Aggressive administration of recombinant oncotoxin AR209 (anti-ErbB-2) in athymic nude mice implanted with orthotopic human non-small cell lung tumours

Abstract

Lung cancer remains a significant public health problem in the U.S.A. and will result in an estimated 160 400 deaths in 1997. This appalling number is due in large part to the lack of adequate treatment for tumours that are refractory to surgery with curable intent, or of an adequate salvage therapy for those patients who recur after surgical resection. Because non-small cell lung cancer is refractory to traditional chemotherapy, non-traditional therapies have been developed to treat patients with this disease. Recombinant oncotoxins have been designed to target cells that express certain proteins as part of their cellular membrane. One such oncotoxin, AR209 (formerly OLX-209 [e23(Fv)PE38KDEL]), has the specificity of an anti-ErbB-2 antibody contained within a single-chain antibody domain (e23v) coupled to a portion of the Pseudomonas exotoxin A (PE38KDEL). Previous studies demonstrate that this drug is capable of significantly reducing the size of orthotopic lung tumour xenografts. However, most of the treated mice developed tumours once therapy was removed. In this study, mice were treated aggressively using one of four drug treatment schedules. Mice were treated with either intravenous or subcutaneous injections of AR209 over a 2 week period. The data indicate that AR209 significantly reduced the size of tumours and upon microscopic analysis at necropsy, some mice were cured. However, despite the treatment schedule used, many mice contained residual tumour. Residual tumours expressed the ErbB-2 protein, indicating that more aggressive treatment with AR209 may have resulted in higher rates of cure.

Introduction

Despite the many advances made in the diagnosis and treatment of most human cancers, several types remain refractory to treatment. Cancers of the lung kill more Americans than any other tumour type, despite higher incidence rates for breast cancer and prostate cancer[1], reflecting the lack of an adequate therapy. Only 13% of people diagnosed with lung cancer will survive for 5 years1, 2. Non-small cell lung cancer (NSCLC, includes adenocarcinoma, squamous cell carcinoma, adenosquamous cell carcinoma, and large cell carcinoma) accounts for approximately 75% of all cancers of the lung[3]. Surgical resection of the tumour is the standard treatment for stage I and II disease, as most NSCLCs are refractory to radiotherapy and chemotherapy[4]. However, the majority of patients present at an advanced stage and are not candidates for a surgical procedure with curative intent[4]. For those patients with advanced stage disease or with recurrent metastatic lesions following surgical resection, palliative radiotherapy often becomes the only option.

Biological therapy has become an attractive alternative for the treatment of tumours that prove refractory to traditional therapies[5]. Immunotoxins and recombinant oncotoxins are an important component of biological therapy. These drugs rely on the specificity of an antibody to direct precisely a toxin that irreversibly inactivates protein synthesis in the tumour cell[6]. One such oncotoxin, AR209 (formerly OLX-209 [e23(Fv)PE38KDEL]), has the specificity of an anti-ErbB-2 antibody contained within a single-chain antibody domain (e23Fv) coupled to a portion of the Pseudomonas exotoxin A (PE38KDEL)[7]. We have previously demonstrated that the recombinant oncotoxin AR209 effectively targets human adenocarcinoma cells in vitro[8]and in vivo in a subcutaneous (s.c.) mouse model[8]and an orthotopic model[9]. The results of these studies indicate that three intravenous (i.v.) injections administered to mice over a 5 day period at 86 μg/kg or 43 μg/kg significantly reduced the size of s.c. tumours, but did not completely eliminate them[8]. Increasing the number of injections to six (three i.v. injections administered to mice over a 5 day period at 86 μg/kg followed by three injections separated by 10 days) resulted in smaller orthotopic tumours, but did not result in ‘cures’ in most mice[9]. Also, the tumours present in AR209-treated animals increased in size once therapy was removed. To evaluate the effect of a more aggressive use of the drug, mice implanted with intrathoracic orthotopic human tumours were treated with one of four drug administration schedules. The lungs of mice treated with AR209 were resected and stained for ErbB-2 oncoprotein using immunohistochemistry.