The handling of biopharmaceuticals used in home-based therapy for cancer patients.

BACKGROUND: Due to their low bioavailability, biopharmaceuticals are typically administered via injection or infusion in a hospital setting. Home-based therapy could be a valuable alternative for cancer patients. However, when exposed to stressors, such as high or low temperatures, particles can form in the biopharmaceutical solution, compromising the safety and efficacy of the biopharmaceutical. AIM: This research investigated current practises with ready-to-administer biopharmaceuticals to determine if it is possible to offer cancer patients home-based therapy with monoclonal antibodies. METHODS: First, a questionnaire was conducted with the survey tool Survalyzer among Amsterdam UMC patients receiving immunoglobulins at home. Secondly, a web-based questionnaire (Survalyzer) was sent to pharmacists throughout Europe with a home-based therapy programme in place. RESULTS: The patient questionnaire (n = 52) showed that the biopharmaceutical is stored outside the recommended temperature range by 38% of the patients. Additionally, 17% of the patients do not recall getting any information on how to store their biopharmaceuticals, and 23% would like more information on the subject. Furthermore, the questionnaire amongst pharmacists (n = 21) showed that there is a lack of resources and logistical challenges when home-based therapy is applied to biopharmaceuticals used in cancer therapy. CONCLUSION: Home-based therapy with monoclonal antibodies for cancer patients is challenging to implement.

Post-translational modifications: Host defence mechanism, pathogenic weapon, and emerged target of anti-infective drugs

Post-translational modifications are changes introduced to proteins after their translation. They are the means to generate molecular diversity, expand protein function, control catalytic activity and trigger quick responses to a wide range of stimuli. Moreover, they regulate numerous biological processes, including pathogen invasion and host defence mechanisms. It is well established that bacteria and viruses utilize post-translational modifications on their own or their host's proteins to advance their pathogenicity. Doing so, they evade immune responses, target signaling pathways and manipulate host cytoskeleton to achieve survival, replication and propagation. Many bacterial species secrete virulence factors into the host and mediate hostpathogen interactions by inducing post-translational modifications that subvert fundamental cellular processes. Viral pathogens also utilize post translational modifications in order to overcome the host defence mechanisms and hijack its cellular machinery for their replication and propagation. For example, many coronavirus proteins are modified to achieve host invasion, evasion of immune responses and utilization of the host translational machinery. PTMs are also considered potential targets for the development of novel therapeutics from natural products with antibiotic properties, like lasso peptides and lantibiotics. The last decade, significant progress was made in understanding the mechanisms that govern PTMs and mediate regulation of protein structure and function. This urges the identification of relevant molecular targets, the design of specific drugs and the discovery of PTM-based medicine. Therefore, PTMs emerge as a highly promising field for the investigation and discovery of new therapeutics for many infectious diseases.Copyright © 2021 Bentham Science Publishers.

Accelerated CMC workflows to enable speed to clinic in the COVID-19 era: A multi-company view from the biopharmaceutical industry.

The COVID-19 pandemic has placed unprecedented pressure on biopharmaceutical companies to develop efficacious preventative and therapeutic treatments, which is unlikely to abate in the coming years. The importance of fast progress to clinical evaluation for treatments, which tackle unmet medical needs puts strain on traditional product development timelines, which can take years from start to finish. Although previous work has been successful in reducing phase 1 timelines for recombinant antibodies, through utilization of the latest technological advances and acceptance of greater business risk or costs, substantially faster development is likely achievable without increased risk to patients during initial clinical evaluation. To optimize lessons learned from the pandemic and maximize multi-stakeholder (i.e., patients, clinicians, companies, regulatory agencies) benefit, we conducted an industry wide benchmarking survey in September/October 2021. The aims of this survey were to: (i) benchmark current technical practices of key process and product development activities related to manufacturing of therapeutic proteins, (ii) understand the impact of changes implemented in COVID-19 accelerated Ab programs, and whether any such changes can be retained as part of sustainable long-term business practices and (iii) understand whether any accelerative action(s) taken have (negatively) impacted the wider development process. This article provides an in-depth analysis of this data, ultimately highlighting an industry perspective of how biopharmaceutical companies can sustainably adopt new approaches to therapeutic protein development and production.

Microalgae as an Efficient Vehicle for the Production and Targeted Delivery of Therapeutic Glycoproteins against SARS-CoV-2 Variants.

Severe acute respiratory syndrome-Coronavirus 2 (SARS-CoV-2) can infect various human organs, including the respiratory, circulatory, nervous, and gastrointestinal ones. The virus is internalized into human cells by binding to the human angiotensin-converting enzyme 2 (ACE2) receptor through its spike protein (S-glycoprotein). As S-glycoprotein is required for the attachment and entry into the human target cells, it is the primary mediator of SARS-CoV-2 infectivity. Currently, this glycoprotein has received considerable attention as a key component for the development of antiviral vaccines or biologics against SARS-CoV-2. Moreover, since the ACE2 receptor constitutes the main entry route for the SARS-CoV-2 virus, its soluble form could be considered as a promising approach for the treatment of coronavirus disease 2019 infection (COVID-19). Both S-glycoprotein and ACE2 are highly glycosylated molecules containing 22 and 7 consensus N-glycosylation sites, respectively. The N-glycan structures attached to these specific sites are required for the folding, conformation, recycling, and biological activity of both glycoproteins. Thus far, recombinant S-glycoprotein and ACE2 have been produced primarily in mammalian cells, which is an expensive process. Therefore, benefiting from a cheaper cell-based biofactory would be a good value added to the development of cost-effective recombinant vaccines and biopharmaceuticals directed against COVID-19. To this end, efficient protein synthesis machinery and the ability to properly impose post-translational modifications make microalgae an eco-friendly platform for the production of pharmaceutical glycoproteins. Notably, several microalgae (e.g., Chlamydomonas reinhardtii, Dunaliella bardawil, and Chlorella species) are already approved by the U.S. Food and Drug Administration (FDA) as safe human food. Because microalgal cells contain a rigid cell wall that could act as a natural encapsulation to protect the recombinant proteins from the aggressive environment of the stomach, this feature could be used for the rapid production and edible targeted delivery of S-glycoprotein and soluble ACE2 for the treatment/inhibition of SARS-CoV-2. Herein, we have reviewed the pathogenesis mechanism of SARS-CoV-2 and then highlighted the potential of microalgae for the treatment/inhibition of COVID-19 infection.

Production and secretion of functional SARS-CoV-2 spike protein in Chlamydomonas reinhardtii.

The spike protein is the major protein on the surface of coronaviruses. It is therefore the prominent target of neutralizing antibodies and consequently the antigen of all currently admitted vaccines against SARS-CoV-2. Since it is a 1,273-amino acids glycoprotein with 22 N-linked glycans, the production of functional, full-length spike protein was limited to higher eukaryotes. Here we report the production of full-length SARS-CoV-2 spike protein - lacking the C-terminal membrane anchor - as a secreted protein in the prefusion-stabilized conformation in the unicellular green alga Chlamydomonas reinhardtii. We show that the spike protein is efficiently cleaved at the furin cleavage site during synthesis in the alga and that cleavage is abolished upon mutation of the multi-basic cleavage site. We could enrich the spike protein from culture medium by ammonium sulfate precipitation and demonstrate its functionality based on its interaction with recombinant ACE2 and ACE2 expressed on human 293T cells. Chlamydomonas reinhardtii is a GRAS organism that can be cultivated at low cost in simple media at a large scale, making it an attractive production platform for recombinant spike protein and other biopharmaceuticals in low-income countries.

Efficacy of hydrogen peroxide wipes for decontamination of AZD1222 adenovirus COVID-19 vaccine strain on pharmaceutical industry materials.

This study aimed to evaluate the performance of accelerated hydrogen peroxide® wipes (HPW) for decontamination of the chimpanzee adenovirus AZD1222 vaccine strain used in the production of recombinant COVID-19 vaccine in a pharmaceutical industry. Two matrices were tested on stainless-steel (SS) and low-density-polyethylene (LDP) surfaces: formulated recombinant COVID-19 vaccine (FCV) and active pharmaceutical ingredient (API). The samples were spiked, dried and the initial inoculum, possible residue effect (RE) and titre reduction after disinfection with HPW were determined. No RE was observed. The disinfection procedure with HPW resulted in complete decontamination the of AZD1222 adenovirus strain in FCV (≥7·46 and ≥7·49 log10 infectious unit [IFU] ml-1 for SS and LDP carriers respectively) and API (≥8·79 and ≥8·78 log10 IFU ml-1 for SS and LDP carriers respectively). In conclusion, virucidal activity of HPW was satisfactory against the AZD1222 adenovirus strain and can be a good option for disinfection processes of SS and LPD surfaces in pharmaceutical industry facilities during recombinant COVID-19 vaccine production. This procedure is simple and can be also applied on safety unit cabins and sampling bags made of LDP as well.

Investigation of the N-Glycosylation of the SARS-CoV-2 S Protein Contained in VLPs Produced in Nicotiana benthamiana.

The emergence of the SARS-CoV-2 coronavirus pandemic in China in late 2019 led to the fast development of efficient therapeutics. Of the major structural proteins encoded by the SARS-CoV-2 genome, the SPIKE (S) protein has attracted considerable research interest because of the central role it plays in virus entry into host cells. Therefore, to date, most immunization strategies aim at inducing neutralizing antibodies against the surface viral S protein. The SARS-CoV-2 S protein is heavily glycosylated with 22 predicted N-glycosylation consensus sites as well as numerous mucin-type O-glycosylation sites. As a consequence, O- and N-glycosylations of this viral protein have received particular attention. Glycans N-linked to the S protein are mainly exposed at the surface and form a shield-masking specific epitope to escape the virus antigenic recognition. In this work, the N-glycosylation status of the S protein within virus-like particles (VLPs) produced in Nicotiana benthamiana (N. benthamiana) was investigated using a glycoproteomic approach. We show that 20 among the 22 predicted N-glycosylation sites are dominated by complex plant N-glycans and one carries oligomannoses. This suggests that the SARS-CoV-2 S protein produced in N. benthamiana adopts an overall 3D structure similar to that of recombinant homologues produced in mammalian cells.

Biologics in COVID-19 So Far: Systematic Review.

This systematic review aimed to reevaluate the available evidence of the use of biologics as treatment candidates for the treatment of severe and advanced COVID-19 disease; what are the rationale for their use, which are the most studied, and what kind of efficacy measures are described? A search through Cochrane, Embase, Pubmed, Medline, medrxiv.org, and Google scholar was performed on the use of biologic interventions in COVID-19/SARS-CoV-2 infection, viral pneumonia, and sepsis, until 11 January 2022. Throughout the research, we identified 4821 records, of which 90 were selected for qualitative analysis. Amongst the results, we identified five popular targets of use: IL6 and IL1 inhibitors, interferons, mesenchymal stem cells treatment, and anti-spike antibodies. None of them offered conclusive evidence of their efficacy with consistency and statistical significance except for some studies with anti-spike antibodies; however, Il6 and IL1 inhibitors as well as interferons show encouraging data in terms of increased survival and favorable clinical course that require further studies with better methodology standardization.

Towards smart vaccine manufacturing: A preliminary study during covid-19

Biopharmaceutical community is devising modern techniques to boost the development, production, and distribution of COVID-19 vaccines in large scale with tremendous speed. This has shifted the focus towards smart manufacturing of vaccines through vaccine platforms. Vaccine platforms have great potential to rapidly generate new vaccines and can overcome the challenges of the traditional vaccine manufacturing approach without compromising on safety and efficacy. This preliminary study compares the traditional and modern vaccine manufacturing techniques, reviews COVID-19 vaccine manufacturing scenarios, and presents a framework to critique on the smartness of the novel platform-based COVID-19 vaccine development and manufacturing. Copyright © 2021 by ASME and The United States Government.

Evaluation of hydrogen peroxide efficacy against AZD1222 chimpanzee adenovirus strain in the recombinant COVID-19 vaccine for application in cleaning validation in a pharmaceutical manufacturing industry.

This study aimed to evaluate the performance of hydrogen peroxide vapour (HPV) to inactivate the chimpanzee adenovirus AZD1222 vaccine strain used in the production of recombinant COVID-19 vaccine for application in cleaning validation in pharmaceutical industries production areas. Two matrixes were tested: formulated recombinant COVID-19 vaccine (FCV) and active pharmaceutical ingredient (API). The samples were dried on stainless steel and exposed to HPV in an isolator. One biological indicator with population >106 Geobacillus stearothermophilus spores was used to validate the HPV decontamination cycle as standard. HPV exposure resulted in complete virus inactivation in FVC (≥5·03 log10 ) and API (≥6·40 log10 ), showing HPV efficacy for reducing chimpanzee adenovirus AZD1222 vaccine strain. However, the optimum concentration and contact time will vary depending on the type of application. Future decontamination studies scaling up the process to the recombinant COVID-19 vaccine manufacturing areas are necessary to evaluate if the HPV will have the same or better virucidal effectivity in each specific production area. In conclusion, HPV showed efficacy for reducing AZD1222 chimpanzee adenovirus strain and can be a good choice for pharmaceutical industries facilities disinfection during recombinant COVID-19 vaccine production.

COVID-19 Crisis: How Can Plant Biotechnology Help?

The emergence of the COVID-19 pandemic has led to significant public health crisis all over the world. The rapid spreading nature and high mortality rate of COVID-19 places a huge pressure on scientists to develop effective diagnostics and therapeutics to control the pandemic. Some scientists working on plant biotechnology together with commercial enterprises for the emergency manufacturing of diagnostics and therapeutics have aimed to fulfill the rapid demand for SARS-CoV-2 protein antigen and antibody through a rapid, scalable technology known as transient/stable expression in plants. Plant biotechnology using transient/stable expression offers a rapid solution to address this crisis through the production of low-cost diagnostics, antiviral drugs, immunotherapy, and vaccines. Transient/stable expression technology for manufacturing plant-based biopharmaceuticals is already established at commercial scale. Here, current opinions regarding how plant biotechnology can help fight against COVID-19 through the production of low-cost diagnostics and therapeutics are discussed.

Affordable oral health care: dental biofilm disruption using chloroplast made enzymes with chewing gum delivery.

Current approaches for oral health care rely on procedures that are unaffordable to impoverished populations, whereas aerosolized droplets in the dental clinic and poor oral hygiene may contribute to spread of several infectious diseases including COVID-19, requiring new solutions for dental biofilm/plaque treatment at home. Plant cells have been used to produce monoclonal antibodies or antimicrobial peptides for topical applications to decrease colonization of pathogenic microbes on dental surface. Therefore, we investigated an affordable method for dental biofilm disruption by expressing lipase, dextranase or mutanase in plant cells via the chloroplast genome. Antibiotic resistance gene used to engineer foreign genes into the chloroplast genome were subsequently removed using direct repeats flanking the aadA gene and enzymes were successfully expressed in marker-free lettuce transplastomic lines. Equivalent enzyme units of plant-derived lipase performed better than purified commercial enzymes against biofilms, specifically targeting fungal hyphae formation. Combination of lipase with dextranase and mutanase suppressed biofilm development by degrading the biofilm matrix, with concomitant reduction of bacterial and fungal accumulation. In chewing gum tablets formulated with freeze-dried plant cells, expressed protein was stable up to 3 years at ambient temperature and was efficiently released in a time-dependent manner using a mechanical chewing simulator device. Development of edible plant cells expressing enzymes eliminates the need for purification and cold-chain transportation, providing a potential translatable therapeutic approach. Biofilm disruption through plant enzymes and chewing gum-based delivery offers an effective and affordable dental biofilm control at home particularly for populations with minimal oral care access.

Insights from Pharmaceutical Biotechnology into Phenolic Biopharmaceuticals against COVID-19.

BACKGROUND: The COVID-19 pandemic had infected more than 3.5M people around the world and more than 250K people died in 187 countries by May 2020. The causal agent of this disease is a coronavirus whose onset of symptoms to death range from 6 to 41 days with a median of 14 days. This period is dependent on several factors such as the presence of comorbidities, age and the efficiency of the innate or adaptive immune responses. METHODS: The effector mechanisms of both types of immune responses depend on the pathogen involved. In the case of a viral infection, the innate immune response may approach the harmful virus through pattern recognition receptors inducing an antiviral state. RESULTS: On the other hand, the adaptive immune response activates antibody production to neutralize or eliminate the virus. Phenolics are plant secondary metabolites with many biological activities for plants and humans against infection. Chemical modification of proteins may enhance their biological properties; thus, a protein of medical interest, for instance, a viral protein can be used as a scaffold to build a biopharmaceutical conjugated or complexated with phenolics exhibiting structural complexity or biological activities to achieve effective phenolic-protein-based therapeutics like vaccine adjuvant complexes, immunogen conjugates, and antiviral conjugates. CONCLUSION: Pharmaceutical biotechnology applies the principles of biotechnology to develop biopharmaceuticals for protein-based therapeutics; such as adjuvants, recombinant proteins, monoclonal antibodies, and antivirals. As neither a vaccine nor a treatment for COVID-19 is currently available, this manuscript focuses on insights from pharmaceutical biotechnology into phenolic biopharmaceuticals against COVID-19.

Potential for Developing Plant-Derived Candidate Vaccines and Biologics against Emerging Coronavirus Infections.

The emerging human coronavirus infections in the 21st century remain a major public health crisis causing worldwide impact and challenging the global health care system. The virus is circulating in several zoonotic hosts and continuously evolving, causing occasional outbreaks due to spill-over events occurring between animals and humans. Hence, the development of effective vaccines or therapeutic interventions is the current global priority in order to reduce disease severity, frequent outbreaks, and to prevent future infections. Vaccine development for newly emerging pathogens takes a long time, which hinders rapid immunization programs. The concept of plant-based pharmaceuticals can be readily applied to meet the recombinant protein demand by means of transient expression. Plants are evolved as an expression platform, and they bring a combination of unique interests in terms of rapid scalability, flexibility, and economy for industrial-scale production of effective vaccines, diagnostic reagents, and other biopharmaceuticals. Plants offer safe biologics to fulfill emergency demands, especially during pandemic situations or outbreaks caused by emerging strains. This review highlights the features of a plant expression platform for producing recombinant biopharmaceuticals to combat coronavirus infections with emphasis on COVID-19 vaccine and biologics development.

Downstream Processing of Therapeutic Peptides by Means of Preparative Liquid Chromatography.

The market of biomolecules with therapeutic scopes, including peptides, is continuously expanding. The interest towards this class of pharmaceuticals is stimulated by the broad range of bioactivities that peptides can trigger in the human body. The main production methods to obtain peptides are enzymatic hydrolysis, microbial fermentation, recombinant approach and, especially, chemical synthesis. None of these methods, however, produce exclusively the target product. Other species represent impurities that, for safety and pharmaceutical quality reasons, must be removed. The remarkable production volumes of peptide mixtures have generated a strong interest towards the purification procedures, particularly due to their relevant impact on the manufacturing costs. The purification method of choice is mainly preparative liquid chromatography, because of its flexibility, which allows one to choose case-by-case the experimental conditions that most suitably fit that particular purification problem. Different modes of chromatography that can cover almost every separation case are reviewed in this article. Additionally, an outlook to a very recent continuous chromatographic process (namely Multicolumn Countercurrent Solvent Gradient Purification, MCSGP) and future perspectives regarding purification strategies will be considered at the end of this review.

Optical biosensors - Illuminating the path to personalized drug dosing.

Optical biosensors are low-cost, sensitive and portable devices that are poised to revolutionize the medical industry. Healthcare monitoring has already been transformed by such devices, with notable recent applications including heart rate monitoring in smartwatches and COVID-19 lateral flow diagnostic test kits. The commercial success and impact of existing optical sensors has galvanized research in expanding its application in numerous disciplines. Drug detection and monitoring seeks to benefit from the fast-approaching wave of optical biosensors, with diverse applications ranging from illicit drug testing, clinical trials, monitoring in advanced drug delivery systems and personalized drug dosing. The latter has the potential to significantly improve patients' lives by minimizing toxicity and maximizing efficacy. To achieve this, the patient's serum drug levels must be frequently measured. Yet, the current method of obtaining such information, namely therapeutic drug monitoring (TDM), is not routinely practiced as it is invasive, expensive, time-consuming and skilled labor-intensive. Certainly, optical sensors possess the capabilities to challenge this convention. This review explores the current state of optical biosensors in personalized dosing with special emphasis on TDM, and provides an appraisal on recent strategies. The strengths and challenges of optical biosensors are critically evaluated, before concluding with perspectives on the future direction of these sensors.

Green giant-a tiny chloroplast genome with mighty power to produce high-value proteins: history and phylogeny.

Free-living cyanobacteria were entrapped by eukaryotic cells ~2 billion years ago, ultimately giving rise to chloroplasts. After a century of debate, the presence of chloroplast DNA was demonstrated in the 1960s. The first chloroplast genomes were sequenced in the 1980s, followed by ~100 vegetable, fruit, cereal, beverage, oil and starch/sugar crop chloroplast genomes in the past three decades. Foreign genes were expressed in isolated chloroplasts or intact plant cells in the late 1980s and stably integrated into chloroplast genomes, with typically maternal inheritance shown in the 1990s. Since then, chloroplast genomes conferred the highest reported levels of tolerance or resistance to biotic or abiotic stress. Although launching products with agronomic traits in important crops using this concept has been elusive, commercial products developed include enzymes used in everyday life from processing fruit juice, to enhancing water absorption of cotton fibre or removal of stains as laundry detergents and in dye removal in the textile industry. Plastid genome sequences have revealed the framework of green plant phylogeny as well as the intricate history of plastid genome transfer events to other eukaryotes. Discordant historical signals among plastid genes suggest possible variable constraints across the plastome and further understanding and mitigation of these constraints may yield new opportunities for bioengineering. In this review, we trace the evolutionary history of chloroplasts, status of autonomy and recent advances in products developed for everyday use or those advanced to the clinic, including treatment of COVID-19 patients and SARS-CoV-2 vaccine.

Emergence of Novel Coronavirus 2019-nCoV: Need for Rapid Vaccine and Biologics Development.

Novel Coronavirus (2019-nCoV) is an emerging pathogen that was first identified in Wuhan, China in late December 2019. This virus is responsible for the ongoing outbreak that causes severe respiratory illness and pneumonia-like infection in humans. Due to the increasing number of cases in China and outside China, the WHO declared coronavirus as a global health emergency. Nearly 35,000 cases were reported and at least 24 other countries or territories have reported coronavirus cases as early on as February. Inter-human transmission was reported in a few countries, including the United States. Neither an effective anti-viral nor a vaccine is currently available to treat this infection. As the virus is a newly emerging pathogen, many questions remain unanswered regarding the virus's reservoirs, pathogenesis, transmissibility, and much more is unknown. The collaborative efforts of researchers are needed to fill the knowledge gaps about this new virus, to develop the proper diagnostic tools, and effective treatment to combat this infection. Recent advancements in plant biotechnology proved that plants have the ability to produce vaccines or biopharmaceuticals rapidly in a short time. In this review, the outbreak of 2019-nCoV in China, the need for rapid vaccine development, and the potential of a plant system for biopharmaceutical development are discussed.

An update on antiviral antibody-based biopharmaceuticals.

Due to the vastness of the science virology, it is no longer an offshoot solely of the microbiology. Viruses have become as the causative agents of major epidemics throughout history. Many therapeutic strategies have been used for these microorganisms, and in this way the recognizing of potential targets of viruses is of particular importance for success. For decades, antibodies and antibody fragments have occupied a significant body of the treatment approaches against infectious diseases. Because of their high affinity, they can be designed and engineered against a variety of purposes, mainly since antibody fragments such as scFv, nanobody, diabody, and bispecific antibody have emerged owing to their small size and interesting properties. In this review, we have discussed the antibody discovery and molecular and biological design of antibody fragments as inspiring therapeutic and diagnostic agents against viral targets.

From conception to COVID-19: an arduous journey of tribulations of racism and triumphs.

Growing up in a densely wooded tropical forest enhanced my curiosity in plants and reading biography of Marie Curie profoundly influenced pursuit of my research career. Early in my career, I developed in vitro functional chloroplasts, capable of expressing foreign genes and this laid the foundation for the chloroplast genetic engineering field. Four decades of research has advanced chloroplast bioreactors for production of industrial enzymes or biopharmaceuticals by small or large companies. Because I experienced firsthand horrors of expensive vaccines or medicines, I devoted most of my career to develop affordable therapeutics. During this long journey, I suffered institutional racial discrimination but was rescued by several guardian angels. This biography gives readers a glimpse of tribulations and triumphs of my journey and recognizes important contributions made by my mentees.