SPBC1604.17c Antibody

What is IL-17C and what biological functions does it serve?

IL-17C is a member of the IL-17 superfamily that primarily functions as a cytokine playing crucial roles in innate immunity of the epithelium. Unlike other members of the IL-17 family that are primarily secreted by T cells, IL-17C is produced by a wide spectrum of cells including macrophages. It stimulates the production of antibacterial peptides and pro-inflammatory molecules for host defense by signaling through the NF-kappa-B and MAPK pathways. IL-17C acts synergistically with IL-22 in inducing the expression of antibacterial peptides, including S100A8, S100A9, REG3A, and REG3G. Similar synergy is observed with TNF and IL1B in inducing DEFB2 from keratinocytes . The cytokine can exhibit both protective and pathogenic properties depending on the type of insult, either by maintaining epithelial homeostasis following inflammatory challenge or by promoting inflammatory phenotypes .

How does IL-17C differ from other members of the IL-17 family?

IL-17C differs from other IL-17 family members primarily in its cellular origin and target cells. While most IL-17 family cytokines are produced by T cells, IL-17C is secreted by a wider spectrum of cells including macrophages, rather than T cells . Research has demonstrated that IL-17C is expressed in CD11b+ MHC class II macrophages, CD11c+MHC class II dendritic cells, and various tissues under inflammatory conditions . Furthermore, normal human tissues barely express IL-17C under physiological conditions, but its expression becomes significant in inflammatory conditions characterized by macrophage infiltration . This distinctive expression pattern suggests specialized functions in tissue-specific inflammatory responses compared to other IL-17 family members.

What are the primary cellular sources of IL-17C in experimental systems?

The primary cellular sources of IL-17C in experimental systems include:

• Macrophages (CD68+ cells) - In aseptic loosening samples, approximately 55.32% ± 10.11% of CD68+ macrophages were found to colocalize with IL-17C

• Synovial fluid mononuclear cells in rheumatoid arthritis patients

• Human keratinocytes, particularly when stimulated with TNF-alpha

• CD11b+ MHC class II+ cells and CD11c+ MHC class II+ cells

• MC3T3-E1 cells and cartilage cells

• CD3+CD4+ cells and CD3+CD19+ cells

These diverse cellular sources make IL-17C distinct from other IL-17 family members and suggest complex roles in different physiological and pathological contexts.

What are the recommended methods for detecting IL-17C expression in tissue samples?

For detecting IL-17C expression in tissue samples, immunofluorescence analysis has been demonstrated to be an effective approach. A validated protocol includes:

• Preparation of paraffin sections (4-μm thick), followed by deparaffinization and rehydration using routine methods

• Antigen retrieval performed by maintaining sections at a moderate boil in citric acid buffer solution (pH 6.0)

• Blocking non-specific binding sites with 10% normal rabbit serum for 40 minutes

• Application of primary antibodies (e.g., goat anti-human IL-17C at 20 μg/ml) with overnight incubation at 4°C

• Washing five times with PBS (5 minutes each)

• Application of appropriate secondary antibodies (e.g., rabbit anti-goat CY3-labeled)

• Counter-staining with DAPI

For co-localization studies to identify cellular sources, double immunofluorescence staining with cell-specific markers (e.g., CD68 for macrophages) can be employed. Semi-quantitative analysis of immunofluorescence can be conducted by measuring mean optical density of staining .

How should researchers design experiments to assess IL-17C's role in inflammatory conditions?

When designing experiments to assess IL-17C's role in inflammatory conditions, researchers should consider:

• Model selection: Choose appropriate disease models where IL-17C may play a role. Studies have successfully used:

  • Rheumatoid arthritis tissue samples as positive controls for IL-17C expression

  • Aseptic loosening models to study IL-17C in non-infectious inflammation

  • Atopic dermatitis models for skin inflammation

• Experimental approaches:

  • Immunofluorescence or immunohistochemistry to detect and localize IL-17C in tissues

  • Co-localization studies with cell-specific markers to identify cellular sources

  • Functional studies using anti-IL-17C antibodies to block IL-17C signaling

  • Comparison between diseased tissues and appropriate controls

• Controls and validation:

  • Include both positive controls (e.g., rheumatoid arthritis tissues) and negative controls

  • Validate findings through multiple methodological approaches

  • Consider temporal aspects of IL-17C expression in disease progression

What considerations are important when designing antibody-based therapies targeting IL-17C?

When designing antibody-based therapies targeting IL-17C, researchers should consider:

• Target validation: Confirm the role of IL-17C in the specific disease pathology through preclinical models and human tissue studies

• Antibody characteristics:

  • Specificity: Ensure antibodies specifically target IL-17C without cross-reactivity with other IL-17 family members

  • Affinity: Develop antibodies with optimal binding affinity for IL-17C

  • Format: Consider different antibody formats (e.g., monoclonal, recombinant) based on application

• Administration routes:

  • Intravenous (i.v.) administration: Studies have used doses between 1-10 mg/kg administered every 2 or 4 weeks

  • Subcutaneous (s.c.) administration: Single doses or multiple doses (e.g., 320 mg every 2 weeks)

• Pharmacokinetic considerations:

  • Bioavailability: Subcutaneous dosing has shown approximately 55% bioavailability

  • Steady-state levels: These are typically achieved within 2-4 weeks of treatment initiation

• Safety monitoring:

  • Design studies to carefully assess safety and tolerability profiles

  • Monitor for adverse events consistent with monoclonal antibody therapies

How should researchers interpret conflicting results regarding IL-17C efficacy in different disease models?

When faced with conflicting results regarding IL-17C efficacy across different disease models, researchers should:

• Contextual analysis:

  • Consider disease-specific contexts: IL-17C may have both protective and pathogenic properties depending on the type of insult

  • Examine temporal aspects: The role of IL-17C may vary during different phases of disease progression

• Methodological considerations:

  • Analyze differences in experimental methodologies that could explain discrepancies

  • Assess variations in antibody specificity, dosing regimens, and routes of administration

  • Review the timing of interventions relative to disease stage

• Translational gaps:

  • Consider differences between preclinical models and clinical outcomes

  • For example, MOR106 (anti-IL-17C antibody) showed promise in preclinical studies but demonstrated ineffectiveness for atopic dermatitis in clinical trials despite favorable safety and pharmacokinetic profiles

• Statistical power and study design:

  • Evaluate sample sizes and statistical methods used across studies

  • Consider whether differences in patient/sample selection criteria might explain conflicting outcomes

• Mechanistic investigations:

  • Conduct additional mechanistic studies to understand why IL-17C targeting may succeed in some contexts but fail in others

  • Consider potential compensatory mechanisms that might limit efficacy in certain disease states

What are the common technical challenges in IL-17C antibody production and validation?

Common technical challenges in IL-17C antibody production and validation include:

• Specificity issues:

  • Ensuring antibodies specifically recognize IL-17C without cross-reactivity with other IL-17 family members

  • Validating specificity through multiple approaches (Western blot, ELISA, immunoprecipitation)

• Sensitivity challenges:

  • Detecting IL-17C in tissues with low expression levels

  • Optimizing signal-to-noise ratio in detection methods

• Validation across applications:

  • Ensuring antibodies work consistently across different applications (immunohistochemistry, flow cytometry, functional blocking)

  • Validating antibodies for use in different species when conducting translational research

• Reproducibility concerns:

  • Batch-to-batch variation in antibody production

  • Standardizing validation protocols across laboratories

• Functional validation:

  • Confirming that anti-IL-17C antibodies can effectively neutralize IL-17C biological activity

  • Developing appropriate functional assays to test antibody efficacy

How do researchers determine the optimal dosing regimen for anti-IL-17C antibodies in experimental studies?

To determine optimal dosing regimens for anti-IL-17C antibodies in experimental studies, researchers should:

• Conduct dose-response studies:

  • Test multiple dose levels to establish dose-response relationships

  • Clinical studies have evaluated intravenous doses between 1-10 mg/kg and subcutaneous doses of 320 mg

• Consider administration frequency:

  • Evaluate different dosing intervals (e.g., every 2 weeks versus every 4 weeks)

  • Base intervals on pharmacokinetic data and half-life of the antibody

• Monitor pharmacokinetic parameters:

  • Measure drug concentrations over time to determine:

    • Bioavailability (approximately 55% for subcutaneous dosing)

    • Time to steady-state (typically 2-4 weeks)

    • Clearance rates and half-life

• Assess target engagement:

  • Determine whether the antibody effectively binds to IL-17C at the chosen dose

  • Measure downstream effects on IL-17C signaling pathways

• Balance efficacy and safety:

  • Identify the minimum effective dose to minimize potential adverse effects

  • Monitor for dose-dependent adverse events or toxicity

How does IL-17C signaling interact with other inflammatory pathways in complex disease models?

IL-17C signaling demonstrates complex interactions with multiple inflammatory pathways:

• Synergistic effects with other cytokines:

  • IL-17C acts synergistically with IL-22 to induce expression of antibacterial peptides (S100A8, S100A9, REG3A, REG3G)

  • Synergy with TNF and IL-1β enhances DEFB2 expression from keratinocytes

  • These synergistic effects amplify inflammatory responses beyond what each cytokine induces alone

• NF-κB and MAPK pathway activation:

  • IL-17C stimulates proinflammatory signaling through NF-κB and MAPK pathways

  • This activation leads to production of antibacterial peptides and inflammatory mediators

• Autocrine feedback mechanisms:

  • Evidence suggests IL-17C can establish positive feedback loops in cells like macrophages

  • This autocrine signaling may contribute to sustained inflammatory responses

• Dual roles in tissue homeostasis:

  • IL-17C can either maintain epithelial homeostasis after inflammatory challenge

  • Or promote inflammatory phenotypes depending on context

  • This dual role complicates therapeutic targeting

• Interplay with innate immune mechanisms:

  • Enhanced IL-17C/IL-17RE signaling may increase susceptibility to autoimmune diseases

  • This suggests cross-talk between IL-17C pathways and mechanisms of self-tolerance

What are the emerging hypotheses regarding IL-17C's role in non-canonical disease processes?

Emerging hypotheses regarding IL-17C's role in non-canonical disease processes include:

• Aseptic loosening of implants:

  • IL-17C has been detected in tissues surrounding aseptically loosened joint implants

  • It potentially contributes to inflammation and osteolysis through:

    • Enhanced expression of TNF-alpha and IL-1beta

    • Direct effects on osteolysis

    • Association with vascular responses in the periprosthetic membrane

• Epithelial homeostasis regulation:

  • IL-17C may form a positive feedback loop with epithelial cells to regulate innate immune function

  • This mechanism might represent a homeostatic function distinct from its proinflammatory role

• Macrophage-driven inflammation:

  • Evidence suggests IL-17C is expressed by CD68+ macrophages in aseptic loosening samples

  • This indicates IL-17C may contribute to macrophage-driven inflammatory processes in conditions beyond classical autoimmune diseases

• Autocrine inflammatory regulation:

  • Given that macrophages can both produce and respond to IL-17C, researchers hypothesize that IL-17C might regulate inflammation through autocrine mechanisms

  • This could represent a novel self-regulatory circuit in inflammation

What methodological approaches can determine the therapeutic potential of IL-17C neutralization in novel disease targets?

To determine the therapeutic potential of IL-17C neutralization in novel disease targets, researchers should employ:

• Comprehensive tissue expression profiling:

  • Analyze IL-17C expression in diseased versus healthy tissues

  • Use immunofluorescence, RNA-sequencing, or protein assays to quantify expression levels

  • Identify cellular sources through co-localization studies (e.g., with CD68 for macrophages)

• Mechanistic studies in relevant models:

  • Develop appropriate disease models that recapitulate key pathological features

  • Test IL-17C neutralization at different disease stages

  • Assess impact on disease-specific endpoints and inflammatory markers

• Translational approach from preclinical to clinical studies:

  • Follow a structured development path similar to that used for MOR106:

    • Phase 1 studies in healthy volunteers to establish safety

    • Phase 2 studies in patient populations to assess efficacy

    • Comprehensive pharmacokinetic profiling

• Pharmacodynamic biomarker development:

  • Identify and validate biomarkers that respond to IL-17C neutralization

  • Use these to monitor target engagement in clinical studies

  • Correlate biomarker changes with clinical outcomes

• Futility analysis framework:

  • Implement interim analyses to assess probability of achieving primary endpoints

  • This approach can prevent unnecessary continuation of studies unlikely to demonstrate efficacy

  • Example: The MOR106 studies were terminated after futility analysis indicated low probability of achieving primary efficacy endpoints in atopic dermatitis