Toxicity assays

Drugs research and development cost every year over $145 billion in US only. One of the key contributors to these rising costs is unacceptable drug safety profile either detected during Phase 1 Clinical Trial (50-60%) or during Phase IV Post marketing surveillance where some severe side effects were unnoticed during the phase I clinical trial. Of note, it has been described that in 30% of the studied cases the animals showed a toxic effect on a different organ compared to human (https://www.gwern.net/docs/dnb/2000-olson.pdf)

Main point of Preclinical testing is to determine the potential drug adverse effects, toxicity and drug-drug interactions before the drug is tested in either animal models or Phase 1 clinical trial. Hepato and renal in vitro toxicity tests are fundamental for pre-screening drug compounds to reduce costs for in vivo testing and Clinical Trials.
Moreover, it should be considered that the animal model might not mirror completely the human model, a good example is the expression of cytochrome P450. Hence the need to identify physiologically-relevant hepatic and kidney cellular models.
We offer a vast selection of 2D and 3D cellular models together with biochemical, metabolic and genetic assays to detect drug toxicity, ADME, pharmacokinetic and toxicogenomics.

Kidney toxicity

Renal proximal tubular epithelial cell: These cells are responsible for active clearance, reabsorption and accumulation of the drug as primary site. They are in direct contact with epithelial cells which can used for coculture assays. Proximal tubular epithelial cells maintain a highly polarized columnar cell morphology in vivo. these cells maintain the apical structure because of the shear stress, the shape is fundamental for enabling active pumps transport for glucose, ions, drugs across the epithelium. The static assays do not reproduce in vivo physiology.
https://www.jci.org
Type of cell Characteristics Assays Outcomes
RPTEC - Renal proximal tubular epithelial cells - Highly polarised columnar cells
- Primarily involved in drugs active clearance, reabsorption and accumulation
- apical morphology is fundamental for active pumps transports
Nephrotoxicity Apoptosis/Necrosis
Cytotoxicity
Cytostaticity/Proliferation
Mitochondrial cytotoxicity
Genototoxicity
    Permeability Cell-cell junctions
Adhesion
    3D Shear stress modulation
    Coculture Motility
Adhesion
       
Mesangial cells - They support the glomerular tuft and synthetize the surrounded mesangial matrix.
- phenotypically similar to vascular smooth muscle cells, able to secret growth factors
- involved in glomerular contraction and hormones release.
Nephrotoxicity

Apoptosis/Necrosis
Cytotoxicity
Cytostaticity/Proliferation
Mitochondrial cytotoxicity
Genotoxicity

    Secretion Growth factors secretion
    Endocytosis Phagocytosis ability
    Adhesion Cell -Matrix interactions
    Coculture Cell - cell cross talk
    Permeability Cell-cell junctions
Adhesion
       
Podocytes

- They are in contact with glomerular capillary
- Stabilise glomerular architecture

- Are able to contract and support the glomerular filtration barrier.
Nephrotoxicity Apoptosis/Necrosis
Cytotoxicity
Cytostaticity/Proliferation
Mitochondrial cytotoxicity
Genotoxicity
    Permeability Cell-cell junctions
Adhesion

Acute kidney disease Chronic kidney disease Nephrotoxicity
Neutrophil gelatinase-associate lipocalin (NGAL) Asymmetric dimethylarginine (ADMA) N-acetyl-β-D-glucosaminidase (NAG)
interleukin-18 (IL18) Neutrophil gelatinase-associate lipocalin (NGAL) Glutathione S-transferases (GST)
Kidney Injury Molecule-1 (KIM1) Kidney Injury Molecule-1 (KIM-1) Gamma-glutamyltransferase (GGT)
Cystatic C (CST3) Liver-type Fatty acid binding protein (L-FABP) Kidney Injury Molecule-1 (KIM1)
Liver-type Fatty acid binding protein (L-FABP)   Lactic dehydrogenase (LDH)
Insulin like growth factor binding protein (IGFBP7)    
Tissue metallopeptidase inhibitor 2 (TIMP-2)    
Kidney injury markers - BioPlex MAGPix technology

Human Mouse Rat
Albumin Clusterin Albumin
Clusterin Cystatin C Calbindin
Collagen IV Epithelial growth factor Clusterin
Cystatin C Interferon gamma-induced protein 10 Cystatin C
Glutathione S-transferases Kidney Injury Molecule-1 Glutathione S-transferases
Interferon gamma-induced protein 10 Lipocalin-2 Interferon gamma-induced protein 10
Kidney Injury Molecule-1 Osteopontin Kidney Injury Molecule-1
Lipocalin-2 Renin Lipocalin-2
Liver-type Fatty acid binding protein β-2-Microglobulin Osteopontin
Macrophage activator protein Tissue metallopeptidase inhibitor 2 Tissue metallopeptidase inhibitor 2
Matrix Metalloproteinases (MMP9)    
Osteoactivin Vascular Endothelial Growth Factor Vascular Endothelial Growth Factor
Osteopontin   β-2-Microglobulin (β2M)
Renin Interleukin 2,6,8,10 (IL2, IL 6, IL8, IL10) Interleukin 2,6,8,10 (IL2, IL 6, IL8, IL10)
Trefoil factor 3   Trefoil factor 3
a-1-Microglobulin    
Heat Shock Protein 70 (HSP70)    
Interleukin 2,6,8,10 (IL2, IL 6, IL8, IL10)    

Biomarkers Target Material required Target species
Alanine aminopeptidase Early marker for glomerulonephritis Serum, plasma, tissue homogenates. Human, rat
Albumin Early marker for kidney damage Urine, serum, plasma, tissue homogenates. Human, rat
Alkaline phosphatase Renal function impairment marker Urine, serum, plasma, tissue homogenates. Human, mouse, rat
Angiotensin-converting enzyme Enzyme involved in renin cascade Serum, plasma, supernatants, saliva Human, mouse, rat
Calbindin Renal function impairment marker Urine Human, rat
Complement C3 Marker for glomerular disease and glomerulopathy Cell culture supernatant, Saliva, Milk, Urine Human, mouse, rat
Clusterin Marker for tubular cells damage Serum, plasma Human, mouse, rat
Creatinine Functional marker for early stage kidney disease Urine, serum, plasma Human, mouse, rat
Cystatin C Biomarker for glomerular filtration Serum, plasma Human, mouse, rat
Endothelin It regulates the glomerular hemodynamic and blood flow. Urine, serum, plasma Human, mouse, rat
Gamma-glutamyl transpeptidase Marker for renal ischemic damage Serum, plasma, culture media. Human, rat
Glutathione S-transferases Marker in acute kidney injury Urine Human, mouse, rat
Kidney Injury Molecule-1 Marker for tubular reabsorbtion and ischemic injury Urine Human, rat
Microalbumin Marker for renal dysfunction and vascular damage (often diabetes related) Urine, serum, plasma, cells Human, mouse, rat
N-acetyl-b-glycosaminidase Marker of nephrotoxicity and early stage proteinuria Urine, serum, plasma, homogenate tissues Human, mouse, rat
Osteopontin Biomarker of renal and glomerular damage Urine Rat
Sodium/hydrogen exchanger isoform Ion transport proteins involved in intracellular pH regulation. Serum, plasma, homogenate tissues Human, mouse, rat
Trefoil factor 3 Marker of acute renal tubular injury Serum, plasma, urine human

Liver toxicity

ASSAY TARGET
XMAP HUMAN LIVER INJURY MAGNETIC BEAD PANEL Circulating marker proteins for kidney injury
CELL MORTALITY To distinguish apoptotic, necrotic and live cells
LIPOFUSCIN ELISA Unsaturated fatty acids oxidation
STEATOSIS COLORIMETRIC ASSAY Lipid accumulation
IMMUNOHISTOCHEMISTRY Liver fibrosis
K8 AND K18 ELISA Mallory-Denk bodies
P62 ELISA Intracellular hyaline bodies

SYSTEM TYPE OF CELL ASSAYS
HOMOGENIX

primary hepatocytes
HepG2

Hepatocytes IP or ES derived
P450, drug-drug interaction, membrane integrity, apoptosis, mitochondrial dysfunction, oxidative stress, oxidative DNA damage
Hepatopac Primary hepatocytes + stromal cells or fibroblasts Inflammation, phase I/II metabolites
Insphero Primary hepatocytes+ Kuppfer cells Bile acid secretion, mitochondrial function, CYP-toxicity, DILI model
Regenemed 3D coculture- stromal, parenchymal, hepatocytes Gene expression, P450, inflammation, apoptosis, metabolic.

Human Rats
Liver-Type Arginase 1 (ARG1) Liver-Type Arginase 1 (ARG1)
Malate dehydrogenase 1 (MDH1) aspartate transaminase 1 (GOT1)
α-glutathione S-transferase (GSTα) α-glutathione S-transferase (GSTα)
Sorbitol Dehydrogenase (SDH) Sorbitol Dehydrogenase (SDH)
5'-Nucleotidase/CD73 (5’-NT) 5'-Nucleotidase (5'-NT/CD73).