[CANCER RESEARCH 61, 7868 –7874, November 1, 2001] Therapeutic Efficacy of Aortic Administration of N-Acetylcysteine as a Chemoprotectant against Bone Marrow Toxicity after Intracarotid Administration of Alkylators, with or without Glutathione Depletion in a Rat Model 1 Edward A. Neuwelt, 2 Michael A. Pagel, Brant P. Hasler, Thomas G. Deloughery, and Leslie L. Muldoon Department of Neurology [E. A. N., L. L. M.], Department of Neurosurgery [E. A. N.], Division of Hematology [T. G. D.], and Department of Cell and Developmental Biology [L. L. M.], Oregon Health Sciences University, Portland, Oregon 97201, and Veterans Administration Medical Center, Portland, Oregon 97201 [E. A. N., M. A. P., B. P. H.] ABSTRACT Modulation of thiol levels may alter both the efficacy and toxicity of chemotherapeutic agents. We investigated cytoenhancement, using L- buthionine-[S,R]-sulfoximine (BSO) to reduce cellular glutathione levels prior to intracarotid alkylator administration. We also evaluated chemo- protection against chemotherapy-induced systemic toxicity when the thiol agents N-acetylcysteine (NAC) and sodium thiosulfate were administered into the descending aorta to limit brain delivery. BSO treatment reduced rat brain and intracerebral tumor glutathione levels by 50 – 65%, equiv- alent to the reduction in liver and s.c. tumor. BSO treatment significantly enhanced the toxicity of chemotherapy with carboplatin, melphalan, and etoposide phosphate against granulocytes, total white cells, and platelets. Intracarotid administration of NAC resulted in high delivery to the brain, whereas infusion via the descending aorta minimized brain delivery. When NAC, with or without sodium thiosulfate, was administered via aortic infusion prior to chemotherapy, the magnitude of the bone marrow toxicity nadir was minimized, even with BSO-enhanced myelosuppression. Thus, BSO depleted brain and brain tumor glutathione but thereby increased chemotherapy-induced myelosuppression. Surprisingly, al- though NAC was found to readily cross the blood-brain barrier when given into the carotid artery, aortic infusion of NAC resulted in minimal exposure to the central nervous system (CNS) vasculature because of rapid clearance. As a result, aortic infusion of NAC to perfuse bone marrow and minimize myelosuppression and toxicity to visceral organs could be performed without interfering with the CNS cytotoxicity of intracarotid alkylators, even after BSO depletion of CNS glutathione. INTRODUCTION The effectiveness of chemotherapy against malignant tumors may be increased by dose intensification (1–3). As an alternative to in- creasing drug dosages, drug cytotoxicity can be enhanced by reducing the intracellular concentration of the endogenous detoxifying tripep- tide glutathione by use of BSO 3 to inhibit glutathione synthesis (4 – 6). BSO treatment increases the effectiveness of chemotherapeutics in vitro (6 –9), in animal models (10, 11), and in patients (12, 13). However, even without dose intensification, chemotherapy has toxic side effects, such as bone marrow toxicity, and this can be enhanced with BSO. It may be possible to reduce the bone marrow toxicity of chemo- therapeutic agents by using sulfur-containing chemoprotective agents (thio, thiol, and thioether compounds) to mimic one or many of the detoxifying activities of glutathione (14), including drug conjugation (15) and antioxidant and free radical scavenging activity (16). Clini- cally relevant compounds that may provide protection against at least some of the systemic toxicities caused by alkylating chemotherapeu- tics include ethyol (17), STS (2, 18, 19), and NAC (20 –22). Chemoprotectants have had relatively limited clinical use because of concerns of impaired chemotherapeutic efficacy. We hypothesize that reduction of tumoricidal effects my be avoided by separating chemoprotectant and chemotherapy treatments in time or space. For example, studies of STS chemoprotection have used two routes of administration (i.a. versus i.v. or i.p.) to minimize interactions of STS with cisplatin in tumor models (23) and patients (2, 24). Preclinical studies (25, 26) and clinical trials (18, 19) have shown that enhanced platinum chemotherapy delivery to the CNS followed by delayed STS chemoprotection, to provide both spatial and temporal separation, results in marked otoprotection without impairing cytotoxicity. The purpose of the present study was to test whether infusion of thiol chemoprotectants into the descending aorta in the rat could safely provide bone marrow protection while limiting NAC delivery to brain. Additionally, we determined the effects of BSO on glutathi- one concentrations in the brain and brain tumors and investigated whether glutathione depletion enhanced chemotherapy-induced bone marrow toxicity in the rat. The results demonstrate a potentially exciting new treatment option to maximize dose intensity in brain or head and neck tumors by use of i.a. (carotid or vertebral) chemother- apy given cephalad, while minimizing systemic toxicity with aortic NAC administration given caudally. MATERIALS AND METHODS Animal studies were performed in accordance with guidelines established by the Oregon Health Sciences University Committee on Animal Care and Use. Tumor Implantation. For assessment of the effects of BSO on glutathione levels, intracerebral and s.c. human small cell lung carcinoma cells were implanted in nude rats from our colony (weight, 220 g; n = 5) as reported previously (25, 27). Osmotic BBBD. BBBD was performed in adult female Long Evans rats (weight, 220 –240 g; n = 6) for assessment of delivery of NAC to the brain. BBBD was performed via the right external carotid artery catheter by infusion of 25% mannitol into the right internal carotid artery as reported previously (25, 27). In animals receiving NAC i.a. without BBBD, 0.9% NaCl was infused instead of mannitol. Aortic Infusion Technique. The left carotid artery (rather than the right carotid to avoid possible first-pass cranial perfusion) was exposed through a ventral neck incision, and a catheter (PE 50) filled with heparinized saline was tied into the left external carotid. To target the descending aorta, the left internal carotid was temporarily occluded with a Biemer vessel clip (9 mm), and the chemoprotectant was administered retrograde down the carotid to the descending aorta via the catheter. The Biemer clip and catheter were then removed, and the external carotid was ligated. The wound was closed with wound clips, and the animal was allowed to recover. Radiolabeled Tracer Studies. Animals for NAC biodistribution studies were evaluated at low (140 mg/kg) and high (1200 mg/kg) NAC doses with Received 2/9/01; accepted 9/4/01. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. 1 Financial support for this work was provided by a Veterans Administration merit review grant and by Grants CA31770 from the National Cancer Institute and NS33618 from the National Institute of Neurological Disorders and Stroke (to E. A. N.). 2 To whom requests for reprints should be addressed, at Oregon Health Sciences University, L603, 3181 S.W. Sam Jackson Park Road, Portland, OR 97201. Phone: (503) 494-5626; Fax: (503) 494-5627; E-mail: neuwelte@ohsu.edu. 3 The abbreviations used are: BSO, L-buthionine-[S,R]-sulfoximine; STS, sodium thiosulfate; NAC, N-acetylcysteine; i.a., intra-arterial; CNS, central nervous system; BBBD, blood-brain barrier disruption. 7868 Research. on December 18, 2015. © 2001 American Association for Cancer cancerres.aacrjournals.org Downloaded from