1 Mayo Clinic and Mayo Foundation, Rochester, MN 55905; 2 Wake Forest University School of Medicine, Winston-Salem, NC 27157; 3 Missouri Valley Cancer Consortium, Omaha, NE 68106; 4 Iowa Oncology Research Association CCOP, Des Moines, IA 50309-1014; 5 Scottsdale CCOP, Scottsdale, AZ 85259-5404, USA
* Correspondence to: Dr A. A. Adjei, Division of Medical Oncology, Mayo Clinic, 200 First St., SW Rochester, MN 55905, USA. Tel: +1-507-538-0548; Fax: +1-507-284-1803; E-mail: adjei.alex{at}mayo.edu
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Abstract |
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Patients and methods: Patients with c-kit-expressing SCLC (1+ by immunohistochemistry) were enrolled in two groups. Arm A included patients with disease progression <3 months and arm B included patients with disease progression
3 months after previous treatment. Imatinib was administered at a dose of 400 mg b.i.d. continuously, with a cycle length of 28 days. A single stage Simon design with a planned interim analysis was used to evaluate the 16-week progression free rate in each arm.
Results: A total of 29 evaluable patients were entered into the study (seven in arm A, median age 68; 22 in arm B, median age 64.5). Median number of treatment cycles was one in both arms. Grade 3+ non-hematologic adverse events were seen in 15 (52%) patients, with nausea, vomiting, dyspnea, fatigue, anorexia and dehydration each occurring in at least 10% of patients. Median survival was 3.9 and 5.3 months and median time to progression was 1 and 1.1 months for arms A and B, respectively. Enrollment to arm A was temporarily suspended prior to reaching interim analysis due to striking early disease progression (29%), early deaths (29%) and patient refusal (42%). No objective responses and no confirmed stable disease 6 weeks were seen in either arm. Accrual was permanently terminated to both arms as only one patient was progression-free at 16 weeks.
Conclusion: Imatinib failed to demonstrate any clinical activity in spite of patient selection for c-kit-expressing SCLC. Our results strengthen the collective evidence that prediction of efficacy of novel therapeutic agents based on target expression, rather than pathway activation (for example, through activating mutations), may not be a valid paradigm for drug development.
Key words: c-kit, imatinib, kinase inhibitors, small-cell lung cancer
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Introduction |
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Autocrine or paracrine activation of growth has been a popular hypothesis used to explain deregulated growth of SCLC. SCLC tumors and cultured cell lines produce a wide variety of peptide hormones and receptors that stimulate growth [46
]. High level of expression of c-kit and its ligand, stem cell factor (SCF), has been widely found in both SCLC tumors and established cell cultures [7
9
]. The role of the c-kit autocrine loop in SCLC has been well studied [10
]. This autocrine loop not only functions cooperatively with other SCLC autocrine loops but, more importantly, seems to confer a tumor survival advantage in SCLC [10
]. More importantly, in vitro treatment with c-kit tyrosine kinase inhibitors reversed apoptosis resistance to growth factor deprivation in H526 cells, a SCLC cell line with co-expression of c-kit and SCF [11
]. The ensuing growth inhibition was well correlated with the inhibition of c-kit tyrosine phosphorylation.
Imatinib mesylate (STI571) is an orally bioavailable promiscuous inhibitor of several tyrosine kinases due to the highly conserved structure of tyrosine kinase at the ATP binding site. Known targets of inhibition include the bcr-abl kinase, PDGF and c-kit. In a pivotal study, Krystal et al. [12] demonstrated that pretreatment of H526 cells with imatinib mesylate inhibited SCF-mediated kit activation. Inhibition of serum-dependent proliferation of multiple SCLC cell lines was established at an approximate IC50 of 5 µmol/l. It was also demonstrated that imatinib mesylate sufficiently blocked the signal transduction cascade triggered by c-kit activation. A separate study of SCLC cell lines documented a dose-dependent inhibition of tyrosine phosphorylation and in vitro kinase activity (at 5 µM) of c-kit using imatinib mesylate [13
].
With the success of imatinib mesylate in the treatment of c-kit expressing gastrointestinal stromal tumor, a rare mesenchymal malignancy previously resistant to all treatment approaches, its use in other c-kit positive tumors such as SCLC presented a novel molecular therapeutic approach. Johnson et al. [14] reported their findings in 19 patients with SCLC, either with chemosensitive relapsed disease or previously untreated extensive stage, who were enrolled in a phase II trial using imatinib mesylate 600 mg daily for up to 12 months. There was no observed antitumor activity in this group of patients. However, the study was inconclusive, as it was weakened by at least two limitations: only 21% of the patients had c-kit positive tumors by immunohistochemstry and 26% of the patients had non-SCLC histology upon an unplanned post-hoc central pathology review. We therefore sought to evaluate clinical activity of imatinib in the treatment of patients with immunohistochemistry-confirmed c-kit + SCLC, utilizing a higher dose of 800 mg/day.
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Patients and methods |
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Patients were excluded from the study if they had a concurrent or past history of another malignancy (except skin cancer or localized prostate cancer) within the preceding 3 years, had uncontrolled symptoms from CNS metastasis (i.e. patients with treated CNS metastasis who were clinically stable were eligible), or had uncontrolled infection or precarious cardiovascular reserve (e.g. unstable angina pectoris, myocardial infarction or class III/IV heart failure within the preceding 3 months). Patients who received prior treatment with imatinib were not allowed to enroll in the study. Patients were also excluded if they had major surgery or preceding systemic chemotherapy within 21 days, or radiation therapy within 14 days prior to treatment. As the study involved an investigational agent whose genotoxic, mutagenic and teratogenic effects on the developing fetus and newborn were unknown, men or women of childbearing potential who were unwilling to employ adequate contraception as well as pregnant or nursing women were excluded.
Pathology and immunohistochemistry
Histology slides were reviewed and diagnosis of small-cell carcinoma confirmed, according to the 2004 World Health Organization Classification of Tumors of the Lung, by one of the authors (M.C.A.) prior to evaluation of c-kit expression in the tumor tissues. Nine cases were deemed ineligible for subsequent registration: six cases due to a change in the histologic diagnosis and three cases due to insufficient tumor tissue for evaluation. Immunohistochemical staining for KIT (CD117) was performed using a 1:250 dilution of the rabbit polyclonal antibody A4502 (DAKO, Carpinteria, CA) with the EnVision detection system. Appropriate positive and negative controls were used. The tumors were scored as 0 if <5% of cells were positive, as 1+ if 5%25% of cells were positive, as 2+ if 26%50% of cells positive and 3+ if >50% of cells were positive (Figure 1).
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At the time of interim analysis, arm A was to be considered ineffective and accrual terminated if only three at most out of the first 17 evaluable patients were progression free at 16 weeks. Similarly, arm B was to be considered ineffective and accrual terminated if only seven at most out of the first 22 evaluable patients were progression free at 16 weeks. Accrual was suspended during the interim analysis evaluation.
The proportion of successes was estimated by the number of successes divided by the total number of evaluable patients and exact binomial confidence intervals for the true treatment success rate were constructed according to the method of Duffy and Santner for each arm [17]. Time to disease progression was defined as the time from registration to the date of first documented disease progression. Patients who died without disease progression were censored at the date of their last evaluation. If a patient died without documentation of disease progression, the patient was considered to have had tumor progression at the time of death, unless there was sufficient documented evidence to conclude that progression did not occur prior to death. Survival time was defined as the time from registration to death due to any cause. The distribution of time to disease progression and survival time was estimated in each arm using the method of KaplanMeier [18
]. All adverse events, regardless of attribution, were summarized (overall and by arm). Presence of c-kit mutations in tumor tissue were to be analyzed and correlated with treatment response if any cohort of patients met the definition for treatment success.
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Results |
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There were no observed responses and no confirmed disease stability (6 weeks). There was one patient on arm B who had stable disease for 31 weeks. He refused further treatment after cycle 1 and was thus not subsequently evaluated for response. All other patients were either not evaluable for the primary end point as they went off study or progressed at their first evaluation (either cycle 1 or cycle 2). The median survival was 3.9 and 5.3 months and median time to progression was 1 and 1.1 months for arms A and B, respectively (Figure 3). There was no statistically significant association between the degree of c-kit expression and overall survival (log rank P value = 0.7). The mutational status of the c-kit receptor was not investigated further since there were no treatment successes, thus making it impossible to draw any relevant conclusions from such testing.
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Discussion |
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c-kit is expressed by a variety of neoplasms, including SCLC [21]. Whether a c-kit tyrosine kinase inhibitor can induce tumor regression or growth inhibition depends largely on the degree of dependence on c-kit activity by the particular tumor cells under study. Single-agent therapy with tyrosine kinase inhibitors, such as imatinib mesylate, will be effective only against malignancies for which proliferation and/or survival are driven predominantly by the tyrosine kinase activity [22
]. In this regard, clinical experience so far corroborates the mounting evidence that the greatest antitumor efficacy is seen in tumor types that have activating mutations of the tyrosine kinase [23
27
]. KIT mutations are most commonly found in cell types (mast cells, germ cells, ICC and hematopoietic stem cells) for which development depends on normal activity of the SCF/KIT axis. On the contrary, the putative cells of origin of SCLC are not developmentally dependent on c-kit [21
] and evidence to support the inference thereby of the lack of activating mutations in c-kit positive SCLC has been reported [28
, 29
]. Moreover, supporting evidence of lack of activity of imatinib in a murine xenograft model has been recently reported, in spite of activity demonstrable in corresponding in vitro studies using the same cell lines from which the xenografts were derived [30
].
A potential limitation of the study is the use of tumor tissue obtained for initial diagnosis (prior to first-line chemotherapy) in the immunohistochemistry analyses. It has been reported that up to 50% of SCLC cases that were c-kit-positive at the time of initial diagnosis were subsequently found to be c-kit-negative using a post-chemotherapy relapse specimen [31]. Moreover, current immunohistochemistry techniques are constrained to merely demonstrating c-kit expression, whereas most preclinical data of imatinib activity in SCLC were in cell lines that co-expressed its cognate ligand SCF [10
12
]. Another condition controlled in the preclinical study of imatinib was serum deprivation [12
, 13
], which self-evidently cannot be achieved in the clinical setting.
Our negative study serves as an important reminder of faulty assumptions that investigators need to be aware of in the design of future studies to test novel agents in solid tumors. Target expression by itself may not be enough to justify the study of kinase inhibitors in particular diseases. Evidence of activation of the relevant pathways need to be demonstrated. A more comprehensive preclinical evaluation must include in vivo studies of targeted agents to avoid waste of limited resources with conducting potentially ineffective trials [32]. Our data support the growing evidence that antitumor response to small molecule kinase inhibitors may not correlate with protein expression. Further understanding of the underlying molecular biology of SCLC is needed to facilitate the identification of appropriate targets for subsequent drug development and clinical testing. The redundancy and lack of dominance among the multiple aberrant growth and survival pathways make any single agent therapeutic intervention unlikely to be successful in SCLC. If imatinib is studied further in this disease, patients with tumors harboring c-kit mutations should be selected for testing.
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Acknowledgements |
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Additional participating institutions include: Medcenter One Health Systems, Bismarck, ND 58506 (E. J. Wos); Wichita Community Clinical Oncology Program, Wichita, KS 672143882 (S. Dakhil); Duluth CCOP, Duluth, MN 55805 (D. Nikcevich); Cedar Rapids Oncology Project CCOP, Cedar Rapids, IA 52403 (M. Wiesenfeld).
Received for publication May 18, 2005. Revision received June 17, 2005. Accepted for publication June 21, 2005.
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