Synthesis of ortho-formylphenylphosphonic acids as covalent probes of active site lysines

Article abstract of DOI:10.1080/10426507.2018.1539996

During the course of an investigation of targeted inhibition of DNA polymerase beta (pol β) lyase activity using small molecules, we observed the formation of an aldimine between (2-formyl)phenylphosphonic acid (2FPP) and butylamine under basic aqueous conditions; complete deprotonation of the phosphonate group was required to stabilize the imine product. Results of computational docking studies suggested that the reaction of Lys-72 on the lyase active site with an aldehyde group could be facilitated by a proximal phosphonate, not only because of the phosphonate’s ability to mimic phosphate interacting with the DNA binding site, but also because of its ability to shield the imine against hydrolysis. Novel pol β lyase inhibitors were thus prepared using a 2FPP analogue with an amine linker; P-C bond formation in synthesis of this intermediate was possible with an unprotected aldehyde using palladium-catalyzed, microwave-assisted Michaelis–Arbuzov chemistry. These compounds, and structurally related derivatives lacking the aldehyde or phosphonate, were evaluated in an assay for pol β to assess their potential for inhibition.

Full text of DOI:10.1080/10426507.2018.1539996

Phosphorus, Sulfur, and Silicon and the Related Elements
ISSN: 1042-6507 (Print) 1563-5325 (Online) Journal homepage: https://www.tandfonline.com/loi/gpss20
Synthesis of ortho-formylphenylphosphonic acids
as covalent probes of active site lysines
Marlon Vincent V. Duro, Khadijeh S. Alnajjar, Joann B. Sweasy, Boris A.
Kashemirov & Charles E. McKenna
To cite this article: Marlon Vincent V. Duro, Khadijeh S. Alnajjar, Joann B. Sweasy, Boris A.
Kashemirov & Charles E. McKenna (2019): Synthesis of ortho-formylphenylphosphonic acids as
covalent probes of active site lysines, Phosphorus, Sulfur, and Silicon and the Related Elements,
DOI: 10.1080/10426507.2018.1539996
To link to this article: https://doi.org/10.1080/10426507.2018.1539996
Published online: 13 Feb 2019.
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PHOSPHORUS, SULFUR, AND SILICON AND THE RELATED ELEMENTS
https://doi.org/10.1080/10426507.2018.1539996
SHORT COMMUNICATION
Synthesis of ortho-formylphenylphosphonic acids as covalent probes of active
site lysines
Marlon Vincent V. Duroa, Khadijeh S. Alnajjarb, Joann B. Sweasyb, Boris A. Kashemirova, and
Charles E. McKennaa
1Department of Chemistry, Dana and David Dornsife College of Letters Arts and Sciences, University of Southern California, Los Angeles,
California, USA;2Department of Therapeutic Radiology, Yale School of Medicine, Yale University, New Haven, Connecticut, USA
ABSTRACT
ARTICLE HISTORY
Received 11 September 2018
Accepted 19 October 2018
During the course of an investigation of targeted inhibition of DNA polymerase beta (pol β) lyase
activity using small molecules, we observed the formation of an aldimine between (2-formyl)phe-
nylphosphonic acid (2FPP) and butylamine under basic aqueous conditions; complete deprotona-
tion of the phosphonate group was required to stabilize the imine product. Results of
computational docking studies suggested that the reaction of Lys-72 on the lyase active site with
an aldehyde group could be facilitated by a proximal phosphonate, not only because of the phos-
phonate’s ability to mimic phosphate interacting with the DNA binding site, but also because of
its ability to shield the imine against hydrolysis. Novel pol β lyase inhibitors were thus prepared
using a 2FPP analogue with an amine linker; P-C bond formation in synthesis of this intermediate
was possible with an unprotected aldehyde using palladium-catalyzed, microwave-assisted
Michaelis–Arbuzov chemistry. These compounds, and structurally related derivatives lacking the
aldehyde or phosphonate, were evaluated in an assay for pol β to assess their potential
for inhibition.
KEYWORDS
DNA polymerase; lyase;
phosphonate; inhibitor
GRAPHICAL ABSTRACT
Results and discussion
unprotected aldehyde. Dealkylation with bromotrimethylsi-
lane (BTMS) yielded 2FPP; all compounds were character-
ized spectroscopically. NMR studies showed that in aqueous
media, an alkaline pH drives formation of a stable aldimine
between 2FPP and butylamine (68% conversion by 31P
NMR in D2O, pD 10.9); the imine concentration is
Based on preliminary computational covalent docking stud-
ies, we hypothesized that incorporating an aldehyde ortho to
a phosphonate on a diphenylamine scaffold might mimic
deoxyribose-5′-phosphate and could provide enhanced
inhibition of DNA polymerase β (pol β) lyase activity
through the formation of a covalent bond with Lys-72 on decreased by dilution or acidification with NaHSO4. The
the active site. To confirm the formation of such imines, we phosphonate ortho to the aldehyde of 2FPP is expected to
performed NMR studies of the reaction between butylamine
and 2-formylphenylphosphonic acid (2FPP). Here, we
describe the first reported synthesis of 2FPP (Scheme 1); the
be fully deprotonated above pH 8, suggesting that its nega-
tive charge may play a role in shielding the imine
against hydrolysis.
key step is
a palladium-catalyzed, microwave-assisted
2FPP-based pol β inhibitors 5 and 6 were synthesized via
Michaelis–Arbuzov-like reaction[1–2] which afforded 2FPP the amino derivative 4 (Scheme 2) from aryl bromide 3.
diethyl ester (1) in good yield even in the presence of an Results of preliminary assays for pol β lyase inhibition show
CONTACT Charles E. McKenna
ß 2018 Taylor & Francis Group, LLC
mckenna@usc.edu

2
M. V. V. DURO ET AL.
Scheme 1. Microwave-assisted synthesis of 2FPP.
Scheme 2. Synthesis of novel pol β inhibitors.
that these “rationally designed” compounds are significantly
more potent than pamoic acid, a known pol β inhibitor.[3]
However, further investigation will be required to determine
the underlying mechanism in terms of a covalent vs. non-
covalent interaction with the enzyme.
References
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[2] Keglevich, G.; Grun, A.; Bolcskei, A.; Drahos, L.; Kraszni, M.;
Balogh, G. T. Synthesis and Proton Dissociation Properties of
Arylphosphonates:
A Microwave-Assisted Catalytic Arbuzov
Funding
Reaction with Aryl Bromides. Heteroatom Chem. 2012, 23, 574–582.
[3] Hu, H. Y.; Horton, J. K.; Gryk, M. R.; Prasad, R.; Naron, J. M.; Sun,
D. A.; Hecht, S. M.; Wilson, S. H.; Mullen, G. P. Identification of
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Chemical Shift Mapping. J. Biol. Chem 2004, 279, 39736–39744.
This research was supported by a grant from the NIH-NCI,
U19CA177547.