This phase I clinical trial in Japan was conducted in accordance with (1) World Medical Association Declaration of Helsinki: Recommendations Guiding Medical Doctors in Biomedical Research Involving Human Subjects (1996), (2) the Japanese Pharmaceutical Affairs Law and Guidelines for Implementation of Clinical Studies for Drugs (Japanese GCP).

Patients were enrolled in this study if they met all of the following criteria: histologic or cytologic diagnosis of NSCLC; no surgery within 2 weeks, no radiotherapy or chemotherapy within 3 weeks prior to study entry (within 6 weeks for nitrosourea or mitomycin C); a measurable or evaluable lesion; a performance status of 0 or 1 on the Eastern Cooperative Oncology Group (ECOG) scale; adequate bone marrow function (leukocyte count, 4,000–120,000/μL; neutrophil count ≥1,500/μL; platelet count ≥100,000/μL), normal hepatic function (bilirubin <1.5 × the upper limit of normal; aspartate amino-transferase (AST) and alanine amino-transferase (ALT) levels <2.5 × the upper limit of normal), and renal function (creatinine ≤ the upper limit of normal and creatinine clearance ≥60 mL/min); between 20 and 74 years of age; written informed consent to the study. Patients were ineligible if they had serious infectious diseases or other severe complications (interstitial pneumonia, uncontrollable diabetes or hypertension); atrial fibrillation, serious arrhythmias, or uncontrollable heart failure in previous 6 months; massive pleural or pericardial effusion; symptomatic brain metastases; active concurrent malignancies; lactating or pregnant women, or those attempting to become pregnant; men not willing to use contraception; hearing impairment or neuropathy; had a history of a drug allergy (alcohol, cremophor, etc.); had other medical problems severe enough to prevent compliance with the protocol. The study was approved in advance by the Institutional Review Board and by the Hospital Ethics Committee.

BNP7787 was supplied from BioNumerik Pharmaceuticals, Inc., San Antonio, TX as a lyophilized powder containing 2 or 10 g of the formulated drug. The starting dose of BNP7787 was 4.1 g/m². This starting dose was based on the analysis and calculation of a BNP7787:cisplatin = 50:1 molar ratio derived from the demonstration of partial cisplatin nephroprotection in rats and elucidating this important relationship to formulate and justify a safe starting dose in humans [⁷], as well as prior safety data from phase I clinical trials. Dose escalation steps of BNP7787 in cohorts of at least three patients each according to the protocol were 8.2, 12.3, 18.4, 27.6, and 41.0 g/m². The highest dose tested was justified based on prior non-clinical GLP safety studies, and phase I clinical trials. In the event of dose-limiting toxicity in one patient at any dose level, a maximum of three additional patients would be treated. BNP7787 was given as a single agent by a 30-min intravenous (iv) infusion 1 week prior to chemotherapy so that pharmacokinetic data on BNP7787 alone could be collected and compared and again on day 1. On treatment day 1, paclitaxel dissolved in 500 mL saline was administered intravenously over 3 h at a dose of 175 mg/m². Immediately following paclitaxel administration, the calculated dosage of BNP7787 for the given dose level (in a minimum volume of 100 mL) was administered intravenously over 30–45 min, which was immediately followed by intravenous cisplatin 75 mg/m² in 500 mL of normal saline administered over 1 h. Study treatment was administered every 3 weeks (one cycle). Prophylactic medications to prevent hypersensitivity reactions to paclitaxel were administered, including iv dexamethasone 20 mg at 12 and 6 h prior to paclitaxel infusion, and diphenhydramine 50 mg po and ranitidine 50 mg iv 30 min before paclitaxel administration for paclitaxel hypersensitivity prophylaxis. Patients received 1.6 L of normal saline and/or electrolyte maintenance fluid prior to BNP7787 administration. Cisplatin was dissolved and infused along a program of forced diuresis that included at least 1.0 L of normal saline and/or electrolyte maintenance fluid, 300 mL of 20% mannitol and furosemide 20 mg iv. Prophylactic antiemetics during and after cisplatin administration consisted of granisetron and dexamethasone.

Patients with evidence of disease progression or those who experienced intolerable toxicity were removed from the study treatment.

The following pharmacokinetic parameters of BNP7787 and mesna were estimated using WinNonlin Version 2.1 (Pharsight Corporation, Mountain View, CA). C_max was the maximum drug concentration, and T_max was the elapsed time to reach C_max after the intravenous infusion. AUC_0-inf was area under the concentration–time curve calculated by global summation of the linear trapezoidal rule from time zero up to the last measurable data point at 24 h post administration with extrapolation to infinity. The elimination rate constant (λz) was determined by log-linear regression analysis of the terminal phase of the plasma concentration–time curves. The terminal half-life (T_1/2) was calculated by the equation: T_1/2 = ln2/λz. The mean residence time (MRT) was calculated by the equation of AUMC (area under the first moment − time curve)/AUC − [infusion time]/2. Clearance (CL) was calculated by dividing the actual dose received by the AUC. The volume of distribution at steady state (Vd_ss) was calculated by the equation of CL × MRT.

Tumors were staged based on a complete medical history and physical examination, routine chest radiography, bone scintiscanning, computed tomography (CT) of the chest and abdomen, whole-brain magnetic resonance imaging (MRI) or CT scan, and fiberoptic bronchoscopy. Staging was performed according to previous TNM criteria [¹⁴]. Prior to the first course of treatment, a complete blood count (including a differential white cell count and platelet count), biochemistry tests (including renal function, hepatic function, and electrolytes), electrocardiogram, and urinalysis were performed. Complete blood count and biochemistry tests were repeated at least twice a week after this initial evaluation, while the other investigations were repeated at least every 6 weeks to evaluate the target lesions. A complete blood count was repeated every day until recovery, when ANC <500/μl, leukocyte count <1,000/μl, or platelet count <10,000/μl was observed during the first cycle of treatment. Adverse events were recorded and graded using the National Cancer Institute Common Toxicity Criteria, Version 2.0. Tumor response was classified in accordance with WHO criteria [¹³].

Plasma samples for BNP7787 were obtained from 22 patients who received BNP7787 alone, and from 21 patients who received BNP7787 in combination with paclitaxel and cisplatin. The plasma concentration–time curves for the different doses of BNP7787 are shown in Fig. 1a, b, and the pharmacokinetic parameters derived from the plotted data are listed in Tables 5 and 6. The plasma concentrations of BNP7787 reached the C_max at the end of 30–45 min infusion, and it increased proportionally by the dose. Plasma elimination curves for BNP7787 alone and in combination with chemotherapy were fitted using a one-compartment model with terminal half-life of 0.62–2.33 h and 0.66–1.67 h, with or without chemotherapy, respectively. Formation of the metabolite mesna was rapidly observed following administration of the parent compound, and the mesna C_max was reached between 0.5 and 1 h after the end of the BNP7787 infusion. The plasma concentration of the mesna metabolite decreased more slowly than that of BNP7787 with the half-life of 0.88–6.45 h and 0.90–6.28 h, respectively (Fig. 2a, b). No significant differences were observed in pharmacokinetic parameters between BNP7787 alone and in combination with chemotherapy. The ratio of C_max of the metabolite mesna to that of BNP7787 ranged from 1.4 to 4.2%; this observation is consistent with prior observations and predicted by the free thiol hypothesis [⁹]. The mean AUC_0-inf of mesna was approximately 6.2% of the AUC_0-inf of BNP7787. The observed BNP7787 plasma half-life, Vd_ss, CL, and MRT also remained relatively constant over the dose ranges studied, regardless of chemotherapy administration. The mean Vd_ss for BNP7787 alone and in combination with chemotherapy were 8.45–17.78 and 9.06–21.72, respectively.

Following careful review of the phase I trial data from this and other studies, a BNP7787 dose of 18.4 g/m² was selected as the clinically recommended dose based on a balance of efficacy and safety observations. The recommended dose was not based on the maximum tolerated dose since no dose-limiting toxicities were observed with the administration of BNP7787 alone or in combination with chemotherapy at any of the dose levels tested.

The selection of 18.4 g/m² as the recommended phase II dose was based on the balance of toxicity, higher grades of neurotoxicity and nephrotoxicity observed with lower doses of BNP7787 (e.g., 4.1–12.3 g/m²), and the increased frequency of local IV site discomfort observed at higher doses of BNP7787 (above 27.6 g/m²).

We also note that the molar ratio of the doses of BNP7787 18.4 g/m² and paclitaxel 175 mg/m² and cisplatin 75 mg/m² are 275:1 and 226:1, respectively. These molar ratios of the drugs are consistent with dose molar ratios that have demonstrated neuroprotection and nephroprotection in preclinical studies.

Twenty-one patients were assessed for objective tumor response. Although there was no complete response, nine patients attained a partial response, with an overall response rate of 43%. Stable disease was observed in eight patients (38%). One patient was non-evaluable because of atelectasis of the lung surrounding and obscuring the primary marker lesion. The remaining three patients (14%) had progressive disease. In 10 patients with prior chemotherapy, four (40%) achieved a partial response. Among 11 patients with no prior chemotherapy, five (46%) had a partial response.

AUC_0-t area under the plasma concentration–time curve up to last measurable point at 24 h from infusion, AUC_0-inf. area under the plasma concentration–time curve extrapolated to infinity, C_max maximum concentration, λz elimination rate constant, T_1/2 half-life in terminal phase, MRT_0-inf. mean residence time extrapolated to infinity, Vd_ss volume of distribution at steady state, CL total body clearance, CT chemotherapy with cisplatin and paclitaxel

* Values are the no. of patients who received BNP7787 alone/no. of patients who had BNP preceding combination of paclitaxel and cisplatin

** Mean ± SD

AUC_0-t area under the plasma concentration–time curve up to last measurable point, AUC_0-inf. area under the plasma concentration–time curve extrapolated to infinity, C_max maximum concentration, T_max time to maximal concentration, T_1/2 half-life in terminal phase, λz elimination rate constant, MRT_0-inf. mean residence time extrapolated to infinity, CT chemotherapy with paclitaxel and cisplatin

* Values are the no. of patients who received BNP alone/no. of patients who had BNP preceding combination of paclitaxel and cisplatin

** Mean ± SD