EDITORIAL

A Curious Link Between Epidermal Growth Factor Receptor Amplification and Survival: Effect of "Allele Dilution" on Gefitinib Sensitivity?

Frederic J. Kaye

Correspondence to: Frederic J. Kaye, MD, Genetics Branch, Center for Cancer Research, NCI and National Naval Medical Center, Bethesda, MD 20889 (e-mail: fkaye{at}helix.nih.gov).

The past few months have seen an increasing number of publications that confirm and extend the discovery in 2004 that specific epidermal growth factor receptor (EGFR) mutations in lung tumors are linked with clinical responsiveness to gefitinib and erlotinib (13). These emerging data are especially exciting, because the genetic, biological, and clinical implications directly interrelate with each other to provide new directions for basic research and new optimism for the management of the major cause of cancer deaths worldwide. Clinicians, however, have to temper their enthusiasm because only a minority of patients may benefit from treatment and there is still uncertainty in how to best select these patients. For example, although EGFR mutational status was the best predictor for overall survival in two recent series of gefitinib-treated lung cancer patients from Japan (P = .0053) (4) and Korea (P<.001) (5), these and other studies (1,3) have shown that objective responses can occur in tumor samples containing an apparently wild-type EGFR sequence and, conversely, tumors carrying EGFR mutations eventually develop resistant disease (6,7). In addition, prolonged survival and clinically meaningful improvements in symptoms may be detected in selected patients with stable disease (8,9) that may not be as tightly associated with the presence of specific EGFR kinase domain mutations.

In this issue of the Journal, Cappuzzo et al. (10) have sought to define an optimal marker for gefitinib drug sensitivity by measuring Akt phosphorylation (P-Akt) as well as EGFR gene copy number (using fluorescence in situ hybridization [FISH]), protein levels, and the presence of mutations in pretreatment tumor biopsy samples collected from 102 patients with non–small-cell lung cancer (NSCLC). The two main observations of this study were that high EGFR gene copy number was a better predictor for survival in gefitinib-treated patients (P = .03) than the presence of somatic EGFR mutations (P = .09) and that 40% of treated patients with lung tumors carrying EGFR mutations showed progressive disease. Although the authors stopped short of concluding that the detection of EGFR mutations by sequence analysis is an unreliable clinical marker, they recommend EGFR FISH analysis, perhaps combined with measurements of EGFR and P-Akt levels, as the optimal test for selecting patients for gefitinib-like therapies.

This study reminds us that much still remains to be learned about this new family of kinase inhibitor drugs in which 1) the biology of objective tumor responses versus stable disease may differ, 2) EGFR protein levels have been an inconsistent marker (11), and 3) assays performed on tumor cell lines, such as ErbB3 expression or gefitinib-induced inhibition of P-Akt, have not yet been extrapolated to primary tumor biopsy samples (1214). Ultimately, as noted by the authors, prospective clinical trials will be required to define the optimal assays for guiding the management of these patients with lung cancer.

The Cappuzzo et al. study, however, raises additional issues that are reminiscent of a curious association between another dominant oncogene and gene amplification. For example, cells carrying mutant K-Ras require either amplification of the mutant allele or loss of the wild-type allele to attain full tumorigenic potential in different murine models and tumor samples (1517). In this context, the oncogene is not "dominant" on a level playing field, and selective amplification of the mutant allele might be required to dilute the pool of wild-type protein. In the special case of mutant EGFR, gefitinib-sensitive tumorigenesis might rely on gene amplification to achieve a relative dilution of not only its wild-type allele but also of a group of heterodimeric family members (Fig. 1). Accordingly, amplification of mutant EGFR has been previously noted in glioblastoma tumors and in several different tumor cell lines (1821). Cappuzzo et al. also noted an association between EGFR FISH+ status and EGFR mutations (P = .01) in the present study (10), and the prevalence of EGFR gene amplification detected in two other publications on lung cancer (approximately 10%) (22,23) overlaps with the frequency of EGFR mutations reported for non-Asian populations. Finally, loss of the wild-type allele and/or gene amplification of the mutant EGFR allele has been observed or inferred indirectly in many lung tumor samples in several recent reports (2,3,21,24).



View larger version (11K):
[in this window]
[in a new window]
 
Fig. 1. A hypothetical mechanism for improved survival of lung cancer patients who receive gefitinib-like therapies through allele dilution. Dilution of the wild-type (wt) allele may be required for the tumorigenicity of other "dominant" oncogenes such as K-ras (15,16), Met (33), and RET (34).

 
Although these observations suggest that the presence of mutant EGFR may, at least partly, explain the high objective response rates seen in FISH+ tumor samples, they do not directly address why 40% of patients with tumors containing EGFR mutations had progressive disease on gefitinib or why FISH+ status was the optimal marker for survival. Before considering these questions, it is perhaps worth mentioning the unknown variable of compliance with an oral agent in individual patients on a clinical trial and that one of the mutations detected by the authors in a gefitinib-refractory tumor was a conservative alteration (V852I) that is of uncertain functional significance. More interestingly, however, the biological basis for gefitinib resistance in four different patients has been recently linked to an additional EGFR missense mutation at codon 790 that is analogous to mutations associated with acquired imatinib resistance (6,7). Because BCR-ABL mutations associated with resistance can be detected in subpopulations of chronic myeloid leukemia cells prior to imatinib therapy (25), it would be interesting to sequence additional EGFR exons, especially codon 790 within exon 20, and other candidate heterodimerization gene partners, in these gefitinib-refractory tumors. In addition, although gene amplification of a mutant EGFR allele arises to promote tumorigenesis, the resulting increased number of gene copies may, paradoxically, serve to improve survival by "buffering" the onset of gefitinib resistance. For example, because additional "resistance" mutations are presumed to arise sequentially within single alleles, the presence of multiple mutant EGFR copies should be predicted to delay the onset of clinical resistance by diluting the biological effect of these "resistance" alleles (Fig. 1). This model, however, proposes that some minimum threshold ratio of the "resistance" receptor is required on the cell membrane to exhibit the phenotype of gefitinib resistance. Again, further data from retrospective and prospective clinical trials will help to define the natural history of EGFR mutant and wild-type lung tumors.

Finally, it is worth acknowledging the power of genetic studies, which have now allowed the tentative proposal of a growing, e.g., EGFR/ERBB2/K-RAS/B-RAF/PIK3CA/PTEN mutational activation pathway for the development of lung adenocarcinoma (24,2631) with the prediction that sensitivity to gefitinib will ultimately depend on which components of the pathway have been targeted and which remain intact. Unfortunately, it appears that it may be easier to accurately predict drug resistance than drug efficacy. For example, it has already been suggested that patients with tumors containing K-Ras mutations are insensitive to gefitinib treatment (27), and it is likely that both the absence of P-Akt activity (10) or constitutive P-Akt+ through other non-EGFR targets, such as loss of PTEN function (13,32), may represent additional examples of gefitinib-refractory disease. Until reliable prognostic tools can be prospectively validated in clinical trials, clinicians will need to balance the desire to minimize false hope while maximizing the ability to offer clinically meaningful benefit to their patients.

REFERENCES

(1) Lynch TJ, Bell DW, Sordella R, Gurubhagavatula S, Okimoto RA, Brannigan BW, et al. Activating mutations in the epidermal growth factor receptor underlying responsiveness of non-small-cell lung cancer to gefitinib. N Engl J Med 2004;350:2129–39.[Abstract/Free Full Text]

(2) Paez JG, Janne PA, Lee JC, Tracy S, Greulich H, Gabriel S, et al. EGFR mutations in lung cancer: correlation with clinical response to gefitinib therapy. Science 2004;304:1497–500.[Abstract/Free Full Text]

(3) Pao W, Miller V, Zakowski M, Doherty J, Politi K, Sarkaria I, et al. EGF receptor gene mutations are common in lung cancers from "never smokers" and are associated with sensitivity of tumors to gefitinib and erlotinib. Proc Natl Acad Sci U S A 2004;101:13306–11.[Abstract/Free Full Text]

(4) Mitsudomi T, Kosaka T, Endoh H, Horio Y, Hida T, Mori S, et al. Mutations of the epidermal growth factor receptor gene predict prolonged survival after gefitinib treatment in patients with non-small-cell lung cancer with postoperative recurrence. J Clin Oncol 2005; [Epub ahead of print].

(5) Han SW, Kim TY, Hwang PG, Jeong S, Kim J, Choi IS, et al. Predictive and prognostic impact of epidermal growth factor receptor mutation in non-small-cell lung cancer patients treated with gefitinib. J Clin Oncol 2005; [Epub ahead of print].

(6) Pao W, Miller VA, Politi KA, Riely GJ, Somwar R, Zakowski MF, et al. Acquired resistance of lung adenocarcinomas to gefitinib or erlotinib is associated with a second mutation in the EGFR kinase domain. PLoS Med 2005;2(3):e73.[CrossRef][Medline]

(7) Kobayashi S, Boggon TJ, Dayaram T, Janne PA, Kocher O, Meyerson M, et al. EGFR mutation and resistance of non-small-cell lung cancer to gefitinib. N Engl J Med 2005;352:786–92.[Abstract/Free Full Text]

(8) Cella D, Herbst RS, Lynch TJ, Prager D, Belani CP, Schiller JH, et al. Clinically meaningful improvement in symptoms and quality of life for patients with non-small-cell lung cancer receiving gefitinib in a randomized controlled trial. J Clin Oncol 2005; [Epub ahead of print].

(9) Kris MG, Natale RB, Herbst RS, Lynch TJ Jr., Prager D, Belani CP, et al. Efficacy of gefitinib, an inhibitor of the epidermal growth factor receptor tyrosine kinase, in symptomatic patients with non-small cell lung cancer: a randomized trial. JAMA 2003;290:2149–58.[Abstract/Free Full Text]

(10) Cappuzzo F, Hirsch FR, Rossi E, Bartolini S, Ceresoli GL, Bemis L, et al. Epidermal growth factor receptor gene and protein and gefitinib sensitivity in non-small-cell lung cancer. J Natl Cancer Inst 2005;97:643–55.[Abstract/Free Full Text]

(11) Parra HS, Cavina R, Latteri F, Zucali PA, Campagnoli E, Morenghi E, et al. Analysis of epidermal growth factor receptor expression as a predictive factor for response to gefitinib (‘Iressa’, ZD1839) in non-small-cell lung cancer. Br J Cancer 2004;91:208–12.[ISI][Medline]

(12) Cappuzzo F, Magrini E, Ceresoli GL, Bartolini S, Rossi E, Ludovini V, et al. Akt phosphorylation and gefitinib efficacy in patients with advanced non-small-cell lung cancer. J Natl Cancer Inst 2004;96:1133–41.[Abstract/Free Full Text]

(13) Engelman JA, Janne PA, Mermel C, Pearlberg J, Mukohara T, Fleet C, et al. ErbB-3 mediates phosphoinositide 3-kinase activity in gefitinib-sensitive non-small cell lung cancer cell lines. Proc Natl Acad Sci U S A 2005;102:3788–93.[Abstract/Free Full Text]

(14) Ono M, Hirata A, Kometani T, Miyagawa M, Ueda S, Kinoshita H, et al. Sensitivity to gefitinib (Iressa, ZD1839) in non-small cell lung cancer cell lines correlates with dependence on the epidermal growth factor (EGF) receptor/extracellular signal-regulated kinase 1/2 and EGF receptor/Akt pathway for proliferation. Mol Cancer Ther 2004;3:465–72.[Abstract/Free Full Text]

(15) Finney RE, Bishop JM. Predisposition to neoplastic transformation caused by gene replacement of H-ras1. Science 1993;260:1524–7.[ISI][Medline]

(16) Zhang Z, Wang Y, Vikis HG, Johnson L, Liu G, Li J, et al. Wildtype Kras2 can inhibit lung carcinogenesis in mice. Nat Genet 2001;29:25–33.[CrossRef][ISI][Medline]

(17) Bremner R, Balmain A. Genetic changes in skin tumor progression: correlation between presence of a mutant ras gene and loss of heterozygosity on mouse chromosome 7. Cell 1990;61:407–17.[CrossRef][ISI][Medline]

(18) Wong AJ, Bigner SH, Bigner DD, Kinzler KW, Hamilton SR, Vogelstein B. Increased expression of the epidermal growth factor gene in malignant gliomas is invariably associated with gene amplification. Proc Natl Acad Sci USA 1987;84:6899–903.[Abstract/Free Full Text]

(19) Libermann TA, Nusbaum HR, Razon N, Kris R, Lax I, Soreq H, et al. Amplification, enhanced expression and possible rearrangement of EGF receptor gene in primary human brain tumours of glial origin. Nature 1985;313:144–7.[CrossRef][ISI][Medline]

(20) Tracy S, Mukohara T, Hansen M, Meyerson M, Johnson BE, Janne PA. Gefitinib induces apoptosis in the EGFRL858R non-small-cell lung cancer cell line H3255. Cancer Res 2004;64:7241–4.[Abstract/Free Full Text]

(21) Amann J, Kalyankrishna S, Massion PP, Ohm JE, Girard L, Shigematsu H, et al. Aberrant epidermal growth factor receptor signaling and enhanced sensitivity to EGFR inhibitors in lung cancer. Cancer Res 2005;65:226–35.[Abstract/Free Full Text]

(22) Shiraishi M, Noguchi M, Shimosato Y, Sekiya T. Amplification of protooncogenes in surgical specimens of human lung carcinomas. Cancer Res 1989;49:6474–9.[Abstract]

(23) Hirsch FR, Varella-Garcia M, Bunn PA Jr., Di Maria MV, Veve R, Bremmes RM, et al. Epidermal growth factor receptor in non-small-cell lung carcinomas: correlation between gene copy number and protein expression and impact on prognosis. J Clin Oncol 2003;21:3798–807.[Abstract/Free Full Text]

(24) Shigematsu H, Lin L, Takahashi T, Nomura M, Suzuki M, Wistuba II, et al. Clinical and biological features associated with epidermal growth factor receptor gene mutations in lung cancers. J Natl Cancer Inst 2005;97:339–46.[Abstract/Free Full Text]

(25) Roche-Lestienne C, Preudhomme C. Mutations in the ABL kinase domain pre-exist the onset of imatinib treatment. Semin Hematol 2003;40 (2 Suppl 2):80–2.[CrossRef][ISI][Medline]

(26) Kosaka T, Yatabe Y, Endoh H, Kuwano H, Takahashi T, Mitsudomi T. Mutations of the epidermal growth factor receptor gene in lung cancer: biological and clinical implications. Cancer Res 2004;64:8919–23.[Abstract/Free Full Text]

(27) Pao W, Wang TY, Riely GJ, Miller VA, Pan Q, Ladanyi M, et al. KRAS Mutations and primary resistance of lung adenocarcinomas to gefitinib or erlotinib. PLoS Med 2005;2:e17.[Medline]

(28) Shigematsu H, Takahashi T, Nomura M, Majmudar K, Suzuki M, Lee H, et al. Somatic mutations of the HER2 kinase domain in lung adenocarcinomas. Cancer Res 2005;65:1642–6.[Abstract/Free Full Text]

(29) Stephens P, Hunter C, Bignell G, Edkins S, Davies H, Teague J, et al. Lung cancer: intragenic ERBB2 kinase mutations in tumours. Nature 2004;431:525–6.

(30) Samuels Y, Wang Z, Bardelli A, Silliman N, Ptak J, Szabo S, et al. High frequency of mutations of the PIK3CA gene in human cancers. Science 2004;304:554.[Free Full Text]

(31) Brose MS, Volpe P, Feldman M, Kumar M, Rishi I, Gerrero R, et al. BRAF and RAS mutations in human lung cancer and melanoma. Cancer Res 2002;62:6997–7000.[Abstract/Free Full Text]

(32) Bianco R, Shin I, Ritter CA, Yakes FM, Basso A, Rosen N, et al. Loss of PTEN/MMAC1/TEP in EGF receptor-expressing tumor cells counteracts the antitumor action of EGFR tyrosine kinase inhibitors. Oncogene 2003;22:2812–22.[CrossRef][ISI][Medline]

(33) Graveel C, Su Y, Koeman J, Wang LM, Tessarollo L, Fiscella M, et al. Activating Met mutations produce unique tumor profiles in mice with selective duplication of the mutant allele. Proc Natl Acad Sci USA 2004;101:17198–203.[Abstract/Free Full Text]

(34) Huang SC, Koch CA, Vortmeyer AO, Pack SD, Lichtenauer UD, Mannan P, et al. Duplication of the mutant RET allele in trisomy 10 or loss of the wild-type allele in multiple endocrine neoplasia type 2–associated pheochromocytomos. Cancer Res 2000;60:6223–26.[Abstract/Free Full Text]



             
Copyright © 2005 Oxford University Press (unless otherwise stated)
Oxford University Press Privacy Policy and Legal Statement