Biopharmaceutical Industry Capability Building in India: Report from a Symposium.

The biopharmaceutical industry is evolving with a shift in focus from recombinant proteins and antibodies towards more complex cell and gene therapies. To be competitive globally, biomanufacturers need to focus on aligning with global standards with regard to drug quality, reducing manufacturing failures and delivering drugs to market quickly. Building these capabilities requires a multifaceted approach that includes improvements in operations, quality compliance, and control strategies. To address these needs, the US Pharmacopeia (USP), the Department of Biotechnology (DBT) India, and the Confederation of Indian Industry (CII) held a symposium to discuss the requirements and gaps in the biotechnology and pharmaceutical sectors in India and other developing countries. A panel of experts from academia, manufacturing, and governmental agencies identified several drivers needed for capability building, including a skilled workforce, public-private partnerships, advanced manufacturing technologies, novel biologics, and favorable policies. This article summarizes the recommendations put forward by this panel.

Building trust in the quality of vaccines.

To date, several COVID-19 vaccines have been authorized for the voluntary immunization of adults. The quick availability of multiple vaccines is a good strategy to achieve herd immunity during a pandemic. However, the fast-track development of vaccines during this pandemic has raised concerns regarding the quality, safety, and effectiveness of vaccines. In response, USP organized a roundtable to discuss challenges and to solicit input on ways to build trust in vaccines. Key discussion points included manufacturing capacity, availability of a skilled workforce, and investment in new technologies that would enable the safety and quality of vaccine products. There was also a consensus that a rigorous and transparent clinical trial design is essential for understanding the safety and effectiveness of vaccines.

Recommendations for Enhancing Quality and Capability of Indian Biopharmaceutical Industry: Summary of a Workshop.

The biopharmaceutical industry is undergoing an evolutionary phase with the rise of advanced manufacturing technologies. The regulatory and customer requirements are shifting towards the development of personalized or targeted medicines. With this changing landscape, industry must evaluate the relevance of quality management systems. Over the past two decades, Indian companies have played a significant role in generating access and reducing costs of medicines. The quality management systems that enable the development and manufacturing of biopharmaceuticals require companies to adapt to regulatory requirements of process development, clinical trials, production, and life cycle management. To better understand the status and potential opportunities to enhance the quality management systems of manufacturing biopharmaceuticals, a workshop was organized by United States Pharmacopeia (USP) and Association of Biotechnology Led Enterprises (ABLE). This paper summarizes the recommendations by the panel and participants of the workshop to industry stakeholders, governance bodies, and policymakers. Following points were proposed to strengthen the culture of quality processes in Indian biopharmaceutical industry: i) Inculcating a culture of quality; ii) Effective training programs on quality processes; iii) Focus on quality beyond compliance; iv) Focus on automation and digitization. v) Enhance processes for pharmacovigilance and product life cycle management. vi) Understanding global regulatory processes.

Novel pyrazole-3-carboxamide derivatives as cannabinoid-1 (CB1) antagonists: journey from non-polar to polar amides.

The synthesis and biological evaluation of novel pyrazole-3-carboxamide derivatives as CB1 antagonists are described. As a part of eastern amide SAR, various chemically diverse motifs were introduced. In general, a range of modifications were well tolerated. Several molecules with high polar surface area were also identified as potent CB1 receptor antagonists. The in vivo proof of principle for weight loss is exemplified with a lead compound from this series.

Pharmacotherapy of obesity: emerging drugs and targets.

BACKGROUND: Obesity and its associated morbidities are the effects of imbalance between energy intake and expenditure. Present drugs either regulate food intake by acting on neural circuits or reduce nutrient absorption from gut. These approaches have shown moderate success, with several safety concerns, leaving an unmet need for effective and safe therapy for obesity. OBJECTIVE: To provide a brief background on obesity, summarize approved drugs and give an overview of emerging therapeutic targets, their potential benefits and disadvantages. METHODS: A review based on information available from medical literature. CONCLUSIONS: Potential anti-obesity targets investigated can be classified into five broad categories: i) decreasing appetite through central action; ii) increasing metabolic rate or affecting metabolism through peripheral action; iii) modulating gut peptide receptors; iv) modulating targets to affect overall cardiometabolic parameters; and v) combination therapies directed against several targets.

Advances in antithrombotic agents.

Thrombosis is the condition where an imbalance in the homeostatic mechanism results in unwanted intravascular thrombus formation. Imbalances in this highly regulated process of coagulation and anticoagulation can lead to a variety of pathophysiological conditions leading to stroke, pulmonary heart attack and other serious conditions. In the western world, thromboembolic diseases are the leading cause of morbidity and mortality. Remarkable progress has occurred over the last decade in the development of antithrombotic drugs, which can be classified into 3 major categories - Anticoagulants, Antiplatelets and thrombolytics. Increased understanding of the pathobiology of thrombotic and vascular disorders has helped researchers to target novel pathways involving the coagulation, thrombolytic, fibrinolytic and integrin systems. Traditionally aspirin and unfractionated heparin was used for myocardial infarction. Newer antiplatelet agents such as, clopidogrel, GP IIb/IIIa inhibitors, low molecular weight heparin, direct thrombin inhibitors and several improved thrombolytic agents have been introduced for clinical use. This review will discuss different important drugs, which have been launched in recent years and also some new targets pursued by different companies.

The role of AMP kinase in diabetes.

Type 2 diabetes is characterized by abnormal metabolism of glucose and fat, due in part to resistance to the actions of insulin in peripheral tissues. If untreated it leads to several complications such as blindness, kidney failure, neuropathy and amputations. The benefit of exercise in diabetic patients is well known and recent research indicates that AMP activated protein kinase (AMPK) plays a major role in this exercise related effect. AMPK is considered as a master switch regulating glucose and lipid metabolism. The AMPK is an enzyme that works as a fuel gauge, being activated in conditions of high energy phosphate depletion. AMPK is also activated robustly by skeletal muscle contraction and myocardial ischaemia, and is involved in the stimulation of glucose transport and fatty acid oxidation produced by these stimuli. In liver, activation of AMPK results in enhanced fatty acid oxidation and decreased production of glucose, cholesterol, and triglycerides. The two leading diabetic drugs namely, metformin and rosiglitazone, show their metabolic effects partially through AMPK. These data, along with evidence from studies showing that chemical activation of AMPK in vivo with 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR) improves blood glucose concentrations and lipid profiles, make this enzyme an attractive pharmacological target for the treatment of type 2 diabetes and other metabolic disorders.

Synthesis, in vitro and in silico evaluation of l-tyrosine containing PPARalpha/gamma dual agonists.

A novel series of l-tyrosine derivatives have been reported with potential PPARalpha/gamma dual agonistic activity. In vitro cell based PPARalpha/gamma transactivation studies have shown compound 4a and compound 4f to be the most potent PPARgamma and PPARalpha activators, respectively. Molecular docking studies performed on these series of compounds have complemented the experimental results and have led to interesting inferences.

Synthesis and evaluation of N-acetyl-L-tyrosine based compounds as PPARalpha selective activators.

The development of type 2 diabetes in obese individuals is linked to lipid accumulation in non-adipose tissues. A series of N-acetyl-L-tyrosine derivatives were synthesized and evaluated for PPAR transactivation. Compounds 4d and 4f were found to show better PPARalpha transactivation as compared to PPARgamma. Molecular docking analysis was carried out to study their important interactions with the active site of PPARalpha.

Biochemical mechanism of insulin sensitization, lipid modulation and anti-atherogenic potential of PPAR alpha/gamma dual agonist: Ragaglitazar.

The current goal in the treatment of diabetes is not only to enhance the glycemic control but also to improve the associated cardiovascular risk factors. Among many of the strategies available, a co-ligand of PPARalpha and gamma in a single molecule which combines the insulin sensitizing potential of PPARgamma and the beneficial lipid modulating properties of PPARalpha agonism, has gained attention in the recent past. Here we report the biochemical mechanism by which a dual PPAR alpha/gamma agonist Ragaglitazar (Raga) achieves this goal. The PPARalpha component of Raga appears to contribute to a significant increase in beta oxidation, ApoA1 secretion and inhibition of TG biosynthesis in HepG2 cells. These effects of Raga at 60 microM were similar to that shown by Fenofibrate (Feno) at 250 microM. The PPARgamma component of Raga showed significant G3PDH activity and TG accumulation with a corresponding increase in aP2 expression in 3T3L1 cells. Significantly reduced levels of IL-6 and TNFalpha were observed in the culture supernatants of Raga treated 3T3L1 cells. Raga resulted in significant insulin dependent glucose uptake in 3T3L1 with a corresponding increase in GLUT4 expression. Further, Raga showed a significant cholesterol efflux with a corresponding increase in ABCA1 protein expression in THP-1 macrophages. In conclusion, Raga activates both PPARalpha and gamma regulated pathway in adipocytes as well as in hepatocytes which together contributes for its insulin sensitizing and lipid lowering activity. In addition the dual activation of PPAR alpha/gamma also shows an athero-protective potential by inducing reverse cholesterol efflux and inhibiting the pro-inflammatory cytokines.

Antihypertensive effect of ragaglitazar: a novel PPARalpha and gamma dual activator.

Ragaglitazar is a novel and potent dual peroxisome proliferators activated receptor (PPAR) alpha and gamma activator. The aim of this study is to investigate the effect of ragaglitazar on blood pressure and endothelial function in insulin resistant animal model and non-insulin resistant hypertensive models. The effects ragaglitazar were tested in Zucker fa/fa, spontaneously hypertensive rats (SHR), 2 kidney 1clip rat (2K1C) and Wistar Kyoto rats (WKY). Pioglitazone was taken as a comparative standard. Ragaglitazar showed significant reduction (P<0.001) of systolic blood pressure (SBP) in insulin resistant fa/fa rats, with concomitant reduction in plasma triglycerides (TG) and insulin levels while pioglitazone (10 mg kg(-1)) showed significant (P<0.05) but comparatively less reduction. Ragaglitazar in contrast to pioglitazone showed significant reduction (P<0.05) of SBP in SHR, 2K1C while the same dose did not have any effect on normotensive WKY. Ragaglitazar also showed significant improvement in acetylcholine-induced relaxation in isolated aorta of Zucker fa/fa, SHR, 2K1C and also potentiated the insulin-induced vasorelaxation in Zucker fa/fa rats. These findings summarize that ragaglitazar shows significant reduction of BP and improvement in endothelial function not only in insulin resistant but also in non-insulin resistant hypertensive models where standard thiazolidinediones are ineffective. These data indicates that dual PPARalpha and gamma activator ragaglitazar can be beneficial for the treatment of hypertension and vascular disease commonly associated with type 2 diabetes.

Antiobesity therapy: emerging drugs and targets.

Obesity and its associated morbidities and mortalities are the effects of imbalance between energy intake and expenditure. The healthcare burden for the treatment of obesity is significantly high, due to increased risk of secondary chronic diseases such as hypertension and associated co-morbidities such as diabetes and cardiovascular disease. Lack of physical activity, high fat diets and sedentary life styles are major factors contributing to obesity. However, genetic predisposition and ethnicity are increasingly found to cause obesity. Till date, approved therapeutics have addressed excess energy intake by acting on central neural circuits that regulate feeding or on peripheral mechanisms to reduce nutrient absorption from the gut. These approaches have met with moderate success; and recently with safety concerns, leaving an unmet medical need for effective and safe pharmacotherapy for obesity thereby posing a significant challenge to pharmaceutical industry. Potential antiobesity drugs, which are being investigated by different companies, can be classified in 4 broad categories: 1) Agents that primarily decrease appetite through central action; 2) Agents that primarily increase metabolic rate or affect metabolism through peripheral action; 3) Agents that act on gastrointestinal tract; and 4) Agents that not only affect obesity but also overall Metabolic Syndrome. The current review will deal mainly with different molecules, which are under development for the above-mentioned targets and also their potential benefits and disadvantages.

Dual PPAR-alpha and -gamma activators derived from novel benzoxazinone containing thiazolidinediones having antidiabetic and hypolipidemic potential.

2,4-Thiazolidinedione derivatives of 1,3-benzoxazinone were synthesized and evaluated for their PPAR-alpha and -gamma dual activation. DRF-2519, a compound obtained through SAR of TZD derivatives of benzoxazinone, has shown potent dual PPAR activation. In ob/ob mice, it showed better efficacy than the comparator molecules. In fat fed rat model, it showed significant improvement in lipid parameters, which was better than fibrates.

Insulin sensitizing property of Indigofera mysorensis extract.

Indigofera mysorens is a shrub used for its antidiabetic activity in rural India. Here, we elucidate the antidiabetic potential of Indigofera mysorensis extract. Ethanolic extract of the whole shrub of Indigofera (EEI) at 300 mg/kg for 10 days, produced a 63% reduction in plasma glucose, 41% reduction in plasma triglyceride and 77% reduction in plasma insulin levels in insulin resistant db/db mice, which is better than insulin sensitizer, troglitazone (400 mg/kg). EEI unlike sulphonylureas failed to show any acute hypoglycemic effect in normoglycemic Swiss albino mice (SAM). Even in a chronic study (10 days) in SAM, EEI (300 mg/kg) like insulin sensitizers showed no effect on plasma glucose, but an 81% reduction in plasma insulin levels. When challenged with 3 gm/kg sucrose, SAM treated with EEI (300 mg/kg, 7 days) failed to show any effect on the absorption of sugar, whereas standard drug, acarbose (10 mg/kg) showed 52% reduction in the area under the plasma glucose curve. EEI failed to show any significant transactivation of PPARgamma, a proposed target of synthetic insulin sensitizers. Taken together, our data indicate that the antidiabetic effect of the ethanolic extract of Indigofera is due to its insulin sensitizing property and is clearly different from that of sulfonylurea or acarbose.

Role of PPAR in cardiovascular diseases.

Cardiovascular disease (CVD) is the most critical global health threat, which contributes more than one third of global morbidity. CVD includes heart disease, vascular disease, atherosclerosis, stroke and hypertension. The most important independent risk factors for CVD include dyslipidemia along with hypertension, obesity, sedentary lifestyle, diabetes and chronic inflammation. These factors are directly regulated by diet, metabolism and physical activity. Diets rich in fat and carbohydrate coupled to sedentary lifestyles have contributed to the increase in dyslipidemia, type 2 diabetes, obesity and CVD in the world. Discovery of Peroxisome Proliferator Activated Receptors (PPARs) as a key regulator of metabolic pathways has led to significant insight into the mechanisms regulating these processes. Three PPAR subtypes, encoded by distinct genes, are designated as PPAR-alpha, PPAR-delta (also know as beta) and PPAR-gamma. PPARs act as nutritional sensors that regulate a variety of homeostatic functions including metabolism, inflammation and development. PPAR-alpha is the main metabolic regulator for catabolism whereas PPAR-gamma regulates anabolism or storage. PPARs are expressed in the cardiovascular system such as endothelial cells, vascular smooth muscle cells and monocytes/macrophages. It has been shown that they play an important role in the modulation of inflammatory, fibrotic and hypertrophic responses. In 1997, a Glaxo patent described that Troglitazone (first PPAR-gamma ligand to reach market) reduced TNF-induced VCAM1 expression in HUVECs indicating the potential benefit in atherosclerosis. A series of patents from Eli Lilly and Dr. Reddy's Laboratories Ltd. between 1999 and 2005 described a variety of PPAR-alpha and -alpha,gamma dual ligands in a number of patents having glucose, triglyceride, cholesterol lowering, HDL elevating and body weight reducing activity. Patents from Metabolex and Tularik in 2001 and 2002 described the beneficial effects of SPPARM molecules for insulin resistance and diabetes, without showing concern on PPAR-gamma related side effects such as edema and body weight. GSK and Takeda described the potential effects of PPAR-delta modulators during 2001 to 2004 in few patents. Several clinical and preclinical studies have demonstrated the beneficial effects of PPAR ligands on various cardiovascular risk factors. This review intends to capture some of the key studies in this area as is described in some recent patents and literature.

Non-insulin dependent diabetes mellitus: present therapies and new drug targets.

Type 2 Diabetes Mellitus (DM) or Non-Insulin Dependent Diabetes Mellitus (NIDDM) accounts for 90-95% of all diabetes cases and has become a major health concern over the years. This disease has assumed frightening proportions due to unhealthy food habits and sedentary life style. About a decade ago, due to the absence of defined molecular targets or an understanding of disease pathophysiology, treatment of this disease was mostly focused on insulin secretion or administration of external insulin. During the past decade however, advent of genomics and proteomics has helped in understanding the molecular alteration characteristics of NIDDM. Untreated type 2 diabetes leads to several complications such as hyperlipidemia, hypertension and atherosclerosis--collectively known as Syndrome X. Though United Kingdom Prospective Diabetes Study (UKPDS) showed that normalization of hyperglycemia could prevent majority of diabetes complications, the available treatment regime does not adequately normalize the blood glucose level in type 2 diabetic patients. Currently, four distinct classes of oral hypoglycemic agents are available, some of which can act as lipid lowering agents as well. The efficacy and side effect profiles of these drugs are still to be optimized, so there is an unmet need for better candidates. Several new targets as well as better drugs for old targets are under investigation across the world. Availability of such drugs, based on the validated targets, may lead to a new therapeutic paradigm for the prevention of diabetes as well as complications arising out of it. The current review will deal with existing oral therapies for type 2 diabetes as well as the emerging therapeutic targets.

Design, synthesis and evaluation of carbazole derivatives as PPAR alpha/gamma dual agonists and antioxidants.

A series of hydroxycarbazole derivatives were synthesized and evaluated for PPAR alpha/gamma dual agonist as well as antioxidant activities. While most compounds showed good antioxidant activity, some compounds were identified as potential PPAR alpha/gamma dual agonists as well. Compounds 10a and 16 were found to be active in animal studies.

Lipopolysaccharide-induced paw edema model for detection of cytokine modulating anti-inflammatory agents.

Cytokines are critical to pathogenesis of inflammatory disorders. So inhibition of their action provides therapeutic benefits in various diseases. Although inhibition of inflammation caused by intraperitoneally administered LPS can identify cytokine modulators, this inflammatory test-agent does not allow one to determine overall anti-inflammatory potential. Functional characteristics of Carrageenan (Cara)-induced edema were valuable for identification of nonsteroidal anti-inflammatory drugs (NSAIDS). Hence, the potential of LPS-induced paw inflammation was investigated and compared to that by Cara. Stimulation of isolated rat peritoneal exudates cells (PEC) with 10 ng/ml LPS, but not Cara, induced IL-6 (3.04+/-0.2 ng/ml) and TNFalpha (1.030.09 ng/ml). At least 100 mg/ml Cara was necessary for detection of IL-6 (2.03+/-0.1 ng/ml) and TNFalpha (0.6+0.09 ng/ml) in PEC. Similar to Cara, subplantar administration of LPS-induced inflammatory paw edema in rats. LPS, but not Cara, induced TNFalpha (2.14+/-0.6 ng/ml) and IL-6 (2.9+/-0.5 ng/ml) in serum at 1 and 3 h, respectively, which returned to basal levels by 5 h. LPS-induced serum TNFalpha (sTNFalpha) levels closely paralleled paw swelling and its neutralization by anti-TNFalpha antibody or inhibition by pentoxifylline and nimesulide correlated with inhibition of inflammation. Similar to earlier reports, rofecoxib induced sTNFalpha at 30 mg/kg and exhibited pro-inflammatory effect by enhancing paw swelling. LPS-induced edema provides a useful functional model for identification of cytokine modulating anti-inflammatory agents.

Antidiabetic and hypolipidemic potential of DRF 2519--a dual activator of PPAR-alpha and PPAR-gamma.

We investigated the biological activity of Dr. Reddy's Research Foundation (DRF) 2519, a benzoxazinone analogue of the thiazolidinedione class of compounds. In the in vitro transactivation assay, DRF 2519 showed interesting dual activation of Peroxisome Proliferator Activated Receptor (PPAR) alpha and gamma. In insulin-resistant ob/ob mouse model, DRF 2519 showed significant alleviation of insulin resistance and dyslipidemia, which is better than rosiglitazone. Fatty Zucker rats treated with DRF 2519 showed better reduction of plasma insulin, triglyceride and free fatty acid levels than those treated with rosiglitazone. In addition, these rats were able to clear plasma lipids better when challenged with exogenous lipid (i.v.). DRF 2519 treatment resulted in improved plasma lipid profiles in high-fat-fed Sprague-Dawley rats. Treated rats showed better plasma lipid clearance and hepatic triglyceride secretion. When compared to DRF 2519, fenofibrate was comparatively less efficacious while rosigltiazone showed no activity in these models. In ex vivo studies, DRF 2519 showed induction of liver acyl CoA oxidase mRNA and increase in lipoprotein lipase (LPL) protein expression and activity in adipose tissue. In the in vitro studies, DRF 2519 inhibited the lipid biosynthesis and secretion of apolipoprotein B from human hepatoma (Hep)G2 cells. It also enhanced insulin-induced relaxation of rat aortic smooth muscle. These results indicate that DRF 2519, a dual activator of PPAR-alpha and gamma, could be an interesting development candidate in the management of metabolic disorders and associated complications.

Studies on some glitazones having pyridine as the linker unit.

Molecular modeling on various well-known glitazones carrying a pyridine ring instead of benzene ring as the middle linker unit showed conformational rigidity as compared to their parent molecules. Blocking the lone pair of electrons on the pyridine N, made them flexible once again. A few representatives of these analogues were synthesized and their efficacy as PPARgamma agonists evaluated.