Authors: Panigrahy D, Edin ML, Lee CR, Huang S, Bielenberg DR, Butterfield CE, Barnés CM, Mammoto A, Mammoto T, Luria A, Benny O, Chaponis DM, Dudley AC, Greene ER, Vergilio JA, Pietramaggiori G, Scherer-Pietramaggiori SS, Short SM, Seth M, Lih FB, Tomer KB, Yang J, Schwendener RA, Hammock BD, Falck JR, Manthati VL, Ingber DE, Kaipainen A, D'Amore PA, Kieran MW, Zeldin DC
Journal: J Clin Invest. 2012 Jan 3;122(1):178-91. doi: 10.1172/JCI58128. Epub 2011 Dec 19
Epoxyeicosatrienoic acids (EETs) are small molecules produced by cytochrome P450 epoxygenases. They are lipid mediators that act as autocrine or paracrine factors to regulate inflammation and vascular tone. As a result, drugs that raise EET levels are in clinical trials for the treatment of hypertension and many other diseases. However, despite their pleiotropic effects on cells, little is known about the role of these epoxyeicosanoids in cancer. Here, using genetic and pharmacological manipulation of endogenous EET levels, we demonstrate that EETs are critical for primary tumor growth and metastasis in a variety of mouse models of cancer. Remarkably, we found that EETs stimulated extensive multiorgan metastasis and escape from tumor dormancy in several tumor models. This systemic metastasis was not caused by excessive primary tumor growth but depended on endothelium-derived EETs at the site of metastasis. Administration of synthetic EETs recapitulated these results, while EET antagonists suppressed tumor growth and metastasis, demonstrating in vivo that pharmacological modulation of EETs can affect cancer growth. Furthermore, inhibitors of soluble epoxide hydrolase (sEH), the enzyme that metabolizes EETs, elevated endogenous EET levels and promoted primary tumor growth and metastasis. Thus, our data indicate a central role for EETs in tumorigenesis, offering a mechanistic link between lipid signaling and cancer and emphasizing the critical importance of considering possible effects of EET-modulating drugs on cancer.
Authors: Goldin E, Zheng W, Motabar O, Southall N, Choi JH, Marugan J, Austin CP, Sidransky E
Journal: PLoS One. 2012;7(1):e29861. Epub 2012 Jan 17.
Gaucher disease (GD), the most common lysosomal storage disorder, results from the inherited deficiency of the lysosomal enzyme glucocerebrosidase (GCase). Previously, wildtype GCase was used for high throughput screening (HTS) of large collections of compounds to identify small molecule chaperones that could be developed as new therapies for GD. However, the compounds identified from HTS usually showed reduced potency later in confirmatory cell-based assays. An alternate strategy is to perform HTS on mutant enzyme to identify different lead compounds, including those enhancing mutant enzyme activities. We developed a new screening assay using enzyme extract prepared from the spleen of a patient with Gaucher disease with genotype N370S/N370S. In tissue extracts, GCase is in a more native physiological environment, and is present with the native activator saposin C and other potential cofactors. Using this assay, we screened a library of 250,000 compounds and identified novel modulators of mutant GCase including 14 new lead inhibitors and 30 lead activators. The activities of some of the primary hits were confirmed in subsequent cell-based assays using patient-derived fibroblasts. These results suggest that primary screening assays using enzyme extracted from tissues is an alternative approach to identify high quality, physiologically relevant lead compounds for drug development.
Authors: Doria-Rose NA, Louder MK, Yang Z, O'Dell S, Nason M, Schmidt SD, McKee K, Seaman MS, Bailer RT, Mascola JR
Journal: J Virol. 2012 Jan 18. [Epub ahead of print]
HIV-1 neutralizing monoclonal antibodies (mAbs) define key targets for vaccine development and are being considered for passive prevention of infection. We analyzed the interaction of mAbs to two independent epitopes on the viral envelope glycoprotein. Potently neutralizing mAbs to the CD4 binding site and V1V2 region displayed no in vitro cross-competition and additive, though not synergistic, neutralization activity. Predicted neutralization coverage of a combination of two mAbs reached 97% on a 208-isolate panel.
Authors: Tyagi M, Hashimoto K, Shoemaker BA, Wuchty S, Panchenko AR
Journal: EMBO Rep. 2012 Jan 20. doi: 10.1038/embor.2011.261. [Epub ahead of print]
Although the identification of protein interactions by high-throughput (HTP) methods progresses at a fast pace, 'interactome' data sets still suffer from high rates of false positives and low coverage. To map the human protein interactome, we describe a new framework that uses experimental evidence on structural complexes, the atomic details of binding interfaces and evolutionary conservation. The structurally inferred interaction network is highly modular and more functionally coherent compared with experimental interaction networks derived from multiple literature citations. Moreover, structurally inferred and high-confidence HTP networks complement each other well, allowing us to construct a merged network to generate testable hypotheses and provide valuable experimental leads.
Authors: Englund EA, Wang D, Fujigaki H, Sakai H, Micklitsch CM, Ghirlando R, Martin-Manso G, Pendrak ML, Roberts DD, Durell SR, Appella DH
Journal: Nat Commun. 2012 Jan 10;3:614. doi: 10.1038/ncomms1629
Multivalent effects dictate the binding affinity of multiple ligands on one molecular entity to receptors. Integrins are receptors that mediate cell attachment through multivalent binding to peptide sequences within the extracellular matrix, and overexpression promotes the metastasis of some cancers. Multivalent display of integrin antagonists enhances their efficacy, but current scaffolds have limited ranges and precision for the display of ligands. Here we present an approach to studying multivalent effects across wide ranges of ligand number, density, and three-dimensional arrangement. Using L-lysine γ-substituted peptide nucleic acids, the multivalent effects of an integrin antagonist were examined over a range of 1-45 ligands. The optimal construct improves the inhibitory activity of the antagonist by two orders of magnitude against the binding of melanoma cells to the extracellular matrix in both in vitro and in vivo models.
Authors: Kim WG, Guigon CJ, Fozzatti L, Park JW, Lu C, Willingham MC, Cheng SY
Journal: Clin Cancer Res. 2012 Jan 23. [Epub ahead of print]
PURPOSE: Src is over-expressed or hyper-activated in a variety of human cancers including thyroid carcinoma. Src is a central mediator in multiple signaling pathways that are important in oncogenesis and cancer progression. In this study, we evaluated the effects of a Src inhibitor, SKI-606 (bosutinib), in a spontaneous metastatic thyroid cancer model with constitutively activated Src (ThrbPV/PVPten+/- mice).
EXPERIMENTAL DESIGN: ThrbPV/PVPten+/- mice were treated with SKI-606 or vehicle controls, beginning at 6 weeks of age until the mice succumbed to thyroid cancer. We assessed the effects of SKI-606 on thyroid cancer progression and analyzed the impact of SKI-606 on aberrant Src-mediated signaling.
RESULTS: SKI-606 effectively inhibited aberrant activation of Src and its downstream targets to markedly inhibit the growth of thyroid tumor, thereby prolonging the survival of treated mice. While Src inhibition did not induce cell apoptosis, it decreased cell proliferation by affecting the expression of key regulators of cell cycle progression. Importantly, SKI-606 dramatically prevented de-differentiation, vascular invasion, and lung metastasis of thyroid cancer cells. These responses were meditated by down-regulation of mitogen-activated protein kinase pathways and inhibition of the epithelial-mesenchymal transition.
CONCLUSIONS: Our findings suggest that Src is critical in the progression of thyroid cancer, making oral SKI-606 a promising treatment strategy for refractory thyroid cancer.
Authors: Ledgerwood JE, Wei CJ, Hu Z, Gordon IJ, Enama ME, Hendel CS, McTamney PM, Pearce MB, Yassine HM, Boyington JC, Bailer R, Tumpey TM, Koup RA, Mascola JR, Nabel GJ, Graham BS; VRC 306 Study Team
Journal: Lancet Infect Dis. 2011 Dec;11(12):916-24
BACKGROUND: Because the general population is largely naive to H5N1 influenza, antibodies generated to H5 allow analysis of novel influenza vaccines independent of background immunity from previous infection. We assessed the safety and immunogenicity of DNA encoding H5 as a priming vaccine to improve antibody responses to inactivated influenza vaccination.
METHODS: In VRC 306 and VRC 310, two sequentially enrolled phase 1, open-label, randomised clinical trials, healthy adults (age 18-60 years) were randomly assigned to receive intramuscular H5 DNA (4 mg) at day 0 or twice, at day 0 and week 4, followed by H5N1 monovalent inactivated vaccine (MIV; 90 μg) at 4 or 24 weeks, and compared with a two-dose regimen of H5N1 MIV with either a 4 or 24 week interval. Antibody responses were assessed by haemagglutination inhibition (HAI), ELISA, neutralisation (ID(80)), and immunoassays for stem-directed antibodies. T cell responses were assessed by intracellular cytokine staining. After enrolment, investigators and individuals were not masked to group assignment. VRC 306 and VRC 310 are registered with ClinicalTrials.gov, numbers NCT00776711 and NCT01086657, respectively.
FINDINGS: In VRC 306, 60 individuals were randomly assigned to the four groups (15 in each) and 59 received the vaccinations. In VRC 310, of the 21 individuals enrolled, 20 received the vaccinations (nine received a two-dose regimen of H5N1 MIV and 11 received H5 DNA at day 0 followed by H5N1 MIV at week 24). H5 DNA priming was safe and enhanced H5-specific antibody titres following an H5N1 MIV boost, especially when the interval between DNA prime and MIV boost was extended to 24 weeks. In the two studies, DNA priming with a 24-week MIV boost interval induced protective HAI titres in 21 (81%) of 26 of individuals, with an increase in geometric mean titre (GMT) of more than four times that of individuals given the MIV-MIV regimen at 4 or 24 weeks (GMT 103-206 vs GMT 27-33). Additionally, neutralising antibodies directed to the conserved stem region of H5 were induced by this prime-boost regimen in several individuals. No vaccine-related serious adverse events were recorded.
INTERPRETATION: DNA priming 24 weeks in advance of influenza vaccine boosting increased the magnitude of protective antibody responses (HAI) and in some cases induced haemagglutinin-stem-specific neutralising antibodies. A DNA-MIV vaccine regimen could enhance the efficacy of H5 or other influenza vaccines and shows that anti-stem antibodies can be elicited by vaccination in man.
FUNDING: National Institutes of Health.
Authors: Wallen GR, Baker K, Stolar M, Miller-Davis C, Ames N, Yates J, Bolle J, Pereira D, Germain DS, Handel D, Berger A.
Journal: Qual Life Res. 2012 Apr;21(3):405-15. Epub 2011 Nov 19.
PURPOSE: To prospectively compare outcomes and processes of hospital-based early palliative care with standard care in surgical oncology patients (N = 152).
METHODS: A randomized, mixed methods, longitudinal study evaluated the effectiveness of a hospital-based Pain and Palliative Care Service (PPCS). Interviews were conducted presurgically and at follow-up visits up to 1 year. Primary outcome measures included the Gracely Pain Intensity and Unpleasantness Scales and the Symptom Distress Scale. Qualitative interviews assessed social support, satisfaction with care, and communication with providers. Survival analysis methods explored factors related to treatment crossover and study discontinuation. Models for repeated measures within subjects over time explored treatment and covariate effects on patient-reported pain and symptom distress.
RESULTS: None of the estimated differences achieved statistical significance; however, for those who remained on study for 12 months, the PPCS group performed better than their standard of care counterparts. Patients identified consistent communication, emotional support, and pain and symptom management as positive contributions delivered by the PPCS.
CONCLUSIONS: It is unclear whether lower pain perceptions despite greater symptom distress were clinically meaningful; however, when coupled with the patients' perceptions of their increased resources and alternatives for pain control, one begins to see the value of an integrated PPCS.
Authors: McLellan JS, Pancera M, Carrico C, Gorman J, Julien JP, Khayat R, Louder R, Pejchal R, Sastry M, Dai K, O'Dell S, Patel N, Shahzad-ul-Hussan S, Yang Y, Zhang B, Zhou T, Zhu J, Boyington JC, Chuang GY, Diwanji D, Georgiev I, Kwon YD, Lee D, Louder MK, Moquin S, Schmidt SD, Yang ZY, Bonsignori M, Crump JA, Kapiga SH, Sam NE, Haynes BF, Burton DR, Koff WC, Walker LM, Phogat S, Wyatt R, Orwenyo J, Wang LX, Arthos J, Bewley CA, Mascola JR, Nabel GJ, Schief WR, Ward AB, Wilson IA, Kwong PD
Journal: Nature. 2011 Nov 23;480(7377):336-43. doi: 10.1038/nature10696
Variable regions 1 and 2 (V1/V2) of human immunodeficiency virus-1 (HIV-1) gp120 envelope glycoprotein are critical for viral evasion of antibody neutralization, and are themselves protected by extraordinary sequence diversity and N-linked glycosylation. Human antibodies such as PG9 nonetheless engage V1/V2 and neutralize 80% of HIV-1 isolates. Here we report the structure of V1/V2 in complex with PG9. V1/V2 forms a four-stranded β-sheet domain, in which sequence diversity and glycosylation are largely segregated to strand-connecting loops. PG9 recognition involves electrostatic, sequence-independent and glycan interactions: the latter account for over half the interactive surface but are of sufficiently weak affinity to avoid autoreactivity. The structures of V1/V2-directed antibodies CH04 and PGT145 indicate that they share a common mode of glycan penetration by extended anionic loops. In addition to structurally defining V1/V2, the results thus identify a paradigm of antibody recognition for highly glycosylated antigens, which-with PG9-involves a site of vulnerability comprising just two glycans and a strand.
Authors: Oh HM, Yu CR, Lee Y, Chan CC, Maminishkis A, Egwuagu CE
Journal: J Immunol. 2011 Sep 15;187(6):3338-46
Organ-specific autoimmune diseases are usually characterized by repeated cycles of remission and recurrent inflammation. However, where the autoreactive memory T cells reside in between episodes of recurrent inflammation is largely unknown. In this study, we have established a mouse model of chronic uveitis characterized by progressive photoreceptor cell loss, retinal degeneration, focal retinitis, retinal vasculitis, multifocal choroiditis, and choroidal neovascularization, providing for the first time to our knowledge a useful model for studying long-term pathological consequences of chronic inflammation of the neuroretina. We show that several months after inception of acute uveitis, autoreactive memory T cells specific to retinal autoantigen, interphotoreceptor retinoid-binding protein (IRBP), relocated to bone marrow (BM). The IRBP-specific memory T cells (IL-7Rα(High)Ly6C(High)CD4(+)) resided in BM in resting state but upon restimulation converted to IL-17/IFN-γ-expressing effectors (IL-7Rα(Low)Ly6C(Low)CD4(+)) that mediated uveitis. We further show that T cells from STAT3-deficient (CD4-STAT3KO) mice are defective in α4β1 and osteopontin expression, defects that correlated with inability of IRBP-specific memory CD4-STAT3KO T cells to traffic into BM. We adoptively transferred uveitis to naive mice using BM cells from wild-type mice with chronic uveitis but not BM cells from CD4-STAT3KO, providing direct evidence that memory T cells that mediate uveitis reside in BM and that STAT3-dependent mechanism may be required for migration into and retention of memory T cells in BM. Identifying BM as a survival niche for T cells that cause uveitis suggests that BM stromal cells that provide survival signals to autoreactive memory T cells and STAT3-dependent mechanisms that mediate their relocation into BM are attractive therapeutic targets that can be exploited to selectively deplete memory T cells that drive chronic inflammation.