In Vitro Efficacy Testing Services for Stargardt Disease

We provide robust and sensitive in vitro screening and characterization platforms for accelerating the discovery and screening of potential therapies for Stargardt disease. Our services are tailored to assess the efficacy and mechanism of action of drug candidates targeting key proteins and pathways involved in Stargardt pathology, including the visual cycle, ABCA4 transporter activity, and the accumulation of toxic retinal byproducts. We focus on evaluating agents that can modulate retinoid metabolism, inhibit toxic bisretinoid formation, and restore retinal health. Our platforms enable the detailed investigation of biochemical and cellular processes underlying photoreceptor degeneration and lipofuscin accumulation associated with Stargardt disease.

We offer a comprehensive suite of in vitro testing methods encompassing bioluminescent, chemiluminescent, fluorescent, radioactivity-based, and ELISA assays to evaluate drug activity and target engagement. These assays enable precise quantification of enzyme activities, binding affinities, and pathway modulations relevant to Stargardt disease. By employing multiple detection formats and substrates, we support robust efficacy and mechanistic studies.

All-trans-retinol as substrate: Used to measure enzymatic activity in retinoid metabolism, providing insights into the effects of compounds on key steps in the visual cycle.

Bioluminescent assay: Employs light-emitting reactions to detect enzymatic activity or molecular interactions with high sensitivity, ideal for high-throughput screening.

Chemiluminescent assay: Uses chemical reactions that emit light to quantitatively assess target activity, enabling sensitive detection of low-abundance analytes.

Displacement of [125I]-MK-351A: Radio-ligand binding assay for determining compound interaction with specific receptors or transporters, useful for affinity and displacement studies.

ELISA assay: Enzyme-linked immunosorbent assay for quantifying proteins, metabolites, or antibodies, facilitating pathway or biomarker analysis.

Fluorescence resonance energy transfer (FRET) assay: Measures molecular interactions or conformational changes via energy transfer between fluorophores, suitable for real-time activity monitoring.

Fluorescent assay: Utilizes fluorescent probes to detect enzymatic activity or molecular binding, allowing for sensitive and quantitative measurements.

Fluorescent polarization assay: Detects binding events or molecular interactions based on changes in fluorescence polarization, providing information on binding kinetics.

Hippuryl-histidyl-leucine as substrate: Specific substrate to assess peptidase or protease activity, supporting mechanistic studies of enzyme modulation.

Homogeneous Time Resolved Fluorescence (HTRF) assay: Combines fluorescence resonance energy transfer with time-resolved detection for high-sensitivity, homogeneous detection of biomolecular interactions.

Radioactivity assay: Uses radiolabeled substrates or ligands to quantify enzymatic activity or binding, offering high sensitivity and specificity.

Retinoic acid 4-hydroxylation assay: Measures cytochrome P450-mediated metabolism of retinoic acid, evaluating drug effects on retinoid pathway enzymes.

Scintillation proximity assay (SPA): Measures molecular interactions or enzymatic activities using radiolabeled molecules and proximity-based detection, enabling rapid, wash-free assays.

We measure key pharmacological parameters including EC-50, IC-50, Kd, and Ki to characterize compound potency, efficacy, and binding properties. These metrics are essential for selecting and optimizing lead candidates by providing quantitative insights into their biological activities. Accurate determination of these parameters ensures informed decision-making throughout the drug development process.

EC-50 (Half maximal effective concentration): Indicates the concentration of a compound required to achieve 50% of its maximal effect, reflecting compound potency.

IC-50 (Half maximal inhibitory concentration): Represents the concentration of an inhibitor needed to reduce target activity by 50%, critical for assessing inhibitor strength.

Kd (Dissociation constant): Measures the affinity between a ligand and its target, with lower values indicating stronger binding, important for evaluating drug-target interactions.

Ki (Inhibition constant): Quantifies the binding affinity of an inhibitor for its target enzyme or receptor, aiding in the comparison and ranking of compound efficacy.

Recommended In Vitro Efficacy Tests

Angiotensin I Converting Enzyme

Angiotensin I Converting Enzyme (ACE) may influence Stargardt disease progression via retinal microvasculature regulation. ACE testing is vital for identifying drug candidates that modulate this pathway. Our service quantifies ACE inhibition using hippuryl-histidyl-leucine substrate assays and [125I]-MK-351A displacement binding. Key parameters measured include Ki (inhibition constant) and IC50 (half-maximal inhibitory concentration), providing critical data for Stargardt disease drug development.

Atp Binding Cassette Subfamily B Member 11

The Atp Binding Cassette Subfamily B Member 11 (ABCB11) is evaluated in Stargardt disease drug development due to its role in retinal lipid transport and potential impact on disease progression. Testing ABCB11 activity helps identify compounds that modulate its function, accelerating therapeutic discovery. Chemiluminescent and bioluminescent assays are employed to quantify transporter activity, with EC-50 values serving as the main parameter to assess compound potency and efficacy.

Complement C5

Complement C5 is implicated in Stargardt disease pathogenesis via complement-mediated retinal damage. Complement C5 testing is crucial for drug development to evaluate therapeutic modulation of this pathway. Key methods include ELISA, Western blot, and immunohistochemistry to quantify C5 protein levels and activation fragments. Main parameters assessed are C5 concentration, C5a generation, and membrane attack complex (MAC) formation, providing insight into complement activity and drug efficacy.

Complement Factor D

Complement Factor D is a pivotal enzyme in the alternative complement pathway, implicated in retinal inflammation in Stargardt disease. Testing Factor D levels and activity is crucial for evaluating complement-targeted therapies. Key methods include ELISA for quantification and functional assays for activity measurement. Main parameters assessed are serum/plasma Factor D concentration and enzymatic activity, providing essential biomarkers for drug efficacy and safety in Stargardt disease clinical research.

Cytochrome P450 Family 26 Subfamily A Member 1

Cytochrome P450 Family 26 Subfamily A Member 1 (CYP26A1) regulates retinoic acid metabolism, which is implicated in Stargardt disease pathogenesis. Testing CYP26A1 activity is crucial for evaluating drug candidates’ effects on retinoic acid clearance, aiding targeted therapy development. Our service utilizes the retinoic acid 4-hydroxylation assay to assess CYP26A1 inhibition, with IC-50 determination as the primary parameter for drug potency and selectivity assessment.

Retinoid Isomerohydrolase Rpe65

Retinoid Isomerohydrolase Rpe65 is critical in the visual cycle and implicated in Stargardt disease pathogenesis. Testing Rpe65 activity supports drug development by enabling assessment of compound effects on this enzyme. Using all-trans-retinol as substrate, our service employs sensitive fluorescent and radioactivity assays to measure Rpe65 function. Key parameters, including Kd and IC-50, are determined to evaluate drug binding affinity and inhibitory potency, facilitating targeted therapy development.

Retinol Binding Protein 4

Retinol Binding Protein 4 (RBP4) transports retinol and is implicated in Stargardt disease by influencing retinal vitamin A metabolism. RBP4 testing is crucial for evaluating drug candidates targeting this pathway. Our service offers IC-50 determination using robust methods—FRET, fluorescent, ELISA, HTRF, and SPA assays—providing precise assessment of compound potency and efficacy, essential for advancing Stargardt disease therapeutics.

Transthyretin

Transthyretin may modulate retinoid transport implicated in Stargardt disease pathology. Testing its activity is crucial for drug development targeting this pathway. Our service employs a fluorescent polarization assay to quantify compound effects on transthyretin-retinoid binding. Key output includes IC-50 values, enabling precise assessment of candidate drug potency and efficacy in modulating transthyretin function relevant to Stargardt disease.