Anti-Human IL-6 (Siltuximab) [Clone CNTO-328] — Fc Muted™

Anti-Human IL-6 (Siltuximab) [Clone CNTO-328] — Fc Muted™

Product No.: I-455

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Product No.I-455
Clone
CNTO-328
Target
IL-6
Product Type
Biosimilar Recombinant Human Monoclonal Antibody
Alternate Names
BSF-2, CDF, Hybridoma growth factor (HPGF), IFN-beta-2, HSF
Isotype
Human IgG1κ
Applications
B
,
ELISA
,
FA
,
IF
,
RIA

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Select Product Size
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Antibody Details

Product Details

Reactive Species
Human
Expression Host
HEK-293 Cells
FC Effector Activity
Muted
Product Concentration
≥ 5.0 mg/ml
Endotoxin Level
< 1.0 EU/mg as determined by the LAL method
Purity
≥95% by SDS Page
≥95% monomer by analytical SEC
Formulation
This biosimilar antibody is aseptically packaged and formulated in 0.01 M phosphate buffered saline (150 mM NaCl) PBS pH 7.2 - 7.4 with no carrier protein, potassium, calcium or preservatives added. Due to inherent biochemical properties of antibodies, certain products may be prone to precipitation over time. Precipitation may be removed by aseptic centrifugation and/or filtration.
State of Matter
Liquid
Product Preparation
Recombinant biosimilar antibodies are manufactured in an animal free facility using only in vitro protein free cell culture techniques and are purified by a multi-step process including the use of protein A or G to assure extremely low levels of endotoxins, leachable protein A or aggregates.
Pathogen Testing
To protect mouse colonies from infection by pathogens and to assure that experimental preclinical data is not affected by such pathogens, all of Leinco’s recombinant biosimilar antibodies are tested and guaranteed to be negative for all pathogens in the IDEXX IMPACT I Mouse Profile.
Storage and Handling
Functional grade preclinical antibodies may be stored sterile as received at 2-8°C for up to one month. For longer term storage, aseptically aliquot in working volumes without diluting and store at ≤ -70°C. Avoid Repeated Freeze Thaw Cycles.
Regulatory Status
Research Use Only
Country of Origin
USA
Shipping
2 – 8° C Wet Ice
Additional Applications Reported In Literature ?
FA,
ELISA,
B,
RIA,
IF
Each investigator should determine their own optimal working dilution for specific applications. See directions on lot specific datasheets, as information may periodically change.

Description

Description

Specificity
This non-therapeutic biosimilar antibody uses the same variable region sequence as the therapeutic antibody Siltuximab. CNTO-328 (Siltuximab) is a neutralizing monoclonal antibody specific against human IL-6.
Background
IL-6 is a pleiotropic 26 kD protein that can act as both a pro-inflammatory cytokine and an anti-inflammatory myokine, a form of cytokine produced in muscle cells that participates in tissue regeneration and repair, maintenance of healthy bodily functioning, and homeostasis within the immune system 1. IL-6 also plays a part in the endocrine, nervous, and hematopoietic systems, bone metabolism, regulation of blood pressure, and inflammation. Furthermore, IL-6 is an important mediator of fever and of the acute phase response which is the body's rapid attempt to restore homeostasis after tissue injury, infection, neoplastic growth, or immunological disturbance. In its role as an anti-inflammatory myokine, IL-6 precedes the appearance of other cytokines in the circulation, is notably elevated with exercise, and is mediated by both its inhibitory effects on TNF-α and IL-1, and activation of IL-1R⍺ and IL-10. IL-6 signals through a cell-surface type I cytokine receptor complex formed by the binding of IL-6 to IL-6R, which in turn combines with GP130 to transduce extracellular signaling via STAT3 activation. Hence, it is thought that blocking the interaction between IL-6 and GP130 may have therapeutic potential via the inhibition of the IL-6/GP130/STAT3 signaling pathway. Moreover, IL-6 initiates inflammatory and auto-immune processes in many diseases, including diabetes, atherosclerosis, depression, Alzheimer's disease, rheumatoid arthritis, and cancer. For example, multicentric Castleman’s disease is a rare lymphoproliferative disorder caused by dysregulation of IL-6 2. Thus, there is an interest in the therapeutic potential of anti-IL-6 mAbs.

CNTO-328 (Siltuximab) is a chimeric monoclonal antibody that was developed for the treatment of IL-6 related disorders 2,3,4,5. Siltuximab is associated with sustained reductions in IL-6 levels along with various other cytokines and markers 2. In vitro studies in ovarian cancer cells show that siltuximab inhibits IL-6 induced STAT3 activation, nuclear translocation, and downstream gene expression 6. Siltuximab also induces apoptosis 2,7 and reduces C-reactive protein levels 2.

Siltuximab has been approved for the treatment of multicentric Castleman’s disease in HIV-negative patients 1. Siltuximab does not bind to virally produced IL-6 (vIL-6).
Antigen Distribution
IL-6 is a pleiotropic cytokine produced by B lymphocytes, T lymphocytes, macrophages, microglia, fibroblasts, keratinocytes, mesangial cells, vascular endothelial cells, mast cells, and dendritic cells. Additionally, osteoblasts secrete IL-6 to stimulate osteoclast formation. Smooth muscle cells in the tunica media of many blood vessels produce IL-6 as a pro-inflammatory cytokine. IL-6 is also released into circulation in response to various stimuli including PAMPs (pathogen-associated molecular patterns) and cortisol, a hormone produced by the human body under psychologically stressful conditions.
Ligand/Receptor
IL6R
NCBI Gene Bank ID
UniProt.org
Research Area
Biosimilars
.
Cell Biology
.
Immunology
.
Inflammatory Disease
.
Innate Immunity
.
Neuroscience
.
Autoimmunity
.
Pro-Inflammatory Cytokines

Leinco Antibody Advisor

Powered by AI: AI is experimental and still learning how to provide the best assistance. It may occasionally generate incorrect or incomplete responses. Please do not rely solely on its recommendations when making purchasing decisions or designing experiments.

Research-grade Siltuximab biosimilars are typically used as calibration standards or reference controls in a pharmacokinetic (PK) bridging ELISA by serving as the analytical standard for establishing the assay’s standard curve and for quantifying both the biosimilar and reference products in serum samples.

Key use and justification:

  • Single Analytical Standard Approach: The consensus best practice is to use a single PK assay that employs a single analytical standard—usually the biosimilar itself—for quantitative measurement of both the biosimilar and the reference product within test samples. This limits variability, increases assay robustness, and removes the need for separate reference curves for each drug source.

  • Assay Calibration and Quantification: The research-grade Siltuximab biosimilar is serially diluted to prepare calibrators (standard curve points) covering the expected concentration range in human serum. During method validation, quality control (QC) samples made with both the biosimilar and the reference product are also quantified against this biosimilar-based calibration curve. This ensures that the assay is precise and accurate for both forms of the drug.

  • PK Bridging Principle: In a PK bridging ELISA, pooling calibration and control across biosimilar and originator products ensures that concentration differences measured are due to PK differences, not analytical variance. The biosimilar standard must show comparable binding and detection characteristics as the reference molecule in the assay’s ligand binding format, which is demonstrated during assay validation.

  • Validation: The validation includes parallelism and recovery experiments in which biosimilar and reference material are spiked into serum and measured across a range of concentrations. Analytical equivalence is confirmed if the biosimilar and reference sample quantitation results fall within a predefined acceptance window (e.g., a 90% confidence interval within 0.8–1.25 ratio).

  • Implementation: For example, in a validated assay, serial dilutions of the Siltuximab biosimilar prepare standards (e.g., 50–12,800 ng/mL), which are run on each ELISA plate alongside unknowns (study serum samples) and QC samples (biosimilar and reference spiked serum at low, mid, high concentrations). The unknowns’ concentrations are interpolated from the biosimilar standard curve.

Summary Table: Use of Siltuximab Biosimilar in PK Bridging ELISA

PurposeApplication
Calibration StandardSerial dilutions establish standard curve for quantification
Reference ControlSpiked QC samples (both biosimilar & reference) monitor assay precision and accuracy
Analytical EquivalenceValidated to ensure single standard curve quantifies both biosimilar & originator accurately

Important context:

  • This strategy aligns with industry/regulatory expectations for biosimilar PK assessment, as it ensures that any measured differences between biosimilar and originator are biological, not analytical.
  • The biosimilar standard’s suitability is rigorously validated so the PK bridging ELISA is considered reliable for comparative PK studies in serum.

If you need guidance on practical ELISA setup (e.g., plate layout, dilution scheme), that can be provided as well.

The primary models in which a research-grade anti-IL-6 antibody is administered in vivo to study tumor growth inhibition and characterize tumor-infiltrating lymphocytes (TILs) are:

  • Human tumor xenograft models in immunodeficient mice
  • Syngeneic mouse tumor models with a fully functional immune system

Essential context and supporting details:

  • Human Tumor Xenografts (in Immunodeficient Mice):

    • These models transplant human tumor cells (often cancer stem cells) into mice lacking a functional adaptive immune system (such as nude or NSG mice). Antibodies like siltuximab (anti-human IL-6) and tocilizumab (humanized anti-IL-6R) are used to block human IL-6 signaling, assessing the resulting effects on tumor growth dynamics.
    • The xenograft system enables administration and evaluation of human-compatible therapeutic antibodies and allows researchers to dissect the impact of tumor-derived IL-6 on cancer cell survival and proliferation.
    • While TILs from the human tumor can sometimes be partially characterized, the utility for immune profiling is limited due to the absence or dysfunction of murine immune cells.
  • Syngeneic Mouse Tumor Models:

    • These models use murine tumor cell lines (such as MC38, CT26, Hepa1-6, and EMT-6) implanted into genetically identical (immunocompetent) mice, preserving a fully functional mouse immune system.
    • Research-grade anti-mouse IL-6 antibodies are used to block the murine IL-6 pathway, enabling assessment of tumor growth inhibition and robust characterization of TIL populations and immunological changes within the tumor microenvironment.
    • Syngeneic models are optimal for studying immune checkpoint inhibition, the effect of IL-6 on immune cell infiltration (such as T cells and macrophages), and the broader impact of immunotherapies on tumor immunity.

Recommended usage for TIL characterization:

  • Syngeneic models are preferred when the goal is detailed analysis of immune cell infiltration (TILs) because the mouse immune system remains intact and responsive.
  • Humanized or xenograft models are necessary when the objective is to evaluate therapeutics that specifically target human IL-6 or to investigate human tumor cell biology. However, immune characterization is limited unless mice are engineered to carry human immune components.

Examples of common models:

  • Syngeneic murine tumor models: MC38, CT26, Hepa1-6, EMT-6.
  • Human xenograft models: Human pancreatic, lung, and head and neck squamous cell carcinoma (HNSCC) cell lines or cancer stem cell populations transplanted into immunodeficient mice.

Model selection depends on:

  • The antibody species specificity (human vs mouse IL-6 target)
  • The aspect of tumor biology or immune function to be characterized (tumor growth inhibition vs immune cell profiling).

In summary, research into anti-IL-6 therapy and its impact on tumor growth and TILs is conducted primarily using either xenograft models (for direct effects on human tumor cells) or syngeneic models (for studying immune responses and TIL dynamics in a functional immune environment).

Researchers use the Siltuximab biosimilar, an anti-IL-6 monoclonal antibody, in combination with other checkpoint inhibitors like anti-CTLA-4 or anti-LAG-3 biosimilars to study synergistic effects in immune-oncology models by targeting distinct but complementary immune pathways.

Context and Supporting Details:

  • Siltuximab biosimilar targets IL-6, a cytokine implicated in cancer progression, immune suppression, and inflammation, making it a valuable tool in immune modulation for research purposes. Its biosimilar versions offer cost-effective and reliable experimental options for non-clinical studies.

  • Checkpoint inhibitors such as anti-CTLA-4 and anti-LAG-3 block inhibitory immune signals and enhance T cell responses. CTLA-4 blockade primarily restores T cell activation in the lymph nodes, while LAG-3 inhibition works alongside PD-1/PD-L1 pathways to synergistically enhance anti-tumor immunity.

  • In complex immune-oncology models, combining Siltuximab biosimilar with checkpoint inhibitors enables the study of how simultaneous IL-6 inhibition and checkpoint blockade can amplify therapeutic efficacy and overcome immune resistance. For instance:

    • In preclinical and translational research, Siltuximab biosimilar can be added to cultures or animal models along with anti-CTLA-4 or anti-LAG-3 agents to observe changes in immune cell infiltration, cytokine profiles, tumor growth, and response rates.
    • Researchers analyze whether IL-6 blockade reduces immunosuppressive tumor microenvironment features, facilitating checkpoint inhibitor activity.
    • Such models help reveal additive or synergistic anti-tumor responses, mechanisms of resistance, and toxicity profiles when disparate immune pathways are inhibited simultaneously.

Additional Relevant Information:

  • Using biosimilars like Siltuximab for research allows broad exploration of combinatorial immunotherapy strategies before clinical translation, supporting mechanistic studies and dosing optimization in complex tumor models.
  • While direct combinatorial data on Siltuximab biosimilar with anti-CTLA-4 or anti-LAG-3 biosimilars is currently limited, the scientific rationale—and parallel findings with other checkpoint/tumor-targeted drug combinations—strongly suggests this approach is intended to maximize immune response diversity and overcome therapy resistance.
  • Researchers typically assess endpoints like tumor regression, progression-free survival, immune activation signatures, and adverse event profiles in preclinical synergy studies.

Summary Table: Combination Rationale and Mechanisms

AgentPathway TargetedPrimary MechanismExpected Synergistic Effect
Siltuximab biosimilarIL-6Reduces inflammation, immune suppressionEnhances efficacy of checkpoint inhibitors by modifying tumor microenvironment
Anti-CTLA-4 biosimilarCTLA-4Promotes T cell activationFurther amplifies immune response at lymph nodes
Anti-LAG-3 biosimilarLAG-3Relieves exhaustion in effector T cellsIncreases cytotoxic T cell activity within tumors

The synergy is tested experimentally in complex tumor immune models to determine the best combinations for future clinical trials.

A Siltuximab biosimilar can be used as both the capture and detection reagent in a bridging ADA ELISA to monitor anti-drug antibodies (ADAs) in patients receiving siltuximab therapy. This method leverages the structural similarity between biosimilar and reference drugs to detect immune responses against the therapeutic.

How it works in bridging ADA ELISA:

  • Capture Reagent: The Siltuximab biosimilar (or siltuximab itself) is immobilized on an ELISA plate, either directly or via biotin-streptavidin interaction. Patient serum containing potential ADAs is added, allowing any anti-siltuximab antibodies present to bind the drug.

  • Detection Reagent: After washing, a labeled form of the Siltuximab biosimilar (commonly HRP-conjugated or biotinylated) is added. This reagent will bind to any ADA already bound to the plate-bound drug, forming a “bridge” through the bivalent nature of antibodies—the ADA binds both the immobilized and the detection drug molecule.

  • Signal Generation: A substrate is added to detect the labeled drug bound via ADA bridging, producing a measurable signal proportional to ADA content in the sample.

Key Points:

  • The biosimilar performs equivalently to the reference drug in bridging ELISA, as it shares the same binding epitopes required for ADA detection.
  • This format is highly sensitive, suitable for detecting low levels of ADAs against siltuximab in patient samples.
  • Both capture and detection reagents must be of high quality and preferably have minimal aggregation or non-specific binding for assay specificity.
  • Use of a biosimilar instead of the original reference drug (siltuximab) is accepted practice, provided characterization and validation show comparable binding behavior.

Limitations:

  • The presence of circulating siltuximab in patient samples may compete with the capture/detection reagents, potentially interfering with ADA detection (“drug interference”).
  • Bridging ELISA is best suited for detecting bivalent antibodies (mainly IgG) and can miss monovalent or low-affinity ADAs.

In summary, a Siltuximab biosimilar in a bridging ADA ELISA acts both as the bait (capture) and the probe (detection), enabling sensitive monitoring of immune responses against the therapeutic drug in patients.

References & Citations

1. Trikha M, Corringham R, Klein B, et al. Clin Cancer Res. 9(13):4653-4665. 2003.
2. Markham A, Patel T. Drugs. 74(10):1147-1152. 2014.
3. van Zaanen HC, Koopmans RP, Aarden LA, et al. J Clin Invest. 98(6):1441-1448. 1996.
4. van Zaanen HC, Lokhorst HM, Aarden LA, et al. Br J Haematol. 102(3):783-790. 1998.
5. van Zaanen HC, Lokhorst HM, Aarden LA, et al. Leuk Lymphoma. 31(5-6):551-558. 1998.
6. Guo Y, Nemeth J, O'Brien C, et al. Clin Cancer Res. 16(23):5759-5769. 2010.
7. Hunsucker SA, Magarotto V, Kuhn DJ, et al. Br J Haematol. 152(5):579-592. 2011.
8. Voorhees PM, Chen Q, Kuhn DJ, et al. Clin Cancer Res. 13(21):6469-6478. 2007.
9. Cavarretta IT, Neuwirt H, Zaki MH, et al. Adv Exp Med Biol. 617:547-555. 2008.
10. Voorhees PM, Chen Q, Small GW, et al. Br J Haematol. 145(4):481-490. 2009.
11. Karkera J, Steiner H, Li W, et al. Prostate. 71(13):1455-1465. 2011.
12. Kurzrock R, Voorhees PM, Casper C, et al. Clin Cancer Res. 19(13):3659-3670. 2013.
13. van Rhee F, Wong RS, Munshi N, et al. Lancet Oncol. 15(9):966-974. 2014.
14. van Rhee F, Rosenthal A, Kanhai K, et al. Blood Adv. 6(16):4773-4781. 2022.
B
Indirect Elisa Protocol
FA
IF
RIA

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Disclaimer AlertProducts are for research use only. Not for use in diagnostic or therapeutic procedures.