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Antibody-mediated complement-dependent cytotoxicity (CDC) on malignant cells is regulated by several complement control proteins, including the inhibitory complement factor H (fH). fH consists of 20 short consensus repeat elements (SCRs) with specific functional domains. Previous research revealed that the fH-derived SCRs 19-20 (SCR1920) can displace full-length fH on the surface of chronic lymphocytic leukemia (CLL) cells, which sensitizes CLL cells for e.g. CD20-targeting therapeutic monoclonal antibody (mAb) induced CDC. Therefore, we constructed lentiviral vectors for the generation of cell lines that stably produce mAb-SCR-fusion variants starting from the clinically approved parental mAbs rituximab, obinutuzumab and ofatumumab, respectively. Flowcytometry revealed that the modification of the mAbs by the SCRs does not impair the binding to CD20. Increased in vitro lysis potency compared to their parental mAbs was corroborated by showing specific and dose dependent target cell elimination by CDC when compared to their parental mAbs. Lysis of CLL cells was not affected by the depletion of NK cells, suggesting that antibodydependent cellular cytotoxicity plays a minor role in this context. Overall, this study emphasizes the crucial role of CDC in the elimination of CLL cells by mAbs and introduces a novel approach for enhancing CDC by directly fusing fH SCR1920 with mAbs. [ABSTRACT FROM AUTHOR]

Introduction: COVID-19 patients can develop autoantibodies against a variety of secreted and membrane proteins, including some expressed on lymphocytes. However, it is unclear what proportion of patients might develop antilymphocyte antibodies (ALAb) and what functional relevance they might have. Methods: We evaluated the presence and lytic function of ALAb in the sera of a cohort of 85 COVID-19 patients (68 unvaccinated and 17 vaccinated) assigned to mild (N=63), or moderate/severe disease (N=22) groups. Thirty-seven patients were followed-up after recovery. We also analyzed in vivo complement deposition on COVID-19 patients' lymphocytes and examined its correlation with lymphocyte numbers during acute disease. Results: Compared with healthy donors (HD), patients had an increased prevalence of IgM ALAb, which was significantly higher in moderate/severe disease patients and persisted after recovery. Sera from IgM ALAb+ patients exhibited complement-dependent cytotoxicity (CDC) against HD lymphocytes. Complement protein C3b deposition on patients' CD4 T cells was inversely correlated with CD4 T cell numbers. This correlation was stronger in moderate/severe disease patients. Discussion: IgM ALAb and complement activation against lymphocytes may contribute to the acute lymphopenia observed in COVID-19 patients. [ABSTRACT FROM AUTHOR]

Antibody-mediated complement-dependent cytotoxicity (CDC) on malignant cells is regulated by several complement control proteins, including the inhibitory complement factor H (fH). fH consists of 20 short consensus repeat elements (SCRs) with specific functional domains. Previous research revealed that the fH-derived SCRs 19–20 (SCR1920) can displace full-length fH on the surface of chronic lymphocytic leukemia (CLL) cells, which sensitizes CLL cells for e.g. CD20-targeting therapeutic monoclonal antibody (mAb) induced CDC. Therefore, we constructed lentiviral vectors for the generation of cell lines that stably produce mAb-SCR-fusion variants starting from the clinically approved parental mAbs rituximab, obinutuzumab and ofatumumab, respectively. Flow-cytometry revealed that the modification of the mAbs by the SCRs does not impair the binding to CD20. Increased in vitro lysis potency compared to their parental mAbs was corroborated by showing specific and dose dependent target cell elimination by CDC when compared to their parental mAbs. Lysis of CLL cells was not affected by the depletion of NK cells, suggesting that antibody-dependent cellular cytotoxicity plays a minor role in this context. Overall, this study emphasizes the crucial role of CDC in the elimination of CLL cells by mAbs and introduces a novel approach for enhancing CDC by directly fusing fH SCR1920 with mAbs.

CD38 is the major NAD+-hydrolyzing ecto-enzyme in most mammals. As a type II transmembrane protein, CD38 is also a promising target for the immunotherapy of multiple myeloma (MM). Nanobodies are single immunoglobulin variable domains from heavy chain antibodies that naturally occur in camelids. Using phage display technology, we isolated 13 mouse CD38-specific nanobodies from immunized llamas and produced these as recombinant chimeric mouse IgG2a heavy chain antibodies (hcAbs). Sequence analysis assigned these hcAbs to five distinct families that bind to three non-overlapping epitopes of CD38. Members of families 4 and 5 inhibit the GDPR-cyclase activity of CD38. Members of families 2, 4 and 5 effectively induce complement-dependent cytotoxicity against CD38-expressing tumor cell lines, while all families effectively induce antibody dependent cellular cytotoxicity. Our hcAbs present unique tools to assess cytotoxicity mechanisms of CD38-specific hcAbs in vivo against tumor cells and potential off-target effects on normal cells expressing CD38 in syngeneic mouse tumor models, i.e. in a fully immunocompetent background.

Haploidentical stem cell transplantation (haplo SCT) in Stage IV neuroblastoma relapsed patients has been proven efficacious, while immunotherapy utilizing the anti-GD2 antibody dinutuximab beta has become a standard treatment for neuroblastoma. The combinatorial therapy of haplo SCT and dinutuximab may potentiate the efficacy of the immunotherapy. To gain further understanding of the synergistic effects, functional immunomonitoring was assessed during the clinical trial CH14.18 1021 Antibody and IL2 After haplo SCT in Children with Relapsed Neuroblastoma (NCT02258815). Rapid immune reconstitution of the lymphoid compartment was confirmed, with clinically relevant dinutuximab serum levels found in all patients over the course of treatment. Only one patient developed human anti-chimeric antibodies (HACAs). In-patient monitoring revealed highly functional NK cell posttransplant capable of antibody-dependent cellular cytotoxicity (ADCC). Degranulation of NK cell subsets revealed a significant response increased by dinutuximab. This was irrespective of the KIR receptor–ligand constellation within the NK subsets, defined by the major KIR receptors CD158a, CD158b, and CD158e. Moreover, complement-dependent cytotoxicity (CDC) was shown to be an extremely potent effector-cell independent mechanism of tumor cell lysis, with a clear positive correlation to GD2 expression on the cancer cells as well as to the dinutuximab concentrations. The ex vivo testing of patient-derived effector cells and the sera collected during dinutuximab therapy demonstrated both high functionality of the newly established lymphoid immune compartment and provided confidence that the antibody dosing regimen was sufficient over the duration of the dinutuximab therapy (up to nine cycles in a 9-month period). During the course of the dinutuximab therapy, proinflammatory cytokines and markers (sIL2R, TNFa, IL6, and C reactive protein) were significantly elevated indicating a strong anti-GD2 immune response. No impact of FcGR polymorphism on event-free and overall survival was found. Collectively, this study has shown that in-patient functional immunomonitoring is feasible and valuable in contributing to the understanding of anti-cancer combinatorial treatments such as haplo SCT and antibody immunotherapy.

CD38 is the major NAD+-hydrolyzing ecto-enzyme in most mammals. As a type II transmembrane protein, CD38 is also a promising target for the immunotherapy of multiple myeloma (MM). Nanobodies are single immunoglobulin variable domains from heavy chain antibodies that naturally occur in camelids. Using phage display technology, we isolated 13 mouse CD38-specific nanobodies from immunized llamas and produced these as recombinant chimeric mouse IgG2a heavy chain antibodies (hcAbs). Sequence analysis assigned these hcAbs to five distinct families that bind to three non-overlapping epitopes of CD38. Members of families 4 and 5 inhibit the GDPR-cyclase activity of CD38. Members of families 2, 4 and 5 effectively induce complement-dependent cytotoxicity against CD38-expressing tumor cell lines, while all families effectively induce antibody dependent cellular cytotoxicity. Our hcAbs present unique tools to assess cytotoxicity mechanisms of CD38-specific hcAbs in vivo against tumor cells and potential off-target effects on normal cells expressing CD38 in syngeneic mouse tumor models, i.e. in a fully immunocompetent background. [ABSTRACT FROM AUTHOR]

Haploidentical stem cell transplantation (haplo SCT) in Stage IV neuroblastoma relapsed patients has been proven efficacious, while immunotherapy utilizing the anti-GD2 antibody dinutuximab beta has become a standard treatment for neuroblastoma. The combinatorial therapy of haplo SCT and dinutuximab may potentiate the efficacy of the immunotherapy. To gain further understanding of the synergistic effects, functional immunomonitoring was assessed during the clinical trial CH14.18 1021 Antibody and IL2 After haplo SCT in Children with Relapsed Neuroblastoma (NCT02258815). Rapid immune reconstitution of the lymphoid compartment was confirmed, with clinically relevant dinutuximab serum levels found in all patients over the course of treatment. Only one patient developed human anti-chimeric antibodies (HACAs). In-patient monitoring revealed highly functional NK cell posttransplant capable of antibody-dependent cellular cytotoxicity (ADCC). Degranulation of NK cell subsets revealed a significant response increased by dinutuximab. This was irrespective of the KIR receptor–ligand constellation within the NK subsets, defined by the major KIR receptors CD158a, CD158b, and CD158e. Moreover, complement-dependent cytotoxicity (CDC) was shown to be an extremely potent effector-cell independent mechanism of tumor cell lysis, with a clear positive correlation to GD2 expression on the cancer cells as well as to the dinutuximab concentrations. The ex vivo testing of patient-derived effector cells and the sera collected during dinutuximab therapy demonstrated both high functionality of the newly established lymphoid immune compartment and provided confidence that the antibody dosing regimen was sufficient over the duration of the dinutuximab therapy (up to nine cycles in a 9-month period). During the course of the dinutuximab therapy, proinflammatory cytokines and markers (sIL2R, TNFa, IL6, and C reactive protein) were significantly elevated indicating a strong anti-GD2 immune response. No impact of FcGR polymorphism on event-free and overall survival was found. Collectively, this study has shown that in-patient functional immunomonitoring is feasible and valuable in contributing to the understanding of anti-cancer combinatorial treatments such as haplo SCT and antibody immunotherapy. [ABSTRACT FROM AUTHOR]

The interactions of cancer cells with components of the complement system are highly complex, leading to an outcome that is either favorable or detrimental to cancer cells. Currently, we perceive only the “tip of the iceberg” of these interactions. In this review, we focus on the complement terminal C5b-9 complex, known also as the complement membrane attack complex (MAC) and discuss the complexity of its interaction with cancer cells, starting with a discussion of its proposed mode of action in mediating cell death, and continuing with a portrayal of the strategies of evasion exhibited by cancer cells, and closing with a proposal of treatment approaches targeted at evasion strategies. Upon intense complement activation and membrane insertion of sufficient C5b-9 complexes, the afflicted cells undergo regulated necrotic cell death with characteristic damage to intracellular organelles, including mitochondria, and perforation of the plasma membrane. Several pro-lytic factors have been proposed, including elevated intracellular calcium ion concentrations and activated JNK, Bid, RIPK1, RIPK3, and MLKL; however, further research is required to fully characterize the effective cell death signals activated by the C5b-9 complexes. Cancer cells over-express a multitude of protective measures which either block complement activation, thus reducing the number of membrane-inserted C5b-9 complexes, or facilitate the elimination of C5b-9 from the cell surface. Concomitantly, cancer cells activate several protective pathways that counteract the death signals. Blockage of complement activation is mediated by the complement membrane regulatory proteins CD46, CD55, and CD59 and by soluble complement regulators, by proteases that cleave complement proteins and by protein kinases, like CK2, which phosphorylate complement proteins. C5b-9 elimination and inhibition of cell death signals are mediated by caveolin and dynamin, by Hsp70 and Hsp90, by the mitochondrial stress protein mortalin, and by the protein kinases PKC and ERK. It is conceivable that various cancers and cancers at different stages of development will utilize distinct patterns of these and other MAC resistance strategies. In order to enhance the impact of antibody-based therapy on cancer, novel precise reagents that block the most effective protective strategies will have to be designed and applied as adjuvants to the therapeutic antibodies.

The interactions of cancer cells with components of the complement system are highly complex, leading to an outcome that is either favorable or detrimental to cancer cells. Currently, we perceive only the "tip of the iceberg" of these interactions. In this review, we focus on the complement terminal C5b-9 complex, known also as the complement membrane attack complex (MAC) and discuss the complexity of its interaction with cancer cells, starting with a discussion of its proposed mode of action in mediating cell death, and continuing with a portrayal of the strategies of evasion exhibited by cancer cells, and closing with a proposal of treatment approaches targeted at evasion strategies. Upon intense complement activation and membrane insertion of sufficient C5b-9 complexes, the afflicted cells undergo regulated necrotic cell death with characteristic damage to intracellular organelles, including mitochondria, and perforation of the plasma membrane. Several pro-lytic factors have been proposed, including elevated intracellular calcium ion concentrations and activated JNK, Bid, RIPK1, RIPK3, and MLKL; however, further research is required to fully characterize the effective cell death signals activated by the C5b-9 complexes. Cancer cells over-express a multitude of protective measures which either block complement activation, thus reducing the number of membrane-inserted C5b-9 complexes, or facilitate the elimination of C5b-9 from the cell surface. Concomitantly, cancer cells activate several protective pathways that counteract the death signals. Blockage of complement activation is mediated by the complement membrane regulatory proteins CD46, CD55, and CD59 and by soluble complement regulators, by proteases that cleave complement proteins and by protein kinases, like CK2, which phosphorylate complement proteins. C5b-9 elimination and inhibition of cell death signals are mediated by caveolin and dynamin, by Hsp70 and Hsp90, by the mitochondrial stress protein mortalin, and by the protein kinases PKC and ERK. It is conceivable that various cancers and cancers at different stages of development will utilize distinct patterns of these and other MAC resistance strategies. In order to enhance the impact of antibody-based therapy on cancer, novel precise reagents that block the most effective protective strategies will have to be designed and applied as adjuvants to the therapeutic antibodies. [ABSTRACT FROM AUTHOR]

CD38 is overexpressed by multiple myeloma cells and has emerged as a target for therapeutic antibodies. Nanobodies are soluble single domain antibody fragments derived from the VHH variable domain of heavy chain antibodies naturally occurring in camelids. We previously identified distinct llama nanobodies that recognize three non-overlapping epitopes of the extracellular domain of CD38. Here, we fused these VHH domains to the hinge, CH2, and CH3 domains of human IgG1, yielding highly soluble chimeric llama/human heavy chain antibodies (hcAbs). We analyzed the capacity of these hcAbs to mediate complement-dependent cytotoxicity (CDC) to CD38-expressing human multiple myeloma and Burkitt lymphoma cell lines. Combinations of two hcAbs that recognize distinct, non-overlapping epitopes of CD38 mediated potent CDC, in contrast to the hcAb monotherapy with only weak CDC capacity. Similarly, combining daratumumab with a hcAb that recognizes a non-overlapping epitope resulted in dramatically enhanced CDC. Further, introducing the E345R HexaBody mutation into the CH3 domain strongly enhanced the CDC potency of hcAbs to CD38-expressing cells. Exploiting their high solubility, we genetically fused two distinct nanobodies into heteromeric dimers via a flexible peptide linker and then fused these nanobody dimers to the hinge, CH2 and CH3 domains of human IgG1, yielding highly soluble, biparatopic hcAbs. These biparatopic hcAbs elicited CDC toward CD38-expressing myeloma cells more effectively than daratumumab. Our results underscore the advantage of nanobodies vs. pairs of VH and VL domains for constructing bispecific antibodies. Moreover, the CD38-specific biparatopic heavy chain antibodies described here represent potential new powerful therapeutics for treatment of multiple myeloma.

Maternal antibodies provide protection for the developing fetus. Transplacental transport of pathogenic autoantibodies might pose a risk for the developing fetus. The transport of antibodies across the placenta to the fetal circulation occurs through the neonatal Fc salvage receptor (FcRn). During gestation, maternal autoantibodies are able to penetrate the embryonic brain before a functional intact blood–brain barrier is established. Brain-reactive antibodies to the water channel protein aquaporin-4 (AQP4) are a hallmark finding in neuromyelitis optica (NMO), a neurological disease that predominantly affects women, many of whom are of childbearing age. AQP4–IgG mediate astrocytic injury in a complement-dependent fashion. Recent studies suggest these antibodies contribute to impaired pregnancy outcome. The aim of the study was to investigate the transplacental transport as well as FcRn binding of a monoclonal AQP4–IgG cloned from an NMO patient (wild-type antibody) compared to five different mutated Fc domain of this antibody containing single amino acid substitutions in the Fc region. All of the Fc-mutated antibodies lack complement-dependent cytotoxicity. Four of the five Fc-mutated antibodies showed limited transplacental transport in vivo. Three mutated Fc with impaired transplacental transport showed persistent binding to rodent FcRn at pH 6 but also at pH 7.2, suggesting that limited transplacental transport could be due to diminished release from FcRn. One mutated Fc with modestly limited transplacental transport showed diminished binding to FcRn at pH 6. This study suggests that mutated Fc with intact transplacental transport may be used to study antibody effector functions and Fc with limited transport may be used as a carrier to deliver therapies to pregnant woman, while sparing the developing fetus.

Astrocyte loss induced by neuromyelitis optica (NMO)-IgG and complement-dependent cytotoxicity (CDC) is the hallmark of NMO pathology. The survival of astrocytes is thought to reflect astrocyte exposure to environmental factors in the CNS and the response of astrocytes to these factors. However, still unclear are how astrocytes respond to NMO-IgG and CDC, and what CNS environmental factors may impact the survival of astrocytes. In a murine model of NMO induced by intracerebral injection of NMO-IgG and human complement, we found dramatic upregulation of IL-15 in astrocytes. To study the role of astrocytic IL-15 in NMO, we generated a transgenic mouse line with targeted expression of IL-15 in astrocytes (IL-15tg), in which the expression of IL-15 is controlled by a glial fibrillary acidic protein promoter. We showed that astrocyte-targeted expression of IL-15 attenuates astrocyte injury and the loss of aquaporin-4 in the brain. Reduced blood–brain barrier leakage and immune cell infiltration are also found in the lesion of IL-15tg mice subjected to NMO induction. IL-15tg astrocytes are less susceptible to NMO-IgG-mediated CDC than their wild-type counterparts. The enhanced resistance of IL-15tg astrocytes to cytotoxicity and cell death involves NF-κB signaling pathway. Our findings suggest that IL-15 reduces astrocyte loss and NMO pathology.

NEO-201 is a novel humanized IgG1 monoclonal antibody that was derived from an immunogenic preparation of tumor-associated antigens from pooled allogeneic colon tumor tissue extracts. It was found to react against a variety of cultured human carcinoma cell lines and was highly reactive against the majority of tumor tissues from many different carcinomas, including colon, pancreatic, stomach, lung, and breast cancers. NEO-201 also exhibited tumor specificity, as the majority of normal tissues were not recognized by this antibody. Functional assays revealed that treatment with NEO-201 is capable of mediating both antibody-dependent cellular cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC) against tumor cells. Furthermore, the growth of human pancreatic xenograft tumors in vivo was largely attenuated by treatment with NEO-201 both alone and in combination with human peripheral blood mononuclear cells as an effector cell source for ADCC. In vivo biodistribution studies in human tumor xenograft-bearing mice revealed that NEO-201 preferentially accumulates in the tumor but not organ tissue. Finally, a single-dose toxicity study in non-human primates demonstrated safety and tolerability of NEO-201, as a transient decrease in circulating neutrophils was the only related adverse effect observed. These findings indicate that NEO-201 warrants clinical testing as both a novel diagnostic and therapeutic agent for the treatment of a broad variety of carcinomas.

CD38 is overexpressed by multiple myeloma cells and has emerged as a target for therapeutic antibodies. Nanobodies are soluble single domain antibody fragments derived from the VHH variable domain of heavy chain antibodies naturally occurring in camelids. We previously identified distinct llama nanobodies that recognize three non-overlapping epitopes of the extracellular domain of CD38. Here, we fused these VHH domains to the hinge, CH2, and CH3 domains of human IgG1, yielding highly soluble chimeric llama/human heavy chain antibodies (hcAbs). We analyzed the capacity of these hcAbs to mediate complement-dependent cytotoxicity (CDC) to CD38-expressing human multiple myeloma and Burkitt lymphoma cell lines. Combinations of two hcAbs that recognize distinct, non-overlapping epitopes of CD38 mediated potent CDC, in contrast to the hcAb monotherapy with only weak CDC capacity. Similarly, combining daratumumab with a hcAb that recognizes a non-overlapping epitope resulted in dramatically enhanced CDC. Further, introducing the E345R HexaBody mutation into the CH3 domain strongly enhanced the CDC potency of hcAbs to CD38-expressing cells. Exploiting their high solubility, we genetically fused two distinct nanobodies into heteromeric dimers via a flexible peptide linker and then fused these nanobody dimers to the hinge, CH2 and CH3 domains of human IgG1, yielding highly soluble, biparatopic hcAbs. These biparatopic hcAbs elicited CDC toward CD38-expressing myeloma cells more effectively than daratumumab. Our results underscore the advantage of nanobodies vs. pairs of VH and VL domains for constructing bispecific antibodies. Moreover, the CD38-specific biparatopic heavy chain antibodies described here represent potential new powerful therapeutics for treatment of multiple myeloma. [ABSTRACT FROM AUTHOR]

NEO-201 is a novel humanized IgG1 monoclonal antibody that was derived from an immunogenic preparation of tumor-associated antigens from pooled allogeneic colon tumor tissue extracts. It was found to react against a variety of cultured human carcinoma cell lines and was highly reactive against the majority of tumor tissues from many different carcinomas, including colon, pancreatic, stomach, lung, and breast cancers. NEO-201 also exhibited tumor specificity, as the majority of normal tissues were not recognized by this antibody. Functional assays revealed that treatment with NEO-201 is capable of mediating both antibody-dependent cellular cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC) against tumor cells. Furthermore, the growth of human pancreatic xenograft tumors in vivo was largely attenuated by treatment with NEO-201 both alone and in combination with human peripheral blood mononuclear cells as an effector cell source for ADCC. In vivo biodistribution studies in human tumor xenograft-bearing mice revealed that NEO-201 preferentially accumulates in the tumor but not organ tissue. Finally, a single-dose toxicity study in non-human primates demonstrated safety and tolerability of NEO-201, as a transient decrease in circulating neutrophils was the only related adverse effect observed. These findings indicate that NEO-201 warrants clinical testing as both a novel diagnostic and therapeutic agent for the treatment of a broad variety of carcinomas. [ABSTRACT FROM AUTHOR]

Maternal antibodies provide protection for the developing fetus. Transplacental transport of pathogenic autoantibodies might pose a risk for the developing fetus. The transport of antibodies across the placenta to the fetal circulation occurs through the neonatal Fc salvage receptor (FcRn). During gestation, maternal autoantibodies are able to penetrate the embryonic brain before a functional intact blood-brain barrier is established. Brain-reactive antibodies to the water channel protein aquaporin-4 (AQP4) are a hallmark finding in neuromyelitis optica (NMO), a neurological disease that predominantly affects women, many of whom are of childbearing age. AQP4-IgG mediate astrocytic injury in a complement-dependent fashion. Recent studies suggest these antibodies contribute to impaired pregnancy outcome. The aim of the study was to investigate the transplacental transport as well as FcRn binding of a monoclonal AQP4-IgG cloned from an NMO patient (wild-type antibody) compared to five different mutated Fc domain of this antibody containing single amino acid substitutions in the Fc region. All of the Fc-mutated antibodies lack complement-dependent cytotoxicity. Four of the five Fc-mutated antibodies showed limited transplacental transport in vivo . Three mutated Fc with impaired transplacental transport showed persistent binding to rodent FcRn at pH 6 but also at pH 7.2, suggesting that limited transplacental transport could be due to diminished release from FcRn. One mutated Fc with modestly limited transplacental transport showed diminished binding to FcRn at pH 6. This study suggests that mutated Fc with intact transplacental transport may be used to study antibody effector functions and Fc with limited transport may be used as a carrier to deliver therapies to pregnant woman, while sparing the developing fetus.

Astrocyte loss induced by neuromyelitis optica (NMO)-IgG and complement-dependent cytotoxicity (CDC) is the hallmark of NMO pathology. The survival of astrocytes is thought to reflect astrocyte exposure to environmental factors in the CNS and the response of astrocytes to these factors. However, still unclear are how astrocytes respond to NMO-IgG and CDC, and what CNS environmental factors may impact the survival of astrocytes. In a murine model of NMO induced by intracerebral injection of NMO-IgG and human complement, we found dramatic upregulation of IL-15 in astrocytes. To study the role of astrocytic IL-15 in NMO, we generated a transgenic mouse line with targeted expression of IL-15 in astrocytes (IL-15 tg ), in which the expression of IL-15 is controlled by a glial fibrillary acidic protein promoter. We showed that astrocyte-targeted expression of IL-15 attenuates astrocyte injury and the loss of aquaporin-4 in the brain. Reduced blood-brain barrier leakage and immune cell infiltration are also found in the lesion of IL-15 tg mice subjected to NMO induction. IL-15 tg astrocytes are less susceptible to NMO-IgG-mediated CDC than their wild-type counterparts. The enhanced resistance of IL-15 tg astrocytes to cytotoxicity and cell death involves NF-κB signaling pathway. Our findings suggest that IL-15 reduces astrocyte loss and NMO pathology.

The classical complement pathway is initiated by the large (~800 kDa) and flexible multimeric C1 complex. Its catalytic function is triggered by the proteases hetero-tetramer C1r2s2, which is associated to the C1q sensing unit, a complex assembly of 18 chains built as a hexamer of heterotrimers. Initial pioneering studies gained insights into the main architectural principles of the C1 complex. A dissection strategy then provided the high-resolution structures of its main functional and/or structural building blocks, as well as structural details on some key protein-protein interactions. These past and current discoveries will be briefly summed up in order to address the question of what is still ill-defined. On a functional point of view, the main molecular determinants of C1 activation and its tight control will be delineated. The current perspective remains to decipher how C1 really works and is controlled in vivo, both in normal and pathological settings.