*2.1.2.2 Phenotypic screening*

Where binding assays are typically focused on identifying target-drug interactions, phenotypic screening takes a more disease-centric approach. Phenotypic assays aim to identify compounds that show effects on disease-relevant outcomes [63]. These are usually performed on cell lines or organelles engineered to function as disease models. Since the assay is target agnostic less, or no, information about specific targets is obtained. However, the fact that it is agnostic also means that there are more potential

targets available within this complex environment, which could lead to the discovery of new targets that would otherwise be left unexplored [63]. There are also additional benefits to this approach in the context of drug repurposing. Since the assays are disease based and compounds are approved drugs or clinical candidates it means that if positive outcomes are obtained the drug already has positive properties and shown efficacy in more complex systems, which is beneficial to real world applications [64].

## *2.1.3 Side effect based or "serendipitous" drug repurposing*

One of the most frequent reasons for drugs failing in (pre)clinical trials is the determination of a side effect that cannot be ignored. Most commonly this is determined to be a dangerous side effect that argues against further clinical investigation of the compound. However, one man's meat is another man's poison. These drugs with unwanted side effects can be given new indications through a drug repurposing strategy. Side effects-based drug repurposing links indications with clinical effect and is one of the common strategies employed for drug repurposing [65, 66]. A key example in this area is Sildenafil, which was originally entered into clinical trials as a drug to treat hypertension and angina [65, 67–70]. Unfortunately, Phase I clinical trials suggested that it had little effect on angina. However, use of Sildenafil causes a significant side effect: marked penile erections. This lead to the discovery that Sildenafil could be used as a treatment for erectile dysfunction (ED) [71]. In 1988, Sildenafil was approved by the FDA for the treatment of ED. Such repurposing approaches could be termed serendipitous repurposing, as the new indication area is revealed during clinical trials. As a result, such repurposing is relatively rare.
