This slide is from a famous paper which reviewed the falling productivity of the pharma industry, well worth a read. It shows the while PK issues were addressed between the nineties and 2000’s, the big problems of efficacy (which occurs mainly as a failure to get through phase II trials but is also a big problem at phase III) and clinical safety/tox didn’t really improve. A more recent analysis by Paul et al (2010) showed the situation hasn’t really improved and that poor prediction of safety and efficacy are the big problems causing clinical attrition, which still shows that a drug entering clinical trials has a less than one in ten chance of getting to market. We believe human tissue research can address the big causes of failure.

In this experiment we see the responses of subcutaneous arteries from both dogs and humans (similar region of chest/abdomen skin) and the fact that there is an enormous difference in the magnitude of the responses to 5-HT (5-hydroxytryptamine, serotonin, a key neurotransmitter and hormone which is /was an important drug target especially in the brain and gut.

In this experiment we see the responses of subcutaneous arteries from both dogs and humans (similar region of chest/abdomen skin) and the fact that there is an enormous difference in the magnitude of the responses to 5-HT (5-hydroxytryptamine, serotonin, a key neurotransmitter and hormone which is /was an important drug target especially in the brain and gut.

In this slide we have exposed major species differences in the expression of nitric oxide synthase enzymes (NOS enzymes). There are three types of NOS enzyme; those in endothelium (eNOS), in neurones (nNOS) and those up-regulated during inflammation (inducible NOS, iNOS). In this test of eNOS activity, we cause vasodilatation of isolated small arteries by adding ACh, which releases NO from the endothelium and causes 80% relaxation in normal vessels. In the presence of the client’s test compound the ability of ACh to relax vessels was severely inhibited, which would have major safety implications for the compound if used as a drug (even a small rise in systemic arterial blood pressure of 4-5 mmHg can be enough to greatly increase risk of cardiovascular disease over long-term use- this drug was a pain med and would be used similarly to ibuprofen). The effect was not observed in rats and this was later confirmed as a species difference in the NOS isoforms, in that the compound was inhibiting human eNOS but not rat eNOS.

Is there a place for human tissue research in the drug approval process? Dr David Bunton CEO, Biopta Ltd

Reasons for drug failures Address through human tissue research

Predicting Human Safety Pharmacology ICH S7A safety pharmacology guidelines; core battery of tests conducted in: CNS Cardiovascular system Respiratory system

“Effects of the test substance on the cardiovascular system should be assessed appropriately. Blood pressure, heart rate and ECG…. In vitro/ex vivo systems should also be considered. …….. Cardiac output, ventricular contractility, vascular resistance …….. Data from humans when available should also be considered.” Safety Pharmacology ICH S7A Guidelines Does this approach work and is human data generated/submitted?

Breakdown of Clinical Safety Problems of Withdrawn drugs 2000-2010

Translation of Animal Experiments to Humans Unexpected cardiovascular observations in Phase II clinical studies; no such effect in preclinical dog studies Biopta compared human and canine blood vessels

Species Differences: Vascular Sensitivity differences to 5-HT in subcutaneous resistance arteries Myography experiment with vessels isolated from healthy human and canine skin

Species Differences: Cardiac New Study Calls Into Question Reliance on Animal Models in Cardiovascular Research From the PharmaLive.com News Archive - Aug. 04, 2011 Human hearts respond differently than mouse hearts to two cardiovascular drugs Fedorov, V.V. et al. Journal of Molecular and Cellular Cardiology Effects of KATP channel openers diazoxide and pinacidil in coronary-perfused atria and ventricles from failing and non-failing human hearts; (2011) 51,2, 215-225

Residual surgical tissue Tissue transported immediately to laboratory Scientist isolates the tissues of interest The “living” tissues are exposed to drugs Small blood vessels isolated from tissue sample

Predicting human myocardial contractility: Ex vivo ventricular and atrial muscle Ventricular or atrial muscle contractility Bundles isolated from left or right side of human hearts Electrically stimulated at 1 Hz (60 bpm)

Case Study Predicting Vascular Adverse Effects Small 200 µm subcutaneous arteries dissected from human skin Predicts the influence of test compounds on human blood pressure Ach - Test compound Ach + Test compound No effect had been detected in earlier in vivo rat experiments

Predicting Vascular Effects: Tegaserod Case Study Coronary Artery Myography - Tegaserod and 5-HT were assessed in large (> 500 µm) and small (< 500 µm) human coronary arteries

A New Model for Predicting CV Risk Factors: Comprehensive Testing in Human Cells and Tissues We propose a new model for the prediction of cardiovascular risks, focused on human models, with reduced dependence on animal models Arrhythmia: HEK cell line- hERG liability; human primary cardiomyocytes- QT prolongation, characterisation of action potentials; human Purkinje fibres Cardiac contractility/cardiac output: ex vivo human ventricular and atrial muscle Blood pressure: peripheral resistance arteries from skin, gut, skeletal muscle; coronary arteries and pulmonary arteries Integrated assessment of heart rate, cardiac output and CNS regulation of CV system: single animal species in vivo administration

Thank You davidbunton@biopta.com