Defining the sensitivity landscape of EGFR variants to tyrosine kinase inhibitors

An, Li; WANG, YUEQIANG; Wu, Guangyao; WANG, ZHENXING; Shi, Zeyuan; liu, chang; Wang, Chunli; Yi, Ming; Niu, Chenguang; Duan, Shaofeng; Li, Xiaodong; Tang, Wenxue; Wu, Kongming; CHEN, SHUQING; Xu, Hongen

doi:10.1016/j.trsl.2022.11.002

Cited by 16 publications

(8 citation statements)

References 45 publications

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“…Another high-throughput method to solve VUSs is deep mutational scanning (DMS) [ 101 ], which is possible due to the development of synthetic DNA technology and the decreasing sequencing costs. DMS technology has been widely used to study the function of variants in clinically actionable genes, such as BRCA1 [ 102 ], PPARG [ 103 ], TP53 [ 104 ], NUDT15 [ 105 ], MSH2 [ 106 ], and EGFR [ 107 ]. DMS includes three parts: the construction of a variant library; the identification and validation of the selection system for the protein function of interest and the introduction of the variant library into the selection system and subjected to selection; and the calculation of a functional score for each variant based on the change in the frequency of the variant during the selection [ 108 ] ( Figure 5 ).…”

Section: The Gap Between Possible and Functionally Studied Missense V...mentioning

confidence: 99%

“…Among the three parts, selection system, i.e., functional assays, needs to be properly designed and validated. Several commonly used functional assays are widely applied in DMS, such as cytotoxicity assays implemented in studying EGFR [ 107 ] and SCN5A variants [ 109 ], protein affinity assay in studying functional interaction sites in CXCR4 and CCR5 [ 110 ], reporter assay in ADRB2 study [ 111 ], and resting membrane potential assay for a K + channel KCNJ2 [ 112 ]. For the GJB2 gene, a library of all possible 4294 missense variants will be created and subjected to the selection system.…”

Section: The Gap Between Possible and Functionally Studied Missense V...mentioning

confidence: 99%

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Molecular Mechanisms and Clinical Phenotypes of GJB2 Missense Variants

Mao

Wang²,

et al. 2023

Biology

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The GJB2 gene is the most common gene responsible for hearing loss (HL) worldwide, and missense variants are the most abundant type. GJB2 pathogenic missense variants cause nonsyndromic HL (autosomal recessive and dominant) and syndromic HL combined with skin diseases. However, the mechanism by which these different missense variants cause the different phenotypes is unknown. Over 2/3 of the GJB2 missense variants have yet to be functionally studied and are currently classified as variants of uncertain significance (VUS). Based on these functionally determined missense variants, we reviewed the clinical phenotypes and investigated the molecular mechanisms that affected hemichannel and gap junction functions, including connexin biosynthesis, trafficking, oligomerization into connexons, permeability, and interactions between other coexpressed connexins. We predict that all possible GJB2 missense variants will be described in the future by deep mutational scanning technology and optimizing computational models. Therefore, the mechanisms by which different missense variants cause different phenotypes will be fully elucidated.

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Section: The Gap Between Possible and Functionally Studied Missense V...mentioning

confidence: 99%

Section: The Gap Between Possible and Functionally Studied Missense V...mentioning

confidence: 99%

Molecular Mechanisms and Clinical Phenotypes of GJB2 Missense Variants

Mao

Wang²,

et al. 2023

Biology

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“…Not surprisingly, most of the identified resistance mutations were at positions associated with drug binding, but additional mutations were located outside the drug-binding pocket of MRTX1257 and sotorasib [76]. Another study explored the sensitivity of EGFR variants to different approved inhibitors by implementing a cytotoxicity screen in the NSCLC cell line PC-9 depleted of endogenous EGFR and transduced with a lentiviral mutant library, and numerous common but also drug-selective variants were reported [77]. Generating a DMS library has high costs, and the screening campaign is labor-intensive.…”

Section: Deep Mutational Scanning (Dms)mentioning

confidence: 99%

Preclinical Anticipation of On- and Off-Target Resistance Mechanisms to Anti-Cancer Drugs: A Systematic Review

Dziubańska-Kusibab,

Nevedomskaya,

Haendler

2024

IJMS

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The advent of targeted therapies has led to tremendous improvements in treatment options and their outcomes in the field of oncology. Yet, many cancers outsmart precision drugs by developing on-target or off-target resistance mechanisms. Gaining the ability to resist treatment is the rule rather than the exception in tumors, and it remains a major healthcare challenge to achieve long-lasting remission in most cancer patients. Here, we discuss emerging strategies that take advantage of innovative high-throughput screening technologies to anticipate on- and off-target resistance mechanisms before they occur in treated cancer patients. We divide the methods into non-systematic approaches, such as random mutagenesis or long-term drug treatment, and systematic approaches, relying on the clustered regularly interspaced short palindromic repeats (CRISPR) system, saturated mutagenesis, or computational methods. All these new developments, especially genome-wide CRISPR-based screening platforms, have significantly accelerated the processes for identification of the mechanisms responsible for cancer drug resistance and opened up new avenues for future treatments.

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“…Although the functional effects of variants could be evaluated by expressing them using cDNA-based transgene libraries 13–16 , this non-physiological approach poses a risk of introducing artifacts from overexpression and does not accurately recapitulate the biology driven by these variants in the genomic context. Thus, the generation of variants at endogenous sites could provide more accurate functional evaluation.…”

Section: Maintextmentioning

confidence: 99%

Saturation resistance profiling of EGFR variants against tyrosine kinase inhibitors using prime editing

Kim,

Oh,

Lee

et al. 2023

Preprint

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Variants of uncertain significance (VUS) hamper the clinical application of genetic information. For example, in treating lung cancer with tyrosine kinase inhibitors (TKIs), many epidermal growth factor receptor (EGFR) variants remain classified as VUS with respect to TKI sensitivity1,2. Such incomplete resistance profiles hinder clinicians from selecting optimal therapeutic agents3,4. A high-throughput approach that can evaluate the functional effects of single nucleotide variants (SNVs) could reduce the number of VUS. Here we introduce SynPrime, a method based on prime editing that enabled the generation and functional evaluation of 2,476 SNVs in theEGFRgene, including 99% of all possible variants in the canonical tyrosine kinase domain (exons 18 to 21). We determined resistance profiles of 95% (= 1,726/1,817) of all possible EGFR protein variants encoded in the whole tyrosine kinase domain (exons 18 to 24) against afatinib, osimertinib, and osimertinib in the presence of the co-occurring mutation T790M, in PC-9 cells. SynPrime, which uses direct sequencing of endogenous regions to identify SNVs, provided more accurate functional evaluations than a guide RNA abundance-based approach. Our study has the potential to substantially improve the precision of therapeutic choices in clinical settings and contribute to addressing the issue of VUS by being applied to other genes.

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Defining the sensitivity landscape of EGFR variants to tyrosine kinase inhibitors

Cited by 16 publications

References 45 publications

Molecular Mechanisms and Clinical Phenotypes of GJB2 Missense Variants

Molecular Mechanisms and Clinical Phenotypes of GJB2 Missense Variants

Preclinical Anticipation of On- and Off-Target Resistance Mechanisms to Anti-Cancer Drugs: A Systematic Review

Saturation resistance profiling of EGFR variants against tyrosine kinase inhibitors using prime editing

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