Reigniting Pharmaceutical Innovation Through Holistic Drug Targeting

Drug Discovery

Reigniting pharmaceutical innovation through holistic drug targeting

Modern drug discovery approaches take too long, are too expensive, have too many clinical failures and uncertain outcomes. There are many reasons for this unsustainable business model, but primarily, the approaches are not comprehensively holistic. Secondly, none of the pharmaceutical companies openly share the reasons for the failure of their clinical candidates in real time to effectively navigate the ‘industry’ from committing the same mistakes. It is time for the pharmaceutical industry to embrace, metaphorically speaking, a community-driven ‘Wikipedia’ or ‘Waze’-type shared-knowledge, openly- accessible innovation model to harvest data and create a crowd-sourced path towards a safer and faster road to the discovery and development of life-saving medicines. This may be a bitter pill for Pharma t o swallow, but one that ought to be given serious consideration. The time is now for a paradigm shift towards multi-target-network polypharmacology drugs exalting symphonic or concert performance with occasional soloists to reignite pharmaceutical innovation.

rom the turn of the 20th century, pharma- and ultra-high throughput workflows enabled By Dr Anuradha Roy, cognosy and ethnopharmacology combined screening of millions of compounds to identify hit Professor Bhushan F with anecdotal clinical evidence accumulat- can didates for lead development. Despite billions Patwardhan and ed over centuries of hands-on knowledge from pri- of dollars spent on R&D, only a fraction of the Dr Rathnam mordial disease management practices, albeit with molecules identified from the screening operations Chaguturu uncertain outcomes, formed the basis for the devel- find their way into clinical trials. Difficulties in opment of drugs. Reverse pharmacology, deep predicting safety profiles, redundancies and effica- rooted in traditional medicine, laid the foundation cies across a genetically-diverse patient population for the emergence and evolution of modern drug contribute to the high attrition rates in clinical tri- discovery approaches as a highly formal and regi- als. Success in clinical trials was found to b e affect- mented science. Advances in genomics, assay and ed, among other reasons, by the poor design of combinatorial chemistry technologies, informatics clinical trials, selected clinical end-points unsuited and robotics led to increased screening operations for the desired outcome and lack of evaluation of significantly compared with traditional discovery pharmacogenomics of the patient population methods. The pharmaceutical industry-driven high selected for the indication. In addition, while a

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vast majority of FDA approved drugs target diseases affecting large populations such as cancers, infectious diseases or cardiovascular diseases, which promises a good return on R&D investment; and hence pursued extensively by big Pharma. Rare dise ases (which affect small populations) or neglected diseases (more prevalent in developing countries) have largely been ignored because of low profitability for the pharmaceutical industry and low affordability for the poor. Market size decisions as well as business portfolios are also known to influence indication selection, population sizes and end-points in clinical trials. The last decade witnessed patent expirations, drug recalls and toxicity-driven withdrawals, all of which neg- atively impacted pharma R&D productivity. Despite significant advances in our knowledge, safety from prolonged use of drugs in the post- FDA approval period remains uncertain. Treatment of complex diseases with single or small combination therapies, while effective in the short term or at early stages of a disease, continues to be insufficient in mitigating advanced or recurrent disease progression. Loss of responsiveness due to long-term administration of single agents is attributable to the robustness of the redundant molecular functions within biological networks. Many diseases are complex, heterogeneous and multifactorial, have several phenotypes, variable risk factors and responses that are also influenced by genetic variations, age, gender and environ- mental factors such as diet, microbiome and lifestyle choices. The ever too often seen resistance majority of the marketed drugs was approved to continued use of drugs and loss in treatment based on experimental data supporting single tar- efficacy are generally mediated by network get selectivity, recent clinical and basic studies have robustness (signalling pathway redundancy and unambiguously shown that many marketed drugs crosstalk) causing compensatory or counter-target lack absolute selectivity. activities or neutralising action. The overall low productivity and innovation of new drug classes, Paradigm shift as well as safety- and efficacy-driven drug recalls, Despite high failure rates of compounds in clinical begs for a fundamental paradigm shift in current settings, target-based drug discovery has contribut- drug discovery approaches. ed to drugs approved for several indications. An overall evaluation of the FDA-approved new Druggability of genome molecular entities reveal that the majority of drugs Of the 19,000 human protein coding genes pre- clustered into previously known classes, and dicted from human genome sequence analysis, encompasses a limited number of molecular targets only about 10% are ‘druggable’ by in silico anal- and diseases. A recent analysis of the DrugBank ysis2. The majority of the ‘products’ from genome, database identified 435 targets modulated by 989 such as the proteins or RNA, still remain function- drugs and include GPCRs, ligand gated ion-chan- ally unclassified and the expansion of functionali- nels, receptor tyrosine kinases and certain other ty may be further influenced by epigenomic events classes of enzymes1. The analysis also highlighted as well as post-transcriptional or post-translation- the fact that only half of the marketed drugs show al regulation3. The challenge lies in generating higher order and number of drug-target interac- high quality data for target identification, drugga- tions within a very limited proteome-scape. The bility via siRNA or chemical perturbation in

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healthy and disease states, confounded by the exis- acterisation and validation. These tools include, tence of functional and physical networks between but are not limited to, protein and RNA microar- molecular players in the same or other signalling rays, bioinformatics-driven protein interaction pathways. The hub proteins such as p53, p27, maps, signalling pathways, phenotypic changes by BRCA1, ubiquitin and calmodulin can have five functional studies using genetic-based technologies or more interactions with other proteins, and may (RNA interference, knockdowns, overexpression, either exhibit a single transient interaction at any genomic mutations using clustered regularly inter- given time or participate in several interactions spaced short palindromic repeats (CRISPR-Cas), simultaneously. The cross-talk and interdepen- and mouse, zebrafish or Caenorhabditis elegans dence of biological networks adds another level of models). The exhaustive information pertaining to complexity in responses to perturbations with the therapeutically-relevant drug target can facili- chemicals, biologics as well as other factors. tate efficient design of assays or selection of appro- Systems analysis of large networks indicates that priate pa nel of cell-lines in targeted or phenotypic targeting upstream events, hub proteins or redun- drug discovery approaches to identify compounds dancies will potentially be more impactful in that can serve as chemical tools to dissect the path- quantitative and qualitative efficiency of cellular ways of interest. response. In addition to proteins and networks, targeting RNA druggability is still under-utilised. Hit to lead strategies All known RNA motifs and their small molecule Once screen actives or hits are identified, it is binders, compiled in the Inforna database4, are essential to experimentally determine the mecha- attractive to interrogate and yield lead molecules nism of action, efficacy and safety of the scaffolds with potential to be validated in disease systems, and target on-off occupancy and engagement as exemplified by the evidence drawn from the time7. The direct binding of the hit molecules to selective binding of 5’-azido-neomycin B to cellular targets can be determined using mass spec- Drosha and Dicer processing sites of miR-525, a trometry on protein samples from cellular thermal microRNA overexpressed in liver cancer5. A tar- shift assays, protein arrays, or compound-affinity get-agnostic approach can thus be effectively used chromatography. The binding affinities to specific to identify in vivo modulators of oncogenic and and off-targets, binding site architecture, co-crys- other disease relevant microRNAs. tallography studies also provide invaluable information on understanding the chemical species. Comprehensive target biology From the start scaffolds may be tested in assays Strong correlative evidence between a target or tar- quantifying potency, selectivity, specificity, gets and a disease is ever more critical for the suc- lipophilicity, molecular mass and early ADME cess of rationale-driven drug discovery. The success (absorption, distribution, metabolism and excre- of designing well-tailored assays for primary and tion). Collaborative effort between AstraZeneca, secondary drug discovery screens, prediction of Eli Lilly, GlaxoSmithKline and Pfizer to under- toxicity profiles, biomarker profiling and ultimate stand the reasons behind drug candidate attrition8 patient population-specific responses are all depen- indicated a strong statistical significance between dent on how much information, both accurate and the physicochemical properties of compounds and reproducible, is available for the disease-relevant, safety failure rates in clinical trials, although no therapeutic target6. It is essential to characterise a rationale emerged for predicting which compounds target in both normal and diseased states for its will be successful in clinical trials. Viable leads functional domain(s) redundancy, alternatively share the properties of high efficacy and potency spliced forms, subcellular localisation, tissue-spe- against a specific target, bind the target directly at cific protein and RNA expression profiles, the same cellular site where the target is expressed, RNA/protein half-life, transcriptional/post-transla- have low off-target effects, no activity against tional regulation and effect of dominant negative undesirable players such as hERG channels and mutant isoforms. Both genetic and chemical vali- ac ceptable toxicity profiles. Target engagement of dation helps define druggability and translational time and activity at the target tissue/organ requires potential of a target. The same target may be mod- designing pharmacokinetics/pharmacodynamics ulated in several distinct cancer types or in several (PK/PD) modelling in both preclinical and clinical se emingly unrelated disorders, and as a result will models. Preclinical drug development that can help expand the indications or design treatments in accurately predict clinical response of drugs can clinical trials. An array of tools has become avail- influence the success rate of lead compounds in late able in recent years for target identification, char- stage drug discovery.

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Relevant animal models from all sources and databases. The last decade There is a growing awareness of the limitations of witnessed an explosion in data sets for the true translation potential of clinical research metabolomics, transcriptomics, genomics, epige- from mouse models; a case in point being that the nomics and proteomics etc. The high-throughput average rate of successful translation from animal ‘omics’ technologies applied to normal and dis- models to clinical cancer trials is less than 8%. eased states have generated large volumes of dis- Differences in genetics and physiology of mouse tinct types of omics data. Each type of data has its and humans add to issues in translational rele- own properties and requires specific analysis vance. Cancer models involving mous e subcuta- methods and tools. The emerging paradigm for neous xenografts may or may not translate well in drug discovery underscores the value of integrat- later stages of drug discovery. Mouse xenograft ing the multi-omics data to build a complete models from primary tumour-derived patient cells model of the processes in biological systems since with distinct driver mutations are often used to val- signalling network cross-talks, functional biologi- idate the clinical translatability of drug candidates, cal redundancies and protein interactomes clearly but are inadequate models for tumour initiation influence the drug activity and safety profiles. A and progression. Several potential alternatives more holistic approach, designing drug discovery include using CRISPR-Cas technology to modify protocols that integrate the knowledge from tradi- the mouse genome specifically and use of large ani- tional medicine to mining omics datasets and mal models (rabbits, dogs, pigs, goats, sheep and designing smarter clinical trials, is certain to non-human primates) to closely replicate the improve the drug discovery process by providing human physiology and disease. new and first-in-class drugs as well as in expand- ing the repertoire of global diseases targeted. Exploiting pan-omics Cumulative knowledge acquired in recent years Genomics has led to an increased emphasis on designing Advances in next-generation sequencing (NGS) treatments that are more personalised, or clus- platforms have led to exponential increase in tered into subtypes based on information collated sequencing data from whole genomes (WGS),

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DNA coding exomes (WES), DNA variants (SNP, single nucleotide polymorphisms, deletions, inser- tions), DNS segments interacting with proteins Biomarkers, challenges (chromatin immunoprecipitation, Chl-IP), mRNAs (transcriptome), long non-coding RNA (lncRNA) and circular RNA (circRNA). Genome- and opportunities wide association studies (GWAS) have the poten- Biomarkers are functional, reliable and measurable biochemical indicators in health tial to identify genetic variations, especially the and disease. In recent years, biomarkers research has played a critical role in phar- single nucleotide polymorphisms, associated with maceutical R&D for creating groundbreaking therapies and companion products. a disease. Data on DNA variations is now avail- Biomarkers use in such discoveries helped in the recognition, validation and thera- able for Crohn’s disease, Type 2 diabetes, prostate peutic intervention in diseases that were considered insurmountable just a few cancer, age-related macular degeneration, obesity decades earlier; development of breast cancer drug Herceptin is such an example. and 50 other human diseases. The potential of Similarly, their widespread clinical use has enabled, along with high technology, devel- establishing associations between disease and opment of ultra-sensitive detection devices and new instrumentation. In the last genetic variations can ultimately influence diag- three decades, research for ever more complex drugs and biotechnology-derived nostics and disease management. products has led to significant adaptation of ultrahigh-throughput automation, minia- turisation and data analytics with advanced computing. Coupling biomarkers Pharmacogenomics research to drug discovery albeit presents a much higher complexity. Technically Precision medicine can improve efficacy scores of challenging detection modalities, needed to keep pace with emerging biology and FDA-approved drugs or new compounds in clini- chemistries, present immense challenges and contrasting opportunities. As most cal trials by integrating information from pharma- biomarkers work is carried out at t he interface of basic biology, clinical and instru- cogenomics. A genetic profile-based identification mentation research, transitioning the laboratory-derived information to clinical of patient population subgroups is helpful in pre- application adds another dimension to the problem. However, potential exists to dicting patient response to drug dosage, mecha- leverage expertise from diverse disciplines and adapt techniques and methods to nism of action, minimising drug-related toxicity enhance productivity, reduce timelines and achieve compelling results. There is a con- and adverse events. Pharmacogenomics data pro- stant need for innovation , with ever increasing cost pressures on pharmaceutical dis- vides information on genetic marker-driven effects covery in a globally challenging healthcare environment. An integrated approach from on drug response, refractiveness of drug treat- the outset thus adds a much-needed aspect to strengthening the traditional, linear ments, as well as functional effects of genomic drug discovery for better clinical outcomes. variants. The concept of pharmaco-metabolomics- Krishna Kodukula, KK Biopharma aided pharmacogenomics helps combine the roles of environment and gut microbiome interactions in interpretation of especially immune mediated disorders. coding RNAs in normal and diseased human tissues and cells9. The epigenomic signature data from var- Epigenomics and transcriptomics ious sources are accessible through IHEC Epigenome encompasses all epigenetic mechanisms (International Human Epigenome Consortium); and that regulate chromatin via DNA methylation and increasingly, more comprehensive global or locus- histone modifications (acetylation, methylation, specific genomic aberrations are being reported for phosphorylation, sumoylation, ubiquitination and many diseases including hypertension, asthma and proline isomerisation). Epigenetics mediates differ- pain. Application of epigenomics can positively ential gene expression, which may be development-, impact clinical therapeutics in multifactorial dis- tissue- or cell-specific, or dysregulated in pathogen- eases and improve early cancer detection, prognosis esis. Numerous technologies, old and new, encom- and prediction of treatment responses (pharmaco- passing high throughput DNA sequencing and chro- epigenetics). matin immunoprecipitation (ChIP), DNA microar- rays (ChIP-chip) are used to profile DNA/histone Biomarkers modifications. Genome sequencing studies have Genomic, imaging, proteomic and metabolomics identified epigenome hotspots in DNA from differ- technologies have unravelled a plethora of novel ent cells and tissues. Epigenomics data from genetic/proteins which can serve as biomarkers Encyclopedia of DNA Elements (ENCODE) and that can help predict disease susceptibility, disease The NIH Roadmap Epigenomics projects, has subtypes and disease progression, as well as pre- helped define functional DNA segments that either dict response to a drug treatment10. If validated possess specific chromatin structures or have protein reproducibly in disease, the biomarkers can be binding sites or directly code for proteins or non- incorporated in preclinical and clinical workflows

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products include drugs against cancer, bacterial, protozoan and fungal infection and inflammation. Natural products as drug The bioactivity evaluation of natural extracts is both labour and time-intensive, requiring several types of extract preparation, reiterative bioassay guided frac- leads: to be or not to be? tionations and isolation of active scaffold (HPLC, Natural products (NPs), evolutionarily optimised by the living organisms for serving Mass spec), target specificity and mechanism of different biological functions, and thus, with inherent diversity and complexity, are a action. In case of low active ingredient yields from a good source of drug leads. However, NP-based drug discovery is rather neglected by source, chromatographic fingerprinting is often per- the pharmaceutical industry: formed to identify richer sources of the active com- • Typical high throughput screening platforms are not readily suitable for bioactivity pound from other related sources. The safety of detection of plant extracts. publically-consumed botanicals, herbal and dietary • NPs are complex structures with multiple stereocentres. Isolation and unambigu- supplements has been addressed by EU hERGscreen ous structure elucidation are time-consuming, and labour and cost-intensive. network (http://hergscreen.univie.ac.at/), a database • Resupply of NPs is critical for hit-to-lead and preclinical studies. Total synthesis of for cardiotoxic risk assessment based on ligand- NPs with multiple chiral centres is rather challenging. based human Ether-à-go-go Related Gene (hERG) • NPs are not necessarily ‘drug-like’, requiring structur al modification for improved channel 3D pharmacophore models. solubility and bioavailability. • NPs are not directly patentable and need significant pharma investment to modify Natural products, the phoenix the structure and claim that the invention (therapeutic use) is ‘significantly different’ From the turn of the 20th century, natural prod- from the natural source. ucts have been a trusted source of a vast majority The Western reductionist approach follows the ‘one target-one compound’ of anti-cancer drugs and anti-infectious agents, and paradigm, however, such entities are not quite effective for multi-factorial diseases. by far the richest source of novel molecular scaf- The concept of polypharmacology and multi-targeted drugs has only recently begun folds. Some famous examples derived from natural to gain recognition. This paradigm shift has reopened the doors towards holistic products over the past 30 years include artemisinin medication, and turned the attention towards the herbal universe. Such ‘natural’ (malaria), colchicine (gout), galanthamine (demen- drugs have a complex chemical composition in a variety of quantities that allows syn- tia) and paclitaxel (cancer). Curcumin is the most ergism between the active substances. Much integrated effort encompassing celebrated example and is currently in 26 clinical metabolomics, chemical genomics, proteomics and network pharmacology is trials for a variety of indications. The therapeutic required for target identification and elucidating the mechanisms of multi-compo- effects of traditional medicine or formulations nent/multi-target herbal medicines. have been documented in various civilisations and György Dorman, Target-Ex continue to be utilised for treatment of several disorders in many countries. Natural product discovery has largely been deprioritised due to complexi- ties in large-scale production, either natural to address toxicity, safety, efficacy and patient sources or via chemical synthesis, among others. selection. The safety and efficacy of investigation Reverse pharmacology (clinic to laboratory) aims drugs can be further improved by including genet- to expand the drug discovery approaches to ic profiling of polymorphisms and drug- include lead identification from traditional formu- metabolising enzymes. lations and extracts. Reverse pharmacology initi- ates from documented clinical and experimental Pharmacognosy observations, utilises modern preclinical and clini- It is a study of ethnobiology, taxonomy, sample col- cal approaches for active principal identification lection, extraction, isolation and defining chemical and isolation, mechanism of action or target iden- and biological properties of naturally occurring, tification as well as formal safety and toxicology medicinally-active substances obtained from plants, clinical study to identify viable leads. microbes and animals, etc. Various herbal and mixed formulations containing minerals and metals Biologics revolution have been documented historically in folk Recombinant protein based therapies (monoclonal medicines. The natural products may be whole antibodies, growth factors, hormones, blood fac- organisms, parts of a plant, animal, secondary tors, enzymes, vaccines, anticoagulants, fusion metabolites or extracts. The rich biodiversity in ter- proteins) have been developed for various diseases. restrial and marine organisms promises to offer The recombinant drugs are produced in bacteria, novel cell permeable chemical scaffolds and novel yeast as well as mammalian cells. Biologics that bioactive compounds. Known bioactive natural stimulate immune response against tumours such

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as oncolytic viruses, antibodies and vaccines, as resources12. Open innovation where ideas, data, well as T-cell mediated therapies hold great failures, reagents and tools are shared between col- promise for tumour immunotherapy. Biologics dis- laborating partners can accelerate the early drug covery helps to diversify a company portfolio, discovery research. Innovation from global collab- shortens the path to discovery and gains from orations will support expansion of the drug discov- restricted competition from biosimilars and gener- ery landscape. Open innovation has led to a boost ics. The increased pharma interest in biologics dis- in the research of neglected and rare diseases. covery is reflected in the 2015 FDA approval of Merck opened up its data and analysis tools to several first-in-class antibodies, proteins and hor- enable complex model building under SAGE mones11. Though target specific, treatment with bionetworks (http://sagebase.org/). In another biologics for chronic disorders are also known to form of compound collaboration, the Eli Lilly-PD2 result in adverse events involving immune reac- Initiative (https://pd2.lilly.com) seeks to test tions and disorders, infections and tumours. The molecules and promising compounds originating long-term treatment with biologics (eg Humira) is from academic research in the Eli Lilly phenotypic also far more costly than the small molecule drugs. discovery platform. The profiling data and the sec- Unlike the generic small molecule drugs, which ondary assay information are shared with the aca- bring down the treatment cost to patients, the demic researchers. Again, we reinforce that it is biosimilar drugs are still associated with high pro- time for the pharmaceutical industry to adopt a duction costs and may not be as efficacious and community-driven ‘Wikipedia’ or ‘Waze’-type safe as the original biologic. shared-knowledge, openly accessible innovation model to harvest data and create a crowd-sourced Stem cell therapy path towards a safer and faster road to the discov- The hype and hope offered by stem cell therapy are ery and development of life saving medicines. undeniable. Stem cells are being used in the treatment of blood and immune disorders, skin grafts Approaches for holistic drug targeting or corneal damage repair (www.eurostemcell.org). Comprehensive disease models can be structured Despite high optimism in the therapeutic potential to define the possible role of individual genetic of the stem cells, the field faces numerous chal- loci, network interactions or post-transcriptional lenges as developing new stem cell treatments events that may contribute to complex disease requires in-depth understanding of human cell lin- development and progression. With the advent of eages, cell markers, niche-dependent potency and personalised medicine, theoretically the sequence processes controlling cell proliferation, differentia- information from a patient can be compared with- tion and functional specialisation. Investigational in the large datasets available from genome drugs can be tested in stem cell models for their sequencing, transcriptome analysis, epigenomics, ability to repair damage in appropriate animal proteomics and metabolomics to create working models under good laboratory practice, though the disease models. The addition of phenome-wide progress has been rather slow. Cellular reprogram- association studies (PheWAS), which links diseases ming efforts by genetic factors to generate induced to genetic variants, is a good starting point for pluripotent- and lineage-specific stem cells from identifying new drug-disease pairs. The person- somatic cell types hold great promise for basic alised model can be used to define a treatment reg- research as well as in clinical applications. A com- imen based on all available FDA-approved drugs, plementary, alternate approach that has been known targets or pathways. In practice, there are equally successful involves the use of small limitations in establishing a direct and specific molecules to maintain the self-renewal potential of causal relationship between the experimental data stem cells and thus target specific epigenetic pro- sets and clinical disease progression. While there is cesses, signalling pathways and the associated cel- an exponential increase in computational algo- lular processes. This approach is quite attractive, rithms and deep mac hine learning protocols for and holds promise for manipulating cell fate to a mining the databases, there is still not enough data desired outcome. validating the bioinformatics datasets with the animal model/clinical data. Collaborative and open innovation Biological systems are complex, circuitous and Partnerships and collaborations across pharmaceu- labyrinthine. Diseases are even more complex. A tical companies and academia can help improve disease may more likely be an eventual phenotypic the drug development process by reducing redun- outcome of a misguided pathway or network. dancy and a more judicious allocation of Developing successful treatment strategies to

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ray crystal structures or models, ligand-based quantitative structure activity relationship (QSAR) Precision medicine similarity prediction of two- or three dimensional The landmark Precision Medicine Initiative, announced by US President Obama in fingerprints of small molecules, binding pocket 2015, heralds an unprecedented paradigm shift in how the stakeholders – similarity of proteins. The predictive power of vir- researchers, physicians, payers and patients – work together towards an individu- tual methods has limited utility unless validated by alised approach for disease treatment and prevention that takes into account indi- experimental science in physiologically relevant vidual variability in genes, environment and lifestyle for each person. It brings an systems where for instance, the protein domains unprecedented improvement in our understanding of underlying disease mecha- are not inflexible and in binding sites/folds change nisms and the detection and treatment options. The cohort programme’s goal is to with conditions. Polypharmacology-driven drug extend precision medicine to all diseases, and in its current form it is US-centric. discovery has the potential for selecting molecules Precision medicine is truly translational in scope and integrates all allied biomedical that have higher efficacy, lower toxicity and less disciplines including global clinical trials in an individualised, patient-centric way. It potential for synergistic effects compared with does not mean, however, the discovery and development of drugs or medical multiple prescriptions administered simultaneous- devices, unique to any given patient. That would be exceedingly cost prohibitive and ly. Polypharmacology that does not involve toxic unrealistic to expect. The power of precision medicine is to bring forth the unique- promiscuity of drugs is predicted to be beneficial ness of each patient for a particular disease at the molecular level and integrates the for the treatment of complex diseases and is of precise treatment needed from the outset. Biomarke rs and diagnostics play a critical great value in multi-target drug discovery. The role. The convergence of panomics and systems biology are integral to link all the drugs with adverse side-effects should be depriori- interacting pathways in addressing the heterogeneity of disease etiology and develop tised early in drug discovery while identifying those an individualised, true combination drug therapy, much like the principle behind the exhibiting potentiation of therapeutic activity by herbal treatments. Routine practice of precision medicine in a clinical setting for all hitting multiple targets followed by characterisa- diseases and all patients may or may not be realised in our lifetime, but it is a worthy tion of pharmacological profiles. approach and a lofty goal for all nations to embrace at least in some form and work collectively together to eradicate diseases that ravage mankind. This is the true holis- Drug repositioning tic approach from ‘bench to bedside’. Drug repositioning or repurposing is one of several endpoints of polypharmacological concept that involves identification of new indications for FDA- approved drugs. Developing an approved drug for ward-off complex diseases requires a comprehen- novel indication is both time and cost-effective as sive understanding of systems biology to integrate the safety, toxicity profiles, formulations and phar- all available knowledge and fill gaps as needed. macology of marketed drugs are already estab- Expecting single drugs to cure diseases is therefore lished and found satisfactory by the regulatory rather simplistic and/or unrealistic. agencies including the FDA. Drug repositioning is a financially viable approach that can help resur- Polypharmacology rect not only the bioactive compounds that previ- Polypharmacology is the property, norm rather ously failed clinical endpoints but could help revive than exception, of bioactive small molecules, natu- expired patents. Approximately 9,000 approved or ral products and other chemical species to interact registered drugs have been reported to be available with multiple targets/pathways in biological sys- for repurposing projects13. Drug repurposing has tems. More than half of ‘single-target’ drugs were historically been anecdotal, based on clinical obser- shown to interact with more than five targets. The vations and chance findings. The most cited exam- polypharmacology of molecules has wide implica- ple is the serendipitous repurposing o f sildenafil for tions ranging from predicting harmful off-target erectile dysfunction, a selective phosphodiesterase effects to designing safe and efficacious drug com- 5 inhibitor, originally developed as an anti-hyper- binations or repositioning drugs for new indica- tensive drug. In addition to chance discovery of tions. Shared functional domains and binding sites new uses for old drugs, other strategies for drug across target families, binding promiscuity of min- repurposing include experimental and in silico- imal scaffolds/fragments of chemical species are based approaches. In the phenotypic screening some of the factors underlying polypharmacology. approach, FDA-approved drugs are tested for Identification of all possible interactions between a activity in assays for cell viability, migration, cas- chemical compound and the biological targets is a pase activity, physiologically-relevant muscle cell challenging task and predictions usually employ contractions, electrophysiology, etc. The activity of integrated approaches utilising bioinformatics min- drugs is compared across a panel of cell lines rep- ing of ‘omics’ datasets, molecular docking using x- resentative of various stages of a disease or across

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cells derived from various cancers. The drugs combinations are known to include two or more active in a selective cell or with pan-panel activity active ingredients against the same indication or a are further optimised for activity in combination formulation of multiple distinct active ingredients screens against a set of standard of care drugs used against di stinct targets. The combinations could in clinical settings. The synergistic pair is tested in target crosstalk between pathways that are activat- animal models before it is advanced for further ed or repressed in disease settings. While clinical clinical investigation. In silico drug repositioning observations define combination prescriptions, utilises bioinformatics tools to define interactions experimental quantitative determination of drug between drugs, targets, text and literature mining combination is more complex as the net effects are of public databases. The PheWAS-driven associa- mathematically grouped under additive, synergistic tion between genetic variants and disease can also or antagonistic based on fractional effects at vari- serve as a starting hypothesis for defining new ous concentrati ons of the two drugs14. The in vitro drug-disease linkages. NIH lists 6,500 rare and profiling is further complicated by translation of neglected diseases (R&N) for which only 250 the effects in pharmacokinetic models where the treatments are available (www.ncats.nih.gov/trnd). two drugs may potentiate or reduce therapeutic The drug repositioning approach is a low-hanging efficacy via modulation of individual ADME char- fruit that can be exploited effectively to address the acteristics. A large number of drug combinations low returns on investment and market-size issues curated from 140,000 clinical studies have been of neglected and rare diseases. With the increase in compiled into a Drug Combination database information on genetics and molecular pathways (DCDB; http://www.cls.zju.edu.cn/dcdb/). As with as well as availability of appropriate animal mod- other approaches, profiling drug activity and side- els for rare and neglected diseases, the last decade effects, network crosstalk and modulation help has seen an increase in the number of repurposed design effective new drug combinations. A large drug approvals for R&N disorders. Examples combination study called the Comprehensive include: mitefosine, an anti-cancer alkylphospho- Undermining of Survival Paths or (CUSP9) was choline used for topical treatment of breast cancer rationally designed for the treatment of recurrent metastasis repurposed for the treatment of glioblastoma (GBM)15. The drug combination Leishmaniasis; arbaclofen, for cerebral palsy man- includes nine FDA approved drugs that block the agement, is recommended for GABAergic treat- activity of 17 distinct targets: artesuanate (PI3 ment of Fragile X syndrome; losartan, an anti- kinase, AKT, TLR2, TNF ), disulfiram (ALDH), hypertensive drug repurposed to alleviate the captopril (ACE, AT1, MMP); celecoxib (carbonic symptoms in subset of populatio n with Marfan anhydrases, COX), aprecipitant (NK-1 receptors), syndrome, a connective tissue disorder which auranofin (thioreductase, STAT3, cathepsin B), affects 1 in 5,000, and difluoromethylornithine itraconzole (P-gp efflux pump, BCRP, Hedgehog, 5 (eflornithine, DFMO), an ornithine decarboxylase Lipooxygease), ritonavir (AKT, mTOR, cyclin D3, inhibitor and anti-cancer drug, is being proposed proteasome) and sertraline (TCTP, AKT, mTOR). for the treatment of facial hirsuitism, neuroblas- The side-effects and concentrations of individual toma as well as sleeping sickness. The drug repur- drugs were also carefully evaluated to minimise posing programmes are being accelerated through risks of side-effects while at the same time targeting effective collaborations and partnerships across multiple effectors of the disease. The HIV/AIDS academia and industry. AstraZeneca, while pre- treatment with HAART (Highly Active Anti- serving its patent rights, has provided access of its Retroviral Therapy) effectively combines two- deprioritised compounds to researchers at MRC nucleoside reverse transcriptase inhibitors lamivu- (UK) under the Mechanism of Disease Initiative dine and zidovudine in presence of a protease programme. The NCATS (NIH) in its Therapeutic inhibitor16. This combination was based on clini- Discovery Pilot programme screens selected pro- cal data showing that lamivudine treatment alone jects using 58 compounds from eight top pharma- triggered resistance due to Met 184 Val (M184V) ceutical partnerships in an effort to reveal new mutation in viral reverse transcriptase (RT) but the mechanisms of action and po tential indications. M184V mutant was still sensitive to zidovudine The progress in repositioning is mitigated by com- and a combination of the two RT inhibitors effec- plex regulatory, legal, pricing and patenting issues. tively suppressed the catalytic activity required for HIV replication. While several combination treat- Combination therapy ments are FDA approved, combination therapies Prescribing combinations of drugs for disease man- add to the healthcare costs, require strict adherence agement is a widely employed clinical practice, and to regimens and the possibility of developing new

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References adverse side-effects with time. Regulatory compli- towards a safer and faster road to the discovery and 1 Rask-Andersen, M, Almén, ance also complicates combination synergistic ther- development of life-saving medicines. MS, Schiöth, HB. 2011. Trends apies as well as drug repurposing especially if two An overall analysis of drugs identified from tar- in the exploitation of novel competing pharmaceutical sources are involved. get-based drug discovery has revealed limitations drug targets. Nat Rev Drug Discov 10:579-90. in drug innovation and proteome targeting. 2 Ezkurdia, I, Juan, D, Multi-target drug discovery Despite high R&D expenditure, there is a lack of Rodriguez, JM, Frankish, A, There has been a recent shift in the drug-discovery significant improvement in therapeutic outcomes Diekhans, M et al. 2014. paradigm from single target-single drug to a multi- and overall patient survival. One of the main rea- Multiple evidence strands target drug discovery approach (MTDD) for the sons for limited therapeutic efficacy of current suggest that there may be as few as 19 000 human protein- identification of single compounds with activity therapies is partial understanding of the disease coding genes. Human against two or more targets. The MTDD approach pathophysiology and overall deficiency in devel- Molecular Genetics 23:5866- has several advantages over combination drug oping therapeutics targeting overlapping dysregu- 78. therapies: (A) the pharmacokinetics and safety pro- lated pathways. Complex heterogeneous poly- 3 Hopkins, AL, Groom, CR. files for single multi-target drugs are easier to eval- genic diseases require more genetics-guided, fine- 2002. The druggable genome. Nat Rev Drug Discov 1:727- uate than predicting potential long term adverse ly-tailored treatment regimens to maximise effec- 30. effects developing with combination drugs, and (B) tiveness. The panomics databases serve as valu- 4 Disney, MD, Winkelsas, AM, a multi-target drug is guaranteed to interact with able tools for extracting diverse underlying disease Velagapudi, SP, Southern, M, its targets in the same tissue/cell compartment and, mechanisms, identifying genetic loci in disease and Fallahi, M, Childs-Disney, JL. unlike the combination drugs, will not have ultimately revealing linkages between drugs and 2016. Inforna 2.0: A Platform for the Sequence-Based adverse drug-drug interactions. Both experimental targets that can serve as starting points for design- Design of Small Molecules screening as well as virtual in silico approaches ing holistic approaches. The evolution of a highly Targeting Structured RNAs. deliberately design the screens to identify com- integrated approach with minimal false positives ACS chemical biology. pounds that are active against multiple targets of requires quality control of data being deposited in 5 Childs-Disney, JL, Disney, interest. Several rational-based designs, computa- public databases, optimisation of standard operat- MD. 2016. Approaches to Validate and Manipulate RNA tional-based docking and virtual screening ing protocols for data-mining, integration with Targets with Small Molecules approaches are available for identifying drugs with patient electronic medical records, supportive reg- in Cells. Annual review of mul tiple functions. Several recent examples attest ulatory protocols and collaborative efforts of pharmacology and toxicology to the attractiveness of this approach, including: (i) patient groups, academia and industry in sharing 56:123-40. successful linking of the ABL kinase inhibitory negative/positive datasets. With an increase in the 6 Anuradha, R, Byron, T, Peter, RM, Ashleigh, P, Rathnam, C. domain of imatinib with the hydroxamic acid/ben- number of studies identifying new and rare vari- 2009. Hit-to-Probe-to-Lead zamide (zinc binding) for anti HDAC activity to ants in a wide spectrum of diseases, the process of Optimization Strategies. In design a fusion molecule inhibiting both ABL and inquiry and data extraction is bound to evolve Handbook of Drug Screening, HDACs17, and (ii) identification of a dual into a highly optimised science with increased Second Edition:21-55: CRC kinase/bromodomain inhibitor through virtual deliverables. The selection of the right combina- Press. Number of 21-55 pp. 7 Bunnage, ME, Gilbert, AM, mining of >six million compounds in the E- tion of targets for a disease of interest is critical in Jones, LH, Hett, EC. 2015. molecules, wherein, of the 908 predicted EGFR multi-target drug discovery and requires good Know your target, know your inhibitors, 108 also docked to BRD4 co-crystal understanding of target-disease associations, path- molecule. Nat Chem Biol structures, eight experimentally inhibited the way-target-drug-disease relationships and adverse 11:368-72. BRD4 bromodomain-activity and one that inhibit- events profiling. Systematic repurposing of 8 Waring, MJ, Arrowsmith, J, Leach, AR, Leeson, PD, ed both BRD4 binding as well as EGFR kinase approved drugs has the potential to bring a trans- Mandrell, S et al. 2015. An activity18. formational therapy to regional breakouts of dis- analysis of the attrition of drug eases such as Zika and Ebola in quick order. candidates from four major Holistic insights Finally, drug discovery will be even more holistic pharmaceutical companies. Nat Science drives technology. Invention leads to inno- if it addresses the global health crisis, enabling Rev Drug Discov 14:475-86. 9 Mullard, A. 2015. The vation. Both scenarios are inherently and funda- patient access to medicines and patient care for Roadmap Epigenomics Project mentally intertwined. For the betterment of human- the treatment and management of chronic, infec- opens new drug development ity, it is imperative upon us, the guardians, to see tious, rare and neglected diseases. With the global avenues. Nat Rev Drug Discov that science-driven inventions ultimately lead to resources reforming health education, drug pro- 14:223-5. technology-based innovations. The era of modern duction costs, intellectual/regulatory systems, 10 Anderson, DC, Kodukula, K. 2014. Biomarkers in drug discovery has gone through revolutionary basic science and innovation and a renewed appre- pharmacology and drug advances, as outlined throughout this discussion, ciation of traditional medicines as a rich source of discovery. Biochemical and it is time to coalesce around a broadly defined novel scaffolds, we must embrace and empower a pharmacology 87:172-88. holistic approach, and to enthusiastically adopt a drug discovery paradigm that will truly be holis- community-driven ‘Wikipedia’ or Wa ze’-type open tic19. The time is now for the drug discovery com- Continued on page 55 innovation model to create a crowd-sourced path munity to focus efforts towards the discovery and

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development of multi-target-network polyphar- non-profit think-tank focused on pharmaceutical Continued from page 54 macology drugs exalting symphonic or concert innovation, and most recently, Deputy Site Head, performance with occasional soloists to reignite Center for Advanced Drug Research, SRI 11 Morrison, C. 2016. Fresh from the biotech pharmaceutical innovation. International. He has more than 35 years of expe- pipeline[mdash]2015. Nat rience in academia and industry, managing new Biotech 34:129-32. Acknowledgements lead discovery projects and forging collaborative 12 Chaguturu, R. 2014 (Ed). We thank Drs György Dorman, Target-Ex, and partnerships with academia, disease foundations, Collaborative Innovation in Krishna Kodukula, KK Biopharma, for sharing their non-profits and government agencies. He is the Drug Discovery: Strategies for Public and Private valuable insight on the relevance of ‘natural prod- Founding President of the International Chemical Partnerships. Wiley. ucts’ and ‘biomarkers’, respectively, in guiding drug Biol ogy Society, a Founding Member of the Society 13 Huang, R, Southall, N, discovery efforts. We are also immensely indebted to for Biomolecular Sciences, and Editor-in-Chief of Wang, Y, Yasgar, A, Shinn, P et al. the authors of our forthcoming book Innovative the journal Combinatorial Chemistry and High 2011. The NCGC Approaches in Drug Discovery: Ethnopharmaco- Throughput Screening. Rathnam passionately pharmaceutical collection: a comprehensive resource of logy, Systems Biology and Holistic Targeting, for advocates the need for innovation and clinically approved drugs shaping our ideas; the viewpoints expressed herein entrepreneurship and the virtues of collaborative enabling repurposing and are our own, however. DDW partnerships in addressing the pharmaceutical chemical genomics. Sci Transl innovation crisis, and aggressively warns the threat Med 3:80ps16. of scientific misconduct in biomedica l sciences. He 14 Chou, TC. 2006. Theoretical basis, experimental design, and Dr Anuradha Roy is the Director, High received his PhD with an award-winning thesis computerized simulation of Throughput Screening Laboratory at the from Sri Venkateswara University, Tirupati, India. synergism and antagonism in University of Kansas, Lawrence, Kansas (USA). drug combination studies. She has more than 20 years of academic and Pharmacological reviews biotech experience in executing HTS campaigns, 58:621-81. 15 Kast, RE, Karpel-Massler, G, and has managed drug discovery projects from tar- Halatsch, M-E. 2014. CUSP9* get identification, validation and hit to lead devel- treatment protocol for opment. As director of the HTS core, her responsi- recurrent glioblastoma: bilities include helping academic investigators aprepitant, artesunate, across state universities through the process of auranofin, captopril, celecoxib, disulfiram, itraconazole, developing their target ideas into screen-ready ritonav ir, sertraline augmenting assays for probe identification. Anuradha led vari- continuous low dose ous aspects of drug discovery at PTC Therapeutics temozolomide. Oncotarget after her postdoctoral work at Cleveland Clinic 5:8052-82. Foundation. She holds a doctorate in 16 Staszewski, S. 1995. Zidovudine and lamivudine: Biochemistry/Molecular Biology from Jawaharlal results of phase III studies. Nehru University (India). Journal of acquired immune deficiency syndromes and Professor Bhushan Patwardhan, Interdisciplinary human retrovirology : official School of Health Sciences, Savitribai Phule Pune publication of the International Retrovirology Association 10 University (India), is an internationally recognised Suppl 1:S57. expert on ethnopharmacology and integrative 17 Ganesan, A. 2016. health. He brings a unique blend of industry- Multitarget Drugs: an academia executive culture in advancing evidence- Epigenetic Epiphany. based Ayurveda. Professor Patwardhan is a Fellow ChemMedChem. 18 Allen, BK, Mehta, S, Ember, of National Academy of Medical Sciences (India), SW, Schonbrunn, E, Ayad, N, Founder and Editor-in-Chief of the Journal of Schurer, SC. 2015. Large-Scale Ayurveda and Integrative Medicine, and serves on Computational Screening the editorial boards of several journals. Bhushan is Identifies First in Class a recipient of the Parkhe Award for Industrial Multitarget Inhibitor of EGFR Kinase and BRD4. Scientific Excellence, Dewang Mehta Award for educational reports 5:16924. excellence, and Sir Ram Nath Chopra Oration. He 19 Patwardhan, B, Chaguturu, received his PhD in Biochemistry from Haffkine R. 2016 (Ed). Innovative Institute (Mumbai, India). Approaches in Drug Discovery: Ethnopharmacology, Systems Dr Rathnam Chaguturu is the Founder & CEO of Biology and Holistic Targeting. iDDPartners (Princeton Junction, NJ, USA), a Elsevier Science (in press).

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