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Longlong Luo,
Qun Luo,
Leiming Guo,
Ming Lv,
Zhou Lin,
Jing Geng,
Xinying Li,
Yan Li,
Beifen Shen,
Chunxia Qiao, Jiannan Feng
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ABSTRACT: Ricin is a highly lethal toxin. Anti-ricin chimeric monoclonal antibody (mAb) C4C13 was prepared in our lab; however, its binding affinity was much weaker than that of the parent antibody 4C13. In this study, based on the computer-guided homology modeling and conformational optimization methods, the 3-D structure of C4C13 variable regions Fv was constructed and optimized. Using molecular docking and dynamics simulation methods, the 3-D complex structure of ricin and C4C13 Fv was obtained. Considering the orientation property, surface electrostatic distribution, residues chemical andphysical character and intermolecular hydrogen bond, the binding mode and key residues were predicted. According to C4C13 Fv fragment and ricin complementary binding surface, electrostatic attraction periphery and van der Waals interaction interface, three mutants (i.e., M1 (N(H102)F, W(H103)Y); M2 (W(H103)Y) and M3 (R(L90)G)) were designed, in which M1 and M2 were predicted to possess higher antigen-binding activity than C4C13, while M3 was weaker. The relative affinity assays by ELISA showed that M1 and M2 mutations had higher affinity (9.6 and 18.3 nmol/L) than C4C13 (130 nmol/L) and M3 had weaker affinity (234.5 nmol/L) than C4C13. The results showed that the modeling complex structure of the antigen (ricin) and antibody (C4C13) is reasonable. Our work offered affinity maturated antibodies by site mutations, which were beneficial for valuable anti-ricin antibody design and preparation in future.
Journal of biomolecular structure & dynamics 03/2013; · 4.99 Impact Factor
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Xiaomei Yang,
Xingwei Jiang,
Guojiang Chen,
Yan Xiao,
Shaoxia Geng,
Chunyan Kang,
Tingting Zhou,
Yurong Li,
Xiaoqin Guo,
He Xiao,
Chunmei Hou,
Renxi Wang,
Zhou Lin,
Xinying Li, Jiannan Feng,
Yuanfang Ma,
Beifen Shen,
Yan Li,
Gencheng Han
[show abstract]
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ABSTRACT: Sepsis is an excessive inflammatory condition with a high mortality rate and limited prediction and therapeutic options. In this study, for the first time, to our knowledge, we found that downregulation and/or blockade of T cell Ig and mucin domain protein 3 (Tim-3), a negative immune regulator, correlated with severity of sepsis, suggesting that Tim-3 plays important roles in maintaining the homeostasis of sepsis in both humans and a mouse model. Blockade and/or downregulation of Tim-3 led to increased macrophage activation, which contributed to the systemic inflammatory response in sepsis, whereas Tim-3 overexpression in macrophages significantly suppressed TLR-mediated proinflammatory cytokine production, indicating that Tim-3 is a negative regulator of TLR-mediated immune responses. Cross-talk between the Tim-3 and TLR4 pathways makes TLR4 an important contributor to Tim-3-mediated negative regulation of the innate immune response. Tim-3 signaling inhibited LPS-TLR4-mediated NF-κB activation by increasing PI3K-AKT phosphorylation and A20 activity. This negative regulatory role of Tim-3 reflects a new adaptive compensatory and protective mechanism in sepsis victims, a finding of potential importance for modulating innate responses in these patients.
The Journal of Immunology 01/2013; · 5.79 Impact Factor
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Fengmin Shi,
Xiaoqin Guo,
Xingwei Jiang,
Ping Zhou,
Yan Xiao,
Tingting Zhou,
Guojiang Chen,
Zhi Zhao,
He Xiao,
Chunmei Hou,
Xinying Li,
Xiaomei Yang,
Renxi Wang, Jiannan Feng,
Beifen Shen,
Yan Li,
Gencheng Han
[show abstract]
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ABSTRACT: The pathogenesis of ulcerative colitis (UC) remains largely unclear. Here we found that T-cell Ig mucin-3 (Tim-3) and its ligand, galectin 9 (Gal-9), were significantly decreased in UC patients and in mice with dextran sodium sulfate (DSS)-induced colitis compared to controls. In addition to an enhanced Th17 response and attenuated regulatory T (Treg) cell response, there was also a significantly decreased Th1 response in UC. Levels of the Th1 cell chemokines CXCL9 and CXCL10 were significantly decreased in UC mice, partially explaining the decreased Th1 cell function in UC. Finally, administration of a putative antagonistic anti-Tim-3 antibody or of recombinant Gal-9 significantly exacerbated or attenuated DSS-induced colitis by altering the balance between different Th cell subsets. Our data suggest that a dysregulated Tim-3/Gal-9 pathway may contribute to the pathogenesis of UC. A better understanding of this pathway may shed new light on the pathogenesis of this disease.
Clinical Immunology 09/2012; 145(3):230-240. · 4.05 Impact Factor
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Yuanyuan Guo,
Wendie Wang,
Jing Wang, Jiannan Feng,
Qingyang Wang,
Jianfeng Jin,
Ming Lv,
Xinying Li,
Yan Li,
Yuanfang Ma,
Beifen Shen,
Jiyan Zhang
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ABSTRACT: c-Jun N-terminal protein kinase (JNK) is a member of the mitogen-activated protein kinase (MAPK) superfamily. The activation of JNK is mediated by sequential protein phosphorylation through a MAPK module, namely, MAPK kinase kinase (MAP3K or MEKK) ? MAPK kinase (MAP2K or MKK) ? MAPK. Elevated levels of JNK activity have been frequently seen in hepatocellular carcinoma (HCC) and have been demonstrated to contribute to HCC growth by promoting HCC cell proliferation and resistance to tumour necrosis factor-related apoptosis inducing ligand (TRAIL)- or Fas-mediated apoptosis. Apparently, chronic inflammation contributes to the up-regulation of JNK activity in HCC. However, it remains unknown whether aberrant JNK activity also results from some cell intrinsic defect(s). Here, we show that receptor for activated C kinase 1 (RACK1), an adaptor protein implicated in the regulation of multiple signaling pathways, could engage in a direct interaction with MKK7, the JNK specific MAP2K, in human HCC cells. The levels of RACK1 protein show correlation with the activity of the JNK pathway in human HCC tissues and cell lines. RACK1 loss-of-function or gain-of-function analyses indicate that RACK1 enhances MKK7/JNK activity in human HCC cells. Further exploration reveals that the interaction of RACK1 with MKK7 is required for the enhancement of MKK7/JNK activity by RACK1. RACK1/MKK7 interaction facilitates the association of MKK7 with MAP3Ks, thereby enhancing MKK7 activity and promoting in vitro HCC cell proliferation and resistance to TRAIL- or Fas-mediated apoptosis as well as in vivo tumor growth. Conclusion: Overexpressed RACK1 augments JNK activity and thereby promotes HCC growth through directly binding to MKK7 and enhancing MKK7 activity. (HEPATOLOGY 2012.).
Hepatology 08/2012; · 11.66 Impact Factor
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Chunxia Qiao,
Meiyun Hu,
Leiming Guo,
Ming Lv,
Zhou Lin,
Jing Geng,
Xiaoling Lang,
Xinying Li,
Yan Li,
Yuanfang Ma, Jiannan Feng,
Beifen Shen
[show abstract]
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ABSTRACT: As a member of the TNF superfamily, TRAIL could induce human tumor cell apoptosis through its cognate death receptors DR4 or DR5, which can induce formation of the death inducing signaling complex (DISC) and activation of the membrane proximal caspases (caspase-8 or caspase-10) and mitochondrial pathway. Some monoclonal antibodies against DR4 or DR5 have been reported to have anti-tumor activity.
In this study, we reported a novel mouse anti-human DR5 monoclonal antibody, named as LaDR5, which could compete with TRAIL to bind DR5 and induce the apoptosis of Jurkat cells in the absence of second cross-linking in vitro. Using computer-guided molecular modeling method, the 3-D structure of LaDR5 Fv fragment was constructed. According to the crystal structure of DR5, the 3-D complex structure of DR5 and LaDR5 was modeled using molecular docking method. Based on distance geometry method and intermolecular hydrogen bonding analysis, the key functional domain in DR5 was predicted and the DR5 mutants were designed. And then, three mutants of DR5 was expressed in prokaryotic system and purified by affinity chromatograph to determine the epitope of DR5 identified by LaDR5, which was consistent with the theoretical results of computer-aided analysis.
Our results demonstrated the specific epitope located in DR5 that plays a crucial role in antibody binding and even antineoplastic bioactivity. Meanwhile, revealed structural features of DR5 may be important to design or screen novel drugs agonist DR5.
BMC Immunology 07/2012; 13:40. · 2.53 Impact Factor
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ABSTRACT: HER2 plays a critical role in the pathogenesis of many cancers and is linked to poor prognosis or cancer metastases. Monoclonal antibodies, such as Herceptin against HER2-overexpressing cancers, have showed satisfactory clinical therapeutic effect. However, they have difficulty to surmount obstacles to enter cells or blood-brain barrier.
In this study, a cell-penetrating peptide Arg9 was linked to the C-terminus of anti-HER2 single chain antibody (MIL5scFv). Flow cytometry, confocal microscopy and electron microscopy analysis all revealed that Arg9 peptide facilitated the penetration of MIL5scFv into HER2-negative cell line NIH3T3 and orientate in mitochondria. More interestingly, Western blot assay showed the potential enhanced bioactivity of MIL5scFv-Arg9 in HER2+ cell line SKOV3, indicating that Arg9 could help large molecules (e.g. antibody) to penetrate into cells and therefore enhance its anti-neoplastic function.
Our work represented an attractive by preliminary strategy to enhance the therapeutic effect of existing antibodies by entering cells easier, or more desirable, surmounting the physical barriers, especially in hard-to-reach cancers such as brain metastases cases.
BMC Research Notes 07/2012; 5:336.
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ABSTRACT: In this study the three-dimensional (3-D) model of the ligand-binding domain (V106-P322) of human interleukin-6 receptor (hIL-6
R) was constructed by computer-guided homology modeling technique using the crystal structure of the ligand-binding domain
(K52-L251) of human growth hormone receptor (hGHR) as templet. Furthermore, the active binding region of the 3-D model of
hIL-6R with the ligand (hIL-6) was predicted. In light of the structural characteristics of the active region, a hydrophobic
pocket shielded by two hydrophilic residues (E115 and E505) of the region was identified by a combination of molecular modelling
and the site-directed or double-site mutation of the twelve crucial residues in the ligand-binding domain of hIL-6R (V106-P322).
We observed and analyzed the effects of these mutants on the spatial conformation of the pocket-like region of hIL-6 R. The
results indicated that any site-directed mutation of the five Cys residues (four conservative Cys residues: Cys121, Cys132,
Cys165, Cys176; near membrane Cys residue: Cys193) or each double-site mutation of the five residues in WSEWS motif of hIL-6R
(V106-P322) makes the corresponding spatial conformation of the pocket region block the linkage between hIL-6 R and hIL-6.
However, the influence of the site-directed mutation of Cys211 and Cys277 individually on the conformation of the pocket region
benefits the interaction between hIL-6R and hIL-6. Our study suggests that the predicted hydrophobic pocket in the 3-D model
of hIL-6R (V106-P322) is the critical molecular basis for the binding of hIL-6R with its ligand, and the active pocket may
be used as a target for designing small hIL-6R-inhibiting molecules in our further study.
Science in China Series C Life Sciences 04/2012; 43(4):425-432. · 1.61 Impact Factor
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ABSTRACT: Based on the spatial conformations of human interleukin-6 (hIL-6) derived from nuclear magnetic resonance analysis and human
interleukin-6 receptor (hIL-6R) modeled with homology modeling method using human growth hormone receptor as template, the
interaction between hIL-6 and its receptor (hIL-6R) is studied with docking program according to the surface electrostatic
potential analysis and spatial conformation complement. The stable region structure composed of hIL-6 and hIL-6R is obtained
on the basis of molecular mechanism optimization and molecular dynamics simulation. The binding domain between hIL-6 and hIL-6R
is predicted theoretically. Furthermore, the especial binding sites that influence the interaction between hIL-6 and hIL-6R
are confirmed. The results lay a theoretical foundation for confirming the active regions of hIL-6 and designing novel antagonist
with computer-guided techniques.
Science in China Series C Life Sciences 04/2012; 43(4):409-417. · 1.61 Impact Factor
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ABSTRACT: Previous studies have shown that different epitopes of HER2 exhibit distinct functions and that the epitope bound by the antibody 2C4 plays a role in formation of hetereodimers between HER2 and other receptors of the HER family. In this study, we used computer modeling to determine that the epitope of HER2 which the C-terminal 79 amino acids of herstatin (named HSTC79) binds is similar to that bound by 2C4. Based on these theoretical results, recombinant HSTC79 fused with GST was expressed in Escherichia coli and purified by affinity chromatography. Experimental analysis showed that HSTC79 did specifically bind to HER2 and that the epitope of HER2 identified by HSTC79 was near that identified by 2C4. Furthermore, HSTC79 inhibited the growth of HER2-overexpressing cells. These results highlight the fact that the binding site architecture and certain key residues of HER2 may be very helpful for understanding the protein's biological role and providing insights for designing novel inhibitors of HER2.
Molecular Biotechnology 12/2011; 51(2):174-82. · 2.17 Impact Factor
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Huawei Wei,
Zhou Lin, Jiannan Feng,
Hui Peng,
Renfeng Guo,
Gencheng Han,
Shusheng Geng,
Xiaoling Lang,
Yingxun Sun,
Beifen Shen,
Yan Li
[show abstract]
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ABSTRACT: Inhibition of C5a by antibodies has been demonstrated to dramatically improve survival in various sepsis models in mice and rats. The structural basis of C5a mediated bioactivity and C5a antibody mediated neutralization are of interesting to be investigated. In the previous study, we obtained a novel functional mouse antibody named as F20. With computer-guided modeling method, the 3-D theoretical structure of F20 Fv fragment was constructed. Using the crystal structure of C5a, the 3-D complex structure of C5a and F20 Fv fragment was modeled with molecular docking method. Based on distance geometry method and intermolecular interaction theory, the key residue Lys(68) in C5a identified by F20 was predicted. The mutant experimental results showed that the residue Lys(68) was the critical residue of C5a for it's bioactivity and F20 binding activity. The present study shed new light on the structural basis of C5a mediated bioactivity. The identification of the critical residue will provide useful information for human complement C5a targeted therapeutic intervention.
Molecular Immunology 07/2011; 48(12-13):1377-83. · 2.90 Impact Factor
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Yurong Li, Jiannan Feng,
Shaoxia Geng,
Shusheng Geng,
Huawei Wei,
Guojiang Chen,
Xinying Li,
Liyan Wang,
Renxi Wang,
Hui Peng,
Gencheng Han,
Beifen Shen,
Yan Li
[show abstract]
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ABSTRACT: By binding to T cell Ig mucin-3 (Tim-3) expressed on different cells, galectin-9 (Gal-9) mediates two important functions, triggering T cell death and activating innate immune cells. The mechanisms by which ligation of the same molecule on different cell types mediates different effects are largely unclear. Gal-9 contains two carbohydrate recognition domains (CRD) in the N- and C-terminal regions (Gal-9-N and Gal-9-C). The N and C terminals of Gal-9 have been shown to have different activities in promoting T cell death. However, whether the differences between two domains account for its dual functions remains to be elucidated. Here we hypothesized that the different functions of Gal-9 in innate immunity and adaptive immunity are mediated by different domains. To test this, we created recombinant Gal-9 (Gal-9-NC) and homodimers containing either the NCRD (Gal-9-N) or the CCRD (Gal-9-C). All these Gal-9 constructs can activate dendritic cells (DCs) and induce T cell death. However, the Gal-9-C was much more potent than the Gal-9-N in inducing T cell death, while the Gal-9-N was much more effective in activating DCs by inducing much higher TNF-α and IL-6 production, greater phosphorylation of p38 and AKT. In both DC and T cells, Gal-9-N but not Gal-9-C stimulation resulted in markedly iκBα degradation. Finally, computer analyses suggested different patterns and affinities for the binding of the Gal-9-N and Gal-9-C to their receptor, Tim-3. Our data suggest that the N- and C-terminal CRDs of Gal-9 contribute differently to its ability to induce T cell death and to activate DCs. Further investigations on the underlying mechanisms will provide new insights into the biochemical basis for the multiple activities of Gal-9.
Molecular Immunology 01/2011; 48(4):670-7. · 2.90 Impact Factor
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Liyan Wang,
Gencheng Han,
Renxi Wang,
Guojiang Chen,
Ruonan Xu,
He Xiao,
Xia Li,
Shaoxia Geng,
Yurong Li,
Xinying Li,
Jianan Wang, Jiannan Feng,
Niels C Riedemann,
Renfeng Guo,
Beifen Shen,
Yan Li
[show abstract]
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ABSTRACT: Excessive complement-activated product complement 5a (C5a) has been implicated in the pathogenesis of sepsis development. Herein, we employed in vitro and in vivo models of sepsis to investigate the functional relationship between overtly produced C5a and IL-8. Our data revealed that C5a could strongly amplify IL-8 expression from human whole blood cells induced by LPS and other types of TLR agonists. ERK1/2 and p38, but not JNK, were mainly participated in signaling pathways for IL-8 production. In the whole blood stimulated by Escherichiacoli, C5a levels were quickly elevated and blockage of C5a significantly decreased E. coli-elicited IL-8 production. In the mouse model of sepsis induced by cecal ligation and puncture (CLP), the markedly increased keratinocyte-derived cytokine (KC) could be strongly suppressed by blockage of C5a. These data suggest that excessive C5a functions as a critical inflammatory mediator to enhance IL-8 production mainly through MAPK signaling pathways.
Clinical Immunology 10/2010; 137(1):157-65. · 4.05 Impact Factor
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ABSTRACT: Rituximab is the first anti-cancer antibody approved by the FDA for the treatment of B-cell lymphoma. However, its efficacy remains variable and often modest. Some patients are initially unresponsive to rituximab or later develop resistance to it, and require alternative therapies. Rituximab activity has been thought to involve antibody-dependent cellular cytotoxicity (ADCC), complement-dependent cytotoxicity (CDC) and apoptosis. Present studies suggest that the patients unresponsive to rituximab may be helped with other CD20 antibodies with enhanced activities. In this study, we characterized a novel anti-CD20 chimeric antibody, TGLA, which binds to various B-cell lines specially and shares an epitope with rituximab. TGLA shows equal activities with rituximab, such as CDC, cell growth arrest and so on. Interestingly, TGLA also shows significant ADCC activity. Immunotherapeutic studies further show that TGLA is far more effective in delaying tumor growth than rituximab. These findings suggest that the ADCC-enhanced anti-CD20 antibody TGLA might be an alternative therapeutic agent for B-cell lymphoma.
Cancer letters 03/2010; 294(1):66-73. · 4.86 Impact Factor
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Jie Dong,
Yaping Gao,
Yu Liu,
Jinxia Shi, Jiannan Feng,
Zhanguo Li,
Heping Pan,
Yanning Xue,
Chuan Liu,
Beifen Shen,
Ningsheng Shao,
Guang Yang
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ABSTRACT: Tumor necrosis factor alpha (TNF-alpha) is a major inflammatory mediator that exhibits actions leading to tissue destruction and hampering recovery from damage. At present, two antibodies against human TNF-alpha (hTNF-alpha) are available, which are widely used for the clinic treatment of certain inflammatory diseases. This work was undertaken to identify a novel functional epitope of hTNF-alpha. We performed screening peptide library against anti-hTNF-alpha antibodies, ELISA and competitive ELISA to obtain the epitope of hTNF-alpha. The key residues of the epitope were identified by means of combinatorial alanine scanning and site-specific mutagenesis. The N terminus (80-91 aa) of hTNF-alpha proved to be a novel epitope (YG1). The two amino acids of YG1, proline and valine, were identified as the key residues, which were important for hTNF-alpha biological function. Furthermore, the function of the epitope was addressed on an animal model of collagen-induced arthritis (CIA). CIA could be suppressed in an animal model by prevaccination with the derivative peptides of YG1. The antibodies of YG1 could also inhibit the cytotoxicity of hTNF-alpha. These results demonstrate that YG1 is a novel epitope associated with the biological function of hTNF-alpha and the antibodies against YG1 can inhibit the development of CIA in animal model, so it would be a potential target of new therapeutic antibodies.
PLoS ONE 01/2010; 5(1):e8920. · 4.09 Impact Factor
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ABSTRACT: Hyperactivation of c-Jun NH2-terminal protein kinase (JNK) has been found in various malignant lymphocytes and inhibition of JNK activity leads to cell cycle arrest and apoptosis. However, the role of JNK activity in the oncogenic growth of T-cell acute lymphoblastic leukemia (T-ALL) cells remains largely unknown. Here, we report that treatment of T-ALL cells with JNK inhibitors led to cell cycle arrest and apoptosis and increased sensitivity to Fas-mediated apoptosis, whereas weak ectopic expression of MKK7-JNK1 fusion protein, which shows constitutive JNK activity, in T-ALL cells resulted in accelerated cell cycle progression and resistance to Fas-mediated apoptosis. The protein levels of c-Myc and Bcl-2 were reduced in the presence of JNK inhibitors but were enhanced with MKK7-JNK1. Small interfering RNA against JNK1, but not JNK2, exhibited similar effects to JNK inhibitors. These findings suggest that targeting JNK, especially JNK1 isoform, may have some important therapeutic implications in the treatment of T-ALL. Further exploration revealed that JNK protein and basal JNK activity in T-ALL cells showed aberrant subcellular localization, but no hyperactivation of JNK was observed. Thus, our work suggests that there might be novel mechanism(s) other than hyperactivation underlying the protumorigenic role of JNK activity.
Molecular Cancer Therapeutics 12/2009; 8(12):3214-22. · 5.23 Impact Factor
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Xia Li,
Guojiang Chen,
Yurong Li,
Renxi Wang,
Liyan Wang,
Zhou Lin,
Xudong Gao, Jiannan Feng,
Yuanfang Ma,
Beifen Shen,
Yan Li,
Gencheng Han
[show abstract]
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ABSTRACT: Augmented intestinal T cells, especially CD4(+)T cells, are involved in the pathogenesis of inflammatory bowel disease (IBD). We used a murine 2, 4, 6-trinitrobenzene sulfonic acid (TNBS)-induced colitis model to investigate whether Tim-3, a negative regulator of CD4(+)T cells, is involved in the suppression of IBD. We found that blocking the Tim-3 signal pathway exacerbated TNBS-induced colitis, as shown by increased weight loss and aggravated tissue injury. Blockade of the Tim-3 pathway resulted in an increase in Tim-3(+)CD4T cells, a biased T effector cell response, and a decrease in Treg cells. It also resulted in an altered profile of co-stimulatory molecules expressed on lymphocytes, which partially explained the biased polarization of different T cell subsets. Our data suggest that the Tim-3 pathway is highly involved in the negative regulation of IBD. A better understanding of this pathway may shed new light on the pathogenesis of this disease.
Clinical Immunology 11/2009; 134(2):169-77. · 4.05 Impact Factor
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ABSTRACT: Previous studies indicated that a partial T-cell receptor signal delivered by non-mitogenic anti-CD3 antibodies is critical for dampening the activated T-cell response. The mini-yCD3 is a novel non-mitogenic anti-CD3 antibody based on a murine anti-human CD3 antibody yCD3. However, the mechanism by which mini-yCD3 suppresses immune responses mediated by activated T-cells remains unknown. To elucidate its mechanism, we examined the effects of the mini-yCD3 on early signaling events in T-cells. Similar to the mitogenic anti-CD3 mAb, mini-yCD3 triggered changes in the T-cell receptor (TCR). However, unlike the mitogenic anti-CD3 stimulation, mini-yCD3 was ineffective at inducing the highly phosphorylated zeta chain and tyrosine phosphorylation of the associated tyrosine kinase ZAP-70. This proximal signaling deficiency failed to mobilize detectable Ca(2+) and translocate NF-AT into the nucleus. Additionally, the non-mitogenic anti-CD3 appeared insufficient for the redistribution of TCRs into an aggregated cap, which correlated with T-cell activation.
International immunopharmacology 11/2009; 10(2):200-6. · 2.21 Impact Factor
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ABSTRACT: One of existing strategies to engineer active antibody is to link V(H) and V(L) domains via a linker peptide. How the composition, length, and conformation of the linker affect antibody activity, however, remains poorly understood. In this study, a dual approach that coordinates molecule modeling, biological measurements, and affinity evaluation was developed to quantify the binding activity of a novel stable miniaturized anti-CD20 antibody or single-chain fragment variable (scFv) with a linker peptide. Upon computer-guided homology modeling, distance geometry analysis, and molecular superimposition and optimization, three new linker peptides PT1, PT2, and PT3 with respective 7, 10, and 15 residues were proposed and three engineered antibodies were then constructed by linking the cloned V(H) and V(L) domains and fusing to a derivative of human IgG1. The binding stability and activity of scFv-Fc chimera to CD20 antigen was quantified using a micropipette adhesion frequency assay and a Scatchard analysis. Our data indicated that the binding affinity was similar for the chimera with PT2 or PT3 and approximately 24-fold higher than that for the chimera with PT1, supporting theoretical predictions in molecular modeling. These results further the understanding in the impact of linker peptide on antibody structure and activity.
Annals of biomedical engineering 10/2009; 38(2):537-49. · 2.41 Impact Factor
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Gencheng Han,
Renxi Wang,
Guojiang Chen,
Jianan Wang,
Ruonan Xu,
Liyan Wang, Jiannan Feng,
Xia Li,
Renfeng Guo,
Li Fu,
Beifen Shen,
Yan Li
[show abstract]
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ABSTRACT: Whether interleukin (IL)-17 promotes a diabetogenic response remains unclear. Here we examined the effects of neutralization of IL-17 on the progress of adoptively transferred diabetes. IL-17-producing cells in non-obese diabetic (NOD) mice were identified and their role in the pathogenesis of diabetes examined using transfer and co-transfer assays. Unexpectedly, we found that in vivo neutralization of IL-17 did not protect NOD-severe combined immunodeficiency (SCID) mice against diabetes transferred by diabetic splenocytes. In NOD mice, gammadelta(+) T cells were dominated by IL-17-producing cells and were found to be the major source of IL-17. Interestingly, these IL-17-producing gammadelta T cells did not exacerbate diabetes in an adoptive transfer model, but had a regulatory effect, protecting NOD mice from diabetes by up-regulating transforming growth factor (TGF)-beta production. Our data suggest that the presence of IL-17 did not increase the chance of the development of diabetes; gammadelta T cells protected NOD mice from diabetes in a TGF-beta-dependent manner, irrespective of their role as major IL-17 producers.
Immunology 08/2009; 129(2):197-206. · 3.32 Impact Factor
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ABSTRACT: p38 is a subfamily of the mitogen-activated protein kinase (MAPK) superfamily with four isoforms. It has been well established that p38 plays a central role in the production of inflammatory molecules and is therefore required for the activation of macrophages in response to inflammatory stimuli. However, little is known about the roles of p38 in macrophage development. The difficulty to get mice deficient in multiple p38 isoforms complicates the study of p38 in macrophage development. With the model of bone marrow-derived murine macrophages and highly selective p38alpha/beta inhibitors SB203580 and SB239063, here we report that macrophage colony-stimulating factor (M-CSF) induces p38 activation during macrophage development. Inhibition of p38 activity showed minor effects on macrophage proliferation or survival, and did not block CD14, F4/80 expression. However, p38 inhibitors resulted in a significant reduction in CD54 expression and impaired phagocytic activity. Taken together, our data suggest that p38 contributes to macrophage development.
Cellular Immunology 02/2009; 256(1-2):6-11. · 1.97 Impact Factor
