A Combination of Oxidative Stress-Induced DNA Damage and NAD⁺ Synthesis Inhibition Induces Synthetic Lethality in A549 Cells

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Nicotinamide adenine dinucleotide (NAD⁺) is an important coenzyme involved in various redox reactions. Further, NAD⁺ is a substrate for poly-ADP-ribose polymerase (PARP). DNA single-strand break (SSB) induces PARP auto-ADP-ribosylation and recruits DNA repair complex. Oxidative stress, due to hydrogen peroxide (H₂O₂), generally induces SSB DNA damage and depletes NAD⁺ via PARP-mediated poly ADP-ribosylation. A low dose of H₂O₂ treatment induces NAD⁺ depletion but not cell death; therefore, we used it to induce synthetic lethality. After inducing DNA damage in A549 cells, a significant decline in NAD⁺ levels was observed at 1 h after H₂O₂ treatment; however, NAD⁺ levels were restored to normal levels at 24 h after the treatment. Next, we investigated how NAD⁺ was resynthesized after the treatment. In particular, the source of ribose moiety in NAD⁺ was unknown. We used a stable isotope-labeled glucose and identified that phosphoribosyl pyrophosphate (PRPP), the source of the ribose moiety in NAD⁺, originated from glucose and not from ADP-ribose, a degradation product of auto-ADP-ribosylated PARP through NUDT5. Then, we examined whether the NAD⁺ resynthesis inhibition by glucose depletion could induce synthetic lethality with the low-dose H₂O₂ treatment. H₂O₂ treatment or glucose depletion did not induce cell death when used separately; however, a combination of both treatments induces the synthetic lethality in A549 cells. These results signify that a combination of oxidative stress and NAD⁺ synthesis inhibition is an optimal and minimal invasive therapeutic option to induce cell death in cancer cells.