DNA double strand breaks as predictor of efficacy of the alpha-particle emitter Ac-225 and the electron emitter Lu-177 for somatostatin receptor targeted radiotherapy

Key biologic effects of the alpha-particle emitter Actinium-225 in comparison to beta-particle emitter Lutetium-177 labeled somatostatin analogue DOTATOC in vitro and in vivo were studied to evaluate the significance of H2AX-foci formation and its downstream effects. To determine relative biological effectiveness (RBE) between the two isotopes somatostatin expressing AR42J cells were incubated with Ac-225-DOTATOC and Lu-177-DOTATOC up to 48 h and viability was analyzed using the MTT assay. DNA double strand breaks were quantified after immunofluorescence staining of H2AX. Cell cycle was analyzed by flow cytometry. In vivo, uptake of both radiolabeled somatostatin-analogues into subcutaneous AR42J tumors and number of cells displaying H2AX-foci were measured. Therapeutic efficacy was assayed by monitoring tumor growth after treatment with activities translated from in vitro cytotoxicity. Ac-225-DOTATOC was synthesized with specific activities between 0.2-0.4 MBq/µg and radiochemical purity of > 90%. ED50 values were 30 kBq/ml after 24 h and 14 kBq/ml after 48 h. Lu-177-DOTATOC displayed radiochemical purity of >95% and ED50 values of 10 MBq/ml after 48 h. Number of DNA double strand breaks increased with increasing concentration of Ac 225 DOTATOC and Lu-177-DOTATOC similarly, if a factor of approximately 700 of Lu-177 activities over Ac-225 activities was applied. Already 24 h after incubation with 2.5, 5, and 10 kBq/ml Ac 225 DOTATOC cell cycle studies showed an increment of the percentage of tumor cells in G2/M phase up to 60%. After 72 h an apoptotic subG1 peak was also detectable. Tumor uptake for both radio peptides at 48 h was identical with 7.5 %ID/g, though overall number of cells with H2AX-foci was higher for tumors treated with 48 kBq Actinium-225-DOTATOC than tumors treated with 30 MBq Lutetium-177-DOTATOC (35% vs. 21%). Tumors with a mean volume of 0.34 ml reached exponential tumor growth after 25 days (44 kBq Ac-225-DOTATOC), after 21 days (34 MBq Lu-177-DOTATOC) and after 5 days (control). Thus H2AX-foci displayed the key parameter after irradiation with similar downstream effects for beta and alpha irradiation.

GRAF Franziska;  FAHRER Jörg;  MAUS Stephan;  MORGENSTERN Alfred;  BRUCHERTSEIFER Frank;  VENKATACHALAM S.;  FOTTNER C.;  WEBER M.M.;  HUELSENBECK Johannes;  SCHRECKENBERGER M;  KAINA B;  MIEDERER Matthias; 

2014-03-10

PUBLIC LIBRARY SCIENCE

JRC83179

1932-6203,   

http://www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0088239,    https://publications.jrc.ec.europa.eu/repository/handle/JRC83179,   

10.1371/journal.pone.0088239,