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RFWD3 Reprograms Nucleotide Metabolism Through PHGDH to Induce Chemoresistance In Osteosarcoma
Wenchao Zhang1,2, Chi Yin1,2, Lin Qi1,2
1Department of Orthopedics, The Second Xiangya Hospital, Central South University, Changsha, 410011, China.
Abstract:
Chemoresistance represents a major challenge for osteosarcoma treatment. Despite the improved knowledge of cancer biology, the core determinants of cisplatin (DDP) resistance in osteosarcoma remain unclear and deserve further exploration. Here, RFWD3 is identified as a key regulator of DDP sensitivity in osteosarcoma using a genome-wide CRISPR screen. It is demonstrated that RFWD3 is overexpressed in post-chemotherapy osteosarcoma tissues compared to pre-chemotherapy tissues. Knocking out RFWD3 increased the sensitivity of osteosarcoma cells to DDP treatment. Mechanistically, RFWD3 bound to and ubiquitinated PHGDH at the Lys137 residue, promoting its degradation and conserving cellular oxidized nicotinamide adenine dinucleotide (NAD+). The resulting surplus of NAD+ enhanced the TCA cycle, leading to increased production of aspartic acid and glutamic acid for de novo nucleotide biosynthesis. In addition, virtual screening techniques are employed to identify Lomitapide as a specific inhibitor of the RFWD3-PHGDH interaction, capable of disrupting the binding between RFWD3 and PHGDH. It is found that Lomitapide exhibits a significant synergistic anti-osteosarcoma effect when combined with DDP. In conclusion, a specific role of RFWD3 in regulating nucleotide metabolism is revealed and comprised of targetable candidates for overcoming chemoresistance in osteosarcoma.
Insights
RFWD3 overexpression drives cisplatin resistance in osteosarcoma by degrading PHGDH, impacting nucleotide synthesis. Inhibiting RFWD3 or its interaction with PHGDH may overcome chemoresistance.
Area of Science:
- Oncology
- Molecular Biology
- Biochemistry
Background:
- Chemoresistance to cisplatin (DDP) is a significant obstacle in osteosarcoma treatment.
- The underlying mechanisms of DDP resistance in osteosarcoma require further investigation.
Purpose of the Study:
- To identify key regulators of DDP sensitivity in osteosarcoma.
- To elucidate the molecular mechanisms by which RFWD3 influences DDP resistance.
- To explore potential therapeutic strategies targeting RFWD3.
Main Methods:
- Genome-wide CRISPR screening to identify DDP sensitivity regulators.
- Western blotting and ubiquitination assays to study protein interactions and degradation.
- Metabolic assays to assess nucleotide biosynthesis and TCA cycle activity.
- Virtual screening and drug combination studies.
Main Results:
- RFWD3 was identified as a crucial regulator of DDP sensitivity and is overexpressed in post-chemotherapy osteosarcoma tissues.
- RFWD3 knockdown sensitized osteosarcoma cells to DDP treatment.
- RFWD3 promotes PHGDH degradation, leading to increased NAD+ levels, enhanced TCA cycle, and nucleotide biosynthesis.
- Lomitapide, an RFWD3-PHGDH interaction inhibitor, showed synergistic effects with DDP against osteosarcoma.
Conclusions:
- RFWD3 plays a critical role in regulating nucleotide metabolism and chemoresistance in osteosarcoma.
- Targeting the RFWD3-PHGDH interaction presents a promising strategy to overcome DDP resistance in osteosarcoma.
