Cellular and molecular properties of (90)Y-labeled cetuximab in combination with radiotherapy on human tumor cells in vitro.

PURPOSE: Anti-EGFR antibody cetuximab (C225) is used in combination with radiotherapy of head and neck squamous cell carcinoma (HNSCC) patients. We investigated whether conjugation of cetuximab with trans-cyclohexyl-diethylene-triamine-pentaacetic acid (CHX-A″-DTPA) and radiolabeling with (90)Yttrium affect the molecular and cellular function of cetuximab and improve its combined effect with external-beam irradiation (EBI). METHODS: The following cell lines were used: HNSCC UT5, SAS, FaDu, as well as A43, Chinese hamster ovary cells (CHO), and human skin fibroblast HSF7. Binding affinity and kinetics, specificity, retention, and the combination of (90)Y-cetuximab with EBI were evaluated. RESULTS: Control cetuximab and CHX-A″-DTPA-cetuximab blocked the proliferation activity of UT5 cells. In combination with EBI, CHX-A″-DTPA-cetuximab increased the radiosensitivity of UT5 to a similar degree as control cetuximab did. In contrast, in SAS and HSF7 cells neither proliferation nor radiosensitivity was affected by either of the antibodies. Binding [(90)Y]Y-CHX-A″-DTPA-cetuximab ((90)Y-cetuximab) to EGFR in HNSCC cells occurred time dependently with a maximum binding at 24 h. Retention of (90)Y-cetuximab was similar in both HNSCC cell lines; 24 h after treatment, approximately 90% of bound activity remained in the cell layer. Competition assays, using cell membranes in the absence of an internalized fraction of cetuximab, showed that the cetuximab affinity is not lost as a result of conjugation with CHX-A″-DTPA. Cetuximab and CHX-A″-DTPA-cetuximab blocked EGF-induced Y1068 phosphorylation of EGFR. The lack of an effect of cetuximab on EGF-induced Akt and ERK1/2 phosphorylation and the inhibition of irradiation (IR)-induced Akt and ERK1/2 phosphorylation by cetuximab were not affected by DTPA conjugation. (90)Y-cetuximab in combination with EBI resulted in a pronounced inhibition of colony formation of HNSCC cells. CONCLUSIONS: Conjugation of CHX-A″-DTPA to cetuximab does not alter the cellular and biological function of cetuximab. (90)Y-labeling of cetuximab in combination with EBI may improve radiotherapy outcome.

An RAPD (random amplified polymorphic DNA) analysis of genetic population structure of Balea biplicata (Gastropoda: Clausiliidae) in fragmented floodplain forests of the Elster/Saale riparian system.

Eight German populations of the land snail Balea biplicata (Mollusca: Clausiliidae) were studied using the randomly amplified polymorphic DNA-polymerase chain reaction and morphometrics (principal component and discriminant analysis) to examine population structure and gene flow patterns in a fragmented landscape mosaic along the Elster/Saale riparian system, Germany. A variety of population genetic analyses targeting either more on the geographic scale of gene flow (genetic distances, F statistics, Mantel test) or on local genotypic structure (heterozygosity, linkage disequilibrium, bottleneck probability) showed that (1) the population system in total is governed by high gene flow independent of geographic distance, (2) genetic structure on the narrower sampling scale is mainly determined by stochastic processes due to genetic drift in small isolated and frequently recolonized populations, and (3) the morphometrical variation of the populations was related neither to habitat nor to genetic heterogeneity. The potentials for active and passive dispersal capacity of the snails and possible environmental impacts on their population structure are discussed.