Potential unreliability of ALK variant allele frequency in the efficacy prediction of targeted therapy in NSCLC
Wei Rao1, Yutao Liu2, Yan Li1, Lei Guo1, Tian Qiu1, Lin Dong1, Jianming Ying1(), Weihua Li1()
1. Department of Pathology, National Cancer Center/National Clinical Research Center for Cancer/Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100021, China 2. Department of Medical Oncology, National Cancer Center/National Clinical Research Center for Cancer/Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100021, China
Anaplastic lymphoma kinase (ALK) is the most common fusion gene involved in non-small cell lung cancer (NSCLC), and remarkable response has been achieved with the use of ALK tyrosine kinase inhibitors (ALK-TKIs). However, the clinical efficacy is highly variable. Pre-existing intratumoral heterogeneity (ITH) has been proven to contribute to the poor treatment response and the resistance to targeted therapies. In this work, we investigated whether the variant allele frequencies (VAFs) of ALK fusions can help assess ITH and predict targeted therapy efficacy. Through the application of next-generation sequencing (NGS), 7.2% (326/4548) of patients were detected to be ALK positive. On the basis of the adjusted VAF (adjVAF, VAF normalization for tumor purity) of four different threshold values (adjVAF < 50%, 40%, 30%, or 20%), the association of ALK subclonality with crizotinib efficacy was assessed. Nonetheless, no statistical association was observed between median progression-free survival (PFS) and ALK subclonality assessed by adjVAF, and a poor correlation of adjVAF with PFS was found among the 85 patients who received first-line crizotinib. Results suggest that the ALK VAF determined by hybrid capture-based NGS is probably unreliable for ITH assessment and targeted therapy efficacy prediction in NSCLC.
. [J]. Frontiers of Medicine, 2023, 17(3): 493-502.
Wei Rao, Yutao Liu, Yan Li, Lei Guo, Tian Qiu, Lin Dong, Jianming Ying, Weihua Li. Potential unreliability of ALK variant allele frequency in the efficacy prediction of targeted therapy in NSCLC. Front. Med., 2023, 17(3): 493-502.
M Soda, YL Choi, M Enomoto, S Takada, Y Yamashita, S Ishikawa, S Fujiwara, H Watanabe, K Kurashina, H Hatanaka, M Bando, S Ohno, Y Ishikawa, H Aburatani, T Niki, Y Sohara, Y Sugiyama, H Mano. Identification of the transforming EML4-ALK fusion gene in non-small-cell lung cancer. Nature 2007; 448(7153): 561–566 https://doi.org/10.1038/nature05945
pmid: 17625570
2
HX Tian, XC Zhang, JJ Yang, WB Guo, ZH Chen, Z Wang, YL Wu. Clinical characteristics and sequence complexity of anaplastic lymphoma kinase gene fusions in Chinese lung cancer patients. Lung Cancer 2017; 114: 90–95 https://doi.org/10.1016/j.lungcan.2017.11.001
pmid: 29173772
3
AT Shaw, DW Kim, K Nakagawa, T Seto, L Crinó, MJ Ahn, Pas T De, B Besse, BJ Solomon, F Blackhall, YL Wu, M Thomas, KJ O’Byrne, D Moro-Sibilot, DR Camidge, T Mok, V Hirsh, GJ Riely, S Iyer, V Tassell, A Polli, KD Wilner, PA Jänne. Crizotinib versus chemotherapy in advanced ALK-positive lung cancer. N Engl J Med 2013; 368(25): 2385–2394 https://doi.org/10.1056/NEJMoa1214886
pmid: 23724913
4
DR Camidge, R Dziadziuszko, S Peters, T Mok, J Noe, M Nowicka, SM Gadgeel, P Cheema, N Pavlakis, Marinis F de, BC Cho, L Zhang, D Moro-Sibilot, T Liu, W Bordogna, B Balas, B Müller, AT Shaw. Updated efficacy and safety data and impact of the EML4-ALK fusion variant on the efficacy of alectinib in untreated ALK-positive advanced non-small cell lung cancer in the global phase III ALEX study. J Thorac Oncol 2019; 14(7): 1233–1243 https://doi.org/10.1016/j.jtho.2019.03.007
pmid: 30902613
5
AT Shaw, TM Kim, L Crinò, C Gridelli, K Kiura, G Liu, S Novello, A Bearz, O Gautschi, T Mok, M Nishio, G Scagliotti, DR Spigel, S Deudon, C Zheng, S Pantano, P Urban, C Massacesi, K Viraswami-Appanna, E Felip. Ceritinib versus chemotherapy in patients with ALK-rearranged non-small-cell lung cancer previously given chemotherapy and crizotinib (ASCEND-5): a randomised, controlled, open-label, phase 3 trial. Lancet Oncol 2017; 18(7): 874–886 https://doi.org/10.1016/S1470-2045(17)30339-X
pmid: 28602779
6
DR Camidge, HR Kim, MJ Ahn, JCH Yang, JY Han, MJ Hochmair, KH Lee, A Delmonte, Campelo MR García, DW Kim, F Griesinger, E Felip, R Califano, A Spira, SN Gettinger, M Tiseo, HM Lin, N Gupta, MJ Hanley, Q Ni, P Zhang, S Popat. Brigatinib versus crizotinib in advanced ALK inhibitor-naive ALK-positive non-small cell lung cancer: second interim analysis of the phase III ALTA-1L trial. J Clin Oncol 2020; 38(31): 3592–3603 https://doi.org/10.1200/JCO.20.00505
pmid: 32780660
7
BJ Solomon, B Besse, TM Bauer, E Felip, RA Soo, DR Camidge, R Chiari, A Bearz, CC Lin, SM Gadgeel, GJ Riely, EH Tan, T Seto, LP James, JS Clancy, A Abbattista, JF Martini, J Chen, G Peltz, H Thurm, SI Ou, AT Shaw. Lorlatinib in patients with ALK-positive non-small-cell lung cancer: results from a global phase 2 study. Lancet Oncol 2018; 19(12): 1654–1667 https://doi.org/10.1016/S1470-2045(18)30649-1
pmid: 30413378
8
B Golding, A Luu, R Jones, AM Viloria-Petit. The function and therapeutic targeting of anaplastic lymphoma kinase (ALK) in non-small cell lung cancer (NSCLC). Mol Cancer 2018; 17(1): 52 https://doi.org/10.1186/s12943-018-0810-4
pmid: 29455675
9
Y Li, T Zhang, J Zhang, W Li, P Yuan, P Xing, Z Zhang, S Chuai, J Li, J Ying. Response to crizotinib in advanced ALK-rearranged non-small cell lung cancers with different ALK-fusion variants. Lung Cancer 2018; 118: 128–133 https://doi.org/10.1016/j.lungcan.2018.01.026
pmid: 29571990
10
P Song, F Zhang, Y Li, G Yang, W Li, J Ying, S Gao. Concomitant TP53 mutations with response to crizotinib treatment in patients with ALK-rearranged non-small-cell lung cancer. Cancer Med 2019; 8(4): 1551–1557 https://doi.org/10.1002/cam4.2043
pmid: 30843662
ZF Lim, PC Ma. Emerging insights of tumor heterogeneity and drug resistance mechanisms in lung cancer targeted therapy. J Hematol Oncol 2019; 12(1): 134 https://doi.org/10.1186/s13045-019-0818-2
pmid: 31815659
13
S Goodwin, JD McPherson, WR McCombie. Coming of age: ten years of next-generation sequencing technologies. Nat Rev Genet 2016; 17(6): 333–351 https://doi.org/10.1038/nrg.2016.49
pmid: 27184599
14
W Li, T Qiu, Y Ling, S Gao, J Ying. Subjecting appropriate lung adenocarcinoma samples to next-generation sequencing-based molecular testing: challenges and possible solutions. Mol Oncol 2018; 12(5): 677–689 https://doi.org/10.1002/1878-0261.12190
pmid: 29518290
15
N Normanno, AM Rachiglio, M Lambiase, E Martinelli, F Fenizia, C Esposito, C Roma, T Troiani, D Rizzi, F Tatangelo, G Botti, E Maiello, G Colucci, F; CAPRI-GOIM investigators Ciardiello. Heterogeneity of KRAS, NRAS, BRAF and PIK3CA mutations in metastatic colorectal cancer and potential effects on therapy in the CAPRI GOIM trial. Ann Oncol 2015; 26(8): 1710–1714 https://doi.org/10.1093/annonc/mdv176
pmid: 25851630
16
B Gieszer, Z Megyesfalvi, V Dulai, J Papay, I Kovalszky, J Timar, J Fillinger, T Harko, O Pipek, V Teglasi, E Regos, G Papp, Z Szallasi, V Laszlo, F Renyi-Vamos, G Galffy, C Bodor, B Dome, J Moldvay. EGFR variant allele frequency predicts EGFR-TKI efficacy in lung adenocarcinoma: a multicenter study. Transl Lung Cancer Res 2021; 10(2): 662–674 https://doi.org/10.21037/tlcr-20-814
pmid: 33718012
17
W Li, L Guo, Y Liu, L Dong, L Yang, L Chen, K Liu, Y Shao, J Ying. Potential unreliability of uncommon ALK, ROS1, and RET genomic breakpoints in predicting the efficacy of targeted therapy in NSCLC. J Thorac Oncol 2021; 16(3): 404–418 https://doi.org/10.1016/j.jtho.2020.10.156
pmid: 33248323
18
W Li, T Qiu, L Guo, J Ying. Major challenges related to tumor biological characteristics in accurate mutation detection of colorectal cancer by next-generation sequencing. Cancer Lett 2017; 410: 92–99 https://doi.org/10.1016/j.canlet.2017.09.014
pmid: 28942013
19
W Li, J Zhang, L Guo, S Chuai, L Shan, J Ying. Combinational analysis of FISH and immunohistochemistry reveals rare genomic events in ALK fusion patterns in NSCLC that responds to crizotinib treatment. J Thorac Oncol 2017; 12(1): 94–101 https://doi.org/10.1016/j.jtho.2016.08.145
pmid: 27614248
20
J Ying, L Guo, T Qiu, L Shan, Y Ling, X Liu, N Lu. Diagnostic value of a novel fully automated immunochemistry assay for detection of ALK rearrangement in primary lung adenocarcinoma. Ann Oncol 2013; 24(10): 2589–2593 https://doi.org/10.1093/annonc/mdt295
pmid: 23904459
21
EA Eisenhauer, P Therasse, J Bogaerts, LH Schwartz, D Sargent, R Ford, J Dancey, S Arbuck, S Gwyther, M Mooney, L Rubinstein, L Shankar, L Dodd, R Kaplan, D Lacombe, J Verweij. New response evaluation criteria in solid tumours: revised RECIST guideline (version 1.1). Eur J Cancer 2009; 45(2): 228–247 https://doi.org/10.1016/j.ejca.2008.10.026
pmid: 19097774
22
DR Camidge, M Theodoro, DA Maxson, M Skokan, T O’Brien, X Lu, RC Doebele, AE Barón, M Varella-Garcia. Correlations between the percentage of tumor cells showing an anaplastic lymphoma kinase (ALK) gene rearrangement, ALK signal copy number, and response to crizotinib therapy in ALK fluorescence in situ hybridization-positive nonsmall cell lung cancer. Cancer 2012; 118(18): 4486–4494 https://doi.org/10.1002/cncr.27411
pmid: 22282074
23
DR Camidge, SA Kono, A Flacco, AC Tan, RC Doebele, Q Zhou, L Crino, WA Franklin, M Varella-Garcia. Optimizing the detection of lung cancer patients harboring anaplastic lymphoma kinase (ALK) gene rearrangements potentially suitable for ALK inhibitor treatment. Clin Cancer Res 2010; 16(22): 5581–5590 https://doi.org/10.1158/1078-0432.CCR-10-0851
pmid: 21062932
24
W Li, J Zhang, Z Wang, L Li, J Ma, X Zhou, J Wang, Z Liang, J Ying. Guidelines for clinical practice of ALK fusion detection in non-small-cell lung cancer: a proposal from the Chinese RATICAL study group. Journal of the National Cancer Center 2021; 1(4): 123–131 https://doi.org/10.1016/j.jncc.2021.07.005
25
MI Ilie, C Bence, V Hofman, E Long-Mira, C Butori, L Bouhlel, S Lalvée, J Mouroux, M Poudenx, J Otto, CH Marquette, P Hofman. Discrepancies between FISH and immunohistochemistry for assessment of the ALK status are associated with ALK ‘borderline’-positive rearrangements or a high copy number: a potential major issue for anti-ALK therapeutic strategies. Ann Oncol 2015; 26(1): 238–244 https://doi.org/10.1093/annonc/mdu484
pmid: 25344360
26
H Wang, L Sun, Y Sang, X Yang, G Tian, Z Wang, J Fang, W Sun, L Zhou, L Jia, MS Tsao, H Shi, D Lin. A study of ALK-positive pulmonary squamous-cell carcinoma: from diagnostic methodologies to clinical efficacy. Lung Cancer 2019; 130: 135–142 https://doi.org/10.1016/j.lungcan.2019.02.015
pmid: 30885334
27
R Benayed, M Offin, K Mullaney, P Sukhadia, K Rios, P Desmeules, R Ptashkin, H Won, J Chang, D Halpenny, AM Schram, CM Rudin, DM Hyman, ME Arcila, MF Berger, A Zehir, MG Kris, A Drilon, M Ladanyi. High yield of RNA sequencing for targetable kinase fusions in lung adenocarcinomas with no mitogenic driver alteration detected by DNA sequencing and low tumor mutation burden. Clin Cancer Res 2019; 25(15): 4712–4722 https://doi.org/10.1158/1078-0432.CCR-19-0225
pmid: 31028088
28
W Li, R Wan, L Guo, G Chang, D Jiang, L Meng, J Ying. Reliability analysis of exonic-breakpoint fusions identified by DNA sequencing for predicting the efficacy of targeted therapy in non-small cell lung cancer. BMC Med 2022; 20(1): 160 https://doi.org/10.1186/s12916-022-02362-9
pmid: 35534835
29
R Raja, M Kuziora, PZ Brohawn, BW Higgs, A Gupta, PA Dennis, K Ranade. Early reduction in ctDNA predicts survival in patients with lung and bladder cancer treated with durvalumab. Clin Cancer Res 2018; 24(24): 6212–6222 https://doi.org/10.1158/1078-0432.CCR-18-0386
pmid: 30093454
30
X Li, W Cai, G Yang, C Su, S Ren, C Zhao, R Hu, X Chen, G Gao, Z Guo, W Li, C Zhou, FR Hirsch. Comprehensive analysis of EGFR-mutant abundance and its effect on efficacy of EGFR TKIs in advanced NSCLC with EGFR mutations. J Thorac Oncol 2017; 12(9): 1388–1397 https://doi.org/10.1016/j.jtho.2017.06.006
pmid: 28624467
31
M Beau-Faller, E Pencreach, C Leduc, H Blons, JP Merlio, PP Bringuier, Fraipont F de, F Escande, A Lemoine, L Ouafik, M Denis, P Hofman, R Lacave, S Melaabi, A Langlais, P Missy, F Morin, D Moro-Sibilot, F Barlesi, J; French Cooperative Thoracic Intergroup (IFCT) Cadranel. Independent prognostic value of ultra-sensitive quantification of tumor pre-treatment T790M subclones in EGFR mutated non-small cell lung cancer (NSCLC) treated by first/second generation TKI, depends on variant allele frequency (VAF): results of the French cooperative thoracic intergroup (IFCT) biomarkers France project. Lung Cancer 2020; 140: 19–26 https://doi.org/10.1016/j.lungcan.2019.10.013
pmid: 31841714
32
E Elez, C Chianese, E Sanz-García, E Martinelli, A Noguerido, FM Mancuso, G Caratù, J Matito, J Grasselli, C Cardone, Abate R Esposito, G Martini, C Santos, T Macarulla, G Argilés, J Capdevila, A Garcia, N Mulet, E Maiello, N Normanno, F Jones, J Tabernero, F Ciardello, R Salazar, A Vivancos. Impact of circulating tumor DNA mutant allele fraction on prognosis in RAS-mutant metastatic colorectal cancer. Mol Oncol 2019; 13(9): 1827–1835 https://doi.org/10.1002/1878-0261.12547
pmid: 31322322
33
A Friedlaender, P Tsantoulis, M Chevallier, C De Vito, A Addeo. The impact of variant allele frequency in EGFR mutated NSCLC patients on targeted therapy. Front Oncol 2021; 11: 644472 https://doi.org/10.3389/fonc.2021.644472
pmid: 33869038
34
GM O’Kane, G Liu, TL Stockley, M Shabir, T Zhang, JH Law, LW Le, A Sacher, FA Shepherd, PA Bradbury, NB Leighl. The presence and variant allele fraction of EGFR mutations in ctDNA and development of resistance. Lung Cancer 2019; 131: 86–89 https://doi.org/10.1016/j.lungcan.2019.03.019
pmid: 31027703
35
SR Head, HK Komori, SA LaMere, T Whisenant, F Van Nieuwerburgh, DR Salomon, P Ordoukhanian. Library construction for next-generation sequencing: overviews and challenges. Biotechniques 2014; 56(2): 61–64, 66, 68 passim doi:10.2144/000114133
pmid: 24502796
36
R Bruno, G Fontanini. Next generation sequencing for gene fusion analysis in lung cancer: a literature review. Diagnostics (Basel) 2020; 10(8): 521 https://doi.org/10.3390/diagnostics10080521
pmid: 32726941
37
T Vaclova, U Grazini, L Ward, D O’Neill, A Markovets, X Huang, J Chmielecki, R Hartmaier, KS Thress, PD Smith, JC Barrett, J Downward, Bruin EC de. Clinical impact of subclonal EGFR T790M mutations in advanced-stage EGFR-mutant non-small-cell lung cancers. Nat Commun 2021; 12(1): 1780 https://doi.org/10.1038/s41467-021-22057-8
pmid: 33741979
38
Q Zheng, S Hong, Y Huang, H Zhao, Y Yang, X Hou, Y Zhao, Y Ma, T Zhou, Y Zhang, W Fang, L Zhang. EGFR T790M relative mutation purity predicts osimertinib treatment efficacy in non-small cell lung cancer patients. Clin Transl Med 2020; 9(1): 17 https://doi.org/10.1186/s40169-020-0269-y
pmid: 32067121
39
X Ai, J Cui, J Zhang, R Chen, W Lin, C Xie, A Liu, J Zhang, W Yang, X Hu, X Hu, Q Zhao, C Rao, YS Zang, R Ning, P Li, L Chang, X Yi, S Lu. Clonal architecture of EGFR mutation predicts the efficacy of EGFR-tyrosine kinase inhibitors in advanced NSCLC: a prospective multicenter study (NCT03059641). Clin Cancer Res 2021; 27(3): 704–712 https://doi.org/10.1158/1078-0432.CCR-20-3063
pmid: 33188140
40
MK Bos, K Nasserinejad, MPHM Jansen, L Angus, PN Atmodimedjo, E de Jonge, WNM Dinjens, RHN van Schaik, M Del Re, HJ Dubbink, S Sleijfer, JWM Martens. Comparison of variant allele frequency and number of mutant molecules as units of measurement for circulating tumor DNA. Mol Oncol 2021; 15(1): 57–66 https://doi.org/10.1002/1878-0261.12827
pmid: 33070443
41
JG Bustamante Alvarez, S Janse, DH Owen, S Kiourtsis, EM Bertino, K He, DP Carbone, GA Otterson. Treatment of non-small-cell lung cancer based on circulating cell-free DNA and impact of variation allele frequency. Clin Lung Cancer 2021; 22(4): e519–e527 https://doi.org/10.1016/j.cllc.2020.11.007
pmid: 33414052
T Qiu, F Zhang, W Li, L Guo, J Ying. Concurrent presence of ALK rearrangement and MET mutation in lung adenocarcinoma. J Thorac Oncol 2019; 14(2): e42–e44 https://doi.org/10.1016/j.jtho.2018.10.149
pmid: 30683300
