Magnetic resonance studies of laryngeal tumors implanted in nude mice: effect of treatment with bleomycin and electroporation Sukhendu B. Dev a,1 , John B. Caban b , Gurvinder S. Nanda a,2 , Susan D. Bleecher a , Dietmar P. Rabussay a , Timothy S. Moerland c , Stephen J. Gibbs d,e , Bruce R. Locke e, * a Genetronics, Inc., San Diego, CA, USA b Institute of Molecular Biophysics, Florida State University, Tallahassee, FL, USA c Department of Biological Sciences, Florida State University, Tallahassee, FL, USA d National High Magnetic Field Laboratory, Florida State University, Tallahassee, FL, USA e Department of Chemical Engineering, Florida State University, Tallahassee, FL, USA Received 22 March 2002; accepted 1 June 2002 Abstract Recently, a new type of cancer treatment has been introduced that combines pulsed electric fields (PEF) with anticancer drugs. The proposed mode of action is that PEF create transient pores in the membranes which allow entry of drugs into the cells. This method increases cytotoxicity of some anticancer drugs like bleomycin (BLM) by 2–3 orders of magnitude, which, in turn, reduces systemic drug dosage without decreasing efficacy. In the present study, magnetic resonance imaging (MRI) was used to determine changes in apparent water self-diffusion coefficients (ADC) and spin-lattice (T 1 ) and spin-spin (T 2 ) relaxation times that occur in an animal laryngeal tumor (HEp-2 cells) model with BLM delivered by PEF. A Bruker 14 Tesla (600 MHz) wide-bore spectrometer with micro-imaging capability was used to generate all the data. Mice carrying 8 mm tumors were treated with several combinations of drug and PEF. All measurements were made on tumor samples excised from mice 24 and 48 hours after treatment with (i) saline, intratumor injection (i.t.), (ii) BLM, i.t., (iii) saline with PEF, and (iv) BLM, i.t., followed by PEF. Although T 1 does not differ between the controls (i, ii, and iii) and full treatment (iv) 6.72  0.20 s vs. 6.31  1.7 s, T 2 for (iv) at 24 hours is significantly different from the controls 52.4  0.91 ms vs. 46.5  1.54 ms. T 2 differences between treatment and controls disappear at 48 hours. ADC increases significantly from 24 to 48 hours (7.31  0.16  10 -6 to 8.28  0.28  10 -6 cm 2 /sec, p = 0.05). Longer T 2 values may reflect early apoptosis and tumor death when the tumor is structurally less dense. Higher ADC’s, associated with the periphery of the tumors and the central region, may indicate loose structural organization and necrosis resulting from the combination treatment. © 2002 Elsevier Science Inc. All rights reserved. Keywords: Cancer treatment; Electroporation; Bleomycin; MRI; Water self-diffusion 1. Introduction A very promising recent development in cancer treat- ment involves the combination of standard anticancer drugs with pulsed electric fields. This technique is known specif- ically as electroporation therapy [EPT], and involves appli- cation of a wide range of therapeutic agents, including drugs, genes, antisense nucleic acids, and proteins. Electro- poration therapy is based on the well-established principle that cell membranes are destabilized under pulsed electric fields thereby causing transient pore formation in the mem- brane [1]. These pores, in turn, allow agents in contact with the membrane [drugs, genes, dyes, or other marker mole- cules] to enter and become trapped in the cell. A large number of studies have been conducted with different can- cer cell lines in vitro, with animal tumor models, and also clinical trials (reviewed by Hofmann et al [2]). In the case of the highly hydrophilic anticancer drug bleomycin, appli- cation of pulsed electric fields allows much lower concen- tration of the drug to be used for effective tumor treatment. For example, in vitro studies of the effectiveness of bleo-  Both the groups at Genetronics and Florida State University contrib- uted equally * Corresponding author. Tel.: +1-850-410-6165; fax: +1-850-410- 6150. E-mail address: locke@eng.fsu.edu (B. Locke). 1 Now at Gene Delivery & Expression Sciences, San Diego, CA 92122 2 Now at Biogenex, Inc., 4600 Norris Canyon Road, San Ramon, CA 94583 Magnetic Resonance Imaging 20 (2002) 389 –394 0730-725X/02/$ – see front matter © 2002 Elsevier Science Inc. All rights reserved. PII: S0730-725X(02)00517-9