2PD - Monitoring of secondary drug resistance mutations in circulating tumor DNA of patients with advanced ALK positive NSCLC

Date 15 April 2016
Event European Lung Cancer Conference 2016 (ELCC) 2016
Session Biomarkers
Topics Translational Research
Non-Small Cell Lung Cancer
Basic Principles in the Management and Treatment (of cancer)
Presenter Paola Bordi
Citation Journal of Thoracic Oncology (2016) 11 (supplement 4): S57-S166. S1556-0864(16)X0004-4
Authors P. Bordi1, M. Del Re2, R. Danesi2, M. Tiseo1
  • 1Medical Oncology Unit, Azienda Ospedaliera di Parma, 43126 - Parma/IT
  • 2Clinical Pharmacology And Pharmacogenetics Unit, Università degli studi di Pisa, 56126 - Pisa/IT



Disease progression in ALK positive NSCLC patients treated with crizotinib occurs after a median of 9–10 months of treatment. Several mechanisms of resistance were identified and include ALK gene mutations and amplification and activation of bypassing signaling pathways like EGFR, KRAS or c-KIT. Second-generation ALK-TKIs demonstrated an enhanced spectrum of activity in crizotinib-resistant patients. However, re-biopsy in NSCLC patients represents a critical issue and analysis of circulating cell-free DNA (cfDNA) has a promising role for the identification of mechanisms of resistance.


Sixteen patients progressed during ALK-TKI were enrolled. After progression, blood was collected and DNA was extracted from plasma using QIAamp circulating nucleic acid kit (Qiagen®) and tested for ALK secondary mutations and KRAS exon 12 mutations using the Digital Droplet PCR (ddPCR – BioRad®).


All patients were stage IV adenocarcinoma; 11 female and 5 male. Nine were never-smokers and 7 former-smokers. Median age was 53 yrs (range 40–81). Fifteen patients received crizotinib and 1 ceritinib. ALK-TKIs was administered mainly as second-line, in 2 cases as first and in the remaining as third-line therapy. Twelve patients had partial response, 3 stable disease, one progressed. Median PFS was 8 months. In 12 cases brain was a site of progression and only 5 patients had a tumor site that could potentially undergo re-biospy. ALK secondary mutations were identified in 4 patients. One showed both p.L1196M and p.G1269A mutations which levels decreased after 2 months of therapy with second generation ALK-TKI, along with tumor response. The second and the third patient had p.L1196M and p.G1269A, respectively. The 4th patient showed p.F1174L after initiation of second generation ALK-TKI. A total of 9 patients KRAS mutations p.G12D or p.G12V appeared in cfDNA at the time of resistance to TKI, 3 of them presented both ALK and KRAS mutations.


ddPCR can detect resistance mutations in cfDNA of ALK+ NSCLC and is an effective alternative to re-biopsy. The assessment of mutant allele burden could be used for response monitoring during treatment. Moreover, KRAS mutations may play a role in resistance to ALK-TKIs.

Clinical trial identification

Not applicable

Legal entity responsible for the study

University of Pisa


Fondazione Cassa di Risparmio di Lucca


All authors have declared no conflicts of interest.