
• Metformin binds to the CYP3A4 active-site heme, identifying it as a biguanide target.
• A novel biguanide, N1-hexyl-N5-benzyl-biguanide (HBB), showed higher affinity for CYP3A4 and potent inhibition of its epoxygenase activity.
• HBB suppressed established ER+ mammary tumor growth and intratumoral mTOR signaling.

Conclusions:

• CYP3A4-mediated mitochondrial eicosanoid synthesis is a key driver of breast cancer progression.
• Biguanides, including metformin and HBB, effectively inhibit CYP3A4's pro-tumorigenic activity.
• Targeting CYP3A4 offers a novel therapeutic strategy for breast cancer, bridging eicosanoid biology and biguanide action.