Publications by authors named "Stephanie D Threatt"

3 Publications

  • Page 1 of 1

In vivo anticancer activity of a rhodium metalloinsertor in the HCT116 xenograft tumor model.

Proc Natl Acad Sci U S A 2020 07 13;117(30):17535-17542. Epub 2020 Jul 13.

Department of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, CA 91125;

Mismatch repair (MMR) deficiencies are a hallmark of various cancers causing accumulation of DNA mutations and mismatches, which often results in chemotherapy resistance. Metalloinsertor complexes, including [Rh(chrysi)(phen)(PPO)]Cl (Rh-PPO), specifically target DNA mismatches and selectively induce cytotoxicity within MMR-deficient cells. Here, we present an in vivo analysis of Rh-PPO, our most potent metalloinsertor. Studies with HCT116 xenograft tumors revealed a 25% reduction in tumor volume and 12% increase in survival with metalloinsertor treatment (1 mg/kg; nine intraperitoneal doses over 20 d). When compared to oxaliplatin, Rh-PPO displays ninefold higher potency at tumor sites. Pharmacokinetic studies revealed rapid absorption of Rh-PPO in plasma with notable accumulation in the liver compared to tumors. Additionally, intratumoral metalloinsertor administration resulted in enhanced anticancer effects, pointing to a need for more selective delivery methods. Overall, these data show that Rh-PPO inhibits xenograft tumor growth, supporting the strategy of using Rh-PPO as a chemotherapeutic targeted to MMR-deficient cancers.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1073/pnas.2006569117DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7395490PMC
July 2020

Cell-Selective Cytotoxicity of a Fluorescent Rhodium Metalloinsertor Conjugate Results from Irreversible DNA Damage at Base Pair Mismatches.

Biochemistry 2020 02 30;59(5):717-726. Epub 2020 Jan 30.

Division of Chemistry and Chemical Engineering , California Institute of Technology , Pasadena , California 91125 , United States.

Up to 20% of solid tumors are characterized by DNA mismatch repair (MMR) deficiency and microsatellite instability that confer resistance to standard of care chemotherapy. MMR-deficient cancers have an increased mutation rate, and DNA mismatches accumulate as part of these cancers. We previously described a class of compounds, rhodium metalloinsertors, that bind DNA mismatches with high specificity and selectivity and have potential as targeted therapy. [Rh(chrysi)(phen)(PPO)] (RhPPO) is the most potent, selective compound in this class and acts by targeting DNA mismatches, resulting in preferential cytotoxicity to MMR-deficient cancers. To explore further the cellular mechanism of action of RhPPO, we conjugated the metal complex to a fluorescent probe, cyanine 3 (Cy3). RhPPO-Cy3 binds DNA mismatches and retains the selectivity and potent cytotoxic activity of RhPPO for MMR-deficient cell lines. RhPPO-Cy3 forms discrete foci in the cell nucleus that overlap with sites of DNA damage, suggesting that the lesions occur at or near DNA mismatch sites. RhPPO-Cy3 foci persist over time, despite initial processing of the lesion and recruitment of repair proteins, consistent with the idea that the complex binding to a mismatch prevents repair. RhPPO-Cy3 binding does not lead to activation of p53 and the apoptotic pathway. Together, these findings support the idea that RhPPO-Cy3 binding leads to irreversible DNA damage at DNA mismatches that enables selective cytotoxicity to MMR-deficient cells.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/acs.biochem.9b01037DOI Listing
February 2020

Perturbation of Critical Prolines in Gloeobacter violaceus Ligand-gated Ion Channel (GLIC) Supports Conserved Gating Motions among Cys-loop Receptors.

J Biol Chem 2016 Mar 14;291(12):6272-80. Epub 2015 Dec 14.

the Department of Biochemistry, University of Cambridge, Tennis Court Road, Cambridge CB2 1GA, United Kingdom.

Gloeobacter violaceus ligand-gated ion channel (GLIC) has served as a valuable structural and functional model for the eukaryotic Cys-loop receptor superfamily. In Cys-loop and other receptors, we have previously demonstrated the crucial roles played by several conserved prolines. Here we explore the role of prolines in the gating transitions of GLIC. As conventional substitutions at some positions resulted in nonfunctional proteins, we used in vivo non-canonical amino acid mutagenesis to determine the specific structural requirements at these sites. Receptors were expressed heterologously in Xenopus laevis oocytes, and whole-cell electrophysiology was used to monitor channel activity. Pro-119 in the Cys-loop, Pro-198 and Pro-203 in the M1 helix, and Pro-299 in the M4 helix were sensitive to substitution, and distinct roles in receptor activity were revealed for each. In the context of the available structural data for GLIC, the behaviors of Pro-119, Pro-203, and Pro-299 mutants are consistent with earlier proline mutagenesis work. However, the Pro-198 site displays a unique phenotype that gives evidence of the importance of the region surrounding this residue for the correct functioning of GLIC.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1074/jbc.M115.694372DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4813548PMC
March 2016