The mGluR5 antagonist, MPEP, potentiated the disruptions in learning and memory induced by PCP. These behavioral data extend previous behavioral findings and further suggest that mGluR5 can modulate NMDA receptor function in vivo.

Fluorocarbons are lipophobic and non-polar molecules that exhibit remarkable bio-compatibility, with applications in liquid ventilation and synthetic blood. The unique properties of these compounds have also enabled mass spectrometry imaging of tissues where the fluorocarbons act as a Teflon-like coating for nanostructured surfaces to assist in desorption/ionization. Here we report fluorinated gold nanoparticles (f-AuNPs) designed to facilitate nanostructure imaging mass spectrometry. Irradiation of f-AuNPs results in the release of the fluorocarbon ligands providing a driving force for analyte desorption. The f-AuNPs allow for the mass spectrometry analysis of both lipophilic and polar (central carbon) metabolites. An important property of AuNPs is that they also act as contrast agents for X-ray microtomography and electron microscopy, a feature we have exploited by infusing f-AuNPs into tissue via fluorocarbon liquids to facilitate multi-modal (molecular and anatomical) imaging.

Age-related Macular Degeneration (AMD) is the leading cause of visual impairment and blindness in the elderly in developed countries. Neovascular/exudative (wet) AMD is the aggressive form of AMD and can involve choroidal neovascularization and vascular leakage. Anti-vascular endothelial growth factor (anti-VEGF) medications have significantly improved treatment of wet-AMD. However, only approximately 40% of patients obtain full benefit from anti-VEGF therapy and the medications are given by intravitreal injection. Axitinib, a small molecule multi-receptor tyrosine kinase inhibitor used for the treatment of advanced renal cell carcinoma, is taken orally and inhibits VEGF activity by blocking VEGF receptors. Axitinib also has the advantage of blocking platelet derived growth factor (PDGF) receptors which play a role in neovascularization. Using in vitro human retinal microvascular endothelial cells (HRMVECs), human brain vascular pericytes (HBVRs), 3D co-culture vessel sprout assay, and in vivo laser induced rat choroidal neovascularization (CNV) models, the effect of axitinib on neovascularization was evaluated. Axitinib inhibited neovascularization better than anti-VEGF and/or anti-hPDGF-B mAb in the in vitro models demonstrating that combined inhibition of both VEGF and PDGF pathways may be synergistic in treating wet-AMD. Additionally, axitinib showed good efficacy at a low dose (0.875 mg/day) in laser-induced CNV model in rats. In conclusion our data shows that axitinib, an inhibitor of VEGF and PDGF-B pathways may be useful in ameliorating wet-AMD therapy.

The role of PI3K and MAPK pathways in tumor initiation and progression is well established; hence, several inhibitors of these pathways are currently in different stages of clinical trials. Recent studies identified a PI3K/mTOR (PF-04691502) and a MEK inhibitor (PD-0325901) with strong potency and efficacy in different cell lines and tumor models. PD-0325901, however, showed adverse effects when administered at or above MTD (maximum tolerated dose) in the clinic. Here, we show in preclinical models that PD-0325901 at doses well below MTD (sub-MTD 1.5 mg/kg SID) is still a potent compound as single agent or in combination with PF-04691502. We first observed that PD-0325901 at 1.5 mg/kg SID and in combination with PF-04691502 (7.5 mg/kg; SID) significantly inhibited growth of H460 (carry Kras and PIK3CA mutations) orthotopic lung tumors. Additionally, we tested efficacy of PD-0325901 in Kras(G12D-LSL) conditional GEMMs (genetically engineered mouse models) which are a valuable tool in translational research to study tumor progression. Intranasal delivery of adenoviruses expressing Cre recombinase (Adeno-Cre) resulted in expression of mutant Kras leading to development of tumor lesions in lungs including adenomatous hyperplasia, large adenoma, and adenocarcinoma. Similar to H460 tumors, PD-0325901 as single agent or in combination with PF-04691502 significantly inhibited growth of tumor lesions in lungs in Kras(G12D-LSL) mice when treatment started at adenocarcinoma stage (at 14 weeks post-Adeno-Cre inhalation). In addition, immunohistochemistry showed inhibition of pS6 (phosphorylated ribosomal S6) in the treated animals particularly in the combination group providing a proof of mechanism for tumor growth inhibition. Finally, m-CT imaging in live Kras(G12D-LSL) mice showed reduction of tumor burdens in PD-0325901-treated animals at sub-MTD dose. In conclusion, our data suggest that PD-0325901 at doses below MTD is still a potent compound capable of tumor growth inhibition where Kras and/or PI3K are drivers of tumor growth and progression.

BackgroundCastration resistant prostate cancer (CRPC) is a leading cause of cancer-related deaths in men. The primary cause of mortality and morbidity in patients is bone metastases and remodeling resulting in osteoblastic and osteolytic lesions. Recently, cabozantinib, a multi-kinase inhibitor (VEGFR2 and c-MET inhibitor), was shown to have efficacy on bone lesions in patients. In this study we tested multi-kinase inhibitors: axitinib (VEGFR inhibitor) and crizotinib (c-MET inhibitor) in a combination trial in mice models.MethodsVCaP-Luc cells were grown as subcutaneous implants in intact and castrated NOD-SCID-gamma (NSG) mice to confirm the androgen dependency. For bone metastasis model two cohorts of NSG mice (castrated and intact) received orthotopic injection of VCaP-Luc cells into the bone marrow cavity of left tibia. Mice were monitored weekly for tumor growth using bioluminescence imaging. Animals were randomized into 4 groups based on the tumor bioluminescence signal: vehicle, crizotinib alone, axitinib alone, crizotinib and axitinib in combination. Animals were imaged weekly by in vivo 2-D X-ray imaging to monitor bone remodeling. At the end of the study animals were euthanized and both tibias were extracted for ex vivo high-resolution 3-D micro-computed tomography (μCT) imaging.ResultsSubcutaneous model showed that androgen stimulation may be helpful but not essential for the growth of VCaP-Luc cells. VCaP-Luc cells grown intra-tibially in intact animals caused extensive remodeling of bone with mixed osteoblastic (bone formation) and osteolytic (bone matrix dissolution) lesions. The osteoblastic lesions were predominant and at times extended beyond the tibial shaft into the surrounding tissue. In contrast, only osteolytic lesions were prominent throughout the study in castrated animals. Treatment with crizotinib alone reduced the osteolytic lesions in castrated animals. Axitinib alone reduced the osteoblastic lesions in the intact animals. Combination therapy with axitinib and crizotinib remarkably inhibited the tibial remodeling by VCaP-Luc cells which resulted in a significant reduction of both osteoblastic and osteolytic lesions.ConclusionOur data show that combined inhibition of c-MET and VEGFR can be beneficial for treatment of metastatic bone disease in CRPC and that the drugs act on two different stages of the disease.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2407-14-742) contains supplementary material, which is available to authorized users.

Introduction: Currently bio-distribution of biologic drugs is evaluated by PET imaging, autoradioraphy using radio-labeled molecules or ex vivo methods. Advances in optical probes and non-invasive imaging technologies have given us an opportunity to conduct such studies without the use of radio-labeled materials or by traditional pharmacokinetic (PK) studies. 5T4 (also known as TPBG or oncofetal antigen) is a transmembrane glycoprotein expressed highly on tumor-initiating cells. Anti-5T4-mcMMAF used in these studies is an anti-5T4-antibody drug conjugate (ADC) that reacts to human, cyno and marmoset orthologs of 5T4. Previously, we showed the efficacy of anti-5T4-ADC in pre-clinical models (Sapra et al.,). In this study we show the utility of Fluorescence Molecular Tomography (FMT) imaging in bio-distribution studies with this ADC using a H1975 non-small cell lung cancer (NSCLC) xenograft model. Methods: For the H1975 xenograft model, five million cells in 50% matrigel were injected into the subcutaneous flanks of the female nu/nu mice, and a biodistribution study was initiated when the tumors reached ∼500 mm3. The anti-5T4-ADC and a control ADC (non-binding) were conjugated with near-IR fluorophore VivoTag680XL. The labeling efficiency and quality was determined by Nanodrop-8000 spectrophotometer and binding assays. FMT imaging was performed longitudinally at 5min, 6hr, 24hr, 48hr, 96hr and 240hrs post injection of labeled ADCs. Ex vivo imaging of organs was performed at intermittent time points after perfusing with PBS. Data was analyzed using TrueQuant software. Plasma and tissues were collected at various time points and analyzed by GyrolabTM workstation and LCMS methods. Results: VivoTag680XL conjugation was efficient and achieved degree of labeling between 2-3. Three-dimensional quantitative analysis of FMT data showed significant specific targeting of anti-5T4-ADC to the tumors (ex vivo and in vivo comparison), relative to the control non-binding ADC. The peak accumulation in tumor was observed at 48hrs post injection and the concentration decreased in later time points. Liver was the major organ for the non-specific accumulation of these antibodies/ADCs, followed by kidneys and lung. Both 5T4-ADC and Control-ADC showed similar accumulation at 48hrs and 96hrs in liver, whereas it decreased significantly at 240hrs. The FMT imaging data was comparative and correlated with the traditional plasma PK profile data. Conclusion: These results show that anti-5T4-ADC targets the tumor better than non-binding ADC. This study also shows the utility of FMT in bio-distribution studies of biologics. Since the fluorophore can be conjugated to any protein/peptide, this novel approach can become a platform technology in conducting biodistribution studies of all biologic drugs. Citation Format: Anand Giddabasappa, Rand Norberg, Mauricio Leal, David Paterson, Kush Lalwani, Ted Levkoff, Stella Rapa, Puja Sapra, Michael Ritche, Joann Wentland, Brian Rago, Jeetendra Eswaraka. Whole-body bio-distribution of anti-5T4-mcMMAF (anti-5T4-ADC) using fluorescence molecular tomography (FMT) imaging in a non-small cell lung cancer mice model. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4293. doi:10.1158/1538-7445.AM2014-4293