Safety and Efficacy of Dacomitinib in Korean Patients with KRAS Wild-Type Advanced Non–Small-Cell Lung Cancer Refractory to Chemotherapy and Erlotinib or Gefitinib: A Phase I/II Trial

IntroductionDacomitinib (PF-00299804), an irreversible pan-human epidermal growth factor receptor ([HER]-1/EGFR, HER-2, and HER-4) tyrosine kinase inhibitor, demonstrated antitumor activity in Western patients with non–small-cell lung cancer (NSCLC) at a dose of 45 mg once daily. We report data from a phase I/II, multicenter, open-label study of Korean patients with refractory KRAS wild-type adenocarcinoma NSCLC (defined as patients with evidence of disease progression during or within 6 months of treatment with chemotherapy and gefitinib or erlotinib).MethodsThe phase I dose-finding portion identified the recommended phase II dose (RP2D) in Korean patients, evaluated safety, and characterized the pharmacokinetics of dacomitinib. In the phase II portion, patients received dacomitinib at the RP2D. The primary end point was progression-free survival at 4 months (PFS4m).ResultsTwelve patients enrolled in phase I, and 43 patients enrolled in phase II at the RP2D of 45 mg once daily. In phase II, PFS4m was 47.2% (95% confidence interval [CI], 31.6–61.3; one-sided p-value = 0.0007). Median PFS was 15.4 weeks (95% CI, 9.7–17.6); median overall survival was 46.3 weeks (95% CI, 32.7–not reached); and the objective response rate was 17.1% (95% CI, 7.2–32.1). Common treatment-related adverse events were dermatitis acneiform, diarrhea, and paronychia; there were no treatment-related grade 4 or 5 adverse events. Pharmacokinetic parameters of dacomitinib in Korean patients were similar to those reported in Western patients. By patient report, NSCLC symptoms “cough” and “pain” showed improvement within 3 weeks of initiating treatment.ConclusionsDacomitinib was well tolerated and had antitumor activity in Korean patients with NSCLC who had previously progressed on chemotherapy and an epidermal growth factor receptor tyrosine kinase inhibitor

Dacomitinib, an irreversible Pan-ErbB inhibitor significantly abrogates growth in head and neck cancer models that exhibit low response to cetuximab.

Aberrant epidermal growth factor (EGF) signaling is associated with tumor growth in squamous cell carcinoma of the head and neck in humans (HNSCC), and is a major focus of targeted therapy. Cetuximab, a monoclonal antibody against EGFR, has been successful at prolonging survival but has only a 10% tumor shrinkage response rate in a clinical setting. The goal of this study was to compare dacomitinib (PF-00299804), a next generation small molecule tyrosine kinase inhibitor that irreversibly blocks multiple HER family receptors (HER-1 (EGFR), HER-2 and HER-4 tyrosine kinases), to cetuximab, the current FDA approved anti-EGFR medication for HNSCC and erlotinib, an EGFR specific small molecule tyrosine kinase inhibitor. Dacomitinib, erlotinib and cetuximab were tested in a panel of 27 HNSCC cell lines. Treatment with 100 ug/ml of cetuximab or 1 uM of erlotinib inhibited growth by at least 50% in 7/27 cell lines, while treatment with 1 uM of dacomitinib had similar growth inhibition in 17/27 lines. Cell lines representing three levels of sensitivity to dacomitinib were further examined using Western blots, cell cycle and apoptosis analysis. Treatment with 100 nM of dacomitinib reduced EGFR activity and downstream AKT and ERK pathways more effectively than treatment with 100 ug/ml of cetuximab in all ten tested lines. Although both compounds induced apoptosis at similar levels, dacomitinib caused greater G0/G1 arrest. Sensitivity to EGFR blockade was associated with levels of EGFR and ERK and was not associated with common oncogenic mutations and copy number variations. Phosphorylated and total EGFR and ERK levels correlate with sensitivity to both cetuximab and dacomitinib. Three of the four lines in the exquisitely sensitive group had the highest levels of phosphorylated and total EGFR and ERK among the ten lines selected, while the three resistant lines collectively had the lowest levels. Neither pAKT nor tAKT was associated with sensitivity

Dacomitinib IC50 g of ten head and neck cancer cell lines representing three different levels of sensitivity to dacomitinib on a log scale.

<p>Four cell lines were selected from the exquisitely sensitive group (IC50 g <10 nM), three from the moderate group (IC50 g10 nM - 1 uM) and three from the resistant group (IC50 g >1 uM). These ten lines are used in the Western blot and flow cytometry experiments. Average IC50 g of the selected sensitive lines is 5 nM; moderate 75 nM; resistant 2186 nM.</p

Growth-inhibitory effects of dacomitinib and cetuximab on head and neck cancer cell lines.

<p>Cells were counted after five days of treatment. SE bars were derived from experiments repeated at least twice. A. Dacomitinib IC50 g values are arranged from lowest to highest IC50 g on a log scale. Cells were treated at concentrations from 0.001 to 10 uM. B. Percentage growth inhibition with cetuximab treatment. Cells were treated at a fixed dose of 100 ug/mL. C. Erlotinib IC50 g values are arranged from lowest to highest IC50 g on a log scale. Cells were treated at concentrations from 0.015 to 10 uM. Red indicates the sensitivity cutoff. For dacomitinib and erlotinib, the sensitivity cutoff is set at 1 uM, and for cetuximab it’s set at 50%.</p

A. Effects of dacomitinib, erlotinib and cetuximab on phosphorylated and total EGFR, ERK, and AKT.

<p>Cells were cultured to log-phase and treated with 100 nM dacomitinib, 100 nM erlotinib or 100 ug/mL cetuximab for one hour. Cells lysates were then harvested and protein was resolved using Western blot analysis. B. Western blot images were quantified using ImageJ software. Protein levels were quantitated for each cell line. Phosphorylated ERK and AKT were normalized to alpha tubulin and presented as a % of the control.</p

Effects of dacomitinib and cetuximab on cell cycle.

<p>Cells were treated with 100 nM dacomitinib or 100 ug/mL cetuximab for five days before analysis using flow cytometry. Data shown is average percentage of cells in the group assignments from <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0056112#pone-0056112-g002" target="_blank">Figure 2</a>. A. Change in percentage of cells in G0/G1 phase. B. Change in percentage of cells in S phase. C. Change in percentage of cells in G2 phase. *. p<0.05, student’s t-test.</p

A. Effects of dacomitinib and cetuximab on phosphorylated and total ERK.

<p>Cells were cultured to log-phase and treated with 100 nM dacomitinib or 100 ug/mL cetuximab for 1 hour, with or without treatment with 10 ng/mL recombinant EGF ligand. Cells lysates were then harvested and protein was resolved using Western blot analysis. B. Western blot images were quantified using ImageJ software. Protein levels were quantitated for each cell line and were averaged by group. Phosphorylated ERK was normalized to total ERK. *. p<0.05, **. p<0.01, student’s t-test.</p

A. Effects of dacomitinib and cetuximab on phosphorylated and total AKT.

<p>Cells were cultured to log-phase and treated with 100 nM dacomitinib or 100 ug/mL cetuximab for 1 hour, with or without treatment with 10 ng/mL recombinant EGF ligand. Cells lysates were then harvested and protein was resolved using Western blot analysis. B. Western blot images were quantified using ImageJ software. Protein levels were quantitated for each cell line and were averaged by group. Phosphorylated AKT was normalized to total AKT. *. p<0.05, student’s t-test.</p

Dacomitinib and cetuximab effect on apoptosis.

<p>A. Changes in % living cells. B. Changes in cell in early apoptosis. C. Changes in cells in late apoptosis. D. Changes in % dead cells.</p

Panel of HNSCC cell lines showing growth-inhibition effects of dacomitinib and cetuximab, mutation status of K-RAS and PIK3CA hotspots (as detected by PCR and sequencing), EGFR amplification status as detected by FISH (presented as ratio of EGFR gene to centromere 7), and anatomical category of original tumor primary site.

<p>Categories encompass the following subsites: oral cavity; front 2/3 of tongue, floor of mouth, alveolar ridge. Hypopharynx; hypopharynx. Larynx; larynx, supraglottis. Oropharynx; base of tongue, tonsil, tonsillar pillar. IC50 g is Dacomitinib IC50 g and Cetux. % Inhib. is Cetuximab percent inhibition.</p>*<p>EGFR:Centromere 7. Dacom.</p>**<p>Ploidy and copy number were variable in these cell lines, and in the table we use the most common copy number.</p