Prof Kiran Kalia, Director and Dr Viral Shah, Assistant Professor, Department of Pharmaceuticals, National Institute of Pharmaceutical Education and Research, in this article, highlight the potential of therapeutics for more efficient and patient-centric treatment against varied diseased conditions
Comprehensive understanding of the pathophysiology of various diseases and precise knowledge of molecular and genetic basis has changed the landscape of management of diseases. Until late 20th century, therapeutic basis relied on the use of drugs that had been developed through empirical approaches with no detailed understanding of the molecular mechanisms involved in the pathologic condition. This approach changed at the end of the 20th century, due to the intensification of interest in the clinical management of diseases, new treatment modalities and potential impact of personalised medicine. A wave of scientific advances and new technologies dramatically changed the discovery mode of new drugs. Greater knowledge of the genetic and molecular level of diseases has allowed researchers to invade new targets for therapy and successfully predict how certain biopharma will affect specific sub populations of patients. The breakthrough innovation of therapeutic biologics to address largely unmet medical needs in varied disease conditions including cancer, diabetes, haemophilia, and immunological impairment made bio-industry as one of the strategic emerging industries (SEI) .
In contrast to small molecular weight drugs, the biologicals as defined in Section 351 of the Public Health Service (PHS) Act are “virus, therapeutic serum, toxin, antitoxin, vaccine, blood, blood component or derivative, allergenic product, or analogous product, applicable to the prevention, treatment, or cure of a disease or condition of human beings.” FDA regulations and policies have established that biological products include blood-derived products, vaccines, in vivo diagnostic allergenic products, immunoglobulin products, products containing cells or microorganisms, and other protein products. 
The categories of therapeutic biological products regulated by Centre for Drug Evaluation and Research (CDER) includes
- Monoclonal antibodies for in vivo use.
- Plant, animal, human, or microorganisms derived therapeutic proteins recombinant versions of these products.
- Immunomodulators (non-vaccine and non-allergenic products intended to treat disease by inhibiting or down-regulating a pre-existing, pathological immune response).
- Monoclonal antibodies, cytokines, and growth factors intended to mobilise, stimulate, decrease or otherwise alter the production of haematopoietic cells in vivo. 
As per the US Food and Drug Administration (FDA), biologicals, represent the cutting-edge of biomedical research and latest scientific discoveries translated into novel therapies. They provide new treatment options for patients and eventually serve as the most effective therapeutics for a variety of medical illnesses and conditions where other treatments are either not available or are most challenging and costly . It is estimated that by 2016, 48 per cent of the top 100 best-selling drugs would consist of biological medicines .
The human genome inside every human body consists of about 25,000 genes, each responsible for a specific protein including various enzymes, hormones, antibodies and other proteins needed to make the body function. The defective and missing genes in the body will not have the proteins for proper functioning and may have many proteins that cause disease. Amalgamation of biotechnology with modern tools of computer technology used to harness scientific progress have helped biopharma scientists to analyse the massive amount of data quickly, determining which genes or proteins are defective and which proteins are being used for treatments across a range of therapeutic areas thereby pushing the frontiers of science.
Currently biologics represent the 30 per cent of licensed products and are the world’s best-selling pharma as depicted in Table 1. The biologics have fewer adverse effects compared to small molecules due to their high affinity for targets. 
The improved manufacturing efficiency, increasing access to biologicals and reducing costs is the need of the hour. Most importantly, with FDA outlining its latest guidance for the future of biological medicines, patient safety must be the primary focus of all stakeholders . In the future, biological therapies are likely to be used more selectively based on personalised benefit/ risk assessment, determined through reliable biomarkers and tissue signatures.
Key challenges of therapeutic biologics industry
Basic difference between therapeutic biological and small drug molecules is depicted in Table 2.
Developing novel biologics is the complex process and requires a thorough understanding of disease state, immune system, target site of immunomodulation and the biologic itself. Due to the complexity in its therapeutic structure biologicals poses several challenges in its development as summarised below:
Multisite manufacture – the hidden challenge of comparability
Logistical hurdles are associated with limited time factor available for transporting harvested patient donor cells to the manufacturing site and their return to the clinical site for administration to the patient. In many cases, product developers are bound to use contract manufacturers where the environmental requirements are usually compliant with GMP. These raised difficulties in manufacturing continuity desired during later phases of development  and makes the transition to commercial-scale production of autologous cell therapies, prohibitively expensive or even infeasible. 
- Pharmacokinetic studies of therapeutic biological: Pharmacokinetic studies are routinely carried out for small therapeutic molecules, whereas for biologics experimental analysis to predetermines underlying mechanisms for clearance and bio-distribution or identify factors that regulate absorption, distribution, metabolism, and excretion (ADME) cannot be determined thoroughly. New technologies to deliver therapeutic biologics using needle-free devices, a formulation with nanoparticles, intranasal and ocular delivery and novel therapeutic approaches such as stem cell therapy and individualised medicine are continually emerging. These advances limit the in vivo disposition assessment of biologics. Recently, unusual PK profiles of mAbs have been reported for many mAbs. [12,13] The mechanistic-based ADME studies are often required to understand possible causes for such unusual PK profiles. Despite of the great clinical and commercial success for some biologic drugs, the rate of clinical success is still a matter of ambiguity.  Recently, significant research efforts have been focused on understanding the correlation between PK, especially drug concentrations at the target site, and pharmacodynamics (PD) to improve clinical trial outcomes.  Three key questions should addressed to select a drug candidate for clinical trials: (1) What is the probability of the compound to reach the target organ(s) at a therapeutic concentration (2) Would the compound bind to the target(s) in vivo and result in required biological activity? (3) Would the compound exert the functional modulation of the target? In-depth investigations of ADME properties and relationship between ADME profiles and pharmacological effects (e.g., biomarker activity) in preclinical and clinical studies would accurately answer above-mentioned questions.
Consideration of other factor for ADME studies of biologics like species difference in target-binding properties and host immune response to a biologic should also be given equal weightage. It is a well established fact that antidrug–antibody mediated clearance for therapeutic biologics is species-dependent. The prediction of ADME profiles of therapeutic biologics in humans may not be relevant based on common animals eg. rodents and dogs. The transgenic animals that express human targets or receptors are useful for the qualitative assessment of potential clearance mechanisms.
- Analytical assay development techniques: Structure similarity between therapeutic biologics and endogenous proteins along with the hydrophilic nature of biologics limits its qualitative and quantitative analytical estimation which is to be followed after extraction and purification of these drugs from in vivo samples. The detection limit of analytical instruments makes the direct measurement of therapeutic proteins in biologic matrices a challenging task. Enzymatic digestion required before qualitative and quantitative analysis of these proteins in vivo samples, limits intact recovery of therapeutic biologics during sample treatment. Interference of endogenous proteins preset in in vivo samples is also a major concern during quantitative estimation of biologics. However, conventional analytical approaches for pharmacokinetic studies of biologics like ligand binding assays, such as the enzyme-linked immunosorbent assay (ELISA) include high specificity and sensitivity, ease of sample handling, and low cost.[18,19] But developing high quality and specific reagents for ELISA methods to quantify therapeutic biologics and antidrug antibodies (ADAs) could be a time-consuming and labour-intensive process. Common imaging technologies are used in in vivo drug bio-distribution studies such as optical imaging with luminescence and fluorescence molecular probes, radiotracer-based single photon emission computed tomography (SPECT) and positron emission tomography (PET) imaging, magnetic resonance imaging (MRI), ultrasound imaging, and X-ray imaging. [20,21] In contrast to the conventional cut-and-count approach or whole-body autoradiography with radiolabeled biologics, imaging studies can be conducted in live animals, which provides the advantage of obtaining real-time dynamics on the biodistribution. 
- Suitable drug delivery systems: With only a few exceptions, almost majority of therapeutic biologics are administered by parenteral routes, which usually lacks patient compliance. New drug delivery systems are need to be explored to increase target-specific delivery, improve safety and convenience for patients, and support for individualised medication. Since biologicals tend to denaturise in harsh gastric environment conditions and also undergo enzymatic degradation so the current need is to develop a efficient dosage form for the oral delivery of such biologicals with high therapeutic efficiency. Some examples of novel strategies investigated for delivery of biologics include polymeric micro-structured arrays for potential delivery of proteins , needle-free injection devices for SC injection,  and nanoparticulate systems. 
- Pharmacovigilance of biologics: Pharmacovigilance is basically checkpoint studies for determining drug’s performance, particularly adverse reactions, after it has been released in market. A variety of natural sources—human, animal or microorganisms are used for obtaining biological so a robust and vigilant traceability system should be in place to facilitate accurate surveillance of marketed therapeutic bilogics. 
The regulatory guidelines for biologics address the manufacturing process and quality aspects, the pre-market regulatory requirements for quality, preclinical and clinical studies and post-market regulatory requirements for similar biologics.
The competent authorities, Central Drugs Standard Control Organization (CDSCO) and the Department of Biotechnology (DBT) are involved in the approval process for biologics and lay down the regulatory pathway for a similar biologic claiming to be similar to an already authorised reference biologic. The Review Committee on Genetic Manipulation (RCGM) functions in the Department of Biotechnology (DBT), Ministry of Science and Technology, Government of India. In the context of similar biologics, Genetic Engineering Appraisal Committee (GEAC) is placed under the Ministry of Environment and Forests (MoEF) as statutory body for review and approval of activities involving large-scale use of genetically engineered organisms and products thereof in research and development, industrial production, environmental release and field applications. CDSCO, responsible for the approval of new drugs, headed by the Drug Controller General (India) DCG(I) is the apex regulatory body under Ministry of Health & Family Welfare (MoHFW), Government of India is also responsible for grant of import/ export license, clinical trial approval and permission for marketing and manufacturing of similar biologics. State Food and Drug Administration (FDA) works with CDSCO in each state and is responsible for issuance of license to manufacture similar biologics in India. Stringent regulatory guidelines as depicted in Table 4 at early stage of developments of new therapeutic biologics creates a hurdle in the growth and development of biologic industry .[28,29]
Most biologics are very complex molecules and are difficult to be fully characterised by existing science. For this reason, they often are characterised by their manufacturing processes. The manufacturing processes are likewise very sensitive and complex. Temperature fluctuations or other factors can impact the final product and affect their efficacy. Changes in the manufacturing process or facility may require clinical studies to demonstrate safety, purity and potency. Batch to batch variability, time length for creating cell lines are some other factors that are needed to be addressed.
With the increase in the number of approved therapeutic biologics, potential drug–drug interactions (DDI) between concurrently administered drugs (small molecule–biologic drugs or biologic–biologics drugs) have become a major concern. The importance of therapeutic protein–drug interactions and guidance that recommends how and when to evaluate such interactions have been published recently by FDA. [31,32] However, preclinical tools and in vitro test systems for assessing drug interaction potential of therapeutic proteins are limited. Thus, DDI assessment is often evaluated as a part of clinical trials.
Blood–brain barrier (BBB) transport
BBB transport for small molecules has been extensively studied which has not been fully explored for biologics. Knowledge of the mechanism of BBB transport for biologics will benefit the safety assessment and the drug discovery for central nervous system (CNS) indications for therapeutic biologics. 
Placental transport of therapeutic biologics
Placental transport of biologic drugs is not studied extensively, because of the common belief that the placental transport is minimal for biologics. Whereas, the placental transport of immunoglobulins has been recognised for more than 50 years .[34,35] A specific FcRn-mediated binding of IgG at the maternal surface of the placenta has been proposed as the first step in the transport mechanism by which IgG is transferred from the mother to the foetus. [36,37] The challenge of drug concentration measurements for foetal tissues limits placental transport studies.
The cost for development, validation, and implementation of these new technologies is significant. Thus cost also serve as a major limiting factor in the growth of bio industry.
Opportunities and growth potential of therapeutic biologics industry
- Value to patients: Therapeutic biologics are leading towards a paradigm shift by significant unmet medical needs for many critical diseases. From recombinant human insulin to interferons to monoclonal antibodies (mAb), biologics have saved patients’ lives and improved quality of life for patients suffering from diabetes, infectious diseases, haemophilia, and cancer.
- Value to industry: Tremendous and noteworthy growth is observed in the global market for therapeutic biological. Bio industries are expected to be a key growth driver in upcoming decade for the global pharma industry.
- Value to the economy: Therapeutic biologics, along with other biotech products in the fields of agriculture and energy, constitute the broader bio-industry that is expected to grow fastest annually.  With the right enabling policies, the broader bio-industry could become one of worlds pillar industries by 2020. The bio-industry if explored correctly will help India to transform from a labour-intensive economy to a knowledge-intensive economy and contribute to the sustainability of future economic growth.
- Biosimilars: The introduction of many biologic therapies to the market has revolutionised the area of therapeutics. However, the patents protecting their manufacture are about to lapse, and follow-on generic products, known as biosimilars, are about to flood the pharma market. The ‘biosimilars’ are different from the generic products of conventional drugs concerning efficacy, safety, and immunogenicity. Biosimilars have attracted great attention with the hope that they will allow wide-spread availability of currently expensive biologic products. Given their structural complexity, multifaceted manufacturing process, and, as a result, challenges for predicting impact of the manufacturing process changes on immunogenicity, biosimilars are not generic alternatives per se and are not interchangeable. Thus, unique regulatory pathways are required for biosimilars. PK and immunogenicity assessments, together with efficacy and toxicology studies, are key components in the development of biosimilars.
- Personalised medicines: The age of ‘personalised medicine’ is predicated to administer the right medicine to the right patient in the right dose at the right time. The cost-centric overshadow the practice of patient-centric medicine, there is a potential negative impact on both patient safety and clinical effectiveness. The future of biological medicines will be bright if patients, physicians, biotechnology companies, and other stakeholders work together to ensure patient safety, which is the foremost priority of the biosimilar policy discussion, likely because of the common belief that the placental transport is minimal for biologics. However, placental transport of immunoglobulins has been recognised for more than 50 years.[34,35] A specific FcRn-mediated binding of IgG at the maternal surface of the placenta has been proposed as the first step in the transport mechanism by which IgG is transferred from the mother to the foetus. [36,37] The role of the second placental barrier, the foetal capillary endothelium, is not yet clear. The challenge of drug concentration measurements for foetal tissues limits placental transport studies. Additional research is required to understand fully the mechanism of placental transfer of various biological modalities and its implications for the safety assessment of therapeutic biologics.
1. Lee YS, Tee YC & Kim DW. Endogenous versus exogenous development:a comparative study of biotechnology industry cluster policies in South Korea and Singapore. Environment and Planning C: Government and Policy 27, 612-631 (2009).
2. Nelson AL, Dhimolea E, Reichert JM. Development trends for human monoclonal antibody therapeutics. Nat. Rev. Drug Discov. 2010;9:767–774.
3. Verdine GL, Hilinski GJ. Stapled peptides for intracellular drug targets. Methods Enzymol. 2012;503:3–33.
4. Gillies SD, Lan Y, Brunkhorst B, Wong WK, Li Y, Lo KM. Bi-functional cytokine fusion proteins for gene therapy and antibody-targeted treatment of cancer. Cancer Immunol, immunother: CII. 2002;51:449–460.
5. Chow SC, Ju C. Assessing biosimilarity and interchangeability of biosimilar products under the Biologics Price Competition and Innovation Act. Generics and Biosimilars Initiative Journal (GaBI Journal). 2013;2(1).
6. Vugmeyster Y, Szklut P, Wensel D, Ross J, Xu X, Awwad M, et al. Complex pharmacokinetics of a humanized antibody against human amyloid beta peptide, anti-abeta Ab2, in nonclinical species. Pharm Res. 2011;28:1696–1706.
7. Vugmeyster Y, Xu X, Theil FP, Khawli L, leach MW. Pharmacokinetics and toxicology of therapeutic proteins: advances and challenges. World J Biol Chem. 2012;3:73–92.
8. Keizer RJ, Huitema ADR, Schellens JHM, Beijnen JH. Clinical pharmacokinetics of the rapeutic monoclonal antibodies. Clin Pharmacokinet. 2010;49:493–507.
9. Wu S, Lourette NM, Tolic N, Zhao R, Robinson EW, Tolmachev AV, et al. An integrated top-down and bottom-up strategy for broadly characterizing protein isoforms and modifications. J Proteome Res. 2009;8:1347–1357.
10. Chirino AJ, Mire-Sluis A. Characterizing biological products and assessing comparability following manufacturing changes. Nat Biotechnol. 2004;22:1383–1391.
11. Lin JH. Pharmacokinetics of biotech drugs: peptides, proteins and monoclonal antibodies. Curr Drug Metabol. 2009;10:661–691.
12. Wang W, Lu P, Fang Y, Hamuro L, Pittman T, Carr B, et al. Monoclonal antibodies with identical Fc sequences can bind to FcRn differentially with pharmacokinetic consequences. Drug Metabol Dispos: Biol Fate Chem. 2011;39:1469–1477.
13. Yeung YA, Leabman MK, Marvin JS, Qiu J, Adams CW, Lien S, et al. Engineering human IgG1 affinity to human neonatal Fc receptor: impact of affinity improvement on pharmacokinetics in primates. J Immunol. 2009;182:7663–7671.
14. Herbertson RA, Tebbutt NC, Lee FT, MacFarlane DJ, Chappell B, Micallef N, et al. Phase I biodistribution and pharmacokinetic study of Lewis Y-targeting immunoconjugate CMD-193 in patients with advanced epithelial cancers. Clin Cancer Res. 2009;15:6709–6715.
15. Lee JI, Zhang L, Men AY, Kenna LA, Huang SM. CYP-mediated therapeutic protein–drug interactions: clinical findings, proposed mechanisms and regulatory implications. Clin Pharmacokinet. 2010;49:295–310.
16. Girish S, Martin SW, Peterson MC, Zhang LK, Zhao H, Balthasar J, et al. AAPS workshop report: strategies to address therapeutic protein–drug interactions during clinical development. AAPS J. 2011;13:405–416.
17. Roman J, Qiu J, Dornadula G, Hamuro L, Bakhtiar R, Verch T. Application of miniaturized immunoassays to discovery pharmacokinetic bioanalysis. J Pharmacol Toxicol Methods. 2011;63:227–235.
18. Ezan E, Dubois M, Becher F. Bioanalysis of recombinant proteins and antibodies by mass spectrometry. Analyst. 2009;134:825–834.
19. Li F, Fast D, Michael S. Absolute quantitation of protein therapeutics in biological matrices by enzymatic digestion and LC-MS. Bioanalysis. 2011;3:2459–2480.
20. Khalil MM, Tremoleda JL, Bayomy TB, Gsell W. Molecular SPECT imaging: an overview. International J Mol Imaging. 2011;24;43-52.
21. Von Schulthess GK, Schlemmer HP. A look ahead: PET/MR versus PET/CT. Eur J Nucl Med Mol Imaging. 2009;36(Suppl 1):S3–S9.
22. Vugmeyster Y, DeFranco D, Szklut P, Wang Q, Xu X. Biodistribution of [125I]-labeled therapeutic proteins: application in protein drug development beyond oncology. J Pharm Sci. 2010;99:1028–1045.
23. Park J-H, Davis S, Yoon Y-K, Prausnitz MR, Allen MG (eds). Micromachined biodegradable microstructures. Proc IEEE Microelectromechanical Systems Conference. 2003; 371–4.
24. Wang MM, Defranco D, Wright K, Quazi S, Spencer-Pierce J, et al. Decreased exposure of Peptide X in Zn formulation after subcutaneous dosing and in vitro metabolism in skin. AAPS J. 2011;13(S1):M1086.
25. Shi J, Votruba AR, Farokhzad OC, Langer R. Nanotechnology in drug delivery and tissue engineering: from discovery to applications. Nano Lett. 2010;10:3223–3230.
26. World Health Organization [homepage on the Internet]. Pharmacovigilance. [cited 2013 Mar 4]. Available from: http://www.who.int/medicines/areas/quality_safety/safety_efficacy/pharmvigi/en
27. Van der Laan JW, Brightwell J, McAnulty P, Ratky J, Stark C. Regulatory acceptability of the minipig in the development of pharmaceuticals, chemicals and other products. J Pharmacol Toxicol Methods. 2010;62:184–195.
28. Minghetti P, Rocco P, Cilurzo F, Del Vecchio L, Locatelli F. The regulatory framework of biosimilars in the European Union. Drug Discov Today. 2012;17:63–70.
29.GaBI Online – Generics and Biosimilars Initiative. FDA’s public hearing on biosimilars draft guidances [www.gabionline.net]. Mol, Belgium: Pro Pharma Communications International; [cited 2013 Mar 4]. Available from: www.gabionline.net/Biosimilars/General/FDA-s-public-hearing-on-biosimilars-draft-guidances
30.Wittrup KD, Thurber GM, Schmidt MM, Rhoden JJ. Practical theoretic guidance for the design of tumor-targeting agents. Methods Enzymol. 2012;503:255–268
31. Ponce R, Abad L, Amaravadi L, Gelzleichter T, Gore E, Green J, et al. Immunogenicity of biologically-derived therapeutics: assessment and interpretation of nonclinical safety studies. Regulatory Toxicol Pharmacol: RTP. 2009;54:164–182
32. De Groot AS, McMurry J, Moise L. Prediction of immunogenicity: in silico paradigms, ex vivo and in vivo correlates. Curr Opin Pharmacol. 2008;8:620–626.
33. Pardridge WM, Boado RJ. Reengineering biopharmaceuticals for targeted delivery across the blood–brain barrier. Methods Enzymol. 2012;503:269–292.
34. Pardridge WM. Biologic TNFalpha-inhibitors that cross the human blood–brain barrier. Bioeng Bugs. 2010;1:231–234.
35. Kane SV, Acquah LA. Placental transport of immunoglobulins: a clinical review for gastroenterologists who prescribe therapeutic monoclonal antibodies to women during conception and pregnancy. Am J Gastroenterol. 2009;104:228–233.
36. Chaparro M, Gisbert JP. Transplacental transfer of immunosuppressants and biologics used for the treatment of inflammatory bowel disease. Curr Pharm Biotechnol. 2011;12:765–773.
37. Pentsuk N, Van der Laan JW. An interspecies comparison of placental antibody transfer: new insights into developmental toxicity testing of monoclonal antibodies. Birth Defects Res Part B, Dev Reprod Toxicol. 2009;86:328–344.
38. Declerck PJ, Simoens S. A European perspective on the market accessibility of biosimilars. Biosimilars. 2012;2:33-40.
39. Declerck PJ. Biologicals and biosimilars: a review of the science and its implications. Generics and Biosimilars Initiative Journal (GaBI Journal). 2012;1(1):13-6.
40. Dranitsaris G, Amir E, Dorward K. Biosimilars of biological drug therapies: regulatory, clinical and commercial considerations. Drugs. 2011;71:1527–1536.
41. Class JN, Langis L. A patient-centered paradigm for the biosimilars market. Generics and Biosimilars Initiative Journal (GaBI Journal). 2012;1(1):17-21.