iLite® Functional Bioassays
- if it binds, it shines

iLite® FUNCTIONAL BIOASSAYS

iLite is a cell-based reporter gene assay and is part of the analytical chemistry that is called bioassays, when utilizing  biological material in the assay or, more commonly, the substance your assaying are biological ones. The iLite Functional Bioassay portfolio is based on a reporter gene system and offers a convenient and powerful way of measuring and quantifying drug potency, detecting neutralizing antibodies (NAbs) and assessing immunogenicity.

Reporter genes cell lines allows for detection of signal that can be tailored and directly proportional to the induced function of your target compound. A report gene, is inserted in the cell under an inducible promotor and the gene is “read” to give a quantifiable readout. The advantages with the iLite technology are that one utilized the existing intra cellular systems better and does not “tamper” so much with the biological settings in the cell. A unique “lock and key” function is developed that makes the signal more specific and amplified, reading in a more sensitive cell and a larger dynamic range.

The iLite Functional Bioassay portfolio consists of cell lines provided in an “assay-ready” format for a rapid and convenient workflow and further reduction in assay variability, enabling their use in antibody screening, characterization, stability and potency studies.

iLite illustration Blue-4-01-1

Functional Bioassays
What are they?

Biological activity is a critical quality attribute for biopharmaceuticals and a functional bioassay will, in many cases, give a more intricate answer than a traditional binding assay since the functionally of the compound can be determined, not just a binding interaction.

Because a bioassay or bioactivity assay can reflect the Mechanism of Action (MoA) of a potential drug, it is an important component of the complete analytical profile and functional Bioassays, Reporter bioassays and cell health assays are used to characterize and develop novel therapeutics all the way from high throughput screening, through CMC lot release testing, to determination of immunogenicity.

iLite® FUNCTIONAL BIOASSAY PORTFOLIO

Valuable tools for the whole drug development continuum

Bioassays are an important component in ensuring regulatory compliance, and effective bioassays require reproducible cell lines. Functional Bioassays based on luminescence are very popular as they have high sensitivity, offer a large dynamic range, are easy to use and quick to perform.

The added advantage of getting these cell lines in an Assay ready format gives superior Ease of use

An iLite Cell Based Assay is run – similar to an ELISA,  but here the cells act as the reagent and are ready‐to-use instantly after thawing. This format gives your assay a very quick turnaround time - all the iLite assays are performed within a workday or at most overnight.

APPLICATIONS for the iLite® Functional Bioassay Portfolio

Accurate functional assessments throughout the drug development process

The iLite functional reporter bioassays are used to characterize and develop novel therapeutics – from high throughput screening, through CMC lot release testing, to determination of immunogenicity.

It is a versatile tool with applications throughout the drug continuum due to its clever engineering and use in accurate functional assessments.

Screening for new drug compounds

At an early stage in the development process, thousands of compounds need to be narrowed down to a small number of the most promising candidates.

In order to identify the best candidates, you should use an assay that as closely as possible reflects the complex processes of the cell, where different proteins or pathways are induced or inhibited depending on the circumstances. It is not sufficient to just determine whether or not the compound binds to its intended target.

Our cell-based iLite assays offer this functionality. In addition to finding out if the candidate drug binds to it target, you get additional information that can help you determine if the compound functions according to its intended mechanism of action.    

Potency assessment of therapeutic antibodies

Determining the potency of a candidate is one of the most essential parts of drug development.

Based on the regulatory guidelines, the potency of the drug candidate often needs to be addressed by a cell-based assay to achieve the biological relevance required. Potency assays also need to be highly reproducible.

The iLite system fits both criteria and can be used for measuring the drug candidate’s potency for Quality Control and as a batch release assay. Typically, it has to be established that the drug gives the same potency (effectiveness) for each batch when tested in a biologically relevant setting,

iLite cell lines can be successfully used to:

  • Measure potency in a biologically relevant setting
  • Get a reproducible, sensitive, and specific cell-based potency assay

Neutralizing Antibodies (NAbs) – Requirements and recommendations

Immunogenicity is an immune response against a therapeutic antigen causing that medication to lose its effectiveness over time and potentially resulting in serious illness. Clearly, immunogenicity is an important factor to consider as manufacturers develop new protein therapeutics.

Because drug development becomes increasingly costly as it moves downstream, it is important to anticipate immunogenicity as far upstream as possible in the drug discovery process.

Customers assessing immunogenicity need an Anti-Drug Antibody (ADA) assay and a NAb assay. While ADAs are most easily assessed by a regular ligand binding assay, detection of NAbs most often requires a functional cell-based assay.

Requirements for NAb assays are:

  • Cell-based assays with high serum tolerance
  • High drug tolerance
  • An assay that can be easily transferred to CROs, since these projects are often outsourced to CROs

This fits iLite assays perfectly. By combining regulatory compliance with excellent reproducibility and assay performance, iLite technology can circumvent many of the hurdles of traditional cell-based assays.

Assessing Biosimilarity in Drug Devlopment

With many of the existing patents of monoclonal antibody block buster drugs set to expire in the next few years, the development of biologic therapeutics similar to the original drug (biosimilars) has become increasingly important.

However, extensive requirements for analytical characterization is needed to show comparability between innovator and biosimilars and it must be proven that “the biological product is highly similar to the reference product notwithstanding minor differences in clinically inactive components” and that “there are no clinically meaningful differences between the biological product and the reference product in terms of the safety, purity, and potency”. This is the biosimilar development approach from the EMA, FDA and WHO.

The most critical evaluation is that of biological function, through assays that replicate the likely mechanism of action in vivo. Here, iLite Functional Bioassay Profiles could be of great value, when assessing and comparing the response of the compounds.

Accurate functional assessments throughout the drug development process

The iLite functional reporter bioassays are used to characterize and develop novel therapeutics – from high throughput screening, through CMC lot release testing, to determination of immunogenicity.

It is a versatile tool with applications throughout the drug continuum due to its clever engineering and use in accurate functional assessments.

Screening for new drug compounds

At an early stage in the development process, thousands of compounds need to be narrowed down to a small number of the most promising candidates.

In order to identify the best candidates, you should use an assay that as closely as possible reflects the complex processes of the cell, where different proteins or pathways are induced or inhibited depending on the circumstances. It is not sufficient to just determine whether or not the compound binds to its intended target.

Our cell-based iLite assays offer this functionality. In addition to finding out if the candidate drug binds to it target, you get additional information that can help you determine if the compound functions according to its intended mechanism of action.    

Potency assessment of therapeutic antibodies

Determining the potency of a candidate is one of the most essential parts of drug development.

Based on the regulatory guidelines, the potency of the drug candidate often needs to be addressed by a cell-based assay to achieve the biological relevance required. Potency assays also need to be highly reproducible.

The iLite system fits both criteria and can be used for measuring the drug candidate’s potency for Quality Control and as a batch release assay. Typically, it has to be established that the drug gives the same potency (effectiveness) for each batch when tested in a biologically relevant setting,

iLite cell lines can be successfully used to:

  • Measure potency in a biologically relevant setting
  • Get a reproducible, sensitive, and specific cell-based potency assay

Neutralizing Antibodies (NAbs) – Requirements and recommendations

Immunogenicity is an immune response against a therapeutic antigen causing that medication to lose its effectiveness over time and potentially resulting in serious illness. Clearly, immunogenicity is an important factor to consider as manufacturers develop new protein therapeutics.

Because drug development becomes increasingly costly as it moves downstream, it is important to anticipate immunogenicity as far upstream as possible in the drug discovery process.

Customers assessing immunogenicity need an Anti-Drug Antibody (ADA) assay and a NAb assay. While ADAs are most easily assessed by a regular ligand binding assay, detection of NAbs most often requires a functional cell-based assay.

Requirements for NAb assays are:

  • Cell-based assays with high serum tolerance
  • High drug tolerance
  • An assay that can be easily transferred to CROs, since these projects are often outsourced to CROs

This fits iLite assays perfectly. By combining regulatory compliance with excellent reproducibility and assay performance, iLite technology can circumvent many of the hurdles of traditional cell-based assays.

Assessing Biosimilarity in Drug Devlopment

With many of the existing patents of monoclonal antibody block buster drugs set to expire in the next few years, the development of biologic therapeutics similar to the original drug (biosimilars) has become increasingly important.

However, extensive requirements for analytical characterization is needed to show comparability between innovator and biosimilars and it must be proven that “the biological product is highly similar to the reference product notwithstanding minor differences in clinically inactive components” and that “there are no clinically meaningful differences between the biological product and the reference product in terms of the safety, purity, and potency”. This is the biosimilar development approach from the EMA, FDA and WHO.

The most critical evaluation is that of biological function, through assays that replicate the likely mechanism of action in vivo. Here, iLite Functional Bioassay Profiles could be of great value, when assessing and comparing the response of the compounds.

ILITE TECHNOLOGY - How does it work?

iLite cell-based technology principle

The iLite® cell-based assays are based on a simple reporter gene technology. Receptors, specific for a certain target or ligand, are expressed on the surface of a cell. Once the ligand binds to the receptor, this will trigger an intra-cellular signaling cascade, which leads down to a promoter region, fused to the reporter gene, in this case the Firefly Luciferase. Activation of the reporter gene along with the addition of a substrate will generate light. The amount of light will be correlated to the amount and activity of ligand that was bound to the receptors.

In the case of the iLite technology, there is a second reporter gene present, here shown by the Renilla Luciferase. This reporter gene is under the control of a constitutive promoter, and as such, will be active all the time in any living cell. In other words, the amount of light generated from this second reporter gene will be correlated to the number of living cells, and can therefore be used for normalizing the results to compensate for differences in cell number.

The iLite technology provides a functional cell-based assay format, with many of the features and benefits normally associated with ligand binding assays. The technology is based on a reporter gene format, which has been modified and adapted for applications during the whole drug development process.

Cell-based Assays vs Ligand Binding Assays

Cell-based assays are powerful tools in drug development as they can be used to determine a drug candidate’s mechanism of action in an in vitro model. The ability of cell-based assays to mimic the environment of the body have made them the preferred choice of regulatory agencies. However, cell-based assays can be challenging to set up and can have drawbacks, like high variance, which make results difficult to interpret.

Ligand binding assays are not as well suited to mimic the mechanism of action of a drug, but have other advantages, such as being drug-tolerant, sensitive and robust. In some cases, use of ligand binding assays may provide valuable information in the early steps of assessing a drug candidate. Often, the best approach is to complement a cell-based assay with a ligand binding assay.    

Regardless of how you choose to proceed, Svar has the capabilities and experience to assist in your immunogenicity project. We can use our inhouse assay development to develop ligand binding assays for your target. In addition, we offer a wide selection of cell-based iLite assays. If your target is not on the list, we can create a custom assay. Both kinds of assay development are offered and run by our Bioanalytical Services.

ADDITIONAL CONTENT - Our cell lines in context

COLONY STIMULATING FACTORS
Their unique biological roles and therapeutic benefits

INTERLEUKINS

A rapidly growing field with major pharmaceutical impact

iLite® RANKL

Ideal for potency measurements of anti-RANKL candidate drugs

iLite® C5a

Introducing a cell based way to study the complement system

Can´t find a perfect match? Let us develop it for you!

In case you haven't got an ideal target cell line and can't find what you’re looking for in our off the shelf solution, you can rely on our extensive expertise to develop the most appropriate one and we have vast experience in designing cell lines ideally fit for purpose!

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