Absence of Preferential Uptake of [125I]Iododihydrorhodamine 123 by Four Human Tumor Xenografts1

The biodistribution of |l2SI]iododihydrorhodamine 123 has been studied over a 96-h period in four human tumor xenograft models: HT-29 colon adenocarcinoma, PC-3 prostate carcinoma, HT-1080 fibrosarcoma, and PaCa-2 pancreatic carcinoma. Elimination of radioactivity in the tumorbearing nude mice was rapid during the first 24 h and slow thereafter. The lack of uptake in the thyroid indicated there was little, if any, deiodination of the molecule. Activity was found mainly in the liver and spleen. Accumulation of radioactivity was low in all four tumors exam ined. At 4 h postinjection, as well as at 24 and 48 h, however, the total radioactive content in each of the four tumors was directly proportional to the weight of the tumor sample. This correlation was independent of tumor type, route of injection (i.v./i.p.) or dose (1.2-6 ¿tCi/mouse). This was not true for any of the normal tissues, suggesting that this accumu lation may be governed by certain intrinsic characteristics of the cancers tested. MATERIALS AND METHODS [125I]IDRwas prepared as previously described (6). The radiolabeled material was taken up in 0.01 M phosphate buffer containing 0.15 M NaCl, pH 7.2 (PBS). Four human tumor xenograft models were used: HT-29 colon ade nocarcinoma, PC-3 prostate adenocarcinoma, HT-1080 fibrosarcoma, and PaCa-2 pancreatic carcinoma. Nude mice (MCr/jVu/TVu/SED) were injected s.c. with 107-108 tumor cells, and when the tumors had grown to palpable size, the animals received a single injection of [I25I]1DR either i.v. through the tail vein or i.p. (see Table 1). At preselected times (1, 4, 8, 24, 48, 72, and 96 h), animals were sacrificed by ether asphyxiation, and various tissues (e.g., tumor, lung, liver, kidney, spleen, heart, skin, muscle, small intestine, large intestine, blood, thyroid, and bone) were dissected out, rinsed in PBS, blotted, and weighed. Urine samples were drawn directly from the bladder whenever possible. The tissues were counted in a Packard Autogamma counter. INTRODUCTION R1233 (Fig. 1) is a lipop hiIir. cationic mitochondria! vital stain that is taken up by living cells (1). Muscle and carcinoma cells have been reported to selectively accumulate and retain this compound while it is being cleared from normal epithelial cells (2). The toxicity of R123 in tumor cells has been demon strated both in vitro and in vivo (3-5). Since a radiolabeled derivative of R123 might, therefore, serve as a cancer imaging or therapeutic agent, we attempted the radioiodination of R123 using a variety of iodinating agents such as chloramine T, iodogen, and peracetic acid (6). These initial attempts were unsuccessful and yielded only the starting material as deter mined by nuclear magnetic resonance, even though other inves tigators have reported success with these reagents (7-9). Since electrophilic iodination could be greatly inhibited by the pres ence of the delocalized positive charge on the xanthene ring system, our results were not unexpected. Subsequently, we prepared an iodinated dihydrorhodamine 123 derivative (IDR) (Fig. 1) and demonstrated that it is taken up by cells, oxidized to a fluorescent form, and localized in the mitochondria. We have also shown that [I25I]IDR is selectively retained in vitro by PC-3 human prostate adenocarcinoma cells but not by Chinese hamster V79 lung fibroblasts (6). Accordingly, we have now studied the biodistribution of [I2SI]IDR in nude mice bearing the PC-3 prostate carcinoma as well as three other human tumor xenografts: HT-29 colon adenocarcinoma, HT-1080 fi brosarcoma, and PaCa-2 pancreatic carcinoma. Two of these cell lines (PC-3 and PaCa-2) were reported to retain R123 preferentially (2), and PaCa-2 was reported to be selectively killed/nv//rabyR123(3). Received 4/3/89; revised 8/1/89; accepted 8/7/89. 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. 1This work was supported by Grant CH-4 from Mallinckrodt. Incorporated. 2To whom requests for reprints should be addressed, at Harvard Medical School, Shields Warren Radiation Laboratory, SO Binney Street, Boston, MA 02115. 'The abbreviations used are: R123, rhodamine 123; IDR, iododihydrorhodamine; %ID/g, percentage of injected dose per gram of tissue; PBS, phosphate buffered saline. RESULTS All four animal groups showed similar biodistribution pat terns over the period examined (from 1 h up to 96 h postinjection). [125I]IDR was not taken up preferentially by any of the four tumors, and target to nontarget ratios were generally less than 1. These ratios improved slightly with time but most of them remained less than 1. As an example, the %ID/g plotted against time for HT-29 tumor-bearing mice is shown in Fig. 2. At early time points (1 h up to 8 h postinjection) most of the activity was found in the urine and the liver. For example, the cumulative dose in the urine was 91.83 ±31.80 %ID/g at 1 h postinjection in mice bearing PaCa-2 tumors and injected i.p.; 43.96 ±4.40 %ID/g at 8 h in mice bearing HT-1080 tumors and injected i.p.; and 96.03 %ID/g (n=l) at 8 h in mice bearing HT-1080 tumors and injected i.v. The liver uptake at 1 h postinjection varied from 9.31 ±2.72 %ID/g (PaCa-2, i.p. route) to 34.15 ±2.07 %ID/g (HT-1080, i.v. route) but de creased rapidly over time. The corresponding uptake in the small intestine was 5.93 ±1.26 %ID/g (PaCa-2, i.p. route) and 27.16 ±23.48 %ID/g (HT-1080, i.v. route). In addition to rapid urinary excretion, these observations suggest additional excretion via the hepatobiliary route. The 24-h biodistribution data in the four groups of tumor-bearing mice, expressed as %ID/g, are shown in Table 2. The highest activities were seen in the liver and the spleen at all times. In contrast, the %ID/g in all four tumors was low and remained so over time. No increase in thyroid uptake was observed over time suggesting that little, if any, dehalogenation of the labeled molecule oc curred. When two routes of injection (i.v. versus i.p.) were compared in nude mice bearing the HT-1080 fibrosarcoma tumor (Table 2), the %ID/g was higher in all tissues following the i.v. injec tion. However, the tumor-to-normal tissue ratio was about the same for the two routes of injection except for the liver and the spleen where it was more favorable for the i.p. injection. When the radioactive content of an individual tumor (nCi/ tumor) was plotted against tumor weight, a direct correlation was found for all time periods examined (Fig. 3, R2 > 0.9). At the 4-, 24-, and 48-h timepoints, the slope of each curve was 5986 on April 16, 2017. © 1989 American Association for Cancer Research. cancerres.aacrjournals.org Downloaded from RADIOIODINATED DIHYDRORHODAMINE