[(64) Cu]-labelled trastuzumab: optimisation of labelling by DOTA and NODAGA conjugation and initial evaluation in mice.

The human epidermal growth factor receptor-2 (HER2) is overexpressed in 20-30% of all breast cancer cases, leading to increased cell proliferation, growth and migration. The monoclonal antibody, trastuzumab, binds to HER2 and is used for treatment of HER2-positive breast cancer. Trastuzumab has previously been labelled with copper-64 by conjugation of a 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) chelator. The aim of this study was to optimise the (64) Cu-labelling of DOTA-trastuzumab and as the first to produce and compare with its 1,4,7-triazacyclononane, 1-glutaric acid-5,7 acetic acid (NODAGA) analogue in a preliminary HER2 tumour mouse model. The chelators were conjugated to trastuzumab using the activated esters DOTA mono-N-hydroxysuccinimide (NHS) and NODAGA-NHS. (64) Cu-labelling of DOTA-trastuzumab was studied by varying the amount of DOTA-trastuzumab used, reaction temperature and time. Full (64) Cu incorporation could be achieved using a minimum of 10-µg DOTA-trastuzumab, but the fastest labelling was obtained after 15 min at room temperature using 25 µg of DOTA-trastuzumab. In comparison, 80% incorporation was achieved for (64) Cu-labelling of NODAGA-trastuzumab. Both [(64) Cu]DOTA-trastuzumab and [(64) Cu]NODAGA-trastuzumab were produced after purification with radiochemical purities of >97%. The tracers were injected into mice with HER2 expressing tumours. The mice were imaged by positron emission tomography and showed high tumour uptake of 3-9% ID/g for both tracers.

Efficient regioselective ring opening of activated aziridine-2-carboxylates with [(18)f]fluoride.

Aziridines can undergo a range of ring-opening reactions with nucleophiles. The regio- and stereochemistry of the products depend on the substituents on the aziridine. Aziridine ring-opening reactions have rarely been used in radiosynthesis. Herein we report the ring opening of activated aziridine-2-carboxylates with [(18)F]fluoride. The aziridine was activated for nucleophilic attack by substitution of various groups on the aziridine nitrogen atom. Fluorine-18 radiolabelling was followed by ester hydrolysis and removal of the activation group. Totally regioselective ring opening and subsequent deprotection was achieved with tert-butyloxycarbonyl- and carboxybenzyl-activated aziridines to give α-[(18)F]fluoro-ß-alanine in good radiochemical yield.

Comparison of two new angiogenesis PET tracers 68Ga-NODAGA-E[c(RGDyK)]2 and (64)Cu-NODAGA-E[c(RGDyK)]2; in vivo imaging studies in human xenograft tumors.

INTRODUCTION: The aim of this study was to synthesize and perform a side-by-side comparison of two new tumor-angiogenesis PET tracers (68)Ga-NODAGA-E[c(RGDyK)](2) and (64)Cu-NODAGA-E[c(RGDyK)](2) in vivo using human xenograft tumors in mice. Human radiation burden was estimated to evaluate potential for future use as clinical PET tracers for imaging of neo-angiogenesis. METHODS: A (68)Ge/(68)Ga generator was used for the synthesis of (68)Ga-NODAGA-E[c(RGDyK)](2). (68)Ga and (64)Cu labeled NODAGA-E[c(RGDyK)](2) tracers were administrated in nude mice bearing either human glioblastoma (U87MG) or human neuroendocrine (H727) xenograft tumors. PET/CT scans at 3 time points were used for calculating the tracer uptake in tumors (%ID/g), integrin αVß3 target specificity was shown by blocking with cold NODAGA-E[c(RGDyK)](2), and biodistribution in normal organs were also examined. From biodistribution data in mice human radiation-absorbed doses were estimated using OLINDA/EXM software. RESULTS: (68)Ga-NODAGA-E[c(RGDyK)](2) was synthesized with a radiochemical purity of 89%-99% and a specific activity (SA) of 16-153 MBq/nmol. (64)Cu-NODAGA-E[c(RGDyK)](2) had a purity of 92%-99% and an SA of 64-78 MBq/nmol. Both tracers showed similar uptake in xenograft tumors 1h after injection (U87MG: 2.23 vs. 2.31%ID/g; H727: 1.53 vs. 1.48%ID/g). Both RGD dimers showed similar tracer uptake in non-tumoral tissues and a human radiation burden of less than 10 mSv with an administered dose of 200 MBq was estimated. CONCLUSION: (68)Ga-NODAGA-E[c(RGDyK)](2) and (64)Cu-NODAGA-E[c(RGDyK)](2) can be easily synthesized and are both promising candidates for PET imaging of integrin αVß3 positive tumor cells. (68)Ga-NODAGA-E[c(RGDyK)](2) showed slightly more stable tumor retention. With the advantage of in-house commercially (68)Ge/(68)Ga generators, (68)Ga-NODAGA-E[c(RGDyK)](2) may be the best choice for future clinical PET imaging in humans.

(64)Cu-NODAGA-c(RGDyK) Is a Promising New Angiogenesis PET Tracer: Correlation between Tumor Uptake and Integrin α(V)ß(3) Expression in Human Neuroendocrine Tumor Xenografts.

Purpose. The purpose of this paper is to evaluate a new PET tracer (64)Cu-NODAGA-c(RGDyK) for imaging of tumor angiogenesis using gene expression of angiogenesis markers as reference and to estimate radiation dosimetry for humans. Procedures. Nude mice with human neuroendocrine tumor xenografts (H727) were administered (64)Cu-NODAGA-c(RGDyK) i.v. for study of biodistribution as well as for dynamic PET. Gene expression of angiogenesis markers integrin α(V), integrin ß(3), and VEGF-A were analyzed using QPCR and correlated to the tracer uptake in the tumors (%ID/g). From biodistribution data human radiation-absorbed doses were estimated using OLINDA/EXM. Results. Tumor uptake was 1.2%ID/g with strong correlations between gene expression and tracer uptake, for integrin α(V) R = 0.76, integrin ß(3) R = 0.75 and VEGF-A R = 0.81 (all P < 0.05). The whole body effective dose for humans was estimated to be 0.038 and 0.029 mSv/MBq for females and males, respectively, with highest absorbed dose in bladder wall. Conclusion. (64)Cu-NODAGA-c(RGDyK) is a promising new angiogenesis PET tracer with potential for human use.