Trophoblast Cell Surface Antigen 2 Expression in Thymic Carcinoma: Brief Report.

Introduction: Trophoblast cell surface antigen 2 (TROP2) is a transmembrane glycoprotein overexpressed in various cancer types. Although TROP2-targeting therapy is currently attracting attention, little is known about TROP2 expression in thymic carcinoma. Methods: TROP2 gene expression in thymic epithelial tumors was analyzed using RNA-sequencing (RNA-seq) data for 122 cases obtained from The Cancer Genome Atlas. Immunohistochemistry (IHC) staining with anti-TROP2 antibody (SP295) was performed in 26 cases of thymic carcinoma tissues surgically resected at Juntendo University. Results: RNA-seq data analysis from The Cancer Genome Atlas revealed that TACSTD2 (gene encoding TROP2) expression was significantly higher in thymic carcinoma than in thymoma (adjusted p = 6.64e-05). There was also a trend of increasing expression in the order of thymoma type B1, B2, B3, and thymic carcinoma. As for IHC in thymic carcinoma, TROP2 expression was localized to the membrane of cancer cells. Intensity 0, 1, and 2 was observed in six (23.1%), 11 (42.3%), and nine (34.6%) cases, respectively, leading to TROP2 positivity in 20 cases (76.9%). The median proportion of TROP2-positive tumor cells and the median H-score were 25.0% (range: 0%-100%) and 25.0 (range: 0-200), respectively. No relevant factors were identified in the analysis of TROP2 expression and patient background. Although not significant, high TROP2 expression (H-score ≥ 50) tended to be associated with shorter survival. Conclusions: TROP2 expression in thymic carcinoma was confirmed by both RNA-seq and IHC, with high expression observed in IHC for intensity (76.9%) and proportion. TROP2 could be a potential target in thymic carcinoma.

Effect of the Xanthine Oxidase Inhibitor, Febuxostat, on WBC Count in Asymptomatic Hyperuricemia: Subanalysis of the Randomized PRIZE Study.

AIMS: The anti-inflammatory effects of the xanthine oxidase inhibitor, febuxostat, a urate-lowering agent, have been reported in animal studies. However, the anti-inflammatory effects of urate-lowering therapy and its associated cardiovascular protective effects have not been fully determined in actual clinical practice. This study aimed to investigate the effect of febuxostat on white blood cell (WBC) count in patients with asymptomatic hyperuricemia and to assess for potential correlations between changes in WBC count and inflammatory biomarkers and atherosclerosis in this patient population. METHODS: This was a post hoc subanalysis of the PRIZE study, a multicenter, prospective, randomized, open-label clinical trial. In the PRIZE study, asymptomatic hyperuricemia patients were randomized to febuxostat group or control group with non-pharmacological therapy and evaluated the effect on vascular. The primary endpoints of this study were the assessment of the time course of WBC count over 24 months and its changes from baseline. Correlations of WBC count with high-sensitivity C-reactive protein (hs-CRP) and mean common carotid artery (CCA)-IMT were also exploratorily examined in the febuxostat group. RESULTS: A total of 444 patients (febuxostat group, n=223; control group, n=221) with WBC measurements available at baseline and at least one of the follow-up time points of 12 or 24 months, were enrolled. Febuxostat modestly, but significantly, reduced WBC counts at 12 and 24 months compared with the baseline levels (P=0.002 and P=0.026, respectively). Notably, the WBC count in the febuxostat group at 12 and 24 months was significantly lower than that in the control group (P=0.007 and P=0.023, respectively). The changes in WBC count were associated with those of hs-CRP (P=0.038), but not with CCA-IMT (P=0.727). CONCLUSIONS: Febuxostat therapy for 24 months modestly, but significantly, decreased WBC count in patients with asymptomatic hyperuricemia. This might potentially reflect a modest anti-inflammatory action of febuxostat in clinical settings.

Long-term effects of ipragliflozin on blood pressure in patients with type 2 diabetes: insights from the randomized PROTECT trial.

Although previous reports have shown that sodium-glucose cotransporter-2 (SGLT2) inhibitors have a blood pressure (BP) lowering effect, relevant long-term data is limited. This study aimed to evaluate the effect of the SGLT2 inhibitor ipragliflozin on BP, and associations between BP reduction and changes in cardiometabolic variables in diabetic patients. This was a sub-analysis of the PROTECT trial, a multicenter, randomized, open-label study to assess if ipragliflozin delays carotid atherosclerosis in patients with type 2 diabetes. Participants were randomized to ipragliflozin and control groups. The primary endpoint of the present sub-analysis was the trajectory of systolic BP over 24 months. Correlations between systolic BP changes and cardiometabolic variables were also evaluated. A total of 232 eligible participants with well-balanced baseline characteristics were included in each study group. Throughout the 24-month study period, mean systolic BP was lower in the ipragliflozin group. At 24 months, a between-group difference (ipragliflozin minus control) in mean systolic BP change from baseline was -3.6 mmHg (95% confidence interval, -6.2 to -1.0 mmHg), and the reduction in systolic BP in the ipragliflozin group was consistent across subgroups examined. Changes in systolic BP significantly correlated with those in body mass index in the ipragliflozin group, while no significant correlations with other cardiometabolic variables tested were observed. In conclusion, ipragliflozin treatment was associated with BP reduction throughout the 24-month follow-up period as compared to control treatment. BP reduction correlated with weight loss, which might be one of the mechanisms for the BP lowering effect of SGLT2 inhibitors.

Body fluid regulation via chronic inhibition of sodium-glucose cotransporter-2 in patients with heart failure: a post hoc analysis of the CANDLE trial.

BACKGROUND: In patients with chronic heart failure (CHF) and type 2 diabetes (T2D), sodium-glucose cotransporter-2 (SGLT2) inhibition improves cardiorenal outcomes, but details of the effects on distinct subsets of body fluid volume remain incomplete. METHODS: This was a post hoc analysis of patients with CHF and T2D in the CANDLE trial (UMIN000017669), an investigator-initiated, multi-center, randomized open-label trial that compared the effect of canagliflozin (100 mg, n = 113) with glimepiride (starting dose: 0.5 mg, n = 120) on changes in N-terminal pro-brain natriuretic peptide. The estimated plasma volume (ePV, calculated with the Straus formula) and estimated extracellular volume (eEV, determined by the body surface area) were compared between treatment groups at weeks 4, 12, and 24. RESULTS: Among 233 patients analyzed, 166 (71.2%) had an ejection fraction (EF) > 50%. Reductions in ePV and eEV were observed only in the canagliflozin group until week 12 (change from baseline at week 12, ePV; - 7.63%; 95% confidence interval [CI], - 10.71 to - 4.55%, p < 0.001, eEV; - 123.15 mL; 95% CI, - 190.38 to - 55.92 mL, p < 0.001). While ePV stopped falling after week 12, eEV continued to fall until week 24 ([change from baseline at week 24] - [change from baseline at week 12], ePV; 1.01%; 95%CI, - 2.30-4.32%, p = 0.549, eEV; - 125.15 mL; 95% CI, - 184.35 to - 65.95 mL, p < 0.001). CONCLUSIONS: Maintenance of a modest reduction in ePV and continuous removal of eEV via chronic SGLT2 inhibition suggests that favorable body fluid regulation contributes to the cardiorenal benefits of SGLT2 inhibitors in patients with CHF, irrespective of EF. TRIAL REGISTRATION: UMIN000017669.

Effect of canagliflozin on white blood cell counts in patients with type 2 diabetes and heart failure: A subanalysis of the randomized CANDLE trial.

AIMS/INTRODUCTION: Clinical evidence is lacking about the influence of sodium-glucose cotransporter 2 inhibitors on white blood cell (WBC) counts, a commonly used and widely available marker of inflammation. The aim of the present analysis was to assess the effect of canagliflozin relative to glimepiride on WBC counts. MATERIALS AND METHODS: This was a post-hoc subanalysis of the CANDLE trial (Effects of Canagliflozin in Patients with Type 2 Diabetes and Chronic Heart Failure: A Randomized Trial; UMIN000017669), an investigator-initiated, multicenter, open-label, randomized, controlled trial. A total of 233 patients with type 2 diabetes and concomitant heart failure were randomly assigned to either canagliflozin (n = 113) or glimepiride (n = 120) treatment for 24 weeks. Overall, patient baseline characteristics were as follows: mean ± standard deviation age, 68.6 ± 10.1 years; hemoglobin A1c, 7.0 ± 0.9%; left ventricular ejection fraction, 56.7 ± 14.4%; and median N-terminal pro-brain natriuretic peptide, 252 pg/mL (interquartile range 96-563 pg/mL). The mean baseline WBC counts were 6704 cells/µL (95% confidence interval 6,362-7,047) in the canagliflozin group and 6322 cells/µL (95% confidence interval 5,991-6,654) in the glimepiride group. There were no significant differences between treatment groups in terms of changes in WBC counts from baseline to weeks 4 and 12. In contrast, a group difference (canagliflozin minus glimepiride) from baseline to week 24 was significant (mean difference - 456 cells/µL [95% confidence interval -774 to -139, P = 0.005]). CONCLUSIONS: Our findings suggest that 24 weeks of treatment with canagliflozin, relative to glimepiride, reduced WBC counts in patients with type 2 diabetes and heart failure.

Comprehensive functional evaluation of variants of fibroblast growth factor receptor genes in cancer.

Various genetic alterations of the fibroblast growth factor receptor (FGFR) family have been detected across a wide range of cancers. However, inhibition of FGFR signaling by kinase inhibitors demonstrated limited clinical effectiveness. Herein, we evaluated the transforming activity and sensitivity of 160 nonsynonymous FGFR mutations and ten fusion genes to seven FGFR tyrosine kinase inhibitors (TKI) using the mixed-all-nominated-in-one (MANO) method, a high-throughput functional assay. The oncogenicity of 71 mutants was newly discovered in this study. The FGFR TKIs showed anti-proliferative activities against the wild-type FGFRs and their fusions, while several hotspot mutants were relatively resistant to those TKIs. The drug sensitivities assessed with the MANO method were well concordant with those evaluated using in vitro and in vivo assays. Comprehensive analysis of published FGFR structures revealed a possible mechanism through which oncogenic FGFR mutations reduce sensitivity to TKIs. It was further revealed that recurrent compound mutations within FGFRs affect the transforming potential and TKI-sensitivity of corresponding kinases. In conclusion, our study suggests the importance of selecting suitable inhibitors against individual FGFR variants. Moreover, it reveals the necessity to develop next-generation FGFR inhibitors, which are effective against all oncogenic FGFR variants.

Highly sensitive fusion detection using plasma cell-free RNA in non-small-cell lung cancers.

ALK, ROS1, and RET kinase fusions are important predictive biomarkers of tyrosine kinase inhibitors (TKIs) in non-small-cell lung cancer (NSCLC). Analysis of cell-free DNA (cfDNA) provides a noninvasive method to identify gene changes in tumor cells. The present study sought to use cfRNA and cfDNA for identifying fusion genes. A reliable protocol was established to detect fusion genes using cfRNA and assessed the analytical validity and clinical usefulness in 30 samples from 20 cases of fusion-positive NSCLC. The results of cfRNA-based assays were compared with tissue biopsy and cfDNA-based liquid biopsy (Guardant360 plasma next-generation sequencing [NGS] assay). The overall sensitivity of the cfRNA-based assay was 26.7% (8/30) and that of cfDNA-based assay was 16.7% (3/18). When analysis was limited to the samples collected at chemo-naïve or progressive disease status and available for both assays, the sensitivity of the cfRNA-based assay was 77.8% (7/9) and that of cfDNA-based assay was 33.3% (3/9). Fusion gene identification in cfRNA was correlated with treatment response. These results suggest that the proposed cfRNA assay is a useful diagnostic test for patients with insufficient tissues to facilitate effective administration of first-line treatment and is a useful tool to monitor the progression of NSCLC for consideration of second-line treatments.

Development of an optimal protocol for molecular profiling of tumor cells in pleural effusions at single-cell level.

Liquid biopsy analyzes the current status of primary tumors and their metastatic regions. We aimed to develop an optimized protocol for single-cell sequencing of floating tumor cells (FTCs) in pleural effusion as a laboratory test. FTCs were enriched using a negative selection of white blood cells by a magnetic-activated cell sorting system, and CD45-negative and cytokeratin-positive selection using a microfluidic cell separation system with a dielectrophoretic array. The enriched tumor cells were subjected to whole-genome amplification (WGA) followed by genome sequencing. The FTC analysis detected an EGFR exon 19 deletion in Case 1 (12/19 cells, 63.2%), and EML4-ALK fusion (17/20 cells, 85%) with an alectinib-resistant mutation of ALK (p.G1202R) in Case 2. To eliminate WGA-associated errors and increase the uniformity of the WGA product, the protocol was revised to sequence multiple single FTCs individually. An analytical pipeline, accurate single-cell mutation detector (ASMD), was developed to identify somatic mutations of FTCs. The large numbers of WGA-associated errors were cleaned up, and the somatic mutations detected in FTCs by ASMD were concordant with those found in tissue specimens. This protocol is applicable to circulating tumor cells analysis of peripheral blood and expands the possibility of utilizing molecular profiling of cancers.

Activation of insulin-like growth factor-1 receptor confers acquired resistance to osimertinib in non-small cell lung cancer with EGFR T790M mutation.

BACKGROUND: Osimertinib (AZD9291) is a third-generation EGFR-tyrosine kinase inhibitor (TKI) that selectively inhibits the activating EGFR mutation and T790M mutation, and is currently used globally to treat EGFR-mutant non-small cell lung cancer (NSCLC). However, acquired resistance to osimertinib is inevitable. METHODS: We established osimertinib-resistant cells (PC9/T790M/AZDR and H1975/AZDR) derived from EGFR-mutant NSCLC cells harboring T790M mutation, and investigated the mechanism of acquired resistance to osimertinib by whole-exome sequencing and multiple phospho-receptor tyrosine kinase (RTK) array. A tumor specimen from an EGFR-mutant NSCLC patient with acquired resistance to osimertinib was also subjected to immunohistochemical analysis. RESULTS: Whole-exome sequencing analysis demonstrated that genetic alterations, such as acquisition of EGFR C797S, loss of T790M mutation, MET amplification, or mutated KRAS, MEK, BRAF, PIK3CA, were not detected. Analysis of phospho-RTK array revealed that insulin-like growth factor-1 receptor (IGF1R) was activated in PC9/T790M/AZDR and H1975/AZDR cells. Knockdown of IGF1R by siRNA as well as inhibition of IGF1R activation by linstinib (IGF1R inhibitor) significantly restored the sensitivity to osimertinib. Immunohistochemical analysis revealed that the expression level of phosphorylated IGF1R was higher in the tumor specimen from the EGFR-mutant NSCLC patient with acquired resistance to osimertinib than in the specimen collected prior to the treatment. CONCLUSIONS: IGF1R activation could occur following treatment with osimertinib in EGFR-mutant NSCLC with T790M mutation, and might be one of the mechanisms underlying osimertinib resistance. Combined treatment of osimertinib and IGF1R inhibitor might be effective in overcoming the acquired resistance to osimertinib induced by IGF1R activation. KEY POINTS: Significant findings of the study: Using osimertinib-resistant cells, we found that IGF1R activation induced by osimertinib treatment in EGFR-mutant NSCLC with T790M mutation is involved in resistance. Increased phosphorylation of IGF1R was observed in the tumor specimen from an EGFR-mutant NSCLC patient with acquired osimertinib resistance. WHAT THIS STUDY ADDS: IGF1R activation might be one of the mechanisms of osimertinib resistance. A combination therapy with osimertinib and an IGF1R inhibitor might be an optimal approach for overcoming the acquired resistance to osimertinib induced by IGF1R activation.

Detection of circulating sarcoma tumor cells using a microfluidic chip-type cell sorter.

Analyses of circulating tumor cells have been shown to be effective for the detection of cancer relapse and prognosis prediction. However, research regarding its utility in sarcoma remains scarce. In this study, the microfluidic chip-type cell sorter On-chip Sort was used to construct a system for detecting circulating sarcoma cells (CSCs). A pilot study using normal fibroblast or sarcoma cell lines was designed to establish a reliable protocol to separate CSCs by On-chip Sort. A single CSC was separated and recovered from 10 ml of whole blood from a patient with locally advanced myxofibrosarcoma. The nonsynonymous mutation for KMT2B p.Ile2602Val identified in the formalin-fixed paraffin-embedded tumor sample was also confirmed in the CSC. Use of the developed protocol may allow CSCs to become an early predictor for metastasis and recurrence of sarcoma. Further, it may aid in optimizing post-operative therapies for patients without metastasis.

ASP2397 Is a Novel Natural Compound That Exhibits Rapid and Potent Fungicidal Activity against Aspergillus Species through a Specific Transporter.

Current therapies against invasive pulmonary aspergillosis (IPA) have a limited cure rate. Given that a delay in treatment initiation may be fatal, a new drug with rapid-onset and potent fungicidal activity is needed. The novel cyclic hexapeptide ASP2397 (currently known as VL-2397) exhibited antifungal activity against Aspergillus fumigatus (including azole-sensitive and azole-resistant isolates), A. terreus, and A. flavus at an MIC range of 1 to 4 µg/ml in human serum. Time-kill curve experiments showed that ASP2397 reduced germinated conidia of A. fumigatus by more than 1 log10 CFU within 6 h. In addition, ASP2397 inhibited hyphal elongation from germinated conidia of A. fumigatus, A. terreus, and A. flavus more rapidly than voriconazole. Under conditions of delayed treatment initiation in an IPA mouse model, ASP2397 had efficacy superior to that of posaconazole, with 100% survival and over 1 log10 CFU/g reduction in lung fungal burden. Histopathological investigation of lungs also showed that ASP2397 markedly suppressed disease progression. To clarify its mechanism of action, we generated a UV-induced mutant of A. fumigatus with low susceptibility to ASP2397. The mutant had a point mutation in the siderophore transporter gene sit1, which is absent in mammalian cells. These findings suggest that ASP2397 may improve clinical treatment options for IPA.

Silkworm fungal infection model for identification of virulence genes in pathogenic fungus and screening of novel antifungal drugs.

The silkworm infection model has the potential to replace conventional animal models for evaluation of the efficacy and toxicity of investigational antifungal agents. Silkworms are relatively inexpensive, can be simply grown in large numbers and can be easily infected with pathogenic fungi, including mutant strains. Antifungal agents can then be injected into the silkworm either via the hemolymph to mimic intravenous administration or directly into the gut for oral administration, and their antifungal effect can be evaluated. Common features regarding the mechanisms of pharmacokinetics between the silkworm and mammals result in consistent therapeutic effectiveness of antifungal agents. ASP2397, a promising new antifungal agent, was discovered using the silkworm model. The conclusion is that silkworms can be a more ethical and less expensive alternative to standard animal models, particularly for the identification and testing of new antifungal agents.

ASP2397: a novel antifungal agent produced by Acremonium persicinum MF-347833.

The novel antifungal agent ASP2397 (Vical's compound ID VL-2397) is produced by the fungal strain MF-347833 that was isolated from Malaysian leaf litter and is identified here as an Acremonium species based on its morphology, physiological properties and 28S ribosomal DNA sequence. Because of its potential importance for producing novel antifungal agents, we determined the taxonomic and biologic properties of MF-347833. We show here that ASP2397 is a cyclic hexapeptide that chelates aluminum ion and is therefore similar to ferrichrome, a hydroxamate siderophore. However, ASP2397 differs structurally from licensed antifungal agents such as amphotericin B, triazoles and echinocandins. To understand the relationship between chemical structure and biological function, we isolated certain ASP2397 derivatives from the culture broth, and we further chemically converted the metal-free form to other derivatives.

Discovery of a new antifungal agent ASP2397 using a silkworm model of Aspergillus fumigatus infection.

Natural products are the major source of currently available drugs. However, screening natural product presents several challenges, including the time-consuming and labor-intensive steps required for the isolation of a drug from crude extracts as well as the differences between the activities of compounds in vitro and in vivo. To address these challenges, we used silkworm larvae infected with Aspergillus fumigatus to screen a natural products library for potent drugs to treat invasive aspergillosis. A rationally designed library was constructed using numerous, geographically diverse fungal species and then screened to collect extracts of microorganisms that had detectable anti-Aspergillus activity. We evaluated this library using cultures of A. fumigatus and a silkworm model system of A. fumigatus infection. With this model, we identified the novel antifungal compound ASP2397 that not only cured infected silkworm larvae but also increased the rates of survival of mice infected with A. fumigatus. These findings strongly support the utility of the silkworm screening system for the simple and rapid isolation of antibiotics from natural products libraries.

Possible effects of glimepiride beyond glycemic control in patients with type 2 diabetes: a preliminary report.

BACKGROUND: The purpose of this study was to elucidate the effects of glimepiride on the levels of biomarkers related to cardiovascular regulation in patients with type 2 diabetes mellitus. METHODS AND RESULTS: Thirty-four patients with type 2 diabetes received glimepiride for 24 weeks. Significant decreases in the levels of glyceraldehyde-derived advanced glycation end products, (glycer-AGE: toxic AGE), eotaxin and fibroblast growth factor (FGF)-2 were recognized after the administration of glimepiride. Moreover, there were trends for there to be increases in the levels of granulocyte-colony stimulating factor (G-CSF) and granulocyte macrophage-colony stimulating factor (GM-CSF), and decreases in the levels of fractalkine, soluble CD40 ligand (sCD40L), macrophage inflammatory protein (MIP)-ß, vascular endothelial growth factor (VEGF) and soluble receptor for AGE (sRAGE). CONCLUSIONS: Glimepiride may have potent anti-oxidative, anti-inflammatory and angiogenic properties and it may potentially repair tissue damage by decreasing the levels of toxic AGE and increasing colony-stimulating factors.

Synergistic antifungal activity of KB425796-C in combination with micafungin against Aspergillus fumigatus and its efficacy in murine infection models.

KB425796-C is a novel antifungal metabolite produced by the newly isolated bacterial strain Paenibacillus sp. No. 530603. This compound is a 40-membered macrocyclic lipopeptidolactone consisting of 12 amino acids and a 3-hydroxy-15-methylpalmitoyl moiety. KB425796-C displayed antifungal activity against micafungin-resistant fungi and was fungicidal to Trichosporon asahii in vitro. In a murine systemic infection model of T. asahii, KB425796-C showed excellent efficacy upon i.p. administration at 32 mg kg(-1). In addition, KB425796-C induced morphological changes in the hyphae of Aspergillus fumigatus and had fungicidal effects in combination with micafungin. In a mouse model of septic A. fumigatus infection, although non-treated mice survived for a maximum of only 6 days, the survival rate of micafungin-treated mice (0.1 mg kg(-1)) increased to 20%, while the survival rate of mice treated with a combination of micafungin (0.1 mg kg(-1)) and KB425796-C (32 mg kg(-1)) increased to 100% during the 31-day post-infection period. Our findings suggest that KB425796-C is a good candidate for the treatment of aspergillosis in combination with micafungin.

Identification of ten KB425796-A congeners from Paenibacillus sp. 530603 using an antifungal assay against Aspergillus fumigatus in combination with micafungin.

The discovery and characterization of natural congeners is one approach for understanding the relationship between chemical structure and biological function. We recently isolated the novel antifungal metabolite KB425796-A produced by the recently isolated bacterium Paenibacillus sp. 530603. On the basis of morphological changes of Aspergillus fumigatus induced by KB425796-A in combination with micafungin, we developed a highly sensitive screening method for the specific detection of KB425796-A congeners. Using this method, we isolated ten congeners of KB425796-A, named KB425796-B, -C, -D, -E, -F, -G, -H, -I, -J and -K, which exhibited diverse antifungal potencies against A. fumigatus. One of the most potent congeners, KB425796-C, had antifungal activities against several micafungin-resistant infectious fungi. KB425796-C can be a potential drug candidate for treating micafungin-resistant fungal infections.

KB425796-A, a novel antifungal antibiotic produced by Paenibacillus sp. 530603.

The novel antifungal macrocyclic lipopeptidolactone, KB425796-A (1), was isolated from the fermentation broth of bacterial strain 530603, which was identified as a new Paenibacillus species based on morphological and physiological characteristics, and 16S rRNA sequences. KB425796-A (1) was isolated as white powder by solvent extraction, HP-20 and ODS-B column chromatography, and lyophilization, and was determined to have the molecular formula C79H115N19O18. KB425796-A (1) showed antifungal activities against Aspergillus fumigatus and the micafungin-resistant infectious fungi Trichosporon asahii, Rhizopus oryzae, Pseudallescheria boydii and Cryptococcus neoformans.

Detection of early stage atherosclerotic plaques using PET and CT fusion imaging targeting P-selectin in low density lipoprotein receptor-deficient mice.

BACKGROUND: Sensitive detection and qualitative analysis of atherosclerotic plaques are in high demand in cardiovascular clinical settings. The leukocyte-endothelial interaction mediated by an adhesion molecule P-selectin participates in arterial wall inflammation and atherosclerosis. METHODS AND RESULTS: A (64)Cu-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid conjugated anti-P-selectin monoclonal antibody ((64)Cu-DOTA-anti-P-selectin mAb) probe was prepared by conjugating an anti-P-selectin monoclonal antibody with DOTA followed by (64)Cu labeling. Thirty-six hours prior to PET and CT fusion imaging, 3MBq of (64)Cu-DOTA-anti-P-selectin mAb was intravenously injected into low density lipoprotein receptor-deficient Ldlr-/- mice. After a 180min PET scan, autoradiography and biodistribution of (64)Cu-DOTA-anti-P-selectin monoclonal antibody was examined using excised aortas. In Ldlr-/- mice fed with a high cholesterol diet for promotion of atherosclerotic plaque development, PET and CT fusion imaging revealed selective and prominent accumulation of the probe in the aortic root. Autoradiography of aortas that demonstrated probe uptake into atherosclerotic plaques was confirmed by Oil red O staining for lipid droplets. In Ldlr-/- mice fed with a chow diet to develop mild atherosclerotic plaques, probe accumulation was barely detectable in the aortic root on PET and CT fusion imaging. Probe biodistribution in aortas was 6.6-fold higher in Ldlr-/- mice fed with a high cholesterol diet than in those fed with a normal chow diet. (64)Cu-DOTA-anti-P-selectin mAb accumulated selectively in aortic atherosclerotic plaques and was detectable by PET and CT fusion imaging in Ldlr-/- mice. CONCLUSIONS: P-selectin is a candidate target molecule for early-phase detection by PET and CT fusion imaging of atherosclerotic plaques.