The radiolanthanides 149Pm, 166Ho, and 177Lu have decay characteristics suitable for radioimmunotherapy (RIT) of cancer. N-Hydroxysulfosuccinimidyl DOTA (DOTA-OSSu) and methoxy-DOTA (MeO-DOTA) were conjugated to the anti-TAG-72 monoclonal antibody CC49 for radiolabeling with 149Pm, 166Ho, and 177Lu. While both DOTA conjugates could be labeled to high specific activity with 177Lu, MeO-DOTA afforded superior conjugate stability, radiolabeling, and radiochemical purity. Pilot biodistributions in nude mice bearing LS174T human colon carcinoma xenografts demonstrated that MeO-DOTA afforded higher tumor uptake and lower kidney retention of 177Lu than DOTA-OSSu. The in vitro stability of 149Pm-, 166Ho-, and 177Lu-MeO-DOTA-CC49 was evaluated using serum and hydroxyapatite assays. Serum stability of radiolanthanide-labeled MeO-DOTA-CC49 followed a trend based on the coordination energies of the radiometals, with 177Lu showing the highest stability after 96 to 168 h at 37 C. In contrast, MeO-DOTA-CC49 labeled with all three radiolanthanides was >92% stable to hydroxyapatite challenge for 168 h at 37 C. Comprehensive biodistributions of 149Pm-, 166Ho-, and 177Lu-MeO-DOTA-CC49 were obtained in LS174T-bearing nude mice. Maximum tumor uptakes were 100.0% ID/g for 149Pm at 96 h, 69.5% ID/g for 166Ho at 96 h, and 132.4% ID/g for 177Lu at 168 h. Normal organ uptakes were generally low, except in the liver, spleen, and kidney at early time points. By 96 to 168 h postinjection, nontarget organ uptake decreased to approximately 7% ID/g (kidney), 12% ID/g (spleen), and 20% ID/g (liver) for each radiolanthanide. When labeled with 149Pm, 166Ho, and 177Lu, MeO-DOTA-CC49 has potential for RIT of colorectal cancer and other carcinomas.

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The therapeutic efficacies of radiolabeled biotin, pretargeted by monoclonal antibody (mAb)-streptavidin fusion protein CC49 scFvSA, were compared to those of radiolabeled mAb CC49, using the three radiolanthanides in an animal model of human colon cancer. The purpose of the present study was to compare antibody pretargeting to conventional radioimmunotherapy using (149)Pm, (166)Ho, or (177)Lu. Nude mice bearing LS174T colon tumors were injected sequentially with CC49 scFvSA, the blood clearing agent biotin-GalNAc(16), and (149)Pm-, (166)Ho-, or (177)Lu-DOTA-biotin. Tumor-bearing mice were alternatively administered (149)Pm-, (166)Ho-, or (177)Lu-MeO-DOTA-CC49. Therapy with pretargeted (149)Pm-,(166)Ho-, and (177)Lu-DOTA-biotin increased the median time of progression to a 1 g tumor to 50, 41, and 50 days post-treatment, respectively. Therapy with (149)Pm-,(166)Ho-, and (177)Lu-MeO-DOTA-CC49 increased the median time to progression to 53, 24, and 67 days post-treatment, respectively. In contrast, saline controls showed a median time to progression of 13 days postinjection. Treatment with pretargeted (149)Pm-, (166)Ho-, and (177)Lu-biotin or (149)Pm-, (166)Ho-, and (177)Lu-CC49 increased tumor doubling time to 18-36 days, compared to 3 days for saline controls. Among treated mice, 23% survived >84 days post-therapy, and 11% survived 6 months, but controls survived <29 days. Long-term survivors showed tumor growth inhibition or partial regression, extensive necrosis in residual masses, and no evidence of nontarget tissue toxicity at necropsy. Both pretargeted and conventional RIT demonstrated considerable efficacy in an extremely aggressive animal model of cancer. Our results identified (177)Lu as an optimal radiolanthanide for future evaluation of these agents in toxicity and multiple-dose therapy studies.

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Copper-64 emits beta(+) and beta(-) particles suitable for positron emission tomography and radioimmunotherapy (RIT) of cancer. Copper-64-labelled antibodies have caused complete responses in laboratory animal RIT studies at far lower radiation doses than traditionally prescribed. The intracellular localization of copper radioisotopes may lead to cytotoxic effects by mechanisms beyond ionizing radiation damage. The purpose of this research was to develop a model using both internalizing and non-internalizing antibodies for direct comparison in future RIT studies using the same animal model of cancer. The monoclonal antibodies, cBR96 and cT84.66, were conjugated with N-hydroxysulfosuccinimidyl DOTA. All conjugates retained high immunoreactivity and labelled efficiently with (64)Cu with high specific activity and radiochemical purity. Twenty-four hour biodistributions determined in LS174T tumour-bearing nude mice demonstrated low organ and high tumour uptakes for both monoclonal antibodies. This model constitutes a promising system for elucidating whether internalization of (64)Cu is responsible for an enhanced tumour cytotoxicity in vivo.

Objectives:  We previously presented preliminary data on pilot biodistributions of 64Cu‐labeled monoclonal antibodies (mAbs).1 The purpose of the present studies was to obtain comprehensive biodistributions using a two‐antibody model for direct comparison of an internalizing and a non‐internalizing mAb in the same animal model of cancer. These studies will allow evaluation of the efficacies of the two 64Cu‐labeled mAbs for radioimmunotherapy (RIT) and are part of a larger, ongoing project examining the unusual tumoricidal properties of internalized 64Cu. Methods:  DOTA (1,4,7,10‐tetraazacyclododecane‐1,4,7,10‐tetraacetic acid) was conjugated to an internalizing antibody, cBR96, and a non‐internalizing antibody, cT84.66. Biodistributions of the 64Cu‐labeled conjugates in nude mice bearing LS174T human colorectal carcinoma xenografts were obtained at time points from 15 min to 48 h. Mouse tumor dosimetry was calculated using a Monte Carlo N‐Particle Transport Code. Results:  The 64Cu‐DOTA‐cBR96 conjugate demonstrated rapid tumor uptake, reaching 20.2% injected dose per gram of tissue (% ID/g) at 3 h and peaking at 35.4% ID/g by 24 h. Tumor accumulation of 64Cu‐DOTA‐cT84.66 was more gradual, 8.19% ID/g at 3 h (p = 0.002), and reached 43.8% ID/g by 24 h, but maximum uptake was not statistically different from 64Cu‐DOTA‐cBR96 (p = 0.05). Liver, spleen, and kidney uptakes were statistically similar at 24 h for both conjugates (p = 0.05). Tumor absorbed radiation doses were estimated to be 1128 rad/mCi for 64Cu‐DOTA‐cBR96 and 1409 rad/mCi for 64Cu‐DOTA‐cT84.66. Conclusion:  The biodistributions and tumor dosimetry of these two radiolabeled antibodies were sufficiently similar for direct comparison of the therapeutic efficacies of internalizing versus non‐internalizing 64Cu RIT agents in the same animal model of cancer. RIT studies using this two‐antibody model are currently in progress to test the hypothesis that internalization of 64Cu is necessary for enhanced cytotoxic properties at low tumor absorbed doses.