SummaryAntibody-blocking studies have demonstrated the role of CD6 in thymocyte-thymic epithelial (TE) cell adhesion. Here we report that CD6 expressed by COS cells mediates adhesion to TE cells and that this interaction is specifically blocked with an anti-CD6 monodonal antibody (mAb) or with a mAb 04-81) that recognized a TE cell antigen. We isolated and expressed a cDNA clone encoding this antigen and show that COS cells transfected with this cDNA bind a CD6 immunoglobulin fusion protein (CD6-Rg). This antigen, which we named ALCAM (activated leukocyte-cell adhesion molecule) because of its expression on activated leukocytes, appears to be the human homologue of the chicken neural adhesion molecule BEN/SC-1/DM-GRASP. The gene was mapped to human chromosome 3q13.1-q13.2 by fluorescence in situ hybridization of cDNA probes to metaphase chromosomes. We prepared an ALCAM-Rg fusion protein and showed that it binds to COS cell transfectants expressing CD6, demonstrating that AI.CAM is a CD6 ligand. The observations that ALCAM is also expressed by activated leukocytes and that both ALCAM and CD6 are expressed in the brain suggest that ALCAM-CD6 interactions may play a role in the binding of T and B cells to activated leukocytes, as well as in interactions between cells of the nervous system.
The negative regulation of T-or B-cell antigen receptor signaling by CD5 was proposed based on studies of thymocytes and peritoneal B-1a cells from CD5-deficient mice. Here, we show that CD5 is constitutively associated with phosphotyrosine phosphatase activity in Jurkat T cells. CD5 was found associated with the Src homology 2 (SH2) domain containing hematopoietic phosphotyrosine phosphatase SHP-1 in both Jurkat cells and normal phytohemagglutinin-expanded T lymphoblasts. This interaction was increased upon T-cell receptor (TCR)-CD3 cell stimulation. CD5 co-cross-linking with the TCR-CD3 complex down-regulated the TCR-CD3-increased Ca 2؉ mobilization in Jurkat cells. In addition, stimulation of Jurkat cells or normal phytohemagglutinin-expanded T lymphoblasts through TCR-CD3 induced rapid tyrosine phosphorylation of several protein substrates, which was substantially diminished after CD5 cross-linking. The CD5-regulated substrates included CD3, ZAP-70, Syk, and phospholipase C␥l but not the Src family tyrosine kinase p56 lck . By mutation of all four CD5 intracellular tyrosine residues to phenylalanine, we found the membrane-proximal tyrosine at position 378, which is located in an immunoreceptor tyrosine-based inhibitory (ITIM)-like motif, crucial for SHP-1 association. The F378 point mutation ablated both SHP-1 binding and the down-regulating activity of CD5 during TCR-CD3 stimulation. These results suggest a critical role of the CD5 ITIM-like motif, which by binding to SHP-1 mediates the down-regulatory activity of this receptor.CD5 is a 67-kDa cell surface glycoprotein expressed on thymocytes, mature peripheral T cells, and a subpopulation of peritoneal B cells (B-1a cells) which are increased in some autoimmune diseases and are associated with the production of autoantibodies (7). Molecular cloning of mouse and human CD5 (mCD5 and hCD5) (16, 17) revealed that it belongs to the scavenger receptor cysteine-rich (SRCR) family group B, which comprises a group of leukocyte membrane or soluble proteins with one or more domains homologous to the aminoterminal domain of type I macrophage SRCR domain (21). Thus far, 10 members of this group of proteins have been identified: CD5, CD6, WC1, M130, Sp␣, Pema-SREG, Ebnerin, CPR-ductin, hensin, and gallbladder mucin (2).Biochemical studies suggest that CD5 is associated with CD3 in the T-cell receptor (TCR)-CD3 complex and with the B-cell receptor (BCR) complex (6,24,32). Two different ligands for CD5 have been reported: CD72, a 42-kDa type II constitutively expressed glycoprotein on B cells (28, 51); and CD5L, an activation antigen expressed on splenocytes (3). The physiologic roles of CD5-CD72 and CD5-CD5L interactions are not known but would be consistent with a potential T-cell-B-cell cooperation during antibody-mediated immune responses (8).Early in vitro studies of T lymphocytes and thymocytes demonstrated that monoclonal antibodies (MAbs) to CD5 were costimulatory for T-cell proliferation (9, 18, 42). However, in vivo studies showed that CD5 down-modulation by spe...
Controlling the biodistribution of nanoparticles upon intravenous injection is the key to achieving target specificity. One of the impediments in nanoparticle-based tumor targeting is the inability to limit the trafficking of nanoparticles to liver and other organs leading to smaller accumulated amounts in tumor tissues, particularly via passive targeting. Here we overcome both these challenges by designing nanoparticles that combine the specificity of antibodies with favorable particle biodistribution profiles, while not exceeding the threshold for renal filtration as a combined vehicle. To that end, ultrasmall silica nanoparticles are functionalized with anti-human epidermal growth factor receptor 2 (HER2) single-chain variable fragments to exhibit high tumor-targeting efficiency and efficient renal clearance. This ultrasmall targeted nanotheranostics/nanotherapeutic platform has broad utility, both for imaging a variety of tumor tissues by suitably adopting the targeting fragment and as a potentially useful drug delivery vehicle.
The pan B-cell surface antigen CD19 is an attractive target for therapeutic monoclonal antibody (mAb) approaches. We have generated a new afucosylated anti-human (hu)CD19 mAb, MEDI-551, with increased affinity to human Fc␥RIIIA and mouse Fc␥RIV and enhanced antibody-dependent cellular cytotoxicity (ADCC). During in vitro ADCC assays with B-cell lines, MEDI-551 is effective at much lower mAb concentrations than the fucosylated parental mAb anti-CD19-2. Furthermore, the afucosylated CD19 mAb MEDI-551 depleted B cells from normal donor peripheral blood mononuclear cell samples in an autologous ADCC assay, as well as blood and tissue B cells in human CD19/CD20 double transgenic (Tg) mice at lower concentrations than that of the positive control mAb rituximab. In huCD19/CD20 Tg mice, both macrophage-mediated phagocytosis and complement-dependent cytotoxicity contribute to depletion with rituximab; MEDI-551 did not require complement for maximal B-cell depletion. Furthermore, extended B-cell depletion from the blood and spleen was achieved with MEDI-551, which is probably explained by bone marrow B-cell depletion in huCD19/CD20 Tg mice relative to the control mAb rituximab. In summary, MEDI-551 has potent B-cell-depleting activity in vitro and in vivo and may be a promising new approach for the treatment of B-cell malignancies and autoimmune diseases.
BackgroundThe prevalence of visual impairment (VI) and dementia increases with age and these conditions may coexist, but few UK data exist on VI among people with dementia.ObjectivesTo measure the prevalence of eye conditions causing VI in people with dementia and to identify/describe reasons for underdetection or inappropriate management.DesignStage 1 – cross-sectional prevalence study. Stage 2 – qualitative research exploring participant, carer and professional perspectives of eye care.SettingStage 1 – 20 NHS sites in six English regions. Stage 2 – six English regions.ParticipantsStage 1 – 708 participants with dementia (aged 60–89 years): 389 lived in the community (group 1) and 319 lived in care homes (group 2). Stage 2 – 119 participants.InterventionsStage 1 gathered eye examination data following domiciliary sight tests complying with General Ophthalmic Services requirements and professional guidelines. Cognitive impairment was assessed using the Standardised Mini-Mental State Examination (sMMSE) test, and functional ability and behaviour were assessed using the Bristol Activities of Daily Living Scale and Cambridge Behavioural Inventory – Revised. Stage 2 involved individual interviews (36 people with dementia and 11 care workers); and separate focus groups (34 optometrists; 38 family and professional carers).Main outcome measures.VI defined by visual acuity (VA) worse than 6/12 or worse than 6/18 measured before and after refraction.ResultsStage 1 – when participants wore their current spectacles, VI prevalence was 32.5% [95% confidence interval (CI) 28.7% to 36.5%] and 16.3% (95% CI 13.5% to 19.6%) for commonly used criteria for VI of VA worse than 6/12 and 6/18, respectively. Of those with VI, 44% (VA < 6/12) and 47% (VA < 6/18) were correctable with new spectacles. Almost 50% of remaining uncorrectable VI (VA < 6/12) was associated with cataract, and was, therefore, potentially remediable, and one-third was associated with macular degeneration. Uncorrected/undercorrected VI prevalence (VA < 6/12) was significantly higher in participants in care homes (odds ratio 2.19, 95% CI 1.30 to 3.73;p < 0.01) when adjusted for age, sex and sMMSE score. VA could not be measured in 2.6% of group 1 and 34.2% of group 2 participants (p < 0.01). The main eye examination elements (excluding visual fields) could be performed in > 80% of participants. There was no evidence that the management of VI in people with dementia differed from that in older people in general. Exploratory analysis suggested significant deficits in some vision-related aspects of function and behaviour in participants with VI. Stage 2 key messages – carers and care workers underestimated how much can be achieved in an eye examination. People with dementia and carers were unaware of domiciliary sight test availability. Improved communication is needed between optometrists and carers; optometrists should be informed of the person’s dementia. Tailoring eye examinations to individual needs includes allowing extra time. Optometrists wanted training and guidance about dementia. Correcting VI may improve the quality of life of people with dementia but should be weighed against the risks and burdens of undergoing examinations and cataract surgery on an individual basis.LimitationsSampling bias is possible owing to quota-sampling and response bias.ConclusionsThe prevalence of VI is disproportionately higher in people with dementia living in care homes. Almost 50% of presenting VI is correctable with spectacles, and more with cataract surgery. Areas for future research are the development of an eye-care pathway for people with dementia; assessment of the benefits of early cataract surgery; and research into the feasibility of specialist optometrists for older people.FundingThe National Institute for Health Research Health Services and Delivery Research programme.
SllmmaryCD6 is a 130-kD glycoprotein expressed on the surface of thymocytes and peripheral blood T cells that is involved in TCK-mediated T cell activation. In thymus, CD6 mediates interactions between thymocytes and thymic epithelial (TE) cells. In indirect immunofluorescence assays, a recombinant CD6-immunoglobulin fusion protein (CD6-Kg) bound to cultured human TE cells and to thymic fibroblasts. CD6-Rg binding to TF and TE cells was trypsin sensitive, and 54 _+ 4% of binding was divalent cation dependent. By screening the blind panel of 479 monoclonal antibodies (mAbs) from the 5th International Workshop on Human Leukocyte Differentiation Antigens for expression on human TE cells and for the ability to block CD6-Rg binding to TE cells, we found one mAb 04-81) that significantly inhibited the binding of CD6-Kg to TE cells (60 _+ 7% inhibition). A second mAb to the surface antigen identified by mAb J4-81, J3-119, enhanced the binding of CD6-Kg to TE cells by 48 + 5%. Using covalent cross-linking and trypsin digestion, we found that mAb J4-81 and CD6-Kg both bound to the same 100-kD glycoprotein (CD6L100) on the surface of TE cells. These data demonstrate that a 100-kD glycoprotein on TE cells detected by mAb J4-81 is a ligand for CD6.
CD6 was established as a marker of T cells more than three decades ago, and recent studies have identified CD6 as a risk gene for multiple sclerosis (MS), a disease in which autoreactive T cells are integrally involved. Nevertheless, the precise role of CD6 in regulating T-cell responses is controversial and its significance in the pathogenesis of various diseases remains elusive, partly due to the lack of animals engineered to alter expression of the CD6 gene. In this report, we found that CD6 KO mice showed decreased pathogenic T-cell responses, reduced spinal cord T-cell infiltration, and attenuated disease severity in experimental autoimmune encephalomyelitis (EAE), an animal model of MS. CD6-deficient T cells exhibited augmented activation, but also significantly reduced survival and proliferation after activation, leading to overall decreased Th1 and Th17 polarization. Activated CD6-deficient T cells also showed impaired infiltration through brain microvascular endothelial cell monolayers. Furthermore, by developing CD6 humanized mice, we identified a mouse anti-human CD6 monoclonal antibody that is highly effective in treating established EAE without depleting T cells. These results suggest that () CD6 is a negative regulator of T-cell activation, () at the same time, CD6 is a positive regulator of activated T-cell survival/proliferation and infiltration; and () CD6 is a potential new target for treating MS and potentially other T-cell-driven autoimmune conditions.
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