Epacadostat is a novel inhibitor of indoleamine-2,3-dioxygenase-1 (IDO1) that suppresses systemic tryptophan catabolism and is currently being evaluated in ongoing clinical trials. We investigated the effects of epacadostat on (a) human dendritic cells (DCs) with respect to maturation and ability to activate human tumor antigen-specific cytotoxic T-cell (CTL) lines, and subsequent T-cell lysis of tumor cells, (b) human regulatory T cells (Tregs), and (c) human peripheral blood mononuclear cells (PBMCs) in vitro. Simultaneous treatment with epacadostat and IFN-γ plus lipopolysaccharide (LPS) did not change the phenotype of matured human DCs, and as expected decreased the tryptophan breakdown and kynurenine production. Peptide-specific T-cell lines stimulated with DCs pulsed with peptide produced significantly more IFN-γ, TNFα, GM-CSF and IL-8 if the DCs were treated with epacadostat. These T cells also displayed higher levels of tumor cell lysis on a per cell basis. Epacadostat also significantly decreased Treg proliferation induced by IDO production from IFN-γ plus LPS matured human DCs, although the Treg phenotype did not change. Multicolor flow cytometry was performed on human PBMCs treated with epacadostat; analysis of 123 discrete immune cell subsets revealed no changes in major immune cell types, an increase in activated CD83+ conventional DCs, and a decrease in immature activated Tim3+ NK cells. These studies show for the first time several effects of epacadostat on human DCs, and subsequent effects on CTL and Tregs, and provide a rationale as to how epacadostat could potentially increase the efficacy of immunotherapeutics, including cancer vaccines.
Refinement of treatment regimens enlisting targeted α-radiation therapy (TAT) is an ongoing effort. Among the variables to consider are the target molecule, radionuclide, dosage, and administration route. The panitumumab F(ab')2 fragment targeting epidermal growth factor receptor tolerated modification with the TCMC chelate as well as radiolabeling with 203Pb or 212Pb. Good specific activity was attained when the immunoconjugate was labeled with 212Pb (9.6 ± 1.4 mCi/mg). Targeting of LS-174T tumor xenografts with the 203Pb-panitumumab F(ab')2 demonstrated comparable amounts of uptake to the similarly radiolabeled panitumumab IgG. A dose escalation study was performed to determine an effective working dose for both intraperitoneal (i.p.) and intravenous (i.v.) injections of 212Pb-panitumumab F(ab')2. Therapeutic efficacy, with modest toxicity, was observed with 30 μCi given i.p. Results for the i.v. administration were not as definitive and the experiment was repeated with a higher dose range. From this study, 20 μCi given i.v. was selected as the effective working dose. A subsequent therapy study combined gemcitabine or paclitaxel with i.v. 212Pb-panitumumab F(ab')2, which increased the median survival (MS) of LS-174T tumor-bearing mice to 208 and 239 d, respectively. Meanwhile, the MS of mice treated with i.v. 212Pb-panitumumab F(ab')2 alone was 61 and 11 d for the untreated group of mice. In conclusion, the panitumumab F(ab')2 fragment whether given by i.p. or i.v. injection, is a viable candidate as a delivery vector for TAT of disseminated i.p. disease.
Identification of the appropriate combination of radionuclide, target and targeting vehicle is critical for successful radioimmunotherapy. For the treatment of disseminated peritoneal diseases such as pancreatic or ovarian cancer, α-emitting radionuclides have been proposed for targeted radiation therapy. This laboratory has taken a systematic approach investigating targeted α-radiation therapy, allowing comparisons to now be made between 211At, 227Th, 213Bi and 212Pb. Herein, trastuzumab radiolabeled with 211At and 227Th was evaluated for therapeutic efficacy in the LS-174T i.p. tumor model. A dose escalation study was conducted with each radioimmunoconjugate (RIC). Therapeutic benefit was realized with 211At-trastuzumab with doses of 20, 30 and 40 μCi. At doses >40 μCi, toxicity was observed with greater weight loss and 2-fold higher decrease in the platelet counts. Following a second study comparing the effect of 20, 30 and 40 μCi of 211At-trastuzumab, 30 μCi was selected as the dose for future studies. A parallel study was performed evaluating 0.25, 0.5, 1.0, 2.0 and 5.0 μCi of 227Th-trastuzumab. The 0.5 and 1.0 μCi injected dose resulted in a therapeutic response; a lower degree of weight loss was experienced by the mice in the 0.5 μCi cohort. When the data is normalized for comparing 211At, 227Th, 213Bi and 212Pb, the choice of radionuclide for RIT is perhaps not entirely based on simple therapeutic efficacy, other factors may play a role in choosing the "right" radionuclide.
Patagonian toothfish(Dissostichus eleginoides) was investigated in the southeastern Atlantic Ocean during the period of 19 March to 7 July 2007 by commercial bottom longliners. The number of 116 hauls were carried out individually in this study area by two bottom longliners during 111 days. The total catches were 89,559kg(9.91kg/100hooks) and 66,758kg(7.38kg/100hooks), respectively. Most Patagonian toothfishes were caught on the $46-48^{\circ}S$, $6-11^{\circ}E$ of southeastern Atlantic Ocean. Patagonian toothfish comprised 67.01-92.49%(by weight) of the catches in the southeastern Atlantic Ocean. No Antarctic toothfish (Dissostichus mawsoni) was caught during the surveys. Patagonian toothfish was found throughout the experimental area, but the density was considerably greater in the layer of 1,300-1,900m, where 49.88% of the fishes were caught. Smaller fishes were generally distributed to the shallow waters and larger fish were more frequently caught with the depth. Mean size of Patagonian toothfish increased with depth of capture. Accordingly economically profitable fishing activities for the Patagonian toothfish fishery requires an understanding of the size distribution of the Patagonian toothfish by depth.
This report describes synthesis and evaluation of novel cationic 99mTc-nitrido complexes, [99mTcN(L)(PNP)]+ (L = ma, ema, tma, etma and mpo; PNP = PNP5, PNP6, and L6), as potential radiotracers for heart imaging. Cationic complexes [99mTcN(L)(PNP)]+ were prepared in two steps. For example, reaction of succinic dihydrazide with 99mTcO4- in the presence of excess stannous chloride and PDTA resulted in the [99mTcN(PDTA)n] intermediate, which then reacted Hmpo and PNP6 at 100 °C for 10−15 min to give [99mTcN(mpo)(PNP6)]+ in >90% yield. It was found that bidentate chelators have a significant impact on lipophilicity, solution stability, biodistribution, and metabolic stability of cationic 99mTc-nitrido complexes. The fact that [99mTcN(ema)(PNP6)]+ decomposes rapidly in the presence of cysteine (1 mg/mL) while [99mTcN(etma)(PNP6)]+ and [99mTcN(mpo)(PNP6)]+ remain stable for >6 h under the same conditions strongly suggests that thione-S donors in bidentate chelators increase the solution stability of their cationic 99mTc-nitrido complexes. Biodistribution studies were performed on four cationic 99mTc-nitrido complexes in Sprague−Dawley rats. [99mTcN(etma)(PNP5)]+ is of particular interest due to its high initial heart uptake (1.81 ± 0.35 %ID/g at 5 min postinjection), and long myocardial retention (1.99 ± 0.47 %ID/g at 120 min postinjection). The heart/liver ratio of [99mTcN(etma)(PNP5)]+ (6.06 ± 1.48) at 30 min postinjection is almost identical that of 99mTcN-DBODC5 (6.01 ± 1.45), and is >2 times better than that of 99mTc-sestamibi (2.90 ± 0.22). Results from metabolism studies show that [99mTcN(etma)(PNP5)]+ has no significant metabolism in the urine, but it does show significant metabolism in feces samples at 120 min postinjection. Planar imaging studies suggest that [99mTcN(etma)(PNP5)]+ might be able to give clinically useful images of the heart as early as 30 min postinjection. [99mTcN(etma)(PNP5)]+ is a very promising candidate for more preclinical evaluations in various animal models.
This report describes the synthesis of two new cyclic RGD (Arg-Gly-Asp) dimers, 3 (E[G(3)-c(RGDfK)](2)) and 4 (G(3)-E[G(3)-c(RGDfK)](2)), and their corresponding conjugates 5 (HYNIC-E[G(3)-c(RGDfK)](2): HYNIC = 6-(2-(2-sulfonatobenzaldehyde)hydrazono)nicotinyl) and 6 (HYNIC-G(3)-E[G(3)-c(RGDfK)](2)). Integrin alpha(v)beta(3) binding affinities of 5 and 6 were determined by displacement of (125)I-echistatin bound to U87MG glioma cells. (99)(m)Tc complexes 7 ([(99m)Tc(5)(tricine)(TPPTS)]: TPPTS = trisodium triphenylphosphine-3,3',3''-trisulfonate) and 8 ([(99m)Tc(6)(tricine)(TPPTS)]) were prepared in high yield and high specific activity. Biodistribution and imaging studies were performed in athymic nude mice bearing U87MG glioma and MDA-MB-435 breast cancer xenografts. It was found that G(3) linkers are particularly useful for increasing integrin alpha(v)beta(3) binding affinity of cyclic RGD dimers and improving the tumor uptake and clearance kinetic of their (99)(m)Tc radiotracers. Complex 8 is a very promising radiotracer for the early detection of integrin alpha(v)beta(3)-positive tumors and may have the potential for noninvasive monitoring of tumor growth or shrinkage during antiangiogenic treatment.
Herein we report the preparation along with the in vivo and in vitro MRI characterization of two generation four and five cystamine core dendrimers loaded with thirty and fifty-eight derivatized Gd-DOTA (G4SS30, G5SS58) respectively. Likewise the development and characterization of two half-dendrimers conjugated to the F(ab′)2 fragment of the monoclonal antibody (mAb) panitumumab functionalized with a maleimide conjugation functional group site (Ab–(G4S15)4, Ab–(G5S29)4) are also described. The in vitro molar relaxivity of the Ab–(G4S15)4 conjugate, measured at pH 7.4, 22 °C, and 3T showed a moderate increase in relaxivity as compared to Magnevist (6.7 vs 4.0 mM–1 s–1) while the Ab–(G5S29)4 conjugate was 2-fold higher (9.1 vs 4.0 mM–1 s–1). The data showed that only a high injection dose (0.050 mmol Gd3+/kg) produced a detectable contrast enhanced contrast for the Ab–(G4S15)4 conjugate while a lower dose (0.035 mmol Gd3+/kg) was sufficient for the Ab–(G5S29)4 conjugate. The antibody–SMCC conjugate was purified by a Sephadex G-100 column, and the antibody-dendrimer-based agents were purified by spin filtration using a Centricon filter (50,000 MCO). The protein assay coupled with cysteine and Ellman's assay indicated an antibody to dendrimer ratio of 1:4. The in vivo blood clearance half-lives of the four agents measured at the jugular vein were ∼12–22 min.
Although the epidermal growth factor receptor (EGFR), also known as HER1, has been studied for over a decade, it continues to be a molecule of great interest and focus of investigators for development of targeted therapies. The marketed monoclonal antibody cetuximab binds to HER1, and thus might serve as the basis for creation of imaging or therapies that target this receptor. The potential of cetuximab as a vehicle for the delivery of α-particle radiation was investigated in an intraperitoneal tumor mouse model. The effective working dose of 10 μCi of 212Pb-cetuximab was determined from a dose (10–50 μCi) escalation study. Toxicity, as indicated by the lack of animal weight loss, was not evident at the 10 μCi dose of 212Pb-cetuximab. A subsequent study demonstrated 212Pb-cetuximab had a therapeutic efficacy similar to that of 212Pb-trastuzumab (p = 0.588). Gemcitabine given 24 h prior to 212Pb-cetuximab increased the median survival from 174 d to 283 d, but carboplatin suppressed the effectiveness of 212Pb-cetuximab. Notably, concurrent treatment of tumor-bearing mice with 212Pb-labeled cetuximab and trastuzumab provided therapeutic benefit that was greater than either antibody alone. In conclusion, cetuximab proved to be an effective vehicle for targeting HER1-expressing tumors with α-radiation for the treatment of disseminated intraperitoneal disease. These studies provide further evidence that the multimodality therapy regimens may have greater efficacy and benefit in the treatment of cancer patients.
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