Skip to main page content
U.S. flag
See More

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2017 Apr 17;30(4):1102-1110.
doi: 10.1021/acs.chemrestox.7b00009. Epub 2017 Mar 24.

Oncometabolites d- and l-2-Hydroxyglutarate Inhibit the AlkB Family DNA Repair Enzymes under Physiological Conditions

Fangyi Chen  1 , Ke Bian  1 , Qi Tang  1 , Bogdan I Fedeles  2 , Vipender Singh  2 , Zachary T Humulock  1 , John M Essigmann  2 , Deyu Li  1
Affiliations

Oncometabolites d- and l-2-Hydroxyglutarate Inhibit the AlkB Family DNA Repair Enzymes under Physiological Conditions

Fangyi Chen et al. Chem Res Toxicol. .

Abstract

Cancer-associated mutations often lead to perturbed cellular energy metabolism and accumulation of potentially harmful oncometabolites. One example is the chiral molecule 2-hydroxyglutarate (2HG); its two stereoisomers (d- and l-2HG) have been found at abnormally high concentrations in tumors featuring anomalous metabolic pathways. 2HG has been demonstrated to competitively inhibit several α-ketoglutarate (αKG)- and non-heme iron-dependent dioxygenases, including some of the AlkB family DNA repair enzymes, such as ALKBH2 and ALKBH3. However, previous studies have only provided the IC50 values of d-2HG on the enzymes, and the results have not been correlated to physiologically relevant concentrations of 2HG and αKG in cancer cells. In this work, we performed detailed kinetic analyses of DNA repair reactions catalyzed by ALKBH2, ALKBH3, and the bacterial AlkB in the presence of d- and l-2HG in both double- and single-stranded DNA contexts. We determined the kinetic parameters of inhibition, including kcat, KM, and Ki. We also correlated the relative concentrations of 2HG and αKG previously measured in tumor cells with the inhibitory effect of 2HG on the AlkB family enzymes. Both d- and l-2HG significantly inhibited the human DNA repair enzymes ALKBH2 and ALKBH3 at pathologically relevant concentrations (73-88% for d-2HG and 31-58% for l-2HG inhibition). This work provides a new perspective that the elevation of the d- or l-2HG concentration in cancer cells may contribute to an increased mutation rate by inhibiting the DNA repair performed by the AlkB family enzymes and thus exacerbate the genesis and progression of tumors.

PubMed Disclaimer

Conflict of interest statement

The authors declare no competing financial interest.

Figures

Figure 1
Figure 1
a) Repair mechanism of the AlkB family enzymes on alkyl DNA lesions. Adduct m1A is used here as an example to show the steps of enzymatic catalysis. b) The generation of D- and L-2HG and mechanisms of inhibition to the AlkB family DNA repair enzymes.
Figure 2
Figure 2
HPLC analyses of the DNA repair reactions. a) Starting material 16mer oligonucleotide containing m1A at the lesion site in ss-DNA reaction. b) ss-Product 16mer oligonucleotide containing A at the “lesion site” in the ss-DNA reaction. c) ds-DNA reaction products of 16mer m1A with 23mer Tcp. The mixture containing ss-16mer m1A, ss-16mer A, ds-16mer m1A:23mer Tcp, and ds-16mer A:23mer Tcp. The latter two species were not fully separable by HPLC. d) ds-DNA reaction products of 16mer m1 A with 23mer Tcp and additional 23mer A, which is fully complementary to 23mer Tcp. The duplex of 23mer Tcp:23mer A was eluted as ds-DNA, thus releasing ss-16mer m1A and ss-16mer A for quantification.
Figure 3
Figure 3
Inhibition of ALKBH2 repair of ml A in ss- and ds-DNA by D- and L-2HG. a) Inhibition of D-2HG on ml A repair in ss-DNA. b) Inhibition of D-2HG on ml A repair in ds-DNA. c) Inhibition of D-2HG on ml A repair under D-2HG:αKG = 373:1 ratio conditions, d) Inhibition of L-2HG on ml A repair in ss-DNA. e) Inhibition of L-2HG on ml A repair in ds-DNA. f) Inhibition of L-2HG on ml A repair under L-2HG:αKG = 28:1 ratio conditions.
Figure 4
Figure 4
Addition of αKG reverses the inhibitory effect of 2-HG toward ALKBH2 repair of ml A. Different concentrations of αKG were added to a fixed concentration of 2HG (10 mM) to recover the repair of ml A by a) ALKBH2, b) ALKBH3 and c) AlkB.
See this image and copyright information in PMC

References

    1. Yan H, Parsons DW, Jin G, McLendon R, Rasheed BA, Yuan W, Kos I, Batinic-Haberle I, Jones S, Riggins GJ, Friedman H, Friedman A, Reardon D, Herndon J, Kinzler KW, Velculescu VE, Vogelstein B, Bigner DD. IDH1 and IDH2 mutations in gliomas. N. Engl. J. Med. 2009;360:765–773. - PMC - PubMed
    1. Parsons DW, Jones S, Zhang X, Lin JC-H, Leary RJ, Angenendt P, Mankoo P, Carter H, Siu I-M, Gallia GL, Olivi A, McLendon R, Rasheed BA, Keir S, Nikolskaya T, Nikolsky Y, Busam DA, Tekleab H, Diaz LA, Hartigan J, Smith DR, Strausberg RL, Marie SKN, Shinjo SMO, Yan H, Riggins GJ, Bigner DD, Karchin R, Papadopoulos N, Parmigiani G, Vogelstein B, Velculescu VE, Kinzler KW. An integrated genomic analysis of human glioblastoma multiforme. Science. 2008;321:1807–1812. - PMC - PubMed
    1. Mardis ER, Ding L, Dooling DJ, Larson DE, McLellan MD, Chen K, Koboldt DC, Fulton RS, Delehaunty KD, McGrath SD, Fulton LA, Locke DP, Magrini VJ, Abbott RM, Vickery TL, Reed JS, Robinson JS, Wylie T, Smith SM, Carmichael L, Eldred JM, Harris CC, Walker J, Peck JB, Du F, Dukes AF, Sanderson GE, Brummett AM, Clark E, McMichael JF, Meyer RJ, Schindler JK, Pohl CS, Wallis JW, Shi X, Lin L, Schmidt H, Tang Y, Haipek C, Wiechert ME, Ivy JV, Kalicki J, Elliott G, Ries RE, Payton JE, Westervelt P, Tomasson MH, Watson MA, Baty J, Heath S, Shannon WD, Nagarajan R, Link DC, Walter MJ, Graubert TA, DiPersio JF, Wilson RK, Ley TJ. Recurring mutations found by sequencing an acute myeloid leukemia genome. N. Engl. J. Med. 2009;361:1058–1066. - PMC - PubMed
    1. Yang H, Ye D, Guan K-L, Xiong Y. IDH1 and IDH2 mutations in tumorigenesis: mechanistic insights and clinical perspectives. Clin. Cancer Res. Off. J. Am. Assoc. Cancer Res. 2012;18:5562–5571. - PMC - PubMed
    1. Cairns RA, Mak TW. Oncogenic isocitrate dehydrogenase mutations: mechanisms, models, and clinical opportunities. Cancer Discov. 2013;3:730–741. - PubMed

MeSH terms

Substances

LinkOut - more resources