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Xue Zhi  Zhao, Ph.D.

Xue Zhi Zhao, Ph.D.

  • Center for Cancer Research
  • National Cancer Institute
Chemical Biology Laboratory

RESEARCH SUMMARY

Dr. Zhao has been a pioneer in the field of anti-AIDS agents, with a particular focus on developing inhibitors of HIV-1 integrase (IN). His recent studies have concentrated on the discovery of inhibitors that retain efficacy against virus harboring mutant forms of IN integrase, which are resistant to current FDA-approved drugs. These efforts are being done in collaboration with the NCI laboratories of Drs. Yves Pommier (Developmental Therapeutics Branch) and Stephen Hughes (HIV Drug Resistance Program). His compounds offer potential leads for further structural variation that may ultimately yield clinical agents capable of overcoming problems associated with the development of resistance. Dr. Zhao has applied oxime-diversification to optimize ligand interactions within a cryptic pocket of the polo-like kinase 1 polo-box domain (Plk1 PBD) and developed a series of peptide ligands. Dr. Zhao has also been developing tyrosyl-DNA phosphodiesterase 1 (TDP1) inhibitors as synergistic combination with current Top1 inhibitors in anti-cancer therapy.

Areas of Expertise

1) medicinal chemistry 2) chemical biology 3) drug discovery and design 4) peptide chemistry 5) HIV integrase inhibitors 6) TDP1 inhibitors

Publications

Selected Key Publications

Small molecule microarray identifies inhibitors of tyrosyl-DNA phosphodiesterase 1 that simultaneously access the catalytic pocket and two substrate binding sites

Zhao XZ, Kiselev E, Lountos GT, Wang W, Tropea JE, Needle D, Hilimire TA, Schneekloth JS, Waugh DS, Pommier Y, Burke TR Jr.
Chemical Science. 12(11): 3876 - 3884, 2021.
Full-Text Article
[ Journal Article ]

Structural basis for strand-transfer inhibitor binding to HIV intasomes.

Passos DO, Li M, Jóźwik IK, Zhao XZ, Santos-Martins D, Yang R, Smith SJ, Jeon Y, Forli S, Hughes SH, Burke TR Jr, Craigie R, Lyumkis D.
Science. 367(6479): 810-814, 2020.
Full-Text Article
[ Journal Article ]

Structure-Guided Optimization of HIV Integrase Strand Transfer Inhibitors

Zhao XZ, Smith SJ, Maskell DP, Métifiot M, Pye VE, Fesen K, Marchand C, Pommier Y, Cherepanov P, Hughes SH, Burke TR Jr.
Journal of Medicinal Chemistry. 60(17): 7315-7332, 2017.
Full-Text Article
[ Journal Article ]

HIV-1 Integrase Strand Transfer Inhibitors with Reduced Susceptibility to Drug Resistant Mutant Integrases

Zhao XZ, Smith SJ, Maskell DP, Metifiot M, Pye VE, Fesen K, Marchand C, Pommier Y, Cherepanov P, Hughes SH, Burke TR Jr.
ACS Chemical Biology. 11(4): 1074-1081, 2016.
Full-Text Article
[ Journal Article ]

Bicyclic 1-hydroxy-2-oxo-1,2-dihydropyridine-3-carboxamide-containing HIV-1 integrase inhibitors having high antiviral potency against cells harboring raltegravir-resistant integrase mutants.

Zhao XZ, Smith SJ, Métifiot M, Johnson BC, Marchand C, Pommier Y, Hughes SH, Burke TR Jr.
Journal of Medicinal Chemistry. 57(4): 1573-1582, 2014.
Full-Text Article
[ Journal Article ]

Covers

INSTIs bound to HIV-1 IN

HIV‑1 Integrase Inhibitors with Modifications That Affect Their Potencies against Drug Resistant Integrase Mutants

Published Date

Integrase strand transfer inhibitors (INSTIs) block the integration step of the retroviral lifecycle and are first-line drugs used for the treatment of HIV-1/AIDS. INSTIs have a polycyclic core with heteroatom triads, chelate the metal ions at the active site, and have a halobenzyl group that interacts with viral DNA attached to the core by a flexible linker. The most broadly effective INSTIs inhibit both wild-type (WT) integrase (IN) and a variety of well-known mutants. However, because there are mutations that reduce the potency of all the available INSTIs, new and better compounds are needed. Models based on recent structures of HIV-1 and red-capped mangabey SIV INs suggest modifications in the INSTI structures that could enhance interactions with the 3′-terminal adenosine of the viral DNA, which could improve performance against INSTI resistant mutants. We designed and tested a series of INSTIs having modifications to their naphthyridine scaffold. One of the new compounds retained good potency against an expanded panel of HIV-1 IN mutants that we tested. Our results suggest the possibility of designing inhibitors that combine the best features of the existing compounds, which could provide additional efficacy against known HIV-1 IN mutants.

Citation

Steven J. Smith, Xue Zhi Zhao, Dario Oliveira Passos, Valerie E. Pye, Peter Cherepanov, Dmitry LyumkisTerrence R. Burke, Jr., and Stephen H. Hughes* in ACS Infectious Diseases, 20217, 1469-1482

Small molecule microarray identifies inhibitors of tyrosyl-DNA phosphodiesterase 1 that simultaneously access the catalytic pock

Small molecule microarray identifies inhibitors of tyrosyl-DNA phosphodiesterase 1 that simultaneously access the catalytic pocket and two substrate binding sites

Published Date

We now report examining a 21 000-member library of drug-like Small Molecules in Microarray (SMM) format for their ability to bind Alexa Fluor 647 (AF647)-labeled Tyrosyl-DNA phosphodiesterase 1 (TDP1). The screen identified structurally similar N,2-diphenylimidazo[1,2-a]pyrazin-3-amines as TDP1 binders and catalytic inhibitors. We then explored the core heterocycle skeleton using one-pot Groebke–Blackburn–Bienayme multicomponent reactions and arrived at analogs having higher inhibitory potencies. Solving TDP1 co-crystal structures of a subset of compounds showed their binding at the TDP1 catalytic site, while mimicking substrate interactions. Although our original fragment screen differed significantly from the current microarray protocol, both methods identified ligand–protein interactions containing highly similar elements. Importantly inhibitors identified through the SMM approach show competitive inhibition against TDP1 and access the catalytic phosphate-binding pocket, while simultaneously providing extensions into both the substrate DNA and peptide-binding channels. As such, they represent a platform for further elaboration of trivalent ligands, that could serve as a new genre of potent TDP1 inhibitors.

Citation

Zhao XZ, Kiselev E, Lountos GT, Wang W, Tropea JE, Needle D, Hilimire TA, Schneekloth JS, Waugh DS, Pommierb Y, Burke TR Jr.  Chemical Science, 2021, 12, 3876 - 3884

Application of Post Solid-Phase Oxime Ligation to Fine-Tune Peptide-Protein Interactions

Application of Post Solid-Phase Oxime Ligation to Fine-Tune Peptide-Protein Interactions

Published Date

Protein-protein interactions (PPIs) represent an extremely attractive class of potential new targets for therapeutic intervention; however, the shallow extended character of many PPIs can render developing inhibitors against them as exceptionally difficult. Yet this problem can be made tractable by taking advantage of the fact that large interacting surfaces are often characterized by confined "hot spot" regions, where interactions contribute disproportionately to overall binding energies. Peptides afford valuable starting points for developing PPI inhibitors because of their high degrees of functional diversity and conformational adaptability. Unfortunately, contacts afforded by the 20 natural amino acids may be suboptimal and inefficient for accessing both canonical binding interactions and transient "cryptic" binding pockets. Oxime ligation represents a class of biocompatible "click" chemistry that allows the structural diversity of libraries of aldehydes to be rapidly evaluated within the context of a parent oxime-containing peptide platform. Importantly, oxime ligation represents a form of post solid-phase diversification, which provides a facile and empirical means of identifying unanticipated protein-peptide interactions that may substantially increase binding affinities and selectivity. The current review will focus on the authors' use of peptide ligation to optimize PPI antagonists directed against several targets, including tumor susceptibility gene 101 (Tsg101), protein tyrosine phosphatases (PTPases) and the polo-like kinase 1 (Plk1). This should provide insights that can be broadly directed against an almost unlimited range of physiologically important PPIs.

Citation

Zhao, X.Z.; Liu, F.; Burke, T.R., Jr. Molecules 202025, 2807. 

Crystal structure of TDP1 bound to one of the small molecule inhibitors

Identification of a ligand binding hot spot and structural motifs replicating aspects of tyrosyl-DNA phosphodiesterase I (TDP1) phosphoryl recognition by crystallographic fragment cocktail screening

Published Date

Tyrosyl DNA-phosphodiesterase I (TDP1) repairs type IB topoisomerase (TOP1) cleavage complexes generated by TOP1 inhibitors commonly used as anticancer agents. TDP1 also removes DNA 3′ end blocking lesions generated by chain-terminating nucleosides and alkylating agents, and base oxidation both in the nuclear and mitochondrial genomes. Combination therapy with TDP1 inhibitors is proposed to synergize with topoisomerase targeting drugs to enhance selectivity against cancer cells exhibiting deficiencies in parallel DNA repair pathways. A crystallographic fragment screening campaign against the catalytic domain of TDP1 was conducted to identify new lead compounds. Crystal structures revealed two fragments that bind to the TDP1 active site and exhibit inhibitory activity against TDP1. These fragments occupy a similar position in the TDP1 active site as seen in prior crystal structures of TDP1 with bound vanadate, a transition state mimic. Using structural insights into fragment binding, several fragment derivatives have been prepared and evaluated in biochemical assays. These results demonstrate that fragment-based methods can be a highly feasible approach toward the discovery of small-molecule chemical scaffolds to target TDP1, and for the first time, we provide co-crystal structures of small molecule inhibitors bound to TDP1, which could serve for the rational development of medicinal TDP1 inhibitors.

Citation

George T. Lountos, Xue Zhi Zhao, Evgeny Kiselev, Joseph E. Tropea, Danielle Needle, Yves Pommier, Terrence R. Burke, Jr. and David S. Waugh in Nucleic Acids Research201947, 10134–10150.

2 inhibitors depicted binding within the hydrophobic pocket on the surface of the PBD..

Development of Highly Selective 1,2,3-Triazole-containing Peptidic Polo-like Kinase 1 Polo-box Domain-binding Inhibitors

Published Date

Cover Story: Two highly selective triazole-containing peptidic polo-like kinase 1 (Plk1) polo-box domain (PBD)-binding inhibitors (3d in green and 4b in orange) are depicted, binding within the hydrophobic “cryptic” binding pocket on the surface of the Plk1 PBD (light blue). The placement was guided by our previously reported X-ray crystal structure of PBD-bound parent peptide (2a) (PDB accession code: 3RQ7), whose binding pocket is highlighted in yellow. These ligands retain the high PBD-binding affinity of the parent peptide, while showing desirable enhanced selectivity for the PBD of Plk1 relative to the PBDs of Plk2 and Plk3.

Members of the polo-like kinase (Plk) family of serine/threonine protein kinases play crucial roles in cell cycle regulation and proliferation. Of five Plks (Plk1 – 5), Plk1 is recognized as an anticancer drug target. Plk1 contains multiple structural components that are important for its proper biological function. These include an N-terminal catalytic domain and a C-terminal non-catalytic polo-box domain (PBD). The PBD binds to phosphothreonine (pT) and phosphoserine-containing sequences. Blocking PBD-dependent interactions offers a potential means of down-regulating Plk1 function that is distinct from targeting its ATP-binding site. Previously, we demonstrated by tethering alkylphenyl chains from the N(π)-position of the His residue in the 5-mer PLHSpT, that we were able to access a hydrophobic "cryptic" binding pocket on the surface of the PBD, and in so doing enhance binding affinities by approximately 1000-fold. More recently, we optimized these PBD-ligand interactions using an oxime ligation-based strategy. Herein, using azide-alkyne cycloaddition reactions, we explore new triazole-containing PBD-binding antagonists. Some of these ligands retain the high PBD-binding affinity of the parent peptide, while showing desirable enhanced selectivity for the PBD of Plk1 relative to the PBDs of Plk2 and Plk3.

Citation

Zhao, X. Z.; Tsuji, K.; Hyman, D. M.; Burke, T. R., Jr. Development of highly selective 1,2,3-triazole-containing peptidic polo-like kinase 1 polo-box domain-binding inhibitors Molecules 2019, 24, 1488 (doi:10.3390/molecules24081488).

A 7-mer peptide based on the shortened Vpr sequence containing a biotin group and a photo-reactive residue

HIV-1 Integrase-Targeted Short Peptides Derived from a Viral Protein R Sequence

Published Date

HIV-1 integrase (IN) inhibitors represent a new class of highly effective anti-AIDS therapeutics. Current FDA-approved IN strand transfer inhibitors (INSTIs) share a common mechanism of action that involves chelation of catalytic divalent metal ions. However, the emergence of IN mutants having reduced sensitivity to these inhibitors underlies efforts to derive agents that antagonize IN function by alternate mechanisms. Integrase along with the 96-residue multifunctional accessory protein, viral protein R (Vpr), are both components of the HIV-1 pre-integration complex (PIC). Coordinated interactions within the PIC are important for viral replication. Herein, we report a 7-mer peptide based on the shortened Vpr (69–75) sequence containing a biotin group and a photo-reactive benzoylphenylalanyl residue, and which exhibits low micromolar IN inhibitory potency. Photo-crosslinking experiments have indicated that the peptide directly binds IN. The peptide does not interfere with IN-DNA interactions or induce higher-order, aberrant IN multimerization, suggesting a mode of action for the peptide that is distinct from clinically used INSTIs and developmental allosteric IN inhibitors. This compact Vpr-derived peptide may serve as a valuable pharmacological tool to identify a potential new pharmacologic site.

Citation

 Xue Zhi Zhao, Mathieu Métifiot, Evgeny Kiselev, Jacques J. Kessi, Kasthuraiah Maddali, Christophe Marchand, Mamuka Kvaratskhelia, Yves Pommier and Terrence R. Burke Jr. in Molecules, 201823 (8), 1858.

Cover of Journal of Medicinal Chemistry September 14, 2017

Structure-Guided Optimization of HIV Integrase Strand Transfer Inhibitors

Published Date

Integrase mutations can reduce the effectiveness of the first-generation FDA-approved integrase strand transfer inhibitors (INSTIs), raltegravir (RAL) and elvitegravir (EVG). The second-generation agent, dolutegravir (DTG), has enjoyed considerable clinical success; however, resistancecausing mutations that diminish the efficacy of DTG have appeared. Our current findings support and extend the substrate envelope concept that broadly effective INSTIs can be designed by filling the envelope defined by the DNA substrates. Previously, we explored 1-hydroxy-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamides as an INSTI scaffold, making a limited set of derivatives, and concluded that broadly effective INSTIs can be developed using this scaffold. Herein, we report an extended investigation of 6-substituents as well the first examples of 7-substituted analogues of this scaffold. While 7-substituents are not well-tolerated, we have identified novel substituents at the 6-position that are highly effective, with the best compound (6p) retaining better efficacy against a broad panel of known INSTI resistant mutants than any analogues we have previously described. View the article.

Citation

Xue Zhi Zhao, Steven J. Smith, Daniel P. Maskell, Mathieu Metífiot, Valerie E. Pye, Katherine Fesen, Christophe Marchand, Yves Pommier, Peter Cherepanov, Stephen H. Hughes, and Terrence R. Burke, Jr. in J. Med. Chem. 201760, 7315-7332.

Cover of Bioorganic & Medicinal Chemistry Letters Oct. 15, 2016

Application of oxime-diversification to optimize ligand interactions within a cryptic pocket of the polo-like kinase 1 polo-box domain

Published Date

By a process involving initial screening of a set of 87 aldehydes using an oxime ligation-based strategy, we were able to achieve a several-fold affinity enhancement over one of the most potent previously known polo-like kinase 1 (Plk1) polo-box domain (PBD) binding inhibitors. This improved binding may result by accessing a newly identified auxiliary region proximal to a key hydrophobic cryptic pocket on the surface of the protein. Our findings could have general applicability to the design of PBD-binding antagonists. 
Cover design by Joseph Myer

Citation

Xue Zhi Zhao, David Hymel and Terrence R. Burke, Jr. in Bioorganic & Medicinal Chemistry Letters2016, 26, 5009-5012. 

ACS Chemical Biology cover - Vol. 11, No. 2, Apr. 15, 2016

HIV‑1 Integrase Strand Transfer Inhibitors with Reduced Susceptibility to Drug Resistant Mutant Integrases

Published Date

On the cover: Mutant forms of HIV-1 IN reduce the therapeutic effectiveness of integrase strand transfer inhibitors (INSTIs). The cover figure shows the IN of prototype foamy virus complexed to a novel INSTI (gold) that retains potency against resistant mutants of HIV-1 IN. Overlain are the host and viral DNA substrates (blue and green, respectively), showing substrate mimicry by the inhibitor.

Cover design by Joseph Myer

Citation

Xue Zhi Zhao, Steven J. Smith, Daniel P. Maskell, Mathieu Metifiot, Valerie E. Pye, Katherine Fesen, Christophe Marchand, Yves Pommier, Peter Cherepanov, Stephen H. Hughes and Terrence R. Burke, Jr. in ACS Chemical Biology, 20165, 11 , 1074-1081.