Summary The Cancer Genome Atlas (TCGA) project has analyzed mRNA expression, miRNA expression, promoter methylation, and DNA copy number in 489 high-grade serous ovarian adenocarcinomas (HGS-OvCa) and the DNA sequences of exons from coding genes in 316 of these tumors. These results show that HGS-OvCa is characterized by TP53 mutations in almost all tumors (96%); low prevalence but statistically recurrent somatic mutations in 9 additional genes including NF1, BRCA1, BRCA2, RB1, and CDK12; 113 significant focal DNA copy number aberrations; and promoter methylation events involving 168 genes. Analyses delineated four ovarian cancer transcriptional subtypes, three miRNA subtypes, four promoter methylation subtypes, a transcriptional signature associated with survival duration and shed new light on the impact on survival of tumors with BRCA1/2 and CCNE1 aberrations. Pathway analyses suggested that homologous recombination is defective in about half of tumors, and that Notch and FOXM1 signaling are involved in serous ovarian cancer pathophysiology.

BackgroundOptical imaging (OI) techniques such as bioluminescence and fluorescence imaging have been widely used to track diseases in a non-invasive manner within living subjects. These techniques generally require bioluminescent and fluorescent probes. Here we demonstrate the feasibility of using radioactive probes for in vivo molecular OI.Methodology/Principal FindingsBy taking the advantages of low energy window of light (1.2–3.1 eV, 400–1000 nm) resulting from radiation, radionuclides that emit charged particles such as β+ and β− can be successfully imaged with an OI instrument. In vivo optical images can be obtained for several radioactive probes including 2-deoxy-2-[18F]fluoro-D-glucose ([18F]FDG), Na18F, Na131I, 90YCl3 and a 90Y labeled peptide that specifically target tumors.Conclusions/SignificanceThese studies demonstrate generalizability of radioactive OI technique. It provides a new molecular imaging strategy and will likely have significant impact on both small animal and clinical imaging.

Epidermal growth factor receptor (EGFR) has become an attractive target for cancer molecular imaging and therapy. Affibody proteins against EGFR have been reported, and thus, we were interested in evaluating their potential for positron emission tomography (PET) imaging of EGFR positive cancer. An Affibody analogue (Ac-Cys-Z(EGFR:1907)) binding to EGFR was made through conventional solid phase peptide synthesis. The purified protein was site-specifically coupled with the 1,4,7,10-tetraazacyclododecane-1,4,7-tris-aceticacid-10-maleimidethylacetamide (maleimido-mono-amide-DOTA) to produce the bioconjugate, DOTA-Z(EGFR:1907). (64)Cu labeled probe (64)Cu-DOTA-Z(EGFR:1907) displayed a moderate specific activity (5-8 MBq/nmol, 22-35 microCi/microg). Cell uptake assays by pre-incubating without or with 300 times excess unlabeled Ac-Cys-Z(EGFR:1907) showed high EGFR-specific uptake (20% applied activity at 0.5 h) in A431 epidermoid carcinoma cancer cells. The affinity (K(D)) of (64)Cu-DOTA-Z(EGFR:1907) as tested by cell saturation analysis was 20 nM. The serum stability test showed excellent stability of the probe with >95% intact after 4 h of incubation in mouse serum. In vivo small-animal PET imaging showed fast tumor targeting, high tumor accumulation (approximately 10% ID/g at 1 h p.i.), and good tumor-to-normal tissue contrast of (64)Cu-DOTA-Z(EGFR:1907) spiked with a wide dose range of Ac-Cys-Z(EGFR:1907). Bio-distribution studies further demonstrated that the probe had high tumor, blood, liver, and kidney uptakes, while blood radioactivity concentration dropped dramatically at increased spiking doses. Co-injection of the probe with 500 microg of Ac-Cys-Z(EGFR:1907) for blocking significantly reduced the tumor uptake. Thus, (64)Cu-DOTA-Z(EGFR:1907) showed potential as a high tumor contrast EGFR PET imaging reagent. The probe spiked with 50 microg of Ac-Cys-Z(EGFR:1907) improved tumor imaging contrast which may have important clinical applications.

This article reports the affibody-based nanoprobes specifically target and image human epidermal growth factor receptor type 2 (HER2)-expressing cells and tumors. The simple, robust, and precise structure of affibody molecules are a promising class of targeting ligands with high affinity. Using near-infrared (NIR) quantum dots (QDs) and iron oxide (IO) nanoparticles as two representative nanomaterials, we designed anti-HER2 affibody molecules with an N-terminus cysteine residue (Cysteine-Z HER2:342 ) and precisely conjugated with maleimide-functionalized nanoparticles to make nanoparticle-affibody conjugates. The in vitro and in vivo study showed the conjugates are highly specific to target and image HER2-expressing cells and tumors. This work indicated the nanoparticle-affibody conjugates may be excellent candidates as targeting probes for molecular imaging and diagnosis.

The present work demonstrates that Cy5.5 conjugated Fe3O4/SiO2 core/shell nanoparticles could allow us to control movement of human natural killer cells (NK-92MI) by an external magnetic field. Required concentration of the nanoparticles for the cell manipulation is as low as ~20 μg Fe/mL. However, the relative ratio of the nanoparticles loaded NK-92MI cells infiltrated into the target tumor site is enhanced by 17-fold by applying magnetic field and their killing activity is still maintained as same as the NK-92MI cells without the nanoparticles. This approach allows us to open alternative clinical treatment with reduced toxicity of the nanoparticles and enhanced infiltration of immunology to the target site.

Dialkylamino-alkyl-benzamide possess affinities for melanin, suggesting that such 18F-labeled benzamides could potentially be used as melanin targeted positron emission tomography (PET) probes to identify melanotic melanoma metastases in vivo with high sensitivity and specificity. In this report, we describe the synthesis, in vitro and in vivo evaluation of 18F-N-[2-(diethylamino)ethyl]-4-fluoro-Benzamide (18F-FBZA) in mouse tumor models. The sigma receptor binding affinity of 19F-FBZA was determined along with the in vitro cellular uptakes of the radiofluorinated 18F-FBZA in B16F10 cells. In vivo distribution and µPET imaging studies using B16F10 melanoma, A375M amelanotic melanoma and U87MG tumor bearing mice were conducted, as well as comparative studies with 2-deoxy-2-18F-fluoro-D-glucose (18F-FDG) PET imaging of the melanoma models is described. In vitro, the uptake of 18F-FBZA was significantly higher in B16F10 cells treated with L-tyrosine (P < 0.001). In vivo, 18F-FBZA displayed significant tumor uptake; 5.95 ± 1.82 %ID/g in B16F10 tumors and only 0.75 ± 0.06 %ID/g and 0.56 ± 0.13 %ID/g was observed at 2 h in the amelanotic A375M and U87MG tumors respectively. Murine-lungs bearing melanotic B16F10 pulmonary metastases displayed a significantly higher lung uptake compared to the normal lungs (P < 0.01). µPET imaging clearly identifies melanotic lesions in both primary and pulmonary metastases B16F10 tumor models. Co-registered µCT with µPET along with biopsies further confirmed the presence of tumor lesions in the mouse lungs. 18F-FBZA specifically targets primary and metastatic melanotic-melanoma lesions with high tumor uptake and may have translational potentials.

Melanocortin type 1 receptor (MC1R), also known as α-melanocyte–stimulating hormone (α-MSH) receptor, is an attractive molecular target for melanoma imaging and therapy. An 18F-labeled linear α-MSH peptide (18F-FB-Ac-Nle-Asp-His-D-Phe-Arg-Trp-Gly-Lys-NH2 [NAPamide]) shows promising melanoma imaging properties but with only moderate tumor uptake and retention. A transition metal rhenium-cyclized α-MSH peptide, ReO[Cys3,4,10,D-Phe7,Arg11] α-MSH3–13 (ReCCMSH(Arg11)), has shown high in vitro binding affinity to MC1R and excellent in vivo melanoma-targeting pro-files when labeled with radiometals. Therefore, we hypothesized that ReCCMSH(Arg11) could be a good platform for the further development of an 18F-labeled probe for PET of MC1R-positive malignant melanoma. Methods In this study, the metallopeptide Ac-D-Lys-ReCCMSH(Arg11) was synthesized using conventional solid-phase peptide synthesis chemistry and a rhenium cyclization reaction. The resulting peptides were then labeled with N-succinimidyl-4-18F-fluorobenzoate (18F-SFB). The 18F-labeled metallopeptides were further tested for their in vitro receptor binding affinities, in vivo biodistribution, and PET imaging properties. Results Both isomers of Ac-D-Lys-ReCCMSH(Arg11), named as RMSH-1 and RMSH-2, were purified and identified by high-performance liquid chromatography. The binding affinities of RMSH-1 and RMSH-2 and their respective 19F-SFB–conjugated peptides (19F-FB-RMSH-1 and 19F-FB-RMSH-2) were all determined to be within nanomolar range. Both 18F-labeled metallopeptides showed good tumor uptake in the B16F10 murine model, with high MC1R expression, but much lower uptake in the A375M human melanoma xenografted in mice, indicating low MC1R expression. 18F-FB-RMSH-1, when compared with 18F-FB-RMSH-2, displayed more favorable in vivo performance in terms of slightly higher tumor uptakes and much lower accumulations in the kidney and liver at 2 h after injection. Small-animal PET of 18F-FB-RMSH-1 and -2 in mice bearing B16F10 tumors at 1 and 2 h showed good tumor imaging quality. As expected, much lower tumor uptakes and poorer tumor–to–normal organ contrasts were observed for the A375M model than for the B16F10 model. 18F-FB-RMSH-1 and -2 showed higher tumor uptake and better tumor retention than did 18F-FB-NAPamide. Conclusion Specific in vivo targeting of 18F-FB-RMSH-1 to malignant melanoma was successfully achieved in preclinical models with high MC1R expression. Thus, the radiofluorinated metallopeptide 18F-FB-RMSH-1 is a promising molecular probe for PET of MC1R-positive tumors.

Peptoids are a rapidly developing class of biomimetic polymers based on oligo-N-substituted glycine backbones, designed to mimic peptides and proteins. Inspired by natural antimicrobial peptides, a group of cationic amphipathic peptoids has been successfully discovered with a potent and broad-spectrum activity against pathogenic bacteria; however, there are limited studies to address the in vivo pharmacokinetics of the peptoids. Herein, 64Cu labeled DOTA conjugates of three different peptoids and two control peptides were synthesized and assayed in vivo by both biodistribution studies and small animal positron emission tomography (PET). The study was designed in a way to assess how structural differences of the peptidomimetics affect in vivo pharmacokinetics. As amphipathic molecules, major uptake of the peptoids occurred at the liver. Increased kidney uptake was observed by deleting one hydrophobic residue in the peptoid, and 64Cu-3 achieved the highest kidney uptake of all the conjugates tested in this study. In comparison to peptides, our data indicated that peptoids had general in vivo properties of higher tissue accumulation, slower elimination, and higher in vivo stability. Different administration routes (intravenous, intraperitoneal, and oral) were investigated with peptoids. When administered orally, the peptoids showed poor bioavailability, reminiscent to that of peptide. But, remarkably longer passage through the gastrointestinal (GI) tract without rapid digestion was observed for peptoids. These unique in vivo properties of peptoids were rationalized by efficient cellular membrane permeability and protease resistance of peptoids. The results observed in the biodistribution studies could be confirmed by the PET imaging, which provides a reliable way to evaluate in vivo pharmacokinetic properties of peptoids noninvasively and in real time. The pharmacokinetic data presented here can provide an insight for further development of the antimicrobial peptoids as pharmaceuticals.