Intermittent Chemotherapy and Erlotinib for Nonsmokers or Light Smokers with Advanced Adenocarcinoma of the Lung: A Phase II Clinical Trial

Background. Intermittent application of chemotherapy and tyrosine kinase inhibitors may avoid antagonism between the two classes of drugs. This hypothesis was tested in a Phase II clinical trial. Patients and Methods. Eligible patients were nonsmokers or light smokers, chemo-naïve, with metastatic adenocarcinoma of the lung. Treatment: 4 to 6 cycles of gemcitabine 1250 mg/m2 on days 1 and 4, cisplatin 75 mg/m2 on day 2, and erlotnib 150 mg daily on days 5–15, followed by erlotinib as maintenance. Results. 24 patients entered the trial. Four pts had grade 3 toxicity. Complete remission (CR) and partial remission (PR) were seen in 5 pts and 9 pts, respectively (response rate 58%). Median time to progression (TTP) was 13.4 months and median overall survival (OS) was 23 months. When compared to patients with negative or unknown status of EGFR mutations, 8 patients with EGFR gene activating mutations had significantly superior experience: 4 CR and 4 PR, with median TTP 21.5 months and OS 24.2 months (P < .05). Conclusions. Intermittent schedule with gemcitabine, cisplatin and erlotinib has mild toxicity. For patients who are positive for EGFR gene activating mutations, this treatment offers excellent response rate, time to progression and survival.


Introduction
To the surprise and deep disappointment of all involved in the treatment of lung cancer, several large trials did not demonstrate any benefit of tyrosine kinase inhibitors (TKIs) as an addition to chemotherapy [1][2][3]. Virtually all further clinical research on combinations of TKIs and chemotherapy was then abandoned. Basic and clinical research then focused on mutations of the gene for epidermal growth factor receptor (EGFR) as a predictive factor for response to monotherapy with TKIs and to development of new compounds with broader and/or irreversible inhibition.
The biological basis for the negative experience with combined treatment was never given proper attention. Gefitinib and erlotinib met all three standard criteria for inclusion in a combination with chemotherapy: activity as monotherapy, different mechanism of action, and different toxicity. Why, then, did the combination not work? As explained in a recent editorial [4], we believe that the cells of tumors sensitive to TKIs are pushed into the G-0 phase of the cell cycle and therefore become resistant to cytotoxic drugs. If antagonism between the two classes of drugs is really the biological basis for the aforementioned negative experience, then an optimal combination of TKIs and chemotherapy should be in an intermittent, rather than a continuous schedule.
This brief report presents a single-institution experience on intermittent chemotherapy and TKI in a small series of patients with advanced adenocarcinoma of the lung. Our hypothesis was that intermittent treatment would lead to superior time to progression, when compared to experience with chemotherapy alone. If confirmed, such a result would be a solid basis for a randomised clinical trial.

Inclusion Criteria.
Patients eligible for the trial were chemonaïve with microscopically confirmed adenocarcinoma of the lung, had stage III B (wet) or IV according to UICC-TNM classification (6th edition), had smoking history of less than 10 packs in years, had an ECOG performance status 0 or 1, and had adequate parameters of hematological, liver, and renal function to receive cisplatinbased chemotherapy. In the absence of neurological symptoms, patients with brain metastases were eligible and were treated with brain irradiation only in case of intracranial progression. All patients were fully informed and gave written consent to participate in the trial.

Initial Diagnostics.
All patients had their diagnosis confirmed by biopsy or cytology. At the time when the trial was initiated, testing for EGFR mutations was not available.
Within three weeks prior to treatment, the precise extent of the disease was determined by chest X-ray and CT scanning of the chest, upper abdomen, and brain. Since 2008, PET-CT scanning has been available and included in the initial diagnostics and in followup. Cycle was repeated on day 22. Patients received 4 to 6 cycles of intermittent treatment. The number of cycles depended on tolerance to cisplatinbased chemotherapy and was determined individually. Immediately after the last cycle, patients continued with erlotinib 150 mg/m 2 daily continuously until progression or unacceptable toxicity.

Monitoring for Response, Time of Progression, and Followup.
Definition of complete response (CR), partial response (PR), stable disease (SD), and progression followed the RECIST criteria [5].
The first evaluation of response was done during the third cycle of intermittent therapy, with confirmation of response during the fifth cycle. After 4 cycles, patients were seen every second month. Control radiological examinations were repeated every 2 months for chest X-ray, every 4 months for CT, and at 6 and 12 months for PET-CT (only patients who had this examination during their initial diagnostics).

Posttreatment Analysis of Archived Bioptic Material.
In October 2010, all biopsy samples were reviewed, and specimens with more than 10% of tumour tissue were analyzed. Genomic DNA was extracted from formalin-fixed, paraffin-embedded tissue sections using QIAAmp DNA FFPE tissue kit (Qiagen, Hilden, Germany) according to the manufacturer's instructions. Quantification of extracted DNA was done on Qubit Fluorometer (Invitrogen, Carlsbad, USA). To detect EGFR gene-activating mutations, we used TheraScreen EGFR29 Mutation Kit (DxS Diagnostics, Qiagen, Manchester, UK). All realtime PCR reactions were performed in a 25 μL final volume on ABI 7500 instrument (Applied Biosystems, Carlsbad, USA).

Endpoints and Statistical
Planning. The primary endpoint was time to progression. Secondary endpoints were response rate, toxicity, and overall survival.
After standard chemotherapy for metastatic nonsmall cell carcinoma, the expected TTP is 5 months. The size of this single-arm nonrandomised trial of intermittent therapy was based on the assumption of 9 months as the median time to progression (TTP). To obtain such a result with a confidence interval of 6-12 months, we planned to recruit 40 patients.

Patient Population.
Between September 2005 and July 2010, 25 patients were recruited into the trial. One patient was later found to have metastatic carcinoma of the pancreas rather than primary lung cancer and was excluded from all further analyses.
With 12 patients each, male and female patients were equally represented. Median age was 50 years (range: 25 to 73 years). Twelve patients were never-smokers, and most were in good general condition (PS 0-1 for 21 patients). With the exception of a single patient with "wet" stage III B, all other patients had stage IV disease. Bone metastases were the most common site of metastatic disease, followed by pleura/pericardium, contralateral lung metastases, and liver. Two or more sites of metastatic disease were documented in 4 and 12 patients, respectively (Table 1).

Treatment.
The actual number of cycles of intermittent therapy was from 1 to 6 cycles (median: 4 cycles). Due to early progression, one patient did not receive erlotinib as maintenance treatment. In October 2010, 7 patients were still on maintenance treatment with erlotinib, and an additional patient stopped treatment with erlotinib after 12 months in PET-CT confirmed complete remission (Figures 1 and  2). For the remaining patients, median total duration of treatment was 10 months.
No patient experienced grade 4 or greater toxicity.

Response to Treatment, Time to Progression, and Survival.
All patients are evaluable for response, and no patient has been lost to followup. For the whole group of 24 patients, complete remission (CR) was seen in 5 pts; partial remission (PR) in 9 pts (response rate 58%), minimal response or stable disease (SD) in 8 pts, and progression in 2 pts. A clear and statistically significant (P < .05) correlation was seen between the presence of activating EGFR mutations and response. Among the 8 patients who were positive for EGFR gene-activating mutations, 4 complete and 4 partial remissions were seen. On the other hand, no CR and only 2 PR were seen among the 10 patients negative for mutations ( Table 2). For the whole group, median time to progression (TTP) was 13.4 months, and median overall survival (OS) was 23 months. Again, patients positive for EGFR gene-activating mutations had-superior experience. Median TTP and OS for this group was 21.5 months and 24.2 months, respectively. For patients without EGFR mutations, TTP was 5 months, and OS was 7 months (Table 2 and Figures 3 and 4).

Discussion and Conclusions
This clinical trial was launched at a time when routine testing for EGFR gene-activating mutations was not yet available. Selection of patients for a combination of chemotherapy and erlotinib was made on the basis of classical histopathology (adenocarcinoma) and smoking status.
Recent developments led to premature closure of our trial. Since testing for EGFR gene mutations is now available, it is clear that patients with activating mutations are those who really benefit from TKIs. In addition, standard firstline treatment for patients with activating EGFR mutations is now monotherapy with a TKI [6,7]. Since continuing a trial with the same selection criteria and without considering the status of EGFR gene activating mutations was not justified, the research group made a decision to close the trial and analyse the experience. In order to get a longer interval for intermittent erlotinib, gemcitabine was given on days 1 and 4 of the cycle. When compared to the standard day 1 and day 8 schedule, this minor modification in timing of cytotoxic drugs did not have any adverse effect on the tolerance to treatment. Clearly, other platin-based schedules which apply chemotherapy on a 3-weekly basis (such as pemetrexed-cisplatin or paclitaxelcarboplatin) can offer an even longer interval for TKIs and might be considered for future trials of intermittent treatment.
Two other groups recently reported promising experience with intermittent chemotherapy and TKIs. In a trial from the USA, two schedules of intermittent treatment were tested [6]. In combination with pemetrexed (500 mg/m 2 on day 1), erlotinib was given either as a pulse application in a high dose (range: 800 to 1400 mg) given on days 2, 9   et al. [7]. This study from Asia compared gemcitabine and either cisplatin or carboplatin to a schedule with addition of intermittent application of erlotinib (150 mg on days 14 to 28 of the cycle) and reported significantly superior TTP with the intermittent schedule. Their experience is most valuable but may not be of direct relevance for the rest of the world, due to the well-known differences in sensitivity of lung cancer to TKIs between Asian and Caucasian patients. Despite its small size, our trial can offer valuable experience for further research on optimisation of treatment with combinations of chemotherapy and TKIs. Looking at the whole series of patients, we can conclude that intermittent chemotherapy and erlotinib is a treatment of very low toxicity. It is also clear that the efficacy of treatment is closely related to the presence or absence of EGFR gene-activating mutations.
The most important finding is the excellent response rate with a substantial proportion of complete responses and prolonged TTP and OS for patients positive for EGFR gene-activating mutations. For many years, the maximal expectation of a patient with metastatic nonsmall cell lung cancer was a partial remission of relatively short duration in the range of 5 to 9 months. With intermittent treatment, we now see durable complete remissions in a subpopulation of patients. While the number of patients in our trial is small and any definitive conclusion would be premature, we nevertheless believe that further research of intermittent therapy for patients positive for EGFR gene-activating mutations is warranted. A randomised trial comparing first-line TKI as monotherapy to the intermittent schedule should clarify the real value of this new approach.