Comparative Evaluation of Anti-HER2 Affibody Molecules Labeled with Cu-64 Using NOTA and NODAGA

Imaging using affi body molecules enables discrimination between breast cancer metastases with high and low expression of HER2, making appropriate therapy selection possible. This study aimed to evaluate if the longer half-life of Cu-64 (T-1/2 = 12.7h) would make Cu-64 a superior nuclide compared to Ga-68 for PET imaging of HER2 expression using affibody molecules. The synthetic ZHER2: S1 affibody molecule was conjugated with the chelators NOTA or NODAGA and labeled with Cu-64. The tumor-targeting properties of Cu-64-NOTA-ZHER2: S1 and Cu-64-NODAGA-ZHER2: S1 were evaluated and compared with the targeting properties of Ga-68-NODAGA-ZHER2: S1 in mice. Both 64 Cu-NOTA-ZHER2: S1 and Cu-64-NODAGA-ZHER2: S1 demonstrated specific targeting of HER2-expressing xenografts. At 2 h after injection of Cu-64-NOTA-ZHER2: S1, Cu-64-NODAGA-ZHER2: S1, and Ga-68-NODAGAZHER2: S1, tumor uptakes did not differ significantly. Renal uptake of Cu-64-labeled conjugateswas dramatically reduced at 6 and 24 h after injection. Notably, radioactivity uptake concomitantly increased in blood, lung, liver, spleen, and intestines, which resulted in decreased tumor-to-organ ratios compared to 2 h postinjection. Organ uptake was lower for Cu-64-NODAGA-ZHER2: S1. The most probable explanation for this biodistribution pattern was the release and redistribution of renal radiometabolites. In conclusion, monoamide derivatives of NOTA and NODAGA may be suboptimal chelators for radiocopper labeling of anti-HER2 affibody molecules and, possibly, other scaffold proteins with high renal uptake

Molecular design of radiocopper-labelled Affibody molecules

The use of long-lived positron emitters Cu-64 or Cu-61 for labelling of Affibody molecules may improve breast cancer patients' stratification for HER-targeted therapy. Previous animal studies have shown that the use of triaza chelators for Cu-64 labelling of synthetic Affibody molecules is suboptimal. In this study, we tested a hypothesis that the use of cross-bridged chelator, CB-TE2A, in combination with Gly-Glu-Glu-Glu spacer for labelling of Affibody molecules with radiocopper would improve imaging contrast. CB-TE2A was coupled to the N-terminus of synthetic Affibody molecules extended either with a glycine (designation CB-TE2A-G-ZHER2:342) or Gly-Glu-Glu-Glu spacer (CB-TE2A-GEEE-ZHER2:342). Biodistribution and targeting properties of Cu-64-CB-TE2A-G-ZHER2:342 and Cu-64-CB-TE2A-GEEE-ZHER2:342 were compared in tumor-bearing mice with the properties of Cu-64-NODAGA-ZHER2:S1, which had the best targeting properties in the previous study. Cu-64-CB-TE2A-GEEE-ZHER2:342 provided appreciably lower uptake in normal tissues and higher tumor-to-organ ratios than Cu-64-CB-TE2A-GZHER2:342 and Cu-64-NODAGA-ZHER2:S1. The most pronounced was a several-fold difference in the hepatic uptake. At the optimal time point, 6 h after injection, the tumor uptake of Cu-64-CB-TE2A-GEEE-ZHER2: 342 was 16 +/- 6% ID/g and tumor-to-blood ratio was 181 +/- 52. In conclusion, a combination of the cross-bridged CB-TE2A chelator and Gly-Glu-Glu-Glu spacer is preferable for radiocopper labelling of Affibody molecules and, possibly, other scaffold proteins having high renal re-absorption

Site-selective protein-modification chemistry for basic biology and drug development.

Nature has produced intricate machinery to covalently diversify the structure of proteins after their synthesis in the ribosome. In an attempt to mimic nature, chemists have developed a large set of reactions that enable post-expression modification of proteins at pre-determined sites. These reactions are now used to selectively install particular modifications on proteins for many biological and therapeutic applications. For example, they provide an opportunity to install post-translational modifications on proteins to determine their exact biological roles. Labelling of proteins in live cells with fluorescent dyes allows protein uptake and intracellular trafficking to be tracked and also enables physiological parameters to be measured optically. Through the conjugation of potent cytotoxicants to antibodies, novel anti-cancer drugs with improved efficacy and reduced side effects may be obtained. In this Perspective, we highlight the most exciting current and future applications of chemical site-selective protein modification and consider which hurdles still need to be overcome for more widespread use.We thank FCT Portugal (FCT Investigator to G.J.L.B.), the EU (Marie-Curie CIG to G.J.L.B. and Marie-Curie IEF to O.B.) and the EPSRC for funding. G.J.L.B. is a Royal Society University Research Fellow.This is the author accepted manuscript. The final version is available from NPG via http://dx.doi.org/10.1038/nchem.239

On the relationship between supplier integration and time-to-market

Recent operations management and innovation management research emphasizes the importance of supplier integration. However, the empirical results as to the relationship between supplier integration and time-to-market are ambivalent. To understand this important relationship, we incorporate two major recent developments. First, the literature has started to redefine supplier integration into two dimensions, supplier product integration and supplier process integration. Second, recent research has begun to examine spillover effects that extend beyond the direct costs and benefits of the supplier contract. Using survey data of 116 firms in the industrials, health care, and information technology industries, the results confirm our hypotheses and show that supplier product integration decelerates time-to-market while supplier process integration accelerates time-to-market. The results also show a positive relationship between supplier integration and the adoption of external technologies, which either decelerates or accelerates time-to-market depending on the level of internal exploration activities. Our research, thus, helps to open the ‘black-box’ of the relationship between supplier integration and time-to-market, and provides a theoretically grounded explanation to the apparent contradictory results in prior research about the influence of supplier integration on time-to-market. In addition, we contribute to research on spillover effects by emphasizing that information technology adoption and assimilation is an important spillover effect of supplier integration

Comparative Evaluation of Anti-HER2 Affibody Molecules Labeled with Cu-64 Using NOTA and NODAGA

Imaging using affi body molecules enables discrimination between breast cancer metastases with high and low expression of HER2, making appropriate therapy selection possible. This study aimed to evaluate if the longer half-life of Cu-64 (T-1/2 = 12.7h) would make Cu-64 a superior nuclide compared to Ga-68 for PET imaging of HER2 expression using affibody molecules. The synthetic ZHER2: S1 affibody molecule was conjugated with the chelators NOTA or NODAGA and labeled with Cu-64. The tumor-targeting properties of Cu-64-NOTA-ZHER2: S1 and Cu-64-NODAGA-ZHER2: S1 were evaluated and compared with the targeting properties of Ga-68-NODAGA-ZHER2: S1 in mice. Both 64 Cu-NOTA-ZHER2: S1 and Cu-64-NODAGA-ZHER2: S1 demonstrated specific targeting of HER2-expressing xenografts. At 2 h after injection of Cu-64-NOTA-ZHER2: S1, Cu-64-NODAGA-ZHER2: S1, and Ga-68-NODAGAZHER2: S1, tumor uptakes did not differ significantly. Renal uptake of Cu-64-labeled conjugateswas dramatically reduced at 6 and 24 h after injection. Notably, radioactivity uptake concomitantly increased in blood, lung, liver, spleen, and intestines, which resulted in decreased tumor-to-organ ratios compared to 2 h postinjection. Organ uptake was lower for Cu-64-NODAGA-ZHER2: S1. The most probable explanation for this biodistribution pattern was the release and redistribution of renal radiometabolites. In conclusion, monoamide derivatives of NOTA and NODAGA may be suboptimal chelators for radiocopper labeling of anti-HER2 affibody molecules and, possibly, other scaffold proteins with high renal uptake