Collateral Sensitivity Research Articles

Transcriptomics, molecular docking, and cross-resistance profiling of nobiletin in cancer cells and synergistic interaction with doxorubicin upon SOX5 transfection

BackgroundNobiletin is a polymethoxylated flavone from citrus fruit peels. Among other bioactivities, it acts antioxidative, anti-inflammatory, neuroprotective, and cardiovascular-protective. Nobiletin exerts profound anticancer activity in vitro and in vivo but the underlying mechanisms are not well understood. PurposeThe aim was to unravel the multiple modes of action against cancer cells by bioinformatic and transcriptomic techniques and their verification by molecular pharmacological methods. MethodsThe in silico methods used were COMPARE analysis of transcriptomic data, signaling pathway analysis, transcription factor binding motif analysis in promoter sequences of target genes, and molecular docking. The in vitro methods used were resazurin assay, isobologram analysis, generation of stably SOX5-tranfected cells, and Western blotting. ResultsNobiletin was cytotoxic against a wide range of cell lines from different tumor types, including diverse phenotypes to established anticancer drugs (e.g., P-glycoprotein, ABCB5, p53, EGFR). Cross-resistance profiling with 83 standard anticancer drugs revealed a correlation to antihormonal anticancer drugs, which can be explained by the phytoestrogenic features of nobiletin. Transcriptomic analysis showed that the responsiveness of tumor cells was predictable by their specific mRNA expression profile. Nobiletin bound to the transcription factor SOX5 in silico. SOX5 conferred resistance to the control drug doxorubicin but collateral sensitivity to nobiletin in HEK293 cells transfected with a lentiviral GFP-tagged pLOCORF-SOX5 vector. The combination of nobiletin and doxorubicin synergistically killed HEK293-SOX5 cells in isobologram analyses, implying attractive new treatment options. ConclusionNobiletin represents an interesting candidate for cancer therapy with broad-spectrum activity and multiple modes of action. The identification of novel targets (i.e., SOX5) may allow its use for targeted tumor therapy in individualized treatment protocols.

The physiology and genetics of bacterial responses to antibiotic combinations.

Several promising strategies based on combining or cycling different antibiotics have been proposed to increase efficacy and counteract resistance evolution, but we still lack a deep understanding of the physiological responses and genetic mechanisms that underlie antibiotic interactions and the clinical applicability of these strategies. In antibiotic-exposed bacteria, the combined effects of physiological stress responses and emerging resistance mutations (occurring at different time scales) generate complex and often unpredictable dynamics. In this Review, we present our current understanding of bacterial cell physiology and genetics of responses to antibiotics. We emphasize recently discovered mechanisms of synergistic and antagonistic drug interactions, hysteresis in temporal interactions between antibiotics that arise from microbial physiology and interactions between antibiotics and resistance mutations that can cause collateral sensitivity or cross-resistance. We discuss possible connections between the different phenomena and indicate relevant research directions. A better and more unified understanding of drug and genetic interactions is likely to advance antibiotic therapy.

Rapid Decline of Ceftazidime Resistance in Antibiotic-Free and Sublethal Environments Is Contingent on Genetic Background.

Trade-offs of antibiotic resistance evolution, such as fitness cost and collateral sensitivity (CS), could be exploited to drive evolution toward antibiotic susceptibility. Decline of resistance may occur when resistance to other drug leads to CS to the first one and when compensatory mutations, or genetic reversion of the original ones, reduce fitness cost. Here we describe the impact of antibiotic-free and sublethal environments on declining ceftazidime resistance in different Pseudomonas aeruginosa resistant mutants. We determined that decline of ceftazidime resistance occurs within 450 generations, which is caused by newly acquired mutations and not by reversion of the original ones, and that the original CS of these mutants is preserved. In addition, we observed that the frequency and degree of this decline is contingent on genetic background. Our results are relevant to implement evolution-based therapeutic approaches, as well as to redefine global policies of antibiotic use, such as drug cycling.

The population genetics of collateral resistance and sensitivity.

Resistance mutations against one drug can elicit collateral sensitivity against other drugs. Multi-drug treatments exploiting such trade-offs can help slow down the evolution of resistance. However, if mutations with diverse collateral effects are available, a treated population may evolve either collateral sensitivity or collateral resistance. How to design treatments robust to such uncertainty is unclear. We show that many resistance mutations in Escherichia coli against various antibiotics indeed have diverse collateral effects. We propose to characterize such diversity with a joint distribution of fitness effects (JDFE) and develop a theory for describing and predicting collateral evolution based on simple statistics of the JDFE. We show how to robustly rank drug pairs to minimize the risk of collateral resistance and how to estimate JDFEs. In addition to practical applications, these results have implications for our understanding of evolution in variable environments.

Targeting multidrug resistance-associated protein 1 (MRP1)-expressing cancers: Beyond pharmacological inhibition.

Resistance to chemotherapy remains one of the most significant obstacles to successful cancer treatment. While inhibiting drug efflux mediated by ATP-binding cassette (ABC) transporters is a seemingly attractive and logical approach to combat multidrug resistance (MDR), small molecule inhibition of ABC transporters has so far failed to confer clinical benefit, despite considerable efforts by medicinal chemists, biologists, and clinicians. The long-sought treatment to eradicate cancers displaying ABC transporter overexpression may therefore lie within alternative targeting strategies. When aberrantly expressed, the ABC transporter multidrug resistance-associated protein 1 (MRP1, ABCC1) confers MDR, but can also shift cellular redox balance, leaving the cell vulnerable to select agents. Here, we explore the physiological roles of MRP1, the rational for targeting this transporter in cancer, the development of small molecule MRP1 inhibitors, and the most recent developments in alternative therapeutic approaches for targeting cancers with MRP1 overexpression. We discuss approaches that extend beyond simple MRP1 inhibition by exploiting the collateral sensitivity to glutathione depletion and ferroptosis, the rationale for targeting the shared transcriptional regulators of both MRP1 and glutathione biosynthesis, advances in gene silencing, and new molecules that modulate transporter activity to the detriment of the cancer cell. These strategies illustrate promising new approaches to address multidrug resistant disease that extend beyond the simple reversal of MDR and offer exciting routes for further research.

Collateral Sensitivity Interactions between Antibiotics Depend on Local Abiotic Conditions.

ABSTRACTMutations conferring resistance to one antibiotic can increase (cross-resistance) or decrease (collateral sensitivity) resistance to others. Antibiotic combinations displaying collateral sensitivity could be used in treatments that slow resistance evolution. However, lab-to-clinic translation requires understanding whether collateral effects are robust across different environmental conditions. Here, we isolated and characterized resistant mutants of Escherichia coli using five antibiotics, before measuring collateral effects on resistance to other paired antibiotics. During both isolation and phenotyping, we varied conditions in ways relevant in nature (pH, temperature, and bile). This revealed that local abiotic conditions modified expression of resistance against both the antibiotic used during isolation and other antibiotics. Consequently, local conditions influenced collateral sensitivity in two ways: by favoring different sets of mutants (with different collateral sensitivities) and by modifying expression of collateral effects for individual mutants. These results place collateral sensitivity in the context of environmental variation, with important implications for translation to real-world applications.IMPORTANCE When bacteria become resistant to an antibiotic, the genetic changes involved sometimes increase (cross-resistance) or decrease (collateral sensitivity) their resistance to other antibiotics. Antibiotic combinations showing repeatable collateral sensitivity could be used in treatment to slow resistance evolution. However, collateral sensitivity interactions may depend on the local environmental conditions that bacteria experience, potentially reducing repeatability and clinical application. Here, we show that variation in local conditions (pH, temperature, and bile salts) can influence collateral sensitivity in two ways: by favoring different sets of mutants during bacterial resistance evolution (with different collateral sensitivities to other antibiotics) and by modifying expression of collateral effects for individual mutants. This suggests that translation from the lab to the clinic of new approaches exploiting collateral sensitivity will be influenced by local abiotic conditions.

New Therapeutic Strategy for Overcoming Multidrug Resistance in Cancer Cells with Pyrazolo[3,4-d]pyrimidine Tyrosine Kinase Inhibitors.

Simple SummaryP-glycoprotein (P-gp) is an ATP-binding cassette transporter whose overexpression in cancer cells is one of the main causes of multidrug resistance (MDR). Tyrosine kinase inhibitors (TKIs) have been reported to interact with ABC transporters and in some cases, increase the susceptibility of cancer cells to chemotherapy. We investigated the potential of novel TKI pyrazolo[3,4-d] pyrimidines and their prodrugs to inhibit P-gp in two MDR cancer cell lines with P-gp overexpression. The tested compounds were able to suppress P-gp by inhibiting its ATPase activity. Interestingly, prodrugs displayed a stronger potential to modulate P-gp and showed higher interaction energies in the docking simulations compared to their parent drugs. Furthermore, prodrugs showed significant potential to inhibit P-gp activity even in prolonged treatment and therefore to enhance the efficacy of doxorubicin and paclitaxel in MDR cancer cells. All of these characteristics imply that the new TKIs could be considered a valuable strategy for combating resistant cancers, especially in combination with other chemotherapeutics.Tyrosine kinase inhibitors (TKIs) often interact with the multidrug resistant (MDR) phenotype of cancer cells. In some cases, TKIs increase the susceptibility of MDR cancer cells to chemotherapy. As the overexpression of membrane transporter P-glycoprotein (P-gp) is the most common alteration in MDR cancer cells, we investigated the effects of TKI pyrazolo[3,4-d]pyrimidines on P-gp inhibition in two cellular models comprising sensitive and corresponding MDR cancer cells (human non-small cell lung carcinoma and colorectal adenocarcinoma). Tested TKIs showed collateral sensitivity by inducing stronger inhibition of MDR cancer cell line viability. Moreover, TKIs directly interacted with P-gp and inhibited its ATPase activity. Their potential P-gp binding site was proposed by molecular docking simulations. TKIs reversed resistance to doxorubicin and paclitaxel in a concentration-dependent manner. The expression studies excluded the indirect effect of TKIs on P-gp through regulation of its expression. A kinetics study showed that TKIs decreased P-gp activity and this effect was sustained for seven days in both MDR models. Therefore, pyrazolo[3,4-d]pyrimidines with potential for reversing P-gp-mediated MDR even in prolonged treatments can be considered a new therapeutic strategy for overcoming cancer MDR.

Diterpenoids from Euphorbia glomerulans with potential reversal activities against P-glycoprotein-mediated multidrug resistance

The development of collateral sensitivity agents that are able to modulate P-glycoprotein (P-gp) is the most promising approaches to overcome multidrug resistance (MDR) in cancer. In this study, eight new diterpenoids of jatrophane and ingenane type, 1–8, and three known ones (9–11) were isolated from Euphorbia glomerulans. Their structures were elucidated by spectroscopic analysis and electronic circular dichroism (ECD) calculations. The MDR reversal activity evaluation of these isolates on breast cancer MCF-7/ADR cells demonstrated the four potent MDR modulators (3, 4, 5, and 9) with great chemoreversal ability and low cytotoxicity. The structure-activity relationship (SAR) analysis indicated that the presence of isobutanoyloxy group at C-8 significantly enhance reversal efficiency. Compound 5 exhibited high efficacy (EC50 = 159.5 nM) in reversing MDR resistance, being stronger than verapamil (EC50 = 302.9 nM). The MDR reversal mechanism assays revealed that 5 could promote the accumulation of Rh123 and DOX in drug-resistant cells in a certain dose-dependent manner, and inhibit P-gp transport function. In addition, the possible recognition mechanism of compound 5 and verapamil (VRP) with P-gp was predicted by molecular docking.

Identification of antibiotic collateral sensitivity and resistance interactions in population surveillance data.

BackgroundCollateral effects of antibiotic resistance occur when resistance to one antibiotic agent leads to increased resistance or increased sensitivity to a second agent, known respectively as collateral resistance (CR) and collateral sensitivity (CS). Collateral effects are relevant to limit impact of antibiotic resistance in design of antibiotic treatments. However, methods to detect antibiotic collateral effects in clinical population surveillance data of antibiotic resistance are lacking.ObjectivesTo develop a methodology to quantify collateral effect directionality and effect size from large-scale antimicrobial resistance population surveillance data.MethodsWe propose a methodology to quantify and test collateral effects in clinical surveillance data based on a conditional t-test. Our methodology was evaluated using MIC data for 419 Escherichia coli strains, containing MIC data for 20 antibiotics, which were obtained from the Pathosystems Resource Integration Center (PATRIC) database.ResultsWe demonstrate that the proposed approach identifies several antibiotic combinations that show symmetrical or non-symmetrical CR and CS. For several of these combinations, collateral effects were previously confirmed in experimental studies. We furthermore provide insight into the power of our method for multiple collateral effect sizes and MIC distributions.ConclusionsOur proposed approach is of relevance as a tool for analysis of large-scale population surveillance studies to provide broad systematic identification of collateral effects related to antibiotic resistance, and is made available to the community as an R package. This method can help mapping CS and CR, which could guide combination therapy and prescribing in the future.

Design principles of collateral sensitivity-based dosing strategies

Collateral sensitivity (CS)-based antibiotic treatments, where increased resistance to one antibiotic leads to increased sensitivity to a second antibiotic, may have the potential to limit the emergence of antimicrobial resistance. However, it remains unclear how to best design CS-based treatment schedules. To address this problem, we use mathematical modelling to study the effects of pathogen- and drug-specific characteristics for different treatment designs on bacterial population dynamics and resistance evolution. We confirm that simultaneous and one-day cycling treatments could supress resistance in the presence of CS. We show that the efficacy of CS-based cycling therapies depends critically on the order of drug administration. Finally, we find that reciprocal CS is not essential to suppress resistance, a result that significantly broadens treatment options given the ubiquity of one-way CS in pathogens. Overall, our analyses identify key design principles of CS-based treatment strategies and provide guidance to develop treatment schedules to suppress resistance.

Chemoreversal Agents from Taiwanofungus Genus and Their More Potent Methyl Derivatives Targeting Signal Transducer and Activator of Transcription 3 (STAT3) Phosphorylation.

Multidrug resistance (MDR), for which the mechanisms are not yet fully clear, is one of the major obstacles to cancer treatment. In recent years, signal transducer and activator of transcription 3 (STAT3) were found to be one of the important MDR mechanism pathways. Based on the previous research, zhankuic acid A, B, and C were found to have collateral sensitivity effects on MDR cancer cells, and MDR inhibitory activity of zhankuic acid methyl ester was found to be better than that of its acid. Therefore, we executed a systematic examination of the structure–activity relationship of zhankuic acid methyl ester derivatives to collateral sensitivity in MDR cancer cells. The results showed that compound 12 is the best in terms of chemoreversal activity, where the reversal fold was 692, and the IC50 value of paclitaxel combined with 10 μM compound 12 treatment was 1.69 nM in MDR KBvin cells. Among all the derivatives, methyl ester compounds were found to be better than their acids, and a detailed discussion of the structure–activity relationships of all of the derivatives is provided in this work. In addition, compounds 8, 12, and 26 were shown to influence the activation of STAT3 in KBvin cells, accounting for part of their chemoreversal effects. Our results may provide a new combined therapy with paclitaxel to treat multidrug-resistant cancers and provide a new therapy option for patients.

Metal- and metalloid-based compounds to target and reverse cancer multidrug resistance.

Drug resistance remains the major cause of cancer treatment failure especially at the late stage of the disease. However, based on their versatile chemistry, metal and metalloid compounds offer the possibility to design fine-tuned drugs to circumvent and even specifically target drug-resistant cancer cells. Based on the paramount importance of platinum drugs in the clinics, two main areas of drug resistance reversal strategies exist: overcoming resistance to platinum drugs as well as multidrug resistance based on ABC efflux pumps. The current review provides an overview of both aspects of drug design and discusses the open questions in the field. The areas of drug resistance covered in this article involve: 1) Altered expression of proteins involved in metal uptake, efflux or intracellular distribution, 2) Enhanced drug efflux via ABC transporters, 3) Altered metabolism in drug-resistant cancer cells, 4) Altered thiol or redox homeostasis, 5) Altered DNA damage recognition and enhanced DNA damage repair, 6) Impaired induction of apoptosis and 7) Altered interaction with the immune system. This review represents the first collection of metal (including platinum, ruthenium, iridium, gold, and copper) and metalloid drugs (e.g. arsenic and selenium) which demonstrated drug resistance reversal activity. A special focus is on compounds characterized by collateral sensitivity of ABC transporter-overexpressing cancer cells. Through this approach, we wish to draw the attention to open research questions in the field. Future investigations are warranted to obtain more insights into the mechanisms of action of the most potent compounds which target specific modalities of drug resistance.

Exploring the Monoterpene Indole Alkaloid Scaffold for Reversing P-Glycoprotein-Mediated Multidrug Resistance in Cancer

Dregamine (1), a major monoterpene indole alkaloid isolated from Tabernaemontana elegans, was submitted to chemical transformation of the ketone function, yielding 19 azines (3–21) and 11 semicarbazones (22–32) bearing aliphatic or aromatic substituents. Their structures were assigned mainly by 1D and 2D NMR (COSY, HMQC, and HMBC) experiments. Compounds 3–32 were evaluated as multidrug resistance (MDR) reversers through functional and chemosensitivity assays in a human ABCB1-transfected mouse T-lymphoma cell model, overexpressing P-glycoprotein. A significant increase of P-gp inhibitory activity was observed for most derivatives, mainly those containing azine moieties with aromatic substituents. Compounds with trimethoxyphenyl (17) or naphthyl motifs (18, 19) were among the most active, exhibiting strong inhibition at 0.2 µM. Moreover, most of the derivatives showed selective antiproliferative effects toward resistant cells, having a collateral sensitivity effect. In drug combination assays, all compounds showed to interact synergistically with doxorubicin. Selected compounds (12, 17, 18, 20, and 29) were evaluated in the ATPase activity assay, in which all compounds but 12 behaved as inhibitors. To gather further insights on drug–receptor interactions, in silico studies were also addressed. A QSAR model allowed us to deduce that compounds bearing bulky and lipophilic substituents were stronger P-gp inhibitors.

The roles of history, chance, and natural selection in the evolution of antibiotic resistance.

History, chance, and selection are the fundamental factors that drive and constrain evolution. We designed evolution experiments to disentangle and quantify effects of these forces on the evolution of antibiotic resistance. Previously, we showed that selection of the pathogen Acinetobacter baumannii in both structured and unstructured environments containing the antibiotic ciprofloxacin produced distinct genotypes and phenotypes, with lower resistance in biofilms as well as collateral sensitivity to β-lactam drugs (Santos-Lopez et al., 2019). Here we study how this prior history influences subsequent evolution in new β-lactam antibiotics. Selection was imposed by increasing concentrations of ceftazidime and imipenem and chance differences arose as random mutations among replicate populations. The effects of history were reduced by increasingly strong selection in new drugs, but not erased, at times revealing important contingencies. A history of selection in structured environments constrained resistance to new drugs and led to frequent loss of resistance to the initial drug by genetic reversions and not compensatory mutations. This research demonstrates that despite strong selective pressures of antibiotics leading to genetic parallelism, history can etch potential vulnerabilities to orthogonal drugs.

Acquired resistance to PRMT5 inhibition induces concomitant collateral sensitivity to paclitaxel

Epigenetic regulators play key roles in cancer and are increasingly being targeted for treatment. However, for many, little is known about mechanisms of resistance to the inhibition of these regulators. We have generated a model of resistance to inhibitors of protein arginine methyltransferase 5 (PRMT5). This study was conducted in KrasG12D;Tp53-null lung adenocarcinoma (LUAD) cell lines. Resistance to PRMT5 inhibitors (PRMT5i) arose rapidly, and barcoding experiments showed that this resulted from a drug-induced transcriptional state switch, not selection of a preexisting population. This resistant state is both stable and conserved across variants arising from distinct LUAD lines. Moreover, it brought with it vulnerabilities to other chemotherapeutics, especially the taxane paclitaxel. This paclitaxel sensitivity depended on the presence of stathmin 2 (STMN2), a microtubule regulator that is specifically expressed in the resistant state. Remarkably, STMN2 was also essential for resistance to PRMT5 inhibition. Thus, a single gene is required for both acquisition of resistance to PRMT5i and collateral sensitivity to paclitaxel in our LUAD cells. Accordingly, the combination of PRMT5i and paclitaxel yielded potent and synergistic killing of the murine LUAD cells. Importantly, the synergy between PRMT5i and paclitaxel also extended to human cancer cell lines. Finally, analysis of The Cancer Genome Atlas patient data showed that high STMN2 levels correlate with complete regression of tumors in response to taxane treatment. Collectively, this study reveals a recurring mechanism of PRMT5i resistance in LUAD and identifies collateral sensitivities that have potential clinical relevance.

Laboratory evolution of Mycobacterium on agar plates for analysis of resistance acquisition and drug sensitivity profiles

Drug-resistant tuberculosis (TB) is a growing public health problem. There is an urgent need for information regarding cross-resistance and collateral sensitivity relationships among drugs and the genetic determinants of anti-TB drug resistance for developing strategies to suppress the emergence of drug-resistant pathogens. To identify mutations that confer resistance to anti-TB drugs in Mycobacterium species, we performed the laboratory evolution of nonpathogenic Mycobacterium smegmatis, which is closely related to Mycobacterium tuberculosis, against ten anti-TB drugs. Next, we performed whole-genome sequencing and quantified the resistance profiles of each drug-resistant strain against 24 drugs. We identified the genes with novel meropenem (MP) and linezolid (LZD) resistance-conferring mutation, which also have orthologs, in M. tuberculosis H37Rv. Among the 240 possible drug combinations, we identified 24 pairs that confer cross-resistance and 18 pairs that confer collateral sensitivity. The acquisition of bedaquiline or linezolid resistance resulted in collateral sensitivity to several drugs, while the acquisition of MP resistance led to multidrug resistance. The MP-evolved strains showed cross-resistance to rifampicin and clarithromycin owing to the acquisition of a mutation in the intergenic region of the Rv2864c ortholog, which encodes a penicillin-binding protein, at an early stage. These results provide a new insight to tackle drug-resistant TB.

Identification of antibiotic pairs that evade concurrent resistance via a retrospective analysis of antimicrobial susceptibility test results

SummaryBackgroundSome antibiotic pairs display a property known as collateral sensitivity in which the evolution of resistance to one antibiotic increases sensitivity to the other. Alternating between collaterally sensitive antibiotics has been proposed as a sustainable solution to the problem of antibiotic resistance. We aimed to identify antibiotic pairs that could be considered for treatment strategies based on alternating antibiotics.MethodsWe did a retrospective analysis of 448 563 antimicrobial susceptibility test results acquired over a 4-year period (Jan 1, 2015, to Dec 31, 2018) from 23 hospitals in the University of Pittsburgh Medical Center (Pittsburgh, PA, USA) hospital system. We used a score based on mutual information to identify pairs of antibiotics displaying disjoint resistance, wherein resistance to one antibiotic is commonly associated with susceptibility to the other and vice versa. We applied this approach to the six most frequently isolated bacterial pathogens (Escherichia coli, Staphylococcus aureus, Klebsiella pneumoniae, Enterococcus faecalis, Pseudomonas aeruginosa, and Proteus mirabilis) and subpopulations of each created by conditioning on resistance to individual antibiotics. To identify higher-order antibiotic interactions, we predicted rates of multidrug resistance for triplets of antibiotics using Markov random fields and compared these to the observed rates.FindingsWe identified 69 antibiotic pairs displaying varying degrees of disjoint resistance for subpopulations of the six bacterial species. However, disjoint resistance was rarely conserved at the species level, with only 6 (0·7%) of 875 antibiotic pairs showing evidence of disjoint resistance. Instead, more than half of antibiotic pairs (465 [53·1%] of 875) exhibited signatures of concurrent resistance, whereby resistance to one antibiotic is associated with resistance to another. We found concurrent resistance to extend to more than two antibiotics, with observed rates of resistance to three antibiotics being higher than predicted from pairwise information alone.InterpretationThe high frequency of concurrent resistance shows that bacteria have means of counteracting multiple antibiotics at a time. The almost complete absence of disjoint resistance at the species level implies that treatment strategies based on alternating between antibiotics might require subspecies level pathogen identification and be limited to a few antibiotic pairings.FundingUS National Institutes of Health.

The phenothiazine, trifluoperazine, is selectively lethal to ABCB1-expressing multidrug resistant cells

P-glycoprotein, member of the B-subfamily of the ATP-binding cassette (ABC) superfamily (e.g., ABCB1), has been demonstrated to confer resistance to clinically relevant anticancer drugs. Paradoxically, ABCB1-expressing multidrug resistant (MDR) cells are hypersensitivity or collateral sensitivity to non-toxic drugs. In this report, we demonstrate the capacity of trifluoperazine (TFP), a calmodulin inhibitor, to confer a collateral sensitivity onto ABCB1-overexpressing MDR cells. We show TFP-induced collateral sensitivity to be linked to ABCB1 expression and ATPase activity, as such phenotype is abolished in ABCB1-knockout MDR cells (CHORC5ΔABCB1 clones A1-A3) or with inhibitors of ABCB1 ATPase. TFP-induced collateral sensitivity is mediated by apoptotic cell death, due to enhanced oxidative stress. The findings in this study show for first time the use TFP as a collateral sensitivity drug, at clinically relevant concentrations. Moreover, given the use of trifluoperazine in the treatment for symptoms of schizophrenia and the role of ABCB1 transporter in tissue blood barriers and other physiologic functions, the finding in this study may have implications beyond cancer chemotherapy.

RASAL2 Confers Collateral MEK/EGFR Dependency in Chemoresistant Triple-Negative Breast Cancer.

While chemotherapy remains the standard treatment for triple-negative breast cancer (TNBC), identifying and managing chemoresistant tumors has proven elusive. We sought to discover hallmarks and therapeutically actionable features of refractory TNBC through molecular analysis of primary chemoresistant TNBC specimens. We performed transcriptional profiling of tumors from a phase II clinical trial of platinum chemotherapy for advanced TNBC (TBCRC-009), revealing a gene expression signature that identified de novo chemorefractory tumors. We then employed pharmacogenomic data mining, proteomic and other molecular studies to define the therapeutic vulnerabilities of these tumors. We reveal the RAS-GTPase-activating protein (RAS-GAP) RASAL2 as an upregulated factor that mediates chemotherapy resistance but also an exquisite collateral sensitivity to combination MAP kinase kinase (MEK1/2) and EGFR inhibitors in TNBC. Mechanistically, RASAL2 GAP activity is required to confer kinase inhibitor sensitivity, as RASAL2-high TNBCs sustain basal RAS activity through suppression of negative feedback regulators SPRY1/2, together with EGFR upregulation. Consequently, RASAL2 expression results in failed feedback compensation upon co-inhibition of MEK1/2 and EGFR that induces synergistic apoptosis in vitro and in vivo. In patients with TNBC, high RASAL2 levels predict clinical chemotherapy response and long-term outcomes, and are associated via direct transcriptional regulation with activated oncogenic Yes-Associated Protein (YAP). Accordingly, chemorefractory patient-derived TNBC models exhibit YAP activation, high RASAL2 expression, and tumor regression in response to MEK/EGFR inhibitor combinations despite well-tolerated intermittent dosing. These findings identify RASAL2 as a mediator of TNBC chemoresistance that rewires MAPK feedback and cross-talk to confer profound collateral sensitivity to combination MEK1/2 and EGFR inhibitors.

Antibiotic Cycling Reverts Extensive Drug Resistance in Burkholderia multivorans.

Antibiotic collateral sensitivity, in which acquired resistance to one drug leads to decreased resistance to a different drug, occurs in Burkholderia multivorans. Here, we observed that treatment of extensively drug-resistant variants evolved from a cystic fibrosis (CF) sputum sample isolate with either meropenem or sulfamethoxazole-trimethoprim, depending on past resistance phenotypes, resulted in increased sensitivity to five different classes of antibiotics. We further identified mutations, including putative resistance-nodulation-division efflux pump regulators and uncharacterized pumps, that may be involved in this phenotype in B. multivorans.