New Lyophilized Kit for Rapid Radiofluorination of Peptides

Radiolabeling compounds with positron-emitting radionuclides often involves a time-consuming, customized process. Herein, we report a simple lyophilized kit formulation for labeling peptides with ¹⁸F, based on the aluminum-fluoride procedure. The prototype kit contains IMP485, a NODA (1,4,7-triazacyclononane-1,4-diacetate)-MPAA (methyl phenylacetic acid)-di-HSG (histamine-succinyl-glycine) hapten-peptide, [NODA-MPAA-d-Lys­(HSG)-d-Tyr-d-Lys­(HSG)-NH₂], used for pretargeting, but we also examined a similar kit formulation for a somatostatin-binding peptide [IMP466, NOTA-d-Phe-<u>Cys-Phe-d-Trp-Lys-Thr-Cys</u>-Throl] bearing a NOTA ligand to determine if the benefits of using a kit can be extended to other AlF-binding peptides. The NODA-MPAA ligand forms a single stable complex with (AlF)²⁺ in high yields. In order to establish suitable conditions for a facile kit, the formulation was optimized for pH, peptide to Al³⁺ ratio, bulking agent, radioprotectant, and the buffer. For optimal labeling, the kit was reconstituted with an aqueous solution of ¹⁸F^– and ethanol (1:1), heated at 100–110 °C for 15 min, and then simply and rapidly purified using one of two equally effective solid-phase extraction (SPE) methods. Al¹⁸F-IMP485 was isolated as a single isomer complex, in high yield (45–97%) and high specific activity (up to 223 GBq/μmol), within 20 min. The labeled product was stable in human serum at 37 °C for 4 h and <i>in vivo</i>, urine samples showed the intact product was eliminated. Tumor targeting of the Al¹⁸F-IMP485 in nude mice bearing human colon cancer xenografts pretargeted with an anti-CEACAM5 bispecific antibody showed very low uptake (0.06% ± 0.02 ID/g) in bone, further illustrating its stability. At 1 h, pretargeted animals had high Al¹⁸F-IMP485 tumor uptake (28.1% ± 4.5 ID/g), with ratios of 9 ± 4, 123 ± 38, 110 ± 43, and 120 ± 108 for kidney, liver, blood and bone, respectively. Tumor uptake remained high at 3 h postinjection, with increased tumor/nontumor ratios. The NOTA-somatostatin-binding peptide also was fluorinated with good yield and high specific activity in the same kit formulation. However, yields were somewhat lower than those achieved with IMP485 containing the NODA-MPAA ligand, likely reflecting this ligand’s superior binding properties over the simple NOTA. These studies indicate that ¹⁸F-labeled peptides can be reproducibly prepared as stable Al–F complexes with good radiochemical yield and high specific activity using a simple, one-step, lyophilized kit followed by a rapid purification by SPE that provides the ¹⁸F-peptide ready for patient injection within 30 min

Data collection and its completeness.

<p>Data collection and its completeness.</p

Cases of disease recurrence with potential explanations.

<p>Cases of disease recurrence with potential explanations.</p

Disease risk factors and their relationship with mortality.

<p>Disease risk factors and their relationship with mortality.</p

The presence of risk factors for disease, stratified by ATSI status.

<p>The presence of risk factors for disease, stratified by ATSI status.</p

Disease severity, outcome and supportive care over the course of the study.

<p>Disease severity, outcome and supportive care over the course of the study.</p

Age at presentation and case-fatality rate.

<p>Age at presentation and case-fatality rate.</p

Improving the Therapeutic Index in Cancer Therapy by Using Antibody–Drug Conjugates Designed with a Moderately Cytotoxic Drug

The antibody–drug conjugate (ADC), IMMU-130, of the moderately cytotoxic topoisomerase I inhibitor, SN-38, and the CEACAM5-targeted humanized antibody (mAb), labetuzumab, was evaluated in model systems of human colon carcinoma and in phase I clinical trials of heavily pretreated patients with metastatic colorectal cancer. The conjugate, designed with a near-homogeneous drug substitution of 7–8 SN-38/mAb and with a linker that released 50% of the drug in ∼20 h, showed significant antitumor effects compared to a nontargeted ADC in human tumor xenografts, which could be augmented in combination with bevacizumab. The advantage of fractionated dosing was demonstrated, with potential implications for the clinical dosing schedule. Biodistribution comparing IMMU-130 with labetuzumab showed that the conjugate cleared somewhat faster from the blood, but this did not affect tumor uptake and retention. The use of an ultrastable linker in the conjugate design abrogated antitumor effects. A tolerability study in rabbits showed a high safety margin, with no-observed-adverse-effect level (NOAEL) corresponding to a cumulative human-equivalent protein dose of 40–60 mg/kg. The preclinical findings appear to be corroborated in two phase I clinical trials, with high tolerability and evidence of antitumor activity, including objective responses. The impact of the ADC design on the utility of IMMU-130, tailored to a poorly internalizing target, is discussed