Coexistence of KRAS mutation with mutant but not wild-type EGFR predicts response to tyrosine-kinase inhibitors in human lung cancer

Choughule, Anuradha; Sharma, Richa; Trivedi, Vaishakhi; Thavamani, Abhishek; Noronha, Vanita; Joshi, Amit; Desai, Saral; Chandrani, Pratik; Sundaram, P; Utture, Sagarika; Jambhekar, Nirmala A.; Gupta, Sudeep; Aich, Jyotirmoi; Prabhash, Kumar; Dutt, Amit

doi:10.1038/bjc.2014.401

Cited by 35 publications

(29 citation statements)

References 18 publications

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“…Nevertheless, the detection of mutations in both plasma and tissue by ddPCR, but not by standard methods, could be due to the higher sensitivity of ddPCR analysis. Two patients, initially diagnosed WT KRAS by standard method, were re-analysed by ddPCR and were found MUT KRAS in the primary biopsy, suggesting that the MUT KRAS clone co-existed with activating MUT EGFR since the beginning, as also demonstrated in previous reports [23, 24]. In these patients, MUT KRAS cannot strictly be considered a mechanisms of resistance but it could be possible that EGFR-TKI treatment may have favored the expansion of MUT KRAS -positive clones.…”

Section: Discussionsupporting

confidence: 65%

Contribution of KRAS mutations and c.2369C > T (p.T790M) EGFR to acquired resistance to EGFR-TKIs in EGFR mutant NSCLC: a study on circulating tumor DNA

Tiseo²,

Bordi³

et al. 2016

Oncotarget

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IntroductionKRAS oncogene mutations (MUTKRAS) drive resistance to EGFR inhibition by providing alternative signaling as demonstrated in colo-rectal cancer. In non-small cell lung cancer (NSCLC), the efficacy of treatment with EGFR tyrosine kinase inhibitors (EGFR-TKIs) depends on activating EGFR mutations (MUTEGFR). However, inhibition of EGFR may select resistant cells displaying alternative signaling, i.e., KRAS, or restoration of EGFR activity due to additional MUTEGFR, i.e., the c.2369C > T (p.T790MEGFR).AimThe aim of this study was to investigate the appearance of MUTKRAS during EGFR-TKI treatment and their contribution to drug resistance.MethodsThis study used cell-free circulating tumor DNA (cftDNA) to evaluate the appearance of codon 12 MUTKRAS and p.T790MEGFR mutations in 33 advanced NSCLC patients progressing after an EGFR-TKI.Resultsp.T790MEGFR was detected in 11 (33.3%) patients, MUTKRAS at codon 12 in 3 (9.1%) while both p.T790MEGFR and MUTKRAS codon 12 were found in 13 (39.4%) patients. Six patients (18.2%) were KRAS wild-type (WTKRAS) and negative for p.T790MEGFR. In 8 subjects paired tumor re-biopsy/plasma samples were available; the percent concordance of tissue/plasma was 62.5% for p.T790MEGFR and 37.5% for MUTKRAS. The analysis of time to progression (TTP) and overall survival (OS) in WTKRAS vs. MUTKRAS were not statistically different, even if there was a better survival with WTKRAS vs. MUTKRAS, i.e., TTP 14.4 vs. 11.4 months (p = 0.97) and OS 40.2 vs. 35.0 months (p = 0.56), respectively.ConclusionsMUTKRAS could be an additional mechanism of escape from EGFR-TKI inhibition and cftDNA is a feasible approach to monitor the molecular development of drug resistance.

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Section: Discussionsupporting

confidence: 65%

Contribution of KRAS mutations and c.2369C > T (p.T790M) EGFR to acquired resistance to EGFR-TKIs in EGFR mutant NSCLC: a study on circulating tumor DNA

Tiseo²,

Bordi³

et al. 2016

Oncotarget

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“…CRE demonstrates the ability to co-amplify all five exons ( KRAS exon 2 and EGFR exon 18-21) in a single multiplex PCR reaction with a limited amount of starting template DNA followed by the enrichment of concatenated product ( Figure 2D) by concatenation PCR using first multiplex PCR product as a template. The concatenated product confirmed EGFR L858R mutation in the FFPE tissues ( Supplementary Figure S2), as reported earlier ( Choughule et al , 2014). Thus our CRE method can be routinely used for the mutational analysis of KRAS and EGFR genes.…”

Section: Resultssupporting

confidence: 86%

“…While EGFR and KRAS mutations largely occur mutually exclusively in non-small cell lung cancer (NSCLC), and predict contrasting response rate to tyrosine-kinase inhibitors (TKI) ( Chougule et al , 2013; Fukuoka et al , 2011; Ihle et al , 2012; Lynch et al , 2004; Mao et al , 2010; Mok et al , 2009), some recent studies, including ours, suggest co-occurrence of EGFR and KRAS mutations in the same patients, albeit at low frequency ( Choughule et al , 2014; Li et al , 2014). While no direct evidence exists as yet, these studies may have implications for carrying out routine KRAS molecular testing along with EGFR mutations for precluding a patient with NSCLC from therapy with EGFR inhibitors, as approved for colorectal cancer ( Lievre et al , 2006).…”

Section: Introductionmentioning

confidence: 78%

“…These activating mutations include point mutations G719S, T790M, L858R, and L861Q in exons 18, 20 and 21 respectively and in-frame deletions/insertions in exon 19 ( Kosaka et al , 2004). The most common mutations that result in an amino acid substitution at position 12 and 13 in KRAS are G12V and G13D ( Choughule et al , 2014). Several screening and target based methods are currently in use for to infer the EGFR and KRAS hot spot mutations, viz; PCR-RFLP (Restriction fragment length polymorphism), Amplification Refractory Mutation System (ARMS), PCR-Invader, TaqMan PCR, allele specific qPCR, high resolution melting analysis and ultra-deep pyrosequencing, SNaPshot analysis and co-amplification at lower denaturation temperature (COLD)-PCR ( Angulo et al , 2012; Borràs et al , 2011; Ellison et al , 2013; Santis et al , 2011; van Eijk et al , 2011; Zinsky et al , 2010).…”

Section: Introductionmentioning

confidence: 99%

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CRE: a cost effective and rapid approach for PCR-mediated concatenation of KRAS and EGFR exons

et al. 2016

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Molecular diagnostics has changed the way lung cancer patients are treated worldwide. Of several different testing methods available, PCR followed by directed sequencing and amplification refractory mutation system (ARMS) are the two most commonly used diagnostic methods worldwide to detect mutations at KRAS exon 2 and EGFR kinase domain exons 18-21 in lung cancer. Compared to ARMS, the PCR followed by directed sequencing approach is relatively inexpensive but more cumbersome to perform. Moreover, with a limiting amount of genomic DNA from clinical formalin-fixed, paraffin-embedded (FFPE) specimens or fine biopsies of lung tumors, multiple rounds of PCR and sequencing reactions often get challenging. Here, we report a cost-effective single multiplex-PCR based method, CRE (for Co-amplification of five K RAS and E GFR exons), followed by concatenation of the PCR product as a single linear fragment for direct sequencing. CRE is a robust protocol that can be adapted for routine use in clinical diagnostics with reduced variability, cost and turnaround time requiring a minimal amount of template DNA extracted from FFPE or fresh frozen tumor samples. As a proof of principle, CRE is able to detect the activating EGFR L858R and T790M EGFR mutations in lung cancer cell line and primary tumors.

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“…Indeed, a recent report evidenced that the expression of inducible KRAS G12V constructs in EGFRmutant PC9 cells results in decreased cell viability after 7 days of selection with doxycyclin (16). Anecdotal reports show evidence of co-occurrence of KRAS and EGFR mutations in lung cancer patients (36,37). More recently Hata and colleagues (14) described two distinct evolutionary paths for the development of resistance to EGFR inhibition.…”

Section: T790mmentioning

confidence: 99%

Heterogeneous Mechanisms of Primary and Acquired Resistance to Third-Generation EGFR Inhibitors

Ortiz-Cuarán

Scheffler

Plenker

et al. 2016

Clinical Cancer Research

227

194

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Purpose: To identify novel mechanisms of resistance to thirdgeneration EGFR inhibitors in patients with lung adenocarcinoma that progressed under therapy with either AZD9291 or rociletinib (CO-1686).Experimental Design: We analyzed tumor biopsies from seven patients obtained before, during, and/or after treatment with AZD9291 or rociletinib (CO-1686). Targeted sequencing and FISH analyses were performed, and the relevance of candidate genes was functionally assessed in in vitro models.Results: We found recurrent amplification of either MET or ERBB2 in tumors that were resistant or developed resistance to third-generation EGFR inhibitors and show that ERBB2 and MET activation can confer resistance to these compounds. Furthermore, we identified a KRAS G12S mutation in a patient with acquired resistance to AZD9291 as a potential driver of acquired resistance. Finally, we show that dual inhibition of EGFR/MEK might be a viable strategy to overcome resistance in EGFR-mutant cells expressing mutant KRAS.Conclusions: Our data suggest that heterogeneous mechanisms of resistance can drive primary and acquired resistance to third-generation EGFR inhibitors and provide a rationale for potential combination strategies.

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Coexistence of KRAS mutation with mutant but not wild-type EGFR predicts response to tyrosine-kinase inhibitors in human lung cancer

Cited by 35 publications

References 18 publications

Contribution of KRAS mutations and c.2369C > T (p.T790M) EGFR to acquired resistance to EGFR-TKIs in EGFR mutant NSCLC: a study on circulating tumor DNA

Contribution of KRAS mutations and c.2369C > T (p.T790M) EGFR to acquired resistance to EGFR-TKIs in EGFR mutant NSCLC: a study on circulating tumor DNA

CRE: a cost effective and rapid approach for PCR-mediated concatenation of KRAS and EGFR exons

Heterogeneous Mechanisms of Primary and Acquired Resistance to Third-Generation EGFR Inhibitors

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