2. Translating Chemoresistant Vulnerabilities—Conceptual and Technological Strategies

The majority of the studies presented in this review harness the concept of collateral sensitivity as a type of synthetic lethality that could be potentially exploited in cancer therapy. The idea that resistance towards a drug must inevitably come accompanied by an increased sensitivity to other compounds has led to the search of these acquired vulnerabilities in cancer cell lines resistant to chemotherapeutic agents, as well as to targeted drugs. Collateral sensitivities have been reported with cancer chemotherapeutics, including an increased sensitivity to microtubule inhibitors, cisplatin or cytarabine in paclitaxel-resistant CHO (Chinese hamster ovary) cells [195]. For example, Jensen et al. screened a panel of SCLC cell lines resistant to different chemotherapy agents and tested their sensitivity to 20 additional drugs [196]. With this approach, they identified cross-resistance as well as collateral sensitivities. Interestingly, vulnerabilities were found for all the resistant cell lines tested, one of them being the previously reported higher sensitivity to cisplatin in paclitaxel-resistant cells. MDR is often caused by the overexpression of drug efflux pumps, such as the ABC transporter MRP1, and this overexpression could itself be exploited as a collateral sensitivity that can be associated with the depletion of critical intracellular compounds such as glutathione, ATP and metals. MRP1-overexpressing cells usually show reduced levels of glutathione (GSH) [197,198,199], which can be additionally decreased using other therapeutic agents, such as verapamil [200,201], that further increase GSH efflux, leading to oxidative stress and apoptosis [202,203]. Oxidative stress induced by high levels of ROS is characteristic of chemotherapy-treated cells [187,204,205], and several redox-regulating enzymes could be potentially targeted, including mitochondrial electron transporters and GSH metabolism-related enzymes (reviewed in [206]). Another approach to exploit this acquired vulnerability is the “one-two-punch” strategy, elaborated by the Bernards’ group [5]. Similar to chemotherapy-resistant cells, melanoma cells resistant to the BRAF inhibitor vemurafenib have increased levels of ROS, which can be additionally increased by using the histone deacetylase inhibitor vorinostat. This sequential treatment leads to much higher levels of ROS in the resistant cells, causing severe DNA damage and apoptosis. The clinical efficacy of the “one-two-punch” hypothesis is currently evaluated with vorinostat treatment in patients with resistant BRAF V600E mutated advanced melanoma [207], and the results will hopefully prove that the concept is clinically relevant.

These discussed studies also demonstrate the usefulness of genetic perturbation screens in the investigation of acquired vulnerabilities in chemotherapy-resistant cancer cells. Many of the hits found to act synthetic lethal with the drugs in the resistant cells pertain to cell proliferation, DNA damage repair or the apoptosis pathways, which makes them potentially targetable with inhibitors of these signaling pathways. For instance, CHEK1 is a commonly found hit in screens that search for genes that could render cells more susceptible to gemcitabine treatment when downregulated. In fact, it has been shown that CHEK1 can be targeted with the inhibitor AZD7762 to sensitize pancreatic cancer cell lines to gemcitabine both in vitro and in vivo [208,209]. Apoptotic pathway alteration is another commonly found feature in chemoresistant cells, usually evidenced by an increased expression of the anti-apoptotic protein Bcl-2 [194,210,211]. Under chemotherapy selective pressure, tumor cells can become “addicted” to Bcl-2 expression for survival, so an obvious strategy to overcome this resistance mechanism may be the use of Bcl-2 inhibitors (reviewed in [212,213]). Pearce et al. showed that the increased expression of Bcl-2 in paclitaxel-resistant lung cancer cells could be exploited by a different approach [214]. Bcl-2 can be converted from an anti-apoptotic protein to a pro-apoptotic one through interaction with Nur77 in the mitochondria. The authors demonstrated that Bcl-2 can be pharmacologically targeted with a peptide that mimics Nur77, preferentially inducing apoptosis in Bcl-2 overexpressing paclitaxel-resistant cells and inhibiting tumor growth in a zebrafish xenograft model.

Most unbiased gene perturbation efforts investigate what would resemble intrinsic resistance mechanisms in the sense that the genetic perturbation is introduced prior to short-term exposure to a given drug. However, searching for targetable collateral sensitivities that emerge as a result of long-term drug exposure might be more beneficial, as these model system likely more accurately mimic the heterogenous chemoresistance profiles found in patients. It is also of great importance to be able to prioritize cancer therapeutic targets, and this should be done on the basis of context-penetrating mechanisms of resistance and/or vulnerabilities. In an effort to link drug response to genomic alterations, Garnett et al. performed a systematic screen in a panel of 639 cancer cell lines with 130 targeted agents and chemotherapeutic drugs [215]. As expected, strong associations were found for oncogenes that are targets of their specific drug, but they also found marked sensitizing genotypes in small subsets of cell lines, highlighting the need for large panels of cell lines to uncover potentially relevant genomic features. To address the above-mentioned aspects, a larger number of drug-resistant cell lines pertaining to different cancer entities should be investigated systematically, with the “The Resistant Cancer Cell Line (RCCL) collection” representing a valuable resource that awaits its comprehensive genetic characterization (https://www.wass-michaelislab.org/rccl.php). Another important aspect is that resistance rarely emerges from a single mechanism. More often, several co-occurring gain-of-function and/or loss-of-function mechanisms synergistically contribute to a patient’s chemoresistance profile. In this context, technologies for the robust performance of genetic multiplexing approaches for the simultaneous perturbation of multiple loci are needed [73,216]. While these combinatorial approaches gain momentum in the identification of synthetic gene interactions, their general application and analysis are far from standardized. However, these approaches hold great potential to uncover druggable vulnerabilities otherwise missed in monogenic screens due to functional buffering effects [217].