Clinical efficacy of the antiplatelet drug clopidogrel is hampered by its variable biotransformation into the active metabolite. The variability in the clinical response to clopidogrel treatment has been attributed to genetic factors, but the specific genes and mechanisms underlying clopidogrel bioactivation remain unclear. Using in vitro metabolomic profiling techniques, we identified paraoxonase-1 (PON1) as the crucial enzyme for clopidogrel bioactivation, with its common Q192R polymorphism determining the rate of active metabolite formation. We tested the clinical relevance of the PON1 Q192R genotype in a population of individuals with coronary artery disease who underwent stent implantation and received clopidogrel therapy. PON1 QQ192 homozygous individuals showed a considerably higher risk than RR192 homozygous individuals of stent thrombosis, lower PON1 plasma activity, lower plasma concentrations of active metabolite and lower platelet inhibition. Thus, we identified PON1 as a key factor for the bioactivation and clinical activity of clopidogrel. These findings have therapeutic implications and may be exploited to prospectively assess the clinical efficacy of clopidogrel.

A.J. performed transthoracic injections. G.A. performed a portion of the CT scans. M.M. and S.T.B. performed IHC staining and analyzed data. M.M. J.T.L. and A.J. performed in vitro 18 FBnTP uptake assays. M.M. did TMRE staining and western blots. M.C.F is a board-certified anatomic pathologist who performed the pathological analysis. L.S. and O.S. performed and guided respirometry experiments. S.S., C.M.W., A.G. and T.H. performed radio-tracer synthesis. D.D. and C.K. performed biochemical analysis of mitochondria. E.S. and H.C. performed metabolic analysis of lung tumors. S.M.D. contributed resources and critical feedback on the project. Data availability Source data for Western blots are provided with the paper as Supplementary Figure 1. Source data for Figures 1b-d; Figures 2d-g; Figures 3b, 3d, 3e; ED Figures 3c-e; ED Figures 4c, 4d; ED Figure 9a are provided with the paper. The data that support the findings of this study are available from the corresponding author upon reasonable request.

New radiolabeled probes for positron-emission tomography (PET) are providing an ever-increasing ability to answer diverse research and clinical questions and to facilitate the discovery, development, and clinical use of drugs in patient care. Despite the high equipment and facility costs to produce PET probes, many radiopharmacies and radiochemistry laboratories use a dedicated radiosynthesizer to produce each probe, even if the equipment is idle much of the time, to avoid the challenges of reconfiguring the system fluidics to switch from one probe to another. To meet growing demand, more cost-efficient approaches are being developed, such as radiosynthesizers based on disposable "cassettes," that do not require reconfiguration to switch among probes. However, most cassette-based systems make sacrifices in synthesis complexity or tolerated reaction conditions, and some do not support custom programming, thereby limiting their generality. In contrast, the design of the ELIXYS FLEX/CHEM cassette-based synthesizer supports higher temperatures and pressures than other systems while also facilitating flexible synthesis development. In this paper, the syntheses of 24 known PET probes are adapted to this system to explore the possibility of using a single radiosynthesizer and hot cell for production of a diverse array of compounds with wide-ranging synthesis requirements, alongside synthesis development efforts. Most probes were produced with yields and synthesis times comparable to literature reports, and because hardware modification was unnecessary, it was convenient to frequently switch among probes based on demand. Although our facility supplies probes for preclinical imaging, the same workflow would be applicable in a clinical setting.

Antibody-based dual-modality (PET/fluorescence) imaging enables both presurgery antigen-specific immuno-PET for noninvasive whole-body evaluation and intraoperative fluorescence for visualization of superficial tissue layers for image-guided surgery. Methods: We developed a universal dual-modality linker (DML) that facilitates site-specific conjugation to a cysteine residuebearing antibody fragment, introduction of a commercially available fluorescent dye (via an amine-reactive prosthetic group), and rapid and efficient radiolabeling via click chemistry with 18 F-labeled trans-cyclooctene (18 F-TCO). To generate a dual-modality antibody fragment-based imaging agent, the DML was labeled with the farred dye sulfonate cyanine 5 (sCy5), site-specifically conjugated to the C-terminal cysteine of the anti-prostate stem cell antigen (PSCA) cys-diabody A2, and subsequently radiolabeled by click chemistry with 18 F-TCO. The new imaging probe was evaluated in a human PSCA-positive prostate cancer xenograft model by sequential immuno-PET and optical imaging. Uptake in target tissues was confirmed by ex vivo biodistribution. Results: We successfully synthesized a DML for conjugation of a fluorescent dye and 18 F. The anti-PSCA cys-diabody A2 was site-specifically conjugated with either DML or sCy5 and radiolabeled via click chemistry with 18 F-TCO. Immuno-PET imaging confirmed in vivo antigen-specific targeting of prostate cancer xenografts as early as 1 h after injection. Rapid renal clearance of the 50-kDa antibody fragment enables same-day imaging. Optical imaging showed antigen-specific fluorescent signal in PSCApositive xenografts and high contrast to surrounding tissue and PSCA-negative xenografts. Conclusion: The DML enables site-specific conjugation away from the antigen-binding site of antibody fragments, with a controlled linker-to-protein ratio, and combines signaling moieties for 2 imaging systems into 1 molecule. Dual-modality imaging could provide both noninvasive whole-body imaging with organ-level biodistribution and fluorescence image-guided identification of tumor margins during surgery.

The trend to inform personalized molecular radiotherapy with molecular imaging diagnostics, a concept referred to as theranostics, has transformed the field of nuclear medicine in recent years. The development of theranostic pairs comprising somatostatin receptor (SSTR)-targeting nuclear imaging probes and therapeutic agents for the treatment of patients with neuroendocrine tumors (NETs) has been a driving force behind this development. With the Neuroendocrine Tumor Therapy (NETTER-1) phase 3 trial reporting encouraging results in the treatment of well-differentiated, metastatic midgut NETs, peptide radioligand therapy (RLT) with the 177 Lu-labeled somatostatin analog (SSA) [ 177 Lu]Lu-DOTA-TATE is now anticipated to become the standard of care. On the diagnostics side, the field is currently dominated by 68 Ga-labeled SSAs for the molecular imaging of NETs with positron emission tomography-computed tomography (PET/CT). PET/CT imaging with SSAs such as [ 68 Ga]Ga-DOTA-TATE, [ 68 Ga]Ga-DOTA-TOC, and [ 68 Ga]Ga-DOTA-NOC allows for NET staging with high accuracy and is used to qualify patients for RLT. Driven by the demand for PET/CT imaging of NETs, a commercial kit for the production of [ 68 Ga]Ga-DOTA-TATE (NETSPOT) was approved by the U.S. Food and Drug Administration (FDA). The synthesis of 68 Ga-labeled SSAs from a 68 Ge/ 68 Ga-generator is straightforward and allows for a decentralized production, but there are economic and logistic difficulties associated with these approaches that warrant the search for a viable, generator-independent alternative. The clinical introduction of an 18 F-labeled SSTR-imaging probe can help mitigate the shortcomings of the generator-based synthesis approach, but despite extensive research efforts, none of the proposed 18 F-labeled SSAs has been translated past prospective first-in-humans studies so far. Here, we review the current state of probe-development from a translational viewpoint and make a case for a clinically viable, 18 F-labeled alternative to the current standard [ 68 Ga]Ga-DOTA-TATE.