Blockchain Papers

Tanuja Manoj Nesari

Animals have always been a companion as well as a crucial part of the lives of human beings for as long as humans have existed. They are not only employed for food and transportation but have been an intrinsic part of biomedical research, including developing novel treatments for infectious and noninfectious diseases, drug testing, and toxicological studies. Apart from these, animal models are used to understand the efficacy, safety, and possible mechanisms for different surgical experiments, various medical treatment approaches, other chemicals, consumer goods, vaccines, and diagnostics. They are also used in different experiments, education, and training modules to understand biological aspects. Ayurveda is one of the world’s oldest surviving medical systems, stretching back over 5000 years. The safety of medicines and therapies has always been a top emphasis in the Ayurvedic system of health care. For ages, Ayurveda has adopted the specific science of toxicity, as one of its eight therapeutic specialties. The roots of experimentation in animals to ensure the safety of food and drugs have been well described in classical texts of Ayurveda. In Ayurveda literature, there is description of different techniques/ experiments to examine food/ drugs, especially in the context of Vishanna (~poisonous food), Virudhanna (~incompatible food) on animals such as pigeons, peacocks, rabbits, monkeys, etc., to assess and establish their safety before their ingestion by humans. The animals have shown an intrinsic connection to human evolution; however, with ample proof of their utilization in ancient Greece, the eighteenth and nineteenth centuries saw the greatest advancement in the use of animal models.[1] The importance of animal models has grown significantly in recent decades due to the remarkable advancements in biomedical research, drug development, and medical technology. The use of animal models for scientific research has been a long-standing practice despite being an issue of debate and raising public concerns in society. With a substantial amount of similarities with humans on anatomical and physiological grounds, they have been developed as an inseparable part of scientific research, paving the way to a wide range of mechanisms to assess the novel therapies in animal models before applying them to humans. In the contemporary era, it is observed that there has been a significant increase in the worldwide use of laboratory animals over the years, with an overall estimate of global animal use in scientific procedures of 79.9 million animals in 2015, signifying a 36.9% increase on the equivalent estimated figure for 2005, of 58.3 million animals. Furthermore, the comprehensive final global figure for the number of animals used for scientific purposes in 2015 was 192.1 million.[2] Although the number of animals used for scientific purposes is increasing, the ethical concern regarding their use has also been growing and being opposed by various organizations, government legislation, researchers, and institutes. Since, “Animals, like people, have intrinsic moral and legal rights,” this implies that the exploitation of animals as pets or for any other reason is unethical and immoral. This restricts the use of animals for human gain as there is a chance that the animals would suffer as a result of such experiments. The suffering, agony, and eventual demise of animals during scientific research have long been debated, and several acts and laws have been passed to control the unethical use of animals. The Royal Society for the Prevention of Cruelty to Animals founded the organization for animal rights in 1824, followed by legislation in 1876 to prevent animal cruelty in the United Kingdom. Numerous laws and regulations are observed to safeguard animals from abuse and neglect. For example, there are specified guidelines for animal housing, breeding, feeding, transportation, and primarily for their use in scientific experiments by organizations such as the International Conference on Harmonization of Technical Requirements for registration of pharmaceuticals for human use, Committee for Control and Supervision of Experiments on Animals, National Institute of Health, and Organization for Economic Cooperation and Development.[3] The concepts of Replacement, Reduction, and Refinement, commonly known as the three Rs or 3Rs, first articulated by Russell and Burch in 1959, serve as a framework for legislation and moral review of protocols for the ethical treatment of animal research and are considered “alternatives” or “alternative methods” for minimizing the potential for animal pain and distress in biomedical research. This method underlines the principle of “Replacement” which means the substitution for conscious living higher animals of insentient material; “Reduction” means a reduction in the numbers of animals used to obtain information of a given amount and precision; and “Refinement” means any decrease in the incidence or severity of inhumane procedures applied to those animals which still have to be used.[4] In simpler terms, “Reduction” encourages the use of fewest possible animals; “Refinement” is meant to limit the amount of pain and anguish experienced by animals during experimentations; and “Refinement” deals with the ideal solutions to use lesser species and alternate techniques in place of higher animals whenever possible. The Food and Drug Administration (FDA) Modernization Act 2.0, which was passed in December 2022, represented a significant attempt to eliminate the requirement of animal testing for drug development. Although the use of animals in scientific study is not prohibited, the law acknowledges its limitations and provides researchers the freedom to apply cutting-edge, nonanimal techniques. The act amends the use of “certain alternatives to animal testing, including cell-based assays and computer models, to obtain an exemption from the FDA to investigate the safety and effectiveness of a drug.”[5] The new law also “removes a requirement to use animal studies as part of the process to obtain a license for a biological product that is biosimilar or interchangeable with another biological product.”[5] The new law also replaces the statute’s reference to “pre-clinical tests (including tests on animals)” with “nonclinical tests” that are to be “conducted in vitro, in silico, or in chemico, or a nonhuman in vivo test, that occurs before or during the clinical trial phase of the investigation of the safety and effectiveness of a drug.” However, the use of animals in research has arguably come with certain drawbacks, such as the need for highly qualified personnel, labor-intensive procedures, expensive expenditure of animal breeding, and housing, along with the extensive protocols required for animal studies, paving the way for various other alternative strategies and models for drug and chemical testing, having the advantages of being economical, time-efficient, and requiring less manpower. Owing to the rising concern of animal use in biomedical research and its limitations, certain alternatives have been developed over the period that can be efficiently applied for drug testing and research without involving animals. These tests may include the following: (1) cell-based assays, tissue cultures (also known as in vitro procedures); (2) sophisticated computer modeling techniques (commonly referred to as in silico models); (3) human biology-based test methods, such as bioprinting; (4) experimental use of higher model vertebrates or lower vertebrates; (5) use of invertebrate organisms; (6) use of microorganisms; and (7) studies involving human volunteers and human–patient simulators.[3,6] With the constant push to decrease the number of animals used during experimentations, several debates have been lasted for decades. There are several other means of experimentation for establishing the safety and effectiveness of drugs which includes cell lines, primary human tissue, cell-based tissue models, miniature cellular and tissue models, such as organs-on-a-chip and three dimensional bioprinting, spheroids, microfluidics, which are being extensively use to replicate the structures and functioning of organs for efficient screening of therapies and medicines. These techniques have added advantages such as reduction in the variables encountered during the study as well as provide platform to understand the drug metabolism, disease processes, and biological phenomena etc. These techniques have the advantages of being less expensive, time-consuming, and simple to use and are frequently used to evaluate the toxicity and efficacy of possible therapeutic compounds and chemicals during preliminary screening.[7] The microtissue models replicate particular parts and functions of the human body and aid in precisely monitoring system activities and developing novel, intelligent, and targeted preclinical testing techniques that can be used in any given human circumstance.[8] Bioprinting is a multidisciplinary and rapidly growing technology that combines engineering principles and life sciences for the fabrication of tissue and organ constructs by selectively depositing biological materials, biochemical, and living cells, typically in a layer-by-layer fashion. It utilizes both bottom-up and media-based cell culture, effectively tackles the intricate parameter space of minimal feature size (resolution), vascularization, perfusion, automation, cost, nutrient and growth factor diffusion, as well as the provision of mechanical and biochemical stimuli, offering a new perspective on the efficacy of patient-specific treatments and is more predictive than animal models.[9] The advancing computer technology and software have made it possible to simulate some parts of the human body and computer models of various organs such as heart, lungs, kidneys, digestive, and musculoskeletal systems have been developed to conduct virtual experiments based on existing mathematical data and information. In addition, these technologies are quick, affordable, and useful as a substitute for experimental assays, also facilitating translation to humans. These in silico approaches are considered potentially important human-based tools for safety pharmacology evaluation. The successful application of in silico modeling in academic, industrial, and regulatory contexts necessitates enhancing the models’ credibility, demonstrating their predictive ability through comparison with current experimental techniques, and encouraging their adoption by offering software that lowers the technological hurdles associated with in silico methods for nonspecialist users.[10] High-throughput screening involves screening a large number of compounds at random to locate hits that exhibit activity or affinity on a chosen target and/or in a model that is considered representative of disease to identify therapeutic compounds, pathways, cell functions, and chemical probes.[11] Several small animal models, including Drosophila melanogaster, zebrafish (Danio rerio), and Caenorhabditis elegans, are gaining popularity as drug discovery screening methods along with the advancing robotic and automated imaging with prime advantages of their genetic amenability, low cost, and culture conditions compatible with large-scale screening, to allow high-throughput screening in a whole animal context. Moreover, there are fewer ethical problems involved in the experimental use of these lower vertebrates. D. melanogaster, a common fruit fly, is a well-researched, very tractable genetic model organism that can be used to study the molecular mechanism of various human diseases, with many shared fundamental biological, physiological, and neurological traits to humans. It has been used extensively for therapeutic discovery through high-throughput screening for small compounds, mainly based on enzymatic assays, receptor binding tests, or in vitro cell culture.[12] The D. rerio, or Zebrafish, is a small freshwater fish with an almost translucent body and visible internal anatomy, facilitates direct observation of developmental stages, identification of phenotypic features during mutagenesis, identification of endpoints of toxicity and in elucidating mechanisms of toxicity, along with monitoring of gene expression through light microscopy. The species offers its wider application in the study of neurological and behavioral disorders, cancer, heart problems, etc. The working space, cost of laboratory solutions, and the manpower involved are reduced comparatively low, and modeling of certain human diseases in zebrafish aids in ameliorating the disease phenotype and malfunctions in organ development.[13] Ayurveda has a well-documented treatise, elaborating the characteristics of animals, birds, and insects with their wider use in the treatment of several illnesses. The Ayurvedic classics extensively document the concept of “Pratitantra siddhanta” (~applying theory specific to a particular school of thought, scripture, or branch of science).[14] This suggests the possibility of rational use and adoption of principles from an alternative branch of science, thereby opening the door to the use of alternative approaches for establishing the safety and efficacy of drugs rather than limiting them to animal experimentation alone. Ayurveda is a timely-tested system of medicines and does not necessarily demand animal experimentation for its drugs’ quality and efficacy. Under the Rule 158 of Drug and Cosmetic Act 1940, and Rules 1945, in the context of issue of licenses for Ayurveda, Siddha, and Unani (ASU) drugs, the experience or evidence of the effectiveness of the ASU drug based on the textual rationale or published literature is sought and the safety and effectiveness study is not necessary for the issuance of license.[15] Similarly, in terms of food safety, it is stipulated by the Food Safety and Standards (Ayurveda Aahara) Regulations, 2022 that no safety data is needed prior to product certification under the category of Ayurveda Aahara, if made in accordance with the authoritative Ayurvedic texts mentioned in the act.[16] This waves off the use of animal experimentation for safety and efficacy studies about Ayurveda classical drugs with textual reference. Although, the drugs and formulations mentioned in Ayurveda classics have been used for over 50 years and have substantial safety profiles, numerous experiments, and clinical trials have been conducted for Ayurveda therapies and drugs just to justify the need of evidence of the larger scientific communities across the globe. In the meanwhile, these investigations are required to determine the precise pharmacokinetics and pharmacodynamics the Ayurveda medications and therapies. Yogya vidhi, or simulation-based learning techniques has been well-described in Ayurveda classics, is aimed to train scholars for the development of precise surgical skills.[17] Such simulation-based learning in the contemporary era includes various models such as cadaveric, animal, bench-top, and virtual reality robotic simulators, etc., which are increasingly used in surgical training. The use of such simulation-based approaches suggests that ancient seers themselves believed in using alternative approaches instead of staking lives. The use of animals in experiments demands a firm and rational moral basis, where animal ethics should be a matter of equal importance to human welfare. Ethical decision-making, as well as evaluation of protocols by researchers and ethical boards, are still challenging when it comes to the use of animal models in biomedical research. Many alternatives to animal testing methods are being considered and employed, in addition to strict implementation of the 3Rs when using animals in laboratories. Furthermore, while animal models must be continuously developed to be more reliable and instructive, animal protection must also be given ongoing concern. All of these alternative models are novel tools for the drug development process that may be utilized in conjunction with the conventional system to meet the demand for large-scale screening while saving effort, resources, funds, and time. More reliable results may be obtained by utilizing a variety of computer models, bioinformatics tools, in vitro cell cultures, enzyme screenings, and model organisms in conjunction with contemporary analytical methods, data collection, and statistical processes, significantly reducing the number of animals used in scientific research, further enhancing health outcomes, without compromising lives of experimental animals.