Synergistic Effect of Endogenous and Exogenous Aldehydes on Doxorubicin Toxicity in Yeast

Abstract

Anthracyclines are frequently used to treat many cancers including triple negative breast cancer, which is commonly observed in African-American women (AA), and tend to be more aggressive, carry worse prognoses, and are harder to manage because they lack molecular targets. Although effective, anthracyclines use can be limited by serious side effects and eventually the development of drug resistance. In S. cerevisiae, mutants of HOM6 display hypersensitivity to doxorubicin. HOM6 is required for synthesis of threonine and interruption of the pathway leads to accumulation of the threonine intermediate L-aspartate-semialdehyde. This intermediate may synergize with doxorubicin to kill the cell. In fact, deleting HOM3 in the first step, preventing the pathway to reach the HOM6 step, rescues the sensitivity of the hom6 strain to doxorubicin. Using several S. cerevisiae strains (wild type, hom6, hom3, hom3hom6, ydj1, siz1, and msh2), we determined their sensitivity to aldehydes and to their combination with doxorubicin, cisplatin, and etoposide. Combination of formaldehyde and doxorubicin was most effective at reducing cell survival by 31-fold-39-fold (in wild type cells) relative to doxorubicin and formaldehyde alone. This effect was dose dependent on doxorubicin. Cotreatment with formaldehyde and doxorubicin also showed increased toxicity in anthracycline-resistant strains siz1 and msh2. The hom6 mutant also showed sensitivity to menadione with a 2.5-fold reduction in cell survival. The potential use of a combination of aldehydes and cytotoxic drugs could potentially lead to applications intended to enhance anthracycline-based therapy.

Figure 1

Defects in threonine biosynthesis sensitize cells to doxorubicin. Inactivated genes in the threonine biosynthetic pathway result in increased toxicity of doxorubicin (a, b). Inactivation of the HOM3 gene rescues the hom6 strain (c). Quantitation of hom3, hom6, and hom6hom3 survival after doxorubicin exposure, 20 μM (d).

Figure 2

Formaldehyde synergizes with doxorubicin to kill wild type cells. (a) Growth of wild type cells tested by spotting onto YPD agar plates. (b) Quantitation of wild type cells treated with formaldehyde and doxorubicin, alone and in combination.

Figure 3

Cotreatment with formaldehyde enhances the toxicity of wild type and the doxorubicin resistant strains siz1 and msh2. (a) Strains growth tested by spotting onto YPD agar plates. (b) Survival rates of strains following formaldehyde and doxorubicin treatment, alone and combined.

Figure 4

The hom6 mutants are sensitive to the generation of ROS. (a) Growth of wild type, sod1, and hom6 mutants after menadione (Men), 6.6 mM exposure. (b) Survival rates following Men, 6.6 mM exposure.

Figure 5

The hom6 mutants are not sensitive to the generation of DNA double strand breaks. (a) Growth of hom6, rad52, and ydj1 mutants after Etoposide (Etopo), 0.5 mM exposure. (b) Survival rates following Etopo, 0.5 mM exposure.