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A phase II study of doxorubicin, cyclophosphamide and vindesine with valproic acid in patients with refractory or relapsing small cell lung cancer after platinum derivatives and etoposide
Writing committee: T. Berghmans, A. Scherpereel, L. Willems, C. Mascaux, J.P. Sculier
Study coordinator : T. Berghmans
Data Manager : N. Leclercq
Statistician : M. Paesmans
Final version : 2 July 2008
Ethical committee : 21 August 2008
Activated : 28 August 2008
4. Study population
5. Study design
7. Chemotherapy treatment plan
8. Trial quality control
9. Drug procurement, preparation and storage
10. Side effects
11. Dose adaptation plan
12. Criteria of evaluation
13. Off treatment regimen and off study definitions and procedures
14. Entry and randomisation procedures
15. Data management and trial coordination
16. Ethical considerations
17. Statistical considerations
18. Publication and authorship
Appendix I : Performance scale (Karnofsky)
Appendix II : WHO toxicity criteria
Appendix III : Staging classification
Appendix IV : The World Medical Association Declaration of Helsinski
Participating centers and members:
Small cell lung carcinomas (SCLC) account for around 15% of newly diagnosed lung cancers. The prognosis of this aggressive disease is poor: 10% to 20% of the patients with limited disease will be cured for less than 5% of those with extensive disease. This tumour is initially chemotherapy-sensitive and first-line treatment is associated with high objective response rates (45% to 75%). However, at progression, results of second-line chemotherapy are disappointing.
Platinum derivatives, mainly cisplatin, and etoposide are the most active drugs for the treatment of SCLC. In a meta-analysis performed by the European Lung Cancer Working Party (ELCWP) (1), we showed that chemotherapy regimen including cisplatin without etoposide, etoposide without cisplatin or both increased survival in comparison with combined chemotherapy not including those drugs with respective hazard ratio (HR) of 0.70 (95% confidence interval [CI] 0.41-1.21), 0.72 (95% CI 0.67-0.78) and 0.57 (95% CI 0.51-0.64). Another French meta-analysis reported that patients treated with a cisplatin-containing regimen had a significant reduction in the risk of death at 6 months and 1 year with respective odds ratio (OR) of 0.87 (95% CI 0.75-0.98, p = 0.03) and 0.80 (95% CI 0.69-0.93, p = 0.002) (2).
There is no definite chemotherapy schedule to be proposed for second-line chemotherapy in SCLC. The prognosis of these patients is poor. The American College of Chest Physicians (ACCP) guidelines (3), recommend that patients who experience relapse or have refractory SCLC should be offered further chemotherapy. No particular combination was proposed. The Cancer Care Ontario guidelines (4), based on a systematic review of the literature, suggest that the selection of patients for second-line therapy should be dependent on the treatment-free interval, the extent of response to first-line therapy, the residual toxicity from first-line therapy, and the performance status of the patient. Patients who relapse < 3 months after first-line therapy are commonly called refractory, and patients who relapse 3 months after therapy are labelled as sensitive. However, no specific chemotherapy schedule is proposed except for administering the initial treatment in sensitive tumours. There is no demonstrated active treatment in refractory patients. The ELCWP guidelines (www.elcwp.org) give the same recommendation.
There are few evidences for chemotherapy activity in second-line setting. Two randomised trials compared chemotherapy to symptomatic care only. In an old British study (5), patients initially randomised between 4 or 8 courses of first-line doxorubicin, cyclophosphamide, vincristine (VAC) were further randomised at relapse between second-line methotrexate plus doxorubicin versus no further treatment. Response rate to salvage chemotherapy was 22.3%. Survival was increased in patients treated with chemotherapy at relapse whatever considering those receiving 4 courses (median survival time [MST] of 20 weeks) or 8 courses (MST of 15 weeks) first-line treatment in comparison with those given symptomatic cares only (MST of 11 and 12 weeks). In another phase III study (6), oral topotecan was compared to best supportive care in patients relapsing at least 45 days after initial response to first-line chemotherapy and unfit for further intravenous chemotherapy. Despite a low response rate (RR) of 7%, topotecan administration resulted in better survival rates (HR 0.64; 95% CI 0.45-0.90, p = 0.01) and a slower decline in quality of life. However, haematological toxicity associated with topotecan was substantial with grade 3/4 neutropenia, thrombopenia and anaemia in 33%, 37% and 25% of the patients, respectively. Diarrhoea (6%) and fatigue (4%) were the most commonly non-haematological adverse events.
There are few available randomised trials allowing determining the best chemotherapy regimen for relapsed SCLC, mainly testing VAC and topotecan. In a phase III trial, von Pawel et al (7) compared VAC to oral topotecan in patients relapsing at least 60 days after first-line chemotherapy. RR and median survival times (MST) were respectively 18.3% and 24.7 weeks and 24.3% and 25 weeks, without any statistically significant difference. More grade 4 thrombocytopenia (57.6% versus 14.9%, p<0.001) and grade 3/4 anaemia (42.3% versus 19.8%, p<0.001) occurred in the topotecan arm. Two other randomised trials compared intravenous to oral topotecan (8;9). No significant difference for RR and MST were found. Haematological toxicity was important in both arms.
According to the currently available evidence, a combination of doxorubicin, cyclophosphamide and vincristine appears to be an acceptable therapeutic option in patients progressing after cisplatin plus etoposide although activity, in terms of response rate and survival, remained limited. Among the four vinca-alcaloids available, vincristine and vindesine are currently used in SCLC. The ELCWP conducted a randomised first-line study comparing etoposide plus vindesine with or without cisplatin (10). Response rates (74% and 55%) and median survival times (40 and 45 weeks) were as expected with these combinations whatever considering limited or extensive diseases. In another randomised phase III study, we combined vindesine to ifosfamide and epirubicin to assess the role of accelerated chemotherapy in non pre-treated patients with extensive-disease SCLC (11). Response rates (between 59% and 76%) and median survival, around 9 months, were comparable to other chemotherapy regimens. We also tested the combination of etoposide and vindesine as maintenance therapy in responding SCLC after induction chemotherapy (12) and the same regimen was given to 25 poor performance status patients in a small size English study (13). In all these studies, toxicity, particularly neurotoxicity, was manageable. In second line, vindesine was added to cisplatin for refractory (14) or recurrent (15) SCLC. In the second study, a 19% response rate was achieved.
Few trials have directly compared vindesine to vincristine. A randomised study compared vindesine to vincristine, both combined with cyclophosphamide as first-line chemotherapy in 116 patients with SCLC (16). Response (47% for both regimens) and survival rates (1-year 47% vs 35%) were similar. Neurotoxicity was comparable with no grade 4 and grade 2-3 observed in 5 and 1 patients for vindesine and vincristine regimens, respectively. A similar toxicity profile was described in a randomised study including 63 patients with non small cell lung cancer treated with vindesine or vincristine monotherapy (17). Less neurotoxicity was reported in breast cancer patients treated by doxorubicin with vindesine than with vincristine (18). In recurrent acute lymphoblastic leukemia, predisone-vincristine was compared to prednisone-vindesine (19). More paresthesias, peripheral neuropathy and ileus were noted with the vindesine combination but it was given twice weekly instead of once weekly for vincristine. Inversely, less neurotoxicity was observed with vindesine than with vincristine in another randomised comparison in haematological patients (20). Overall, the effectiveness and the toxicity profile of vincristine and vindesine appeared similar across these randomised trials.
Based on these considerations and the limited effectiveness of available chemotherapy regimens, new treatments including drugs with original activity and/or new mode of administration are needed in order to ameliorate the prognosis of patients with refractory or relapsing SCLC after platinum derivatives and etoposide. Here, we propose to evaluate a novel approach based on the concept of a gene activation therapy. Our working hypothesis relies on the regulation of cell homeostasis, which is the result of a clinical balance between cell proliferation and death. Our tenet is that this lack of death of tumour cells is the result of cell quiescence correlating with an absence of gene expression and, we propose to relieve this silencing block by using a histone deacetylase inhibitor. Due to their ability to remove acetyl groups from the histone acetyl-lysine residues, histone deacetylases (HDAC) are major regulators of gene expression, acting via the local chromatin remodelling. Regulation of gene expression further affects proper cell functioning as well as differentiation and proliferation. HDAC inhibitors exert profound modifications of the cell biology and some of these molecules have been proposed as potential anticancer agents (21). HDAC inhibitors include a variety of compounds belonging to several structural classes: hydroxamic acids (TSA or trichostatin A, SAHA or suberoylanilide hydroxamic acid); cyclic peptides (trapoxin, FR901228 also called depsipeptide), benzamides (MS-27-275) and short-chain fatty acids (butyric acid and valproic acid). These molecules might exhibit pleiotropic effects and modulate different metabolic pathways (22;23).
In particular, valproic acid has demonstrated interesting inhibiting activity on different cell lines and tumour xenografts in mice. Growth inhibition and apoptosis were reported in haematological (leukaemia, lymphoma, multiple myeloma) (24-27) and solid tumours like thyroid, neuroblastoma, melanoma, breast, colon, EBV-positive tumours, prostate, medulloblastoma, endometria, hepatoma and glioma cancer cell lines, either alone or with potentialisation by mitomycin, paclitaxel, doxorubicin or tamoxifene (28-38). Furthermore, among all HDAC inhibitors, valproic acid harbours major advantages (39). Used since decades in the therapy of epilepsy, its pharmacology and toxicology are very well characterized. Its toxicologic profile allows generally a good therapeutic index. Valproic acid has proved to possess considerable teratogenic potential, due in part to some anti-tumour properties.
Some small phase I and II studies have been reported testing the concept of epigenetic modulation with valproic acid in patients. They have mainly been performed in haematological patients (40-46). Valproic acid was administered by the oral route, at different dosages. Action on HDAC was documented at serum therapeutic levels of valproic acid (concentrations used for the treatment of epilepsy) without higher activity in supra-therapeutic levels (40;41;43;46). Three phase I studies were performed in patients with solid tumours. In two studies, increasing intravenous high doses of valproic acid (until 120 to 160 mg/kg) were administered on a short period of time either alone (47) or in combination with epirubicin (48). In the first study (47), neurotoxicity was the dose-limiting toxicity among 26 patients. Induction of histone hyperacetylation and downregulation of HDAC2 in peripheral blood lymphocytes were observed in most patients. No objective response was documented. In the second study (48), histone acetylation was also reported. Nine objective responses were documented (22%). Four patients with SCLC were included of which one had a partial response and another had a stable disease. In a third phase I study (49), oral valproic acid was given for five days at doses of 20-40 mg/kg to twelve untreated women with cervical cancer. There was hyperacetylation of histone H3 and H4 in the tumours of nine and seven patients, respectively without any correlation between acetylation levels and serum levels of valproic acid. Lastly, the ELCWP is currently testing a combination of doxorubicin and valproic acid for relapsing mesothelioma (www.elcwp.org) in a two-step phase II study. The first step was successful with at least one response among the first 16 eligible patients allowing us to continue inclusion as required in the protocol.
We obtained very promising preliminary in vitro data demonstrating the pro-apoptotic potential of valproic acid on tumour cell lines. Four SCLC cell lines (NCI-H146, NCI-H526, NCI-H69 and NCI-N417) were cultivated in presence of 1 mM valproic acid alone or in combination with vindesine (100nM), doxorubicin (100nM) and bis(2-chloroethyl)amine (50µM), the active metabolite of cyclophosphamide (CEA). CEA is a DNA alkylating agent used to mimic nitrogen mustard metabolites. After 24 hours of culture, the rates of apoptosis were determined by flow cytometry after ethanol fixation and propidium iodide staining (sub-G1 peak). The data illustrated in figure 1 are the mean apoptotic rates (± standard deviation) of triplicate analyses. We conclude that, in all SCLC cell lines, valproic acid increments apoptosis induced by the combination of vindesine, CEA and doxorubicin.
There could be some concerns regarding the interaction between valproic acid, cancer and chemotherapy. In a recent article, Singh et al (50) reviewed the carcinoma promoting effects of antiepileptic drugs. They found that phenobarbital demonstrated carcinogenic properties among rodents and was implicated in the development of hepatocellular carcinoma and possibly lung cancer in humans. Phenytoin was suggested to favour the occurrence of lymphoma, myeloma and neuroblastoma. As the evidences remain limited in humans, these drugs are only considered as possible carcinogenics. In contrast, there are indirect evidences for cancer protective effect of valproic acid (50). Furthermore, there is no evidence that valproic acid is promoting cancer in humans, despite its wide use in people with cancers (50-56). A second concern is the possible relationship between antiepileptic drugs and chemotherapy because these drugs are potent inducers or inhibitors of the hepatic cytochrome P450 (CYP). Valproic acid is an important inhibitor of the CYP isoenzymes, the uridinediphosphate glucuronyltransferase and the epoxide hydrolase. A few interactions have been reported. Increased haematological toxicity was noted when valproic acid was added to a regimen combining fotemustine, cisplatin and etoposide (51). More hypersensitivity reactions to procarbazine were observed among patients receiving simultaneously valproic acid (55). Activity of valproic acid was reduced when administered concomitantly with methotrexate (57) or the combination of cisplatin and doxorubicin (56). Based on the literature, Vecht et al recommended that prescription of enzyme-inducing antiepileptic drugs such as phenobarbital and phenytoin should be avoided in patients with cancer, but that valproic acid would be the agent of first choice (58).
Together, these data support a possible use of valproic acid as an anti-cancer drug. Our working hypothesis is based on the ability of HDAC inhibitors to trigger gene expression and subsequently to relieve cells from quiescence. Our tenet supposes that cellular gene activation would reactivate the apoptotic pathways which are frequently blocked in tumour cells. Furthermore, the doses to which valproic acid are effective in our in vitro experiments and in clinical studies (40-46) are those achieved with conventional oral doses for the treatment of epilepsy. As the toxicological profile of valproic acid in humans is well known at these doses, we propose to use valproic acid at the doses recommended for the treatment of epilepsy. Based on our in vitro data and on our clinical experiences (10-12), we will chose the combination of doxorubicin, cyclophosphamide and vindesine (VdsAC) to be combined with valproic acid. The doses of each drug are those commonly used in first and second-line treatment, 45 mg/m², 1 g/m² and 3 mg/m², respectively. The primary aim of this study is to determine if the addition of valproic acid to VdsAC could increase progression-free survival in patients relapsing after first-line chemotherapy including platinum derivatives, cisplatin or carboplatin, and etoposide.
In addition, we aim to assess some molecular biology involved in the growth of solid tumours, mainly angiogenesis and the microenvironment (stroma) (59;60). In lung cancer, neoangiogenesis is associated with stromal inflammatory reaction and local secretion of proteases, pro-inflammatory mediators (chemokines and cytokines) and pro-angiogenic factors (VEGF, FGF-2,…) and their receptors (mostly the VEGF-receptor 2 for the VEGF) (61-67). The different steps of tissue remodeling involve the vascular endothelium, a main actor in carcinogenesis. This tumor angiogenesis is under control of growth factors (VEGF, FGF…) secreted by tumor and stromal cells. In response to these factors, the endothelium releases mediators such as proteoglycanes, which may interact with growth, cytokines and adhesion molecules (integrins) and modulate tumor growth. Thus, the interactions host - tumor are a key in the cancer development. The study of some of the biomolecules involved in this process may help to better understand the carcinogenesis and to develop some new lung cancer biomarkers and perhaps some therapeutic targets. Therefore we would like to evaluate during the trial the following potential biomarkers:
- Endocan is a 50 kDa pulmonary endothelial proteoglycan that binds the integrin LFA-1 on leukocytes surface and may inhibit the interaction between LFA-1 and its ligand ICAM-1 (68). This suggests that endocan may inhibit the migration and/or the LFA-1-dependent activation of leukocytes. In vitro, the glycan part of endocan binds growth factors such as HGF/SF (Hepatocyte Growth Factor / Scatter Factor) and potentiates their mitogenic action on human cells. We confirmed these potential properties of endocan in mouse tumor models (69). In humans, elevated mRNA levels found in some tumor tissues suggested endocan as a gene of bad prognosis in kidney, prostate (70), breast (71) and lung cancers (72). Finally, in a preliminary study, we found elevated tissue and blood levels in patients with lung cancer (73).
- CCL18 is an original chemokine produced specifically in the lung by antigens presenting cells. CCL18 is involved in the recruitment of NK cells, NKT cells and T regulatory cells (Treg), which play a role in the anti-tumour immune response.
- The ADAMs (A Disintegrin And Metalloprotease) are membrane metalloproteases involved in the proteolysis of components of the extracellular matrix, the regulation of intercellular exchanges by the activation of mediators (cytokines, chemokines and growth factors) or the cleavage of receptors, the cell fusion, and the cell adhesion to the matrix. They play a key role in the inflammatory reaction, the tissue remodelling and probably the carcinogenesis. Because the ADAMs are membrane metalloproteases, these proteins cannot be directly assessed and their enzymatic activity is difficult to quantify in biological fluids. Therefore we plan to assess the amount of released ADAMs prodomain, directly reflecting the activated ADAMs.
Figure 1. In vitro experiments in small cell lung cancer lines of doxorubicin, vindesine, bis(2-chloroethyl)amine (CEA) alone or in combination with or without valproic acid
3. Study objectives
3.1. Primary endpoint
To determine 6 months progression-free survival of the combination of doxorubicin, cyclophosphamide and vindesine (VdsAC) plus valproic acid in patients with refractory SCLC after treatment with platinum derivatives, cisplatin or carboplatin, and etoposide.
3.2. Secondary endpoints
- to determine overall survival of the combination of VdsAC plus valproic acid
- to determine response rate of the combination of VdsAC plus valproic acid
- to assess the toxicity of the combination of VdsAC plus valproic acid
- to assess HDAC activity in blood peripheral lymphocytes in patients treated with VdsAC plus valproic acid
- to assess new potential blood and tissue markers of lung cancer pathogenesis or prognosis (endocan, VEGF, VEGF-R2, CCL18, ADAMs, …) in patients treated with VdsAC plus valproic acid
4. Study population
4.1 Criteria of eligibility include :
- Histological or cytological diagnosis of small-cell lung cancer (SCLC)
- SCLC refractory to prior chemotherapy regimen including platinum derivatives (cisplatin or carboplatin) and etoposide, either primary refractory (immediate progression or recurrence less than 3 months after the end of previous chemotherapy) or secondary refractory (sensitive patients to platinum plus etoposide in first-line, progressing or recurring less than 3 months after reintroduction of the same chemotherapy).
- At least one evaluable or measurable lesion
- Availability for participating in the detailed follow-up of the protocol
- Signed informed consent.
4.2 Criteria of ineligibility include :
- Patient who were previously treated with anthracyclin or vinca-alcaloid derivatives or cyclophosphamide
- Performance status < 60 on the Karnofsky scale
- A history of prior malignant tumour, except non-melanoma skin cancer or in situ carcinoma of the cervix or of the bladder or cured malignant tumour (more than 5-year disease free interval)
- A history of prior HIV infection
- Polynuclear cells < 2,000/mm³
- Platelet cells < 100,000/mm³
- Abnormal coagulation tests (aPTT, PTT, prothrombin time) and/or decreased fibrinogen
- Serum bilirubin >1.5 mg/100 ml
- Transaminases more than twice the normal range
- Serum creatinine > 1.5 mg/100 ml
- Recent myocardial infarction (less than 3 months prior to date of diagnosis)
- Congestive cardiac failure (ejection fraction of the left ventricle < 50%) or uncontrolled cardiac arrhythmia
- Uncontrolled infectious disease
- Active epilepsy needing a specific treatment
- Concomitant treatment with IMAO, carbamazepine, mefloquine, phenobarbital, primidone, phenytoïn, lamotrigine, zidovudine
- Pregnancy or refusal to use active contraception
- A known allergy to valproic acid and/or doxorubicin, cyclophosphamide, vindesine
- Serious medical or psychological factors which may prevent adherence to the treatment schedule.
5. Study design
After registration, eligible patients will receive doxorubicin (45 mg/m² day 1) plus cyclophosphamide (1 g/m² day 1) and vindesine (3 mg/m² day 1) every 3 weeks plus valproic acid. Valproic acid will be administered orally at the dose of 20 to 30 mg/kg/day during meals in order to obtain serum concentration in the range of the recommended values for the treatment of epilepsy (50-100 mg/ml). The administration of valproic acid will begin one week before first administration of VdsAC. Chemotherapy will be given every 3 weeks and valproic acid continued during the whole treatment.
Response will be assessed after 3 courses and responders will continue to receive treatment until best response is achieved, unacceptable toxicity or cumulative dose of doxorubicin > 500 mg/m².
At progression, the choice of the treatment will be left at the discretion of the treating physicians.
Patient receive orally valproate acid at the dose of 20 to 30 mg/kg/day in order to obtain serum concentration between 50-100 mg/ml
When stable valproate therapeutic concentration is obtained,
continue oral valproate acid at the same dosage unless serum concentration is under or above 50-100 mg/ml and administer
doxorubicin 45 mg/m², cyclophosphamide 1g/m² and vindesine 3 mg/m² every 3 weeks
After 3 courses of chemotherapy
Continue until best response, excessive toxicity or cumulative dose of doxorubicin > 500 mg/m²
3 further courses of chemotherapy
6.1. Investigations required for chemotherapy
6.1.1. Initial investigations
a. History and physical examination
b. Weight, height, surface area and record of performance status.
c. Biological tests including haemoglobin, white blood cell count, differential count, platelet count; serum urea; serum creatinine, serum bilirubin, alkaline phosphatase, SGOT; SGPT, LDH, calcium, uric acid, Mg, electrolytes (Na, K, Cl, HCO3), aPTT, PTT, prothrombin time and fibrinogen; plasma level of valproic acid will be performed before first administration of doxorubicin.
d. Chest X-ray (P.A. and lateral) and Chest CT scan
e. Electrocardiogram and Isotopic or echographic left ventricular fraction assessment
f. Bone isotopic scintigraphy (with X rays of suspected lesions).
g. CT Scan or echography of the liver and adrenals.
If a PET scan is performed, bone isotopic scintigraphy can be avoided and abdominal CT scan or echography will be performed in case of positive lesions.
h. Brain NMR or CT Scan.
i. Bronchoscopy with biopsy (optional).
6.1.2. During treatment
a. Between cycles: haemoglobin, white blood cell count, differential count, platelet count; uraemia and serum creatinine. This analysis will be performed weekly between the first and second cycles and, in absence of excessive toxicity, once between subsequent cycles of chemotherapy
b. Before each new course of chemotherapy: history and physical examination; weight, height, surface area and record of performance status; biological tests as initially; plasma level of valproic acid, chest X-ray (P.A. and lateral) and electrocardiogram.
c. After the 3rd course and every 3 subsequent courses of chemotherapy: same investigations as initially, including chest CT scan.
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