Effective RNA Knockdown Using CRISPR-Cas13a and Molecular Targeting of the EML4-ALK Transcript in H3122 Lung Cancer Cells

RNAi technology has significant potential as a future therapeutic and could theoretically be used to knock down disease-specific RNAs. However, due to frequent off-target effects, low efficiency, and limited accessibility of nuclear transcripts, the clinical application of the technology remains challenging. In this study, we first assessed the stability of Cas13a mRNA and guide RNA. Next, we titrated Cas13a and guide RNA vectors to achieve effective knockdown of firefly luciferase (FLuc) RNA, used as a target transcript. The interference specificity of Cas13a on guide RNA design was next explored. Subsequently, we targeted the EML4-ALK v1 transcript in H3122 lung cancer cells. As determined by FLuc assay, Cas13a exhibited activity only toward the orientation of the crRNA–guide RNA complex residing at the 5′ of the crRNA. The activity of Cas13a was maximal for guide RNAs 24–30 bp in length, with relatively low mismatch tolerance. After knockdown of the EML4-ALK transcript, cell viabili..... READ ARTICLE

International Journal of Molecular Sciences DOI:10.3390/ijms21238904

Authors: Saifullah, Sakari M, Suzuki T, Yano S, Tsukahara T.

Pre-ClinicalKirk SmithNovember 24, 2020

Discovery of CJ-2360 as a Potent and Orally Active Inhibitor of Anaplastic Lymphoma Kinase Capable of Achieving Complete Tumor Regression

We report herein the discovery of a class of potent small-molecule inhibitors of anaplastic lymphoma kinase (ALK) containing a fused indoloquinoline scaffold. The most promising compound CJ-2360 has an IC50 value of 2.2 nM against wild-type ALK and low-nanomolar potency against several clinically reported ALK mutants. This compound is capable of achieving complete tumor regression in the ALK-positive KARPAS-299 xenograft model with oral administration in mice. CJ-2360 represents a promising ALK inhibitor for advanced preclinical development. READ ARTICLE

Journal of Medicinal Chemistry DOI:10.1021/acs.jmedchem.0c01550

Authors: Jianyong Chen, Yunlong Zhou, Xuyuan Dong, Liu Liu, Longchuan Bai, Donna McEachern, Sally Przybranowski, Chao-Yie Yang, Jeanne Stuckey, Xiaoqin Li, Bo Wen, Ting Zhao, Siwei Sun, Duxin Sun, Lingling Jiao, Yu Jing, Ming Guo, Dajun Yang, and Shaomeng Wang

Pre-ClinicalKirk SmithNovember 13, 2020Rodent models, Inhibitors, Inhibition, Peptides and proteins, Tumors

Longitudinal therapy monitoring of ALK-positive lung cancer by combined copy number and targeted mutation profiling of cell-free DNA

Combined copy number and targeted mutation profiling could improve monitoring of ALK+ NSCLC. Potential advantages include the identification of treatment failure, in particular for patients without detectable mutations, and broader detection of genomic changes acquired during therapy, especially in later treatment lines and in high-risk patients. READ ARTICLE

EBioMedicine DOI: 10.1016/j.ebiom.2020.103103

Authors: Steffen Dietz, Petros Christopoulos, Zhao Yuan, Arlou Kristina Angeles, Lisa Gu, Anna-Lena Volckmar, Simon J. Ogrodnik, Florian Janke, Chiara Dalle Fratte, Tomasz Zemojtel, Marc A. Schneider, Daniel Kazdal, Volker Endris, Michael Meister, Thomas Muley, Erika Cecchin, Martin Reck, Matthias Schlesner, Michael Thomas, Albrecht Stenzinger, Holger Sültmann,

Dual Mechanisms of Novel CD73-Targeted Antibody and Antibody–Drug Conjugate in Inhibiting Lung Tumor Growth and Promoting Antitumor Immune-Effector Function

We found elevated CD73 expression to be prevalent in non–small cell lung cancer (NSCLC) including those harboring the RAS- or RTK (EGFR, EML4-ALK) oncogenes. CD73 expression is enriched closely with the transcriptome signature of epithelial–mesenchymal transition and the immune-tolerant tumor microenvironment, which are increasingly relevant for disease progression and therapy resistance. We developed two novel series of CD73 antibody, Ab001/Ab002 and humanized version Hu001/Hu002, which demonstrated high CD73 binding affinity, potent enzyme inhibition, and efficiently protected effector T lymphocyte function from adenosine/cancer-imposed toxicity. Hu001/Hu002 inhibited growth of RAS-mutant NSCLC tumors in mice via enhanced antibody-dependent cell-mediated cytotoxicity and multifaceted remodeling of the tumor immune environment, reflecting diminished levels of tumor-associated macrophages, myeloid-derived suppressor cells, and tumor vasculature. A novel MMAE-conjugated CD73-ADC (Hu001–..... READ ARTICLE

Molecular Cancer Therapeutics DOI: 10.1158/1535-7163.MCT-20-0076

Authors: Rui Jin, Liang Liu, Yun Xing, Tao Meng, Lanping Ma, Jinpeng Pei, Ying Cong, Xuesai Zhang, Zhiqiang Ren, Xin Wang, Jingkang Shen and Ker Yu

Pre-ClinicalKirk SmithNovember 1, 2020Antibody-Drug Conjugates (ADC)

Dissecting the landscape of CAF-mediated drug resistance mechanisms in ALK-rearranged NSCLC

Cancer-associated fibroblasts (CAFs) are predominant stromal cells associated with cancer development and drug resistance. Due to the complexity of the crosstalk between CAFs and cancer cells, the underlying mechanism often remains elusive. Here, we aim to elucidate the complex, bidirectional signaling network that governs CAF-mediated resistance to targeted drugs in EML4-ALK-rearranged non-small-cell lung cancer (NSCLC) cells. READ ARTICLE

European Journal of Cancer DOI:10.1016/S0959-8049(20)31205-3

Authors: Q. Hu, L.L. Remsing Rix, X. Li, E.A. Welsh, B. Fang, S. Yun, J. Kroeger, H.R. Lawrence, A. Marusyk, J.M. Koomen, E.B. Haura, U. Rix,

Pre-ClinicalKirk SmithOctober 26, 2020CAF-mediated resistance, ALK, NSCLC

Fragment-based modification of 2,4-diarylaminopyrimidine derivatives as ALK and ROS1 dual inhibitors to overcome secondary mutants

In order to explore novel ALK and ROS1 dual inhibitors capable of overcoming crizotinib-resistant mutants, two series of 2,4-diarylaminopyrimidine derivatives were designed, synthesized and evaluated for their in vitro cytotoxic activity. In this work, we retained the 2,4-diarylaminopyrimidine scaffold and derivatize the DAAP scaffold with sulfonyl and acrylamide moieties to extend the structure–activity relationship (SAR) study. To our delight, some compounds exhibited excellent inhibitory activity with a double-digit nanomolar level in MTT assay. Four compounds were selected for enzymic assays further, the results led to the identification of a potent ALK and ROS1 dual inhibitor X-17, with IC50 values of 3.7 nM, 2.3 nM, 8.9 nM and 1.9 nM against ALK, ALKL1196M, ALKG1202R and ROS1, respectively. Ultimately, the molecular docking studies on X-17 clearly disclosed reasonable and optimal binding interactions with ALK. READ ARTICLE

Bioorganic & Medicinal Chemistry DOI:10.1016/j.bmc.2020.115719

Authors: Minglin Zhu, Wei Li, Tianming Zhao, Yuxiang Chen, Tong Li, Shangfei Wei, Ming Guo, Xin Zhai

Pre-ClinicalKirk SmithOctober 15, 20202, 4-Diarylaminopyrimidine, Dual inhibitor, ALK, ROS1, Crizotinib-resistant

Pharmacological inhibition of the ALK axis elicits therapeutic potential in Consensus Molecular Subtype 1 colon cancer patients

In the last years, several efforts have been made to classify colorectal cancer (CRC) into well-defined molecular subgroups, representing the intrinsic inter-patient heterogeneity, known as Consensus Molecular Subtypes (CMSs). In this work, we performed a meta-analysis of 1700 CRC patients stratified into four CMSs. We identified a negative correlation between a high level of anaplastic lymphoma kinase (ALK) expression and relapse-free survival, exclusively in CMS1 subtype. Stemming from this observation, we tested several CMSs in vitro models with crizotinib (CZB) or alectinib (ALC), potent ALK inhibitors, already approved for clinical use. ALK interception strongly inhibits cell proliferation already at nanomolar doses, specifically in CMS1 cell lines, while no effect was found in CMS2/3/4 groups. Furthermore, in vivo imaging identified a role for ALK in the dynamic formation of 3D spheroids, which was impaired by the pharmacological inhibition of ALK. Consistently, CZB was responsib..... READ ARTICLE

BioRxiv DOI:10.1101/2020.10.07.307991

Authors: Martina Mazzeschi, Michela Sgarzi, Donatella Romaniello, Valerio Gelfo, Carola Cavallo, Spartaco Santi, Michelangelo Fiorentino, Gabriele D’Uva, Balázs Győrffy, Ruth Palmer, Mattia Lauriola

Prevalence and potential biological role ofTERTamplifications inALKtranslocated adenocarcinoma of the lung

The advent of specific ALK-targeting drugshas radically changed the outcome of patients withALKtranslocated non-small-cell lung cancer(NSCLC). However, emerging resistance to treatmentwithALKinhibitors in these patients remains amajor concern. In previous studies, we analysed twoALK+patient cohorts (TP53wild-type/TP53mutated) in terms of copy number alterations. Allpatients belonging to theTP53wild-type group hadmainly genetically stable genomes, with one excep-tion showing chromosomal instability and amplifica-tions of several gene loci, includingTERT. Here, weaimed to determine the prevalence ofTERTamplifi-cations in theseALK+lung cancer patients by ana-lysing an independent cohort of 109ALKtranslocated cases. We further analysed the copy numbers of numerous cancer-relevant genes andother genetic aberrations. The prevalence ofTERTamplifi-cations was determined by means of FISH analyses.Copy numbers of 87 cancer-relevant genes were deter-mined by NanoString nCounterâtechnolo..... READ ARTICLE

Histopathology DOI:10.1111/his.14256

Authors: Alidousty C, Duerbaum N, Wagener-Ryczek S, Baar T, Martelotto L G, Heydt C, Siemanowski J, Holz B, Binot E,Fassunke J, Merkelbach-Bruse S, Wolf J, Kron A, Buettner R, Schultheis A M

Pre-ClinicalKirk SmithSeptember 18, 2020ALK, cancer, lung, TERT, genetic stability

ALK Inhibitors Do Not Increase Sensitivity to Radiation in EML4-ALK Non-small Cell Lung Cancer

ALK inhibitors like Crizotinib, Ceritinib and Alectinib are targeted therapies used in patients with anaplastic lymphoma kinase (ALK)-positive, advanced non-small cell lung cancer (NSCLC). Since in this tumor entity radiotherapy is employed sequentially or concomitantly, potential synergistic effects were investigated, which may support the hypothesis of induced radiosensitization by using ALK inhibitors. Two cell lines expressing wild-type (WT) or echinoderm microtubule-associated protein-like 4 (EML4)-ALK were treated with ALK inhibitors, followed by irradiation. Cell survival, cell death, cell cycle and phosphorylation of H2A histone family, member X (H2AX) were examined. Combined treatment with ALK-inhibitors plus 10 Gy-irradiation led to effects similar to those of sole radiotherapy, but was more effective than sole drug treatment. There is no clear evidence of sensitization to radiation by treating EML4-ALK mutated cells with ALK inhibitors. READ ARTICLE

Anticancer Research DOI:10.21873/anticanres.14497

Authors: Kathrin Fleschutz, Lisa Walter, Tumo Leistner, Lucie Heinzerling

Therapy-Induced Evolution of Human Lung Cancer Revealed by Single-Cell RNA Sequencing

Lung cancer, the leading cause of cancer mortality, exhibits heterogeneity that enables adaptability, limits therapeutic success, and remains incompletely understood. Single-cell RNA sequencing (scRNA-seq) of metastatic lung cancer was performed using 49 clinical biopsies obtained from 30 patients before and during targeted therapy. Over 20,000 cancer and tumor microenvironment (TME) single-cell profiles exposed a rich and dynamic tumor ecosystem. scRNA-seq of cancer cells illuminated targetable oncogenes beyond those detected clinically. Cancer cells surviving therapy as residual disease (RD) expressed an alveolar-regenerative cell signature suggesting a therapy-induced primitive cell-state transition, whereas those present at on-therapy progressive disease (PD) upregulated kynurenine, plasminogen, and gap-junction pathways. Active T-lymphocytes and decreased macrophages were present at RD and immunosuppressive cell states characterized PD. Biological features revealed by scRNA-seq were biomarkers of clinical outcomes in independent cohorts. This study highlights how therapy-induced adaptation of the multi-cellular ecosystem of metastatic cancer shapes clinical outcomes. READ ARTICLE

Cell DOI:10.1016/j.cell.2020.07.017

Authors: Ashley Maynard, Caroline E. McCoach, Julia K. Rotow, Lincoln Harris, Franziska Haderk, D. Lucas Kerr, Elizabeth A. Yu, Erin L. Schenk, Weilun Tan, Alexander Zee, Michelle Tan, Philippe Gui, Tasha Lea, Wei Wu, Anatoly Urisman, Kirk Jones, Rene Sit, Pallav K. Kolli, Eric Seeley, Yaron Gesthalter, Daniel D. Le, Kevin A. Yamauchi, David M. Naeger, Sourav Bandyopadhyay, Khyati Shah, Lauren Cech, Nicholas J. Thomas, Anshal Gupta, Mayra Gonzalez, Hien Do, Lisa Tan, Bianca Bacaltos, Rafael Gomez-Sjoberg, Matthew Gubens, Thierry Jahan, Johannes R. Kratz, David Jablons, Norma Neff, Robert C. Doebele, Jonathan Weissman, Collin M. Blakely, Spyros Darmanis, Trever G. Bivona

Pre-ClinicalKirk SmithAugust 20, 2020single-cell RNA, lung cancer, ALK, EGFR, Targeted therapy

Abstract 1865: Phosphoproteomic analyses identify EGFR as a critical cooperative kinase for ALK

In order to interrogate the contribution of EGFR to downstream signaling under ALK inhibition, we conducted phosphoproteomics for two ALK-driven cell lines, DFCI032 and H3122, using the ALK inhibitor crizotinib or in combination with the EGFR inhibitor gefitinib. Kinase-Substrate Enrichment Analysis (KSEA) and Phosphoproteomics Dissection Using Networks (PHOTON) analysis were used to identify whether a signaling node was functionally regulated based on substrate phosphorylation or on protein-protein interaction partner phosphorylation, respectively. Strikingly, both methods showed that EGFR was functionally downregulated by ALK inhibition. ALK was also considered functionally more downregulated under EGFR/ALK dual inhibition compared to ALK inhibition alone by KSEA analysis. These data suggest ALK and EGFR engage in overlapping signaling pathways. Collectively, EGFR and ALK dual inhibition can significantly delay or prevent the emergence of ALK TKI resistance by suppressing critical overlapping signaling pathways and promotes signaling reprogramming in cancer cells. READ ARTICLE

AACR abstract DOI: 10.1158/1538-7445.AM2020-1865

Authors: Nan Chen, Anh T. Le, Andrea E. Doak, Guolin Zhang, Bin Fang, Eric B. Haura and Robert C. Doebele

Pre-ClinicalKirk SmithAugust 1, 2020ALK, EGFR, proteomics

Genome-Scale CRISPR Screening reveals novel factors regulating Wnt-dependent regeneration of mouse gastric organoids

An ability to safely harness the powerful regenerative potential of adult stem cells for clinical applications is critically dependent on a comprehensive understanding of the underlying mechanisms regulating their activity. Epithelial organoid cultures accurately recapitulate many features of in vivo stem cell-driven epithelial regeneration, providing an excellent ex vivo platform for interrogation of key regulatory mechanisms. Here, we employed a Genome-Scale CRISPR Knock-Out (GeCKO) screening assay using mouse gastric epithelial organoids to identify novel modulators of Wnt-driven stem cell-dependent epithelial regeneration in the gastric mucosa. In addition to known Wnt pathway regulators such as Apc, we found that knock-out (KO) of Alk, Bclaf3 or Prkra supports the Wnt independent self-renewal of gastric epithelial cells ex vivo. In adult mice, expression of these factors is predominantly restricted to non Lgr5-expressing stem cell zones above the gland base, implicating a critical..... READ ARTICLE

BioRxiv DOI:10.1101/2020.07.17.208496

Authors: Kazuhiro Murakami, Yumi Terakado, Kikue Saito, Yoshie Jomen, Haruna Takeda, Masanobu Oshima, Nick Barker

Forced degradation of Ceritinib under stress conditions: Structural interpretation of novel degradants using HR-MS/MS and NMR

The current work describes the preparative isolation and structural interpretation of acid degradation product of ceritinib (CRB). The drug is exposed to different stress conditions (acid, base hydrolysis, oxidation, thermal and photolytic) as suggested by International Conference on Harmonisation guidelines (ICH). The degradation of drug (CRB) is observed when exposed to acidic condition whereas the drug is stable in remaining stress conditions. The separation between drug and acid degradant was attained on a C18 BEH UPLC column (50 mm × 2.1 mm, 1.7 μm) in a gradient separation mode. The mobile phase comprises of acetonitrile and 0.05% formic acid buffer at a flow rate of 0.6 mL/min and UV wavelength monitored at 215 nm. One novel degradation product formed in acidic condition was isolated by Preparative HPLC (Prep-HPLC). Structural interpretation and characterisation done by HR-MS/MS and NMR studies which is not yet described in the literature. READ ARTICLE

Materials Today: Proceedings DOI:10.1016/j.matpr.2020.01.588.

Authors: Soujanya Vajjha, Vijay Bommuluri, Chidananda Swamy Rumalla, Muralidharan Kaliyaperumal, Raghu Babu Korupolu, Vidyasagar chopella

Pre-ClinicalKirk SmithJuly 12, 2020Ceritinib, Drug degradation, Structural elucidation

ZX-29, a novel ALK inhibitor, induces apoptosis via ER stress in ALK rearrangement NSCLC cells and overcomes cell resistance caused by an ALK mutation

Although anaplastic lymphoma kinase (ALK) inhibitors have good clinical efficacy, the inevitable development of drug resistance is the most common obstacle to their clinical application. There is an urgent need to develop more effective and selective ALK inhibitors to overcome the problem of drug resistance. Here, we screened a series of ALK inhibitors and found that ZX-29 displayed potent cytotoxic activity against ALK rearrangement non-small cell lung cancer (NSCLC) NCI-H2228 cells. Then, we investigated the antitumor effects of ZX-29. We demonstrated that ZX-29 time- and dose-dependently inhibited the viability of NCI-H2228 cells, induced cell cycle arrest in the G1 phase, and then they subsequently progressed into cell death. The type of cell death induced by ZX-29 was apoptosis through endoplasmic reticulum (ER) stress. Interestingly, ZX-29 induced protective autophagy, and inhibiting autophagy could enhance the antitumor effect of ZX-29. Furthermore, ZX-29 suppressed tumor growth..... READ ARTICLE

Biochimica et Biophysica Acta (BBA) - Molecular Cell Research DOI:10.1016/j.bbamcr.2020.118712

Authors: Wenfeng Gou, Zengqiang Li, Xiaobo Xu, Jiwei Shen, Ming Guo, Xuejiao Zhou, Xiaoning Zhang, Yingliang Wu, Xin Zhai, Daiying Zuo

Pre-ClinicalKirk SmithJuly 1, 2020ALK, NSCLC, ER stress, Apoptosis, Drug resistance

Vulnerability of drug-resistant EML4-ALK rearranged lung cancer to transcriptional inhibition

A subset of lung adenocarcinomas is driven by the EML4-ALK translocation. Even though ALK inhibitors in the clinic lead to excellent initial responses, acquired resistance to these inhibitors due to on-target mutations or parallel pathway alterations is a major clinical challenge. Exploring these mechanisms of resistance, we found that EML4-ALK cells parental or resistant to crizotinib, ceritinib or alectinib are remarkably sensitive to inhibition of CDK7/12 with THZ1 and CDK9 with alvocidib or dinaciclib. These compounds robustly induce apoptosis through transcriptional inhibition and downregulation of anti-apoptotic genes. Importantly, alvocidib reduced tumour progression in xenograft mouse models. In summary, our study takes advantage of the transcriptional addiction hypothesis to propose a new treatment strategy for a subset of patients with acquired resistance to first-, second- and third-generation ALK inhibitors. READ ARTICLE

EMBO Molecular Medicine DOI:10.15252/emmm.201911099

Authors: Athanasios R Paliouras, Marta Buzzetti, Lei Shi, Ian J Donaldson, Peter Magee, Sudhakar Sahoo, Hui-Sun Leong, Matteo Fassan, Matthew Carter, Gianpiero Di Leva, Matthew G Krebs, Fiona Blackhall, Christine M Lovly, Michela Garofalo

Pre-ClinicalKirk SmithJune 17, 2020ALK/EML4 translocation, ALKi, CDKi, NSCLC, drug resistance

MET Alterations Are a Recurring and Actionable Resistance Mechanism in ALK-Positive Lung Cancer

MET amplification was detected in 15% of tumor biopsies from patients relapsing on next-generation ALK inhibitors, including 12% and 22% of biopsies from patients progressing on second-generation inhibitors or lorlatinib, respectively. Patients treated with a second-generation ALK inhibitor in the first-line setting were more likely to develop MET amplification than those who had received next-generation ALK inhibitors after crizotinib (P = 0.019). Two tumor specimens harbored an identical ST7-MET rearrangement, one of which had concurrent MET amplification. Expressing ST7-MET in the sensitive H3122 ALK-positive cell line induced resistance to ALK inhibitors that was reversed with dual ALK/MET inhibition. MET inhibition resensitized a patient-derived cell line harboring both ST7-MET and MET amplification to ALK inhibitors. Two patients with ALK-positive lung cancer and acquired MET alterations achieved rapid responses to ALK/MET combination therapy. Treatment with next-generation ALK inhibitors, particularly in the first-line setting, may lead to MET-driven resistance. Patients with acquired MET alterations may derive clinical benefit from therapies that target both ALK and MET. READ ARTICLE

Clinical Cancer Research DOI: 10.1158/1078-0432.CCR-19-3906

Authors: Ibiayi Dagogo-Jack, Satoshi Yoda, Jochen K. Lennerz, Adam Langenbucher, Jessica J. Lin, Marguerite M. Rooney, Kylie Prutisto-Chang, Audris Oh, Nathaniel A. Adams, Beow Y. Yeap, Emily Chin, Andrew Do, Hetal D. Marble, Sara E. Stevens, Subba R. Digumarthy, Ashish Saxena, Rebecca J. Nagy, Cyril H. Benes, Christopher G. Azzoli, Michael S. Lawrence, Justin F. Gainor, Alice T. Shaw, Aaron N. Hata

Pre-ClinicalKirk SmithJune 1, 2020MET, ALK+ lung cancer

HDAC inhibition synergizes with ALK inhibitors to overcome resistance in a novel ALK mutated lung adenocarcinoma model

Objectives: Somatic chromosomal rearrangements resulting in ALK fusion oncogenes are observed in 3–7 % of lung adenocarcinomas. ALK tyrosine kinase inhibitors (ALKi) induce initially response, however, various resistance mechanisms limit their efficacy. Novel therapeutic approaches are of utmost importance to tailor these targeted therapies... Conclusion: Using a patient-derived ALKi resistant lung cancer model we demonstrated the synergism of HDAC and ALK inhibition. Furthermore, our findings provide strong evidence for intratumoral heterogeneity under targeted therapy and highlight the importance of site-specific mutational analysis. READ ARTICLE

Lung Cancer DOI:10.1016/j.lungcan.2020.04.002

Authors: Paul Stockhammer, Cassandra Su Lyn Ho, Luca Hegedus, Gabor Lotz, Eszter Molnár, Agnes Bankfalvi, Thomas Herold, Stavros Kalbourtzis, Till Ploenes, Wilfried E. E. Eberhardt, Martin Schuler, Clemens Aigner, Alexander Schramm, Balazs Hegedus

Comprehensive analyses of immunodynamics and immunoreactivity in response to treatment in ALK-positive non-small-cell lung cancer

ALK-positive tumors progressing on ceritinib is not immunogenic enough to respond to immune checkpoint inhibitors. READ ARTICLE

Journal for ImmunoTherapy of Cancer DOI: 10.1136/jitc-2020-000970

Authors: Kyoung-Ho Pyo, Sun Min Lim, Chae-Won Park, Ha-Ni Jo, Jae Hwan Kim, Mi-Ran Yun, Dohee Kim, Chun-Feng Xin, Wongeun Lee, Bianca Gheorghiu, Min Hee Hong, Hye Ryun Kim, Hyo Sup Shim, Mi Jang, Sung Sook Lee, Byoung Chul Cho

Pre-ClinicalKirk SmithJune 1, 2020Pd-1 inhibitor, ceritinib

Resistance to targeted therapies as a multifactorial, gradual adaptation to inhibitor specific selective pressures

Despite high initial efficacy, targeted therapies eventually fail in advanced cancers, as tumors develop resistance and relapse. In contrast to the substantial body of research on the molecular mechanisms of resistance, understanding of how resistance evolves remains limited. Using an experimental model of ALK positive NSCLC, we explored the evolution of resistance to different clinical ALK inhibitors. We found that resistance can originate from heterogeneous, weakly resistant subpopulations with variable sensitivity to different ALK inhibitors. Instead of the commonly assumed stochastic single hit (epi) mutational transition, or drug-induced reprogramming, we found evidence for a hybrid scenario involving the gradual, multifactorial adaptation to the inhibitors through acquisition of multiple cooperating genetic and epigenetic adaptive changes. Additionally, we found that during this adaptation tumor cells might present unique, temporally restricted collateral sensitivities, absent in therapy naïve or fully resistant cells, suggesting the potential for new therapeutic interventions, directed against evolving resistance. READ ARTICLE

Nature Communications DOI:10.1038/s41467-020-16212-w

Authors: Robert Vander Velde, Nara Yoon, Viktoriya Marusyk, Arda Durmaz, Andrew Dhawan, Daria Miroshnychenko, Diego Lozano-Peral, Bina Desai, Olena Balynska, Jan Poleszhuk, Liu Kenian, Mingxiang Teng, Mohamed Abazeed, Omar Mian, Aik Choon Tan, Eric Haura, Jacob Scott & Andriy Marusyk

EML4–ALK V3 oncogenic fusion proteins promote microtubule stabilization and accelerated migration through NEK9 and NEK7

EML4–ALK is an oncogenic fusion present in ∼5% of non-small cell lung cancers. However, alternative breakpoints in the EML4 gene lead to distinct variants of EML4–ALK with different patient outcomes. Here, we show that, in cell models, EML4–ALK variant 3 (V3), which is linked to accelerated metastatic spread, causes microtubule stabilization, formation of extended cytoplasmic protrusions and increased cell migration. EML4–ALK V3 also recruits the NEK9 and NEK7 kinases to microtubules via the N-terminal EML4 microtubule-binding region. Overexpression of wild-type EML4, as well as constitutive activation of NEK9, also perturbs cell morphology and accelerates migration in a microtubule-dependent manner that requires the downstream kinase NEK7 but does not require ALK activity. Strikingly, elevated NEK9 expression is associated with reduced progression-free survival in EML4–ALK patients. Hence, we propose that EML4–ALK V3 promotes microtubule stabilization through NEK9 and NEK7, leading to..... READ ARTICLE

Journal of Cell Science DOI:10.1242/jcs.241505

Authors: Laura O'Regan, Giancarlo Barone, Rozita Adib, Chang Gok Woo, Hui Jeong Jeong, Emily L. Richardson, Mark W. Richards, Patricia A. J. Muller, Spencer J. Collis, Dean A. Fennell, Jene Choi, Richard Bayliss, Andrew M. Fry

Pre-ClinicalKirk SmithMay 11, 2020EML4–ALK, EML4, NEK9, NEK7, NSCLC, Microtubules, Cell migration, Metastasis, versions