Biology subjects

Singleton, K. R.

Publications and source records attributed to Singleton, K. R..

3 recordsLinked to original sources

bioRxiv2023

MCB-613 exploits a collateral sensitivity in drug-resistant EGFR-mutant non-small cell lung cancer through covalent inhibition of KEAP1

Targeted therapies have revolutionized cancer chemotherapy. Unfortunately, most patients develop multifocal resistance to these drugs within a matter of months. Here, we used a high-throughput phenotypic small molecule screen to identify MCB-613 as a compound that selectively targets EGFR-mutant, EGFR inhibitor-resistant non-small cell lung cancer (NSCLC) cells harboring diverse resistance mechanisms. Subsequent proteomic and functional genomic screens involving MCB-613 identified its target in this context to be KEAP1, revealing that this gene is selectively essential in the setting of EGFR inhibitor resistance. In-depth molecular characterization demonstrated that (1) MCB-613 binds KEAP1 covalently; (2) a single molecule of MCB-613 is capable of bridging two KEAP1 monomers together; and, (3) this modification interferes with the degradation of canonical KEAP1 substrates such as NRF2. Surprisingly, NRF2 knockout sensitizes cells to MCB-613, suggesting that the drug functions through modulation of an alternative KEAP1 substrate. Together, these findings advance MCB-613 as a new tool for exploiting the selective essentiality of KEAP1 in drug-resistant, EGFR-mutant NSCLC cells.

cancer biology↗

bioRxiv2022

Drug dependence in cancer is exploitable by optimally constructed treatment holidays

Recent work in cell culture models, animal models, and human patients indicates that cancers with acquired resistance to a drug can become simultaneously dependent upon the presence of that drug for survival. This drug dependence offers a potential avenue for improving treatments aimed at slowing resistance, yet relatively little is known about the frequency with which drug dependence arises, the mechanisms underlying that dependence, and how drug schedules might be tuned to optimally exploit drug dependence. In this work, we address these open questions using a combination of laboratory evolution, in vitro experiments, and simple mathematical models. First, we used laboratory evolution to select more than 100 resistant BRAF mutant melanoma cell lines with acquired resistance to BRAF, MEK, or ERK inhibitors. We found that nearly half of these lines exhibit drug dependence, and the dependency response is associated with EGFR-driven senescence induction, but not apoptosis, following drug withdrawal. Then, using melanoma populations with evolved resistance to the BRAF inhibitor PLX4720, we showed that drug dependence can be leveraged to dramatically reduce population growth when treatment strategies include optimally chosen drug-free "holidays". On short timescales, the duration of these holidays depends sensitively on the composition of the population, but for sufficiently long treatments it depends only on a single dimensionless parameter ({gamma}) that describes how the growth rates of each cell type depend on the different treatment environments. Experiments confirm that the optimal holiday duration changes in time-with holidays of different durations leading to optimized treatments on different timescales. Furthermore, we find that the presence of "non-dependent" resistant cells does not change the optimal treatment schedule but leads to a net increase in population size. Finally, we show that even in the absence of detailed information about the composition and growth characteristics of cellular clones within a population, a simple adaptive therapy protocol can produce near-optimal outcomes using only measurements of total population size, at least when these measurements are sufficiently frequent. As a whole, these results may provide a stepping-stone toward the eventual development of evolution-inspired treatment strategies for drug dependent cancers.

cancer biology↗

bioRxiv2019

Evolved resistance to GAPDH inhibition results in loss of the Warburg Effect but retains a different state of glycolysis

Aerobic glycolysis or the Warburg Effect (WE) is characterized by increased glucose uptake and incomplete oxidation to lactate. Although ubiquitous, the biological role of the WE remains controversial and whether glucose metabolism is functionally different during fully oxidative glycolysis or during the WE is unknown. To investigate this question, we evolved resistance to koningic acid (KA), a natural product shown to be a specific inhibitor of glyceraldehyde-3-phosphate dehydrogenase (GAPDH), a rate-controlling glycolytic enzyme during the WE. We find that KA-resistant cells lose the WE but conduct glycolysis and surprisingly remain dependent on glucose and central carbon metabolism. Consequentially this altered state of glycolysis leads to differential metabolic activity and requirements including emergent activities in and dependencies on fatty acid metabolism. Together, these findings reveal that, contrary to some recent reports, aerobic glycolysis is a functionally distinct entity from conventional glucose metabolism and leads to distinct metabolic requirements and biological functions.

systems biology↗