Generation Of Vaccines Research Articles

MONKEYPOX AND ITS CURRENT REALITY

Introduction: Monkeypox or "Monkeypox" is a species of poxvirus, which possesses double-stranded DNA. This species has the potential to be used as bioterrorism; and currently has re-emerged in non-endemic countries. Objective: To detail the current information related to monkeypox and its epidemiological repercussions; in addition, to analyze the management and prevention of the contagion of this disease. Methodology: a search was carried out in PubMed, Elsevier and Google Scholar, with the terms Monkeypox, Ape Pox and Monkeypox. Some articles were rejected due to lack of relevance. Results: Monkeypox is transmitted by nasopharyngeal, oropharyngeal or intradermal routes; it has an incubation period of 5 to 21 days. Its diagnosis is made by laboratory tests such as PCR-RT and biopsy of skin lesions. Prevention is carried out with vaccines (especially the Ankara vaccine), which is 85% effective. Other drugs can be used in case of infection by this virus; however, their efficacy has not been determined so far. Currently, third generation vaccines are being developed. Conclusions: Monkeypox had its outbreak in the 1970s and re-emerged in 1990. In 2020, 5257 suspected patients were found. It has now been discovered in the United Kingdom, Israel, United States, Singapore, among others. Current management is symptomatic and prevention with vaccines. The administration of Tecovirimat and Brincidofovir have controversial efficacy. Vaccines have shown greater effectiveness. KEY WORDS: Monkeypox, orthopoxvirus, smallpox, Polymerase Chain Reaction, Poxviridae infections.

Vaccines against new coronavirus infection in Russia and the world

The article is devoted to the review of vaccines against new coronavirus infection in the world and Russia.
 Vaccination is an affordable and cost-effective way to reduce morbidity and mortality from COVID-19. The importance of vaccination is beyond doubt and is a method of creating active artificial immunity based on the formed immunological memory to an infectious agent. Conventionally, vaccines can be divided into two groups: classical (recombinant, peptide and virusinactivated), and gene vaccines (vector and mRNA vaccines). Classical vaccines are based on the introduction of ready-made antigens into the body, which can be purified viral proteins, fragments of viral proteins (peptides) or whole inactivated (killed) viral particles. Inactivated whole-virion vaccines contain a weakened or inactivated virus that promotes the development of antiviral immunity. Subunit vaccines contain only surface antigens (specific fragments subunits), which reduces the amount of protein in the vaccine and thereby reduces its allergenicity. Vector vaccines use safe viruses that are unable to reproduce in the human body (vectors), in which a gene is embedded ― a small section of the coronavirus genome. Vector vaccines effectively cause a cellular and humoral immune response, since the vector, entering the cell, is perceived by the body as a viral infection. Gene vaccines differ significantly from classical ones and produce an immune response at the gene level, since they contain not the virus or protein itself, but the genetic material of the SARS-CoV-2 coronavirus. An RNA-based vaccine delivers a specific set of instructions to the bodys cells for the synthesis of a specific protein, to which the bodys immune system must give an immune response.
 The criterion for the effectiveness of the vaccine is the immune response that it creates. The more pronounced and stable it is, the higher the protection against infection. So far, no drugs have been developed that have 100% protection against infection, since the immune response depends on numerous reasons, including individual characteristics of the body, general health, age, etc.
 At the present stage, the tasks of vaccination are complicated by such factors of the epidemiological process as the emergence of new highly pathogenic strains of coronavirus.

Monkeypox: Virology, Pathophysiology, Clinical Characteristics, Epidemiology, Vaccines, Diagnosis, and Treatments.

The World Health Organization, has declared the recent multiregional outbreak of monkeypox, a global public health emergency. Monkeypox is a zoonotic viral infection endemic to the west and central Africa. It belongs to the Poxviridae family, the Chordopoxvirinae subfamily, and the Orthopoxvirus genus. The Poxviridae family generally consists of complex, large, enveloped, and linear double-stranded DNA viruses. The initial clinical symptoms of monkeypox are often fever, severe headache, lymphadenopathy, myalgia, and fatigue. The skin lesions typically erupt within 1-3 days of the onset of fever. The rash tends to be more localized on the face and extremities than on the trunk. Monkeypox is often a self-limiting infection, and symptoms last from 2 to 4 weeks. It is isolated from various species, but the exact natural host is uncertain. Monkeypox is transmitted by close contact with infected humans or animals. Currently, no specific medication is available for monkeypox, and the existing therapeutics are the anti-viral agents approved for smallpox infection, including tecovirimat, cidofovir, and brincidofovir. Additionally, the U.S. Food and Drug Administration has approved Vaccinia Immune Globulin Intravenous for treating vaccination complications. It is diagnosed by PCR. There are currently two vaccines licensed by the U.S. Food and Drug Administration. According to the WHO guidance, the first-generation smallpox vaccines held in national reserves of some countries are not recommended as they do not meet the current safety and manufacturing standards. The interim guidance indicates that new and safer (second- and third generation) vaccines for smallpox, may be beneficial for monkeypox prevention, including JYNNEOS, which has been approved for the prevention of monkeypox. Human monkeypox was first reported in 1970. Since then, it has caused several outbreaks, mainly in central and west Africa. The first monkeypox outbreak outside of Africa occurred in the United States in 2003, linked to contact with infected pet prairie dogs. More recently (2018-2021), monkeypox cases have been reported in travelers from Nigeria to the United Kingdom, Israel, Singapore, and the US. Since May 2022, multiple monkeypox cases have been confirmed in several non-endemic countries, raising the concern of an emerging global pandemic. This review is an updated overview of our current state of knowledge regarding monkeypox virology, pathophysiology, clinical characteristics, epidemiology, vaccines, diagnosis, and treatment options.

Immunology to Immunotherapeutics of SARS-CoV-2: Identification of Immunogenic Epitopes for Vaccine Development.

The emergence of COVID19 pandemic caused by SARS-CoV-2 virus has created a global public health and socio-economic crisis. Immunoinformatics-based approaches to investigate the potential antigens is the fastest way to move towards a multiepitope-based vaccine development. This review encompasses the underlying mechanisms of pathogenesis, innate and adaptive immune signaling along with evasion pathways of SARS-CoV-2. Furthermore, it compiles the promiscuous peptides from in silico studies which are subjected to prediction of cytokine milieu using web-based servers. Out of the 434 peptides retrieved from all studies, we have identified 33 most promising T cell vaccine candidates. This review presents a list of the most potential epitopes from several proteins of the virus based on their immunogenicity, homology, conservancy and population coverage studies. These epitopes can form a basis of second generation of vaccine development as the first generation vaccines in various stages of trials mostly focus only on Spike protein. We therefore, propose them as most potential candidates which can be taken up immediately for confirmation by experimental studies.Supplementary InformationThe online version contains supplementary material available at 10.1007/s00284-022-03003-3.

Plasmid DNA for Therapeutic Applications in Cancer.

Recently, the interest in using nucleic acids for therapeutic applications has been increasing. DNA molecules can be manipulated to express a gene of interest for gene therapy applications or vaccine development. Plasmid DNA can be developed to treat different diseases, such as infections and cancer. In most cancers, the immune system is limited or suppressed, allowing cancer cells to grow. DNA vaccination has demonstrated its capacity to stimulate the immune system to fight against cancer cells. Furthermore, plasmids for cancer gene therapy can direct the expression of proteins with different functions, such as enzymes, toxins, and cytotoxic or proapoptotic proteins, to directly kill cancer cells. The progress and promising results reported in animal models in recent years have led to interesting clinical results. These DNA strategies are expected to be approved for cancer treatment in the near future. This review discusses the main strategies, challenges, and future perspectives of using plasmid DNA for cancer treatment.

Monkeypox Vaccines: Current Update

Monkeypox belonging to the genus Orthopoxvirus (OPV) of the family poxviridiae poses a major human threat even after 30 years after eradicating smallpox. OPV family includes other viruses like vaccinia virus (VACV), variola virus and cowpox virus. VACV smallpox vaccine provided cross protective antibody response against other orthopox viruses.1 The ongoing monkeypox outbreak has affected 31,800 people in more than 89 different countries. India has documented 9 confirmed cases with one death reported.2 Stockpile of smallpox vaccines that contain live vaccinia virus is maintained at two locations, one at Centre for Disease Control laboratory at Atlanta, Georgia and the other at VECTOR institute, Koltsovo, Russia. Data shows effectiveness of Smallpox vaccines against monkeypox to be 85%.3 Vaccine used in smallpox eradication were first generation vaccines which produced multiple side effects like eczema, progressive vaccinia and myopericarditis. This fear of adverse effects and safety concerns had led to the production of second and third generation vaccines like imvanex or ACAM2000.4 The interim guidelines for monkeypox vaccination does not urge the need for mass vaccination considering the side effects and the vaccine production. Countries like Canada, United Kingdom and USA are using ring vaccination to prevent monkeypox. The two vaccines currently being used against monkeypox are Jynneos vaccine and ACAM2000.5 JYNNEOS vaccine (Imvamune in Canada /Imvanex in Europe developed by Bavarian Nordiac) for adults aged 18 years and older was approved in Europe and U.S. The vaccine contains a replication deficient live vaccinia virus (Modified vaccinia Ankara-MVA BN) that is given as subcutaneous injection of 0.5 ml, two doses four weeks apart. The vaccine is used for post exposure prophylaxis against monkeypox virus. CDC recommends the intermediate or high risk group as potential group to receive post exposure propylaxis of the vaccine (Unprotected contact with the body fluid of infected person or contact with less 6 meter distance with the infected person). Vaccine is available in countries like Canada, Europe and Britain.6 ACAM2OOO, containing live replicating vaccinia virus that can replicate as compared to jynneos vaccine. Owning to the side effects associated with the vaccine its only used for laboratory workers handling pox viruses or for military personnel.4,7,8 WHO has stated there is no need for mass vaccination and decision regarding vaccination should be based on the individual case accessing the risk and benefits.5 In India, the Serum Institute of India has confirmed its ongoing research to find vaccine against monkeypox. Monkeypox vaccine pipelines include 1. Tonix Pharmaceuticals Holding Corp TNX-8011, a preclinical live virus, which uses horsepox virus 2. EpiVax, Inc’s-Epitome driven vaccine 3. VennVax, DNA prime peptide boost multi-T cell epitome Pox virus vaccine.9

COVID-19 Infection and Response to Vaccination in Chronic Kidney Disease and Renal Transplantation: A Brief Presentation.

Chronic kidney disease (CKD) is associated with phenotypic and functional changes in the immune system, followed by detrimental clinical consequences, such as severe infections and defective response to vaccination. Two years of the pandemic, due to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), have undoubtedly changed the world; however, all efforts to confront infection and provide new generation vaccines tremendously improved our understanding of the mechanisms of the immune response against infections and after vaccination. Humoral and cellular responses to vaccines, including mRNA vaccines, are apparently affected in CKD patients, as elimination of recent thymic emigrant and naïve lymphocytes and regulatory T-cells, together with contraction of T-cell repertoire and homeostatic proliferation rate, which characterized CKD patients are responsible for impaired immune activation. Successful renal transplantation will restore some of these changes, although several epigenetic changes are irreversible and even accelerated by the induction of immunosuppression. Response to vaccination is definitely impaired among both CKD and RT patients. In the present review, we analyzed the differences in immune response after vaccination between these patients and healthy individuals and depicted specific parameters, such as alterations in the immune system, predisposing to this deficient response.

Stability and Dissociation of Adeno-Associated Viral Capsids by Variable Temperature-Charge Detection-Mass Spectrometry.

Adeno-associated viral (AAV) vectors have emerged as gene therapy and vaccine delivery systems. Differential scanning fluorimetry or differential scanning calorimetry is commonly used to measure the thermal stability of AAVs, but these global methods are unable to distinguish the stabilities of different AAV subpopulations in the same sample. To address this challenge, we combined charge detection-mass spectrometry (CD-MS) with a variable temperature (VT) electrospray source that controls the temperature of the solution prior to electrospray. Using VT-CD-MS, we measured the thermal stabilities of empty and filled capsids. We found that filled AAVs ejected their cargo first and formed intermediate empty capsids before completely dissociating. Finally, we observed that pH stress caused a major decrease in thermal stability. This new approach better characterizes the thermal dissociation of AAVs, providing the simultaneous measurement of the stabilities and dissociation pathways of different subpopulations.

The role of cell-mediated immunity against influenza and its implications for vaccine evaluation

Influenza vaccines remain the most effective tools to prevent flu and its complications. Trivalent or quadrivalent inactivated influenza vaccines primarily elicit antibodies towards haemagglutinin and neuraminidase. These vaccines fail to induce high protective efficacy, in particular in older adults and immunocompromised individuals and require annual updates to keep up with evolving influenza strains (antigenic drift). Vaccine efficacy declines when there is a mismatch between its content and circulating strains. Current correlates of protection are merely based on serological parameters determined by haemagglutination inhibition or single radial haemolysis assays. However, there is ample evidence showing that these serological correlates of protection can both over- or underestimate the protective efficacy of influenza vaccines. Next-generation universal influenza vaccines that induce cross-reactive cellular immune responses (CD4+ and/or CD8+ T-cell responses) against conserved epitopes may overcome some of the shortcomings of the current inactivated vaccines by eliciting broader protection that lasts for several influenza seasons and potentially enhances pandemic preparedness. Assessment of cellular immune responses in clinical trials that evaluate the immunogenicity of these new generation vaccines is thus of utmost importance. Moreover, studies are needed to examine whether these cross-reactive cellular immune responses can be considered as new or complementary correlates of protection in the evaluation of traditional and next-generation influenza vaccines. An overview of the assays that can be applied to measure cell-mediated immune responses to influenza with their strengths and weaknesses is provided here.

Cryo-EM analysis workflow for viral vector-based gene therapy and vaccine development

Cryo-EM analysis workflow for viral vector-based gene therapy and vaccine development

Protective immunity of plant-produced African horse sickness virus serotype 5 chimaeric virus-like particles (VLPs) and viral protein 2 (VP2) vaccines in IFNAR-/- mice

Next generation vaccines have the capability to contribute to and revolutionise the veterinary vaccine industry. African horse sickness (AHS) is caused by an arbovirus infection and is characterised by respiratory distress and/or cardiovascular failure and is lethal to horses. Mandatory annual vaccination in endemic areas curtails disease occurrence and severity. However, development of a next generation AHSV vaccine, which is both safe and efficacious, has been an objective globally for years. In this study, both AHSV serotype 5 chimaeric virus-like particles (VLPs) and soluble viral protein 2 (VP2) were successfully produced in Nicotiana benthamiana ΔXT/FT plants, partially purified and validated by gel electrophoresis, transmission electron microscopy and liquid chromatography-mass spectrometry (LC-MS/MS) based peptide sequencing before vaccine formulation. IFNAR-/- mice vaccinated with the adjuvanted VLPs or VP2 antigens in a 10 µg prime-boost regime resulted in high titres of antibodies confirmed by both serum neutralising tests (SNTs) and enzyme-linked immunosorbent assays (ELISA). Although previous studies reported high titres of antibodies in horses when vaccinated with plant-produced AHS homogenous VLPs, this is the first study demonstrating the protective efficacy of both AHSV serotype 5 chimaeric VLPs and soluble AHSV-5 VP2 as vaccine candidates. Complementary to this, coating ELISA plates with the soluble VP2 has the potential to underpin serotype-specific serological assays.

Expression of mycobacterium tuberculosis induced SOCS3 and STAT3 and the implications on innate immunity in TB patients vs healthy contacts in high TB/HIV endemic setting: A cross-sectional analytical study.

BackgroundMycobacterium tuberculosis (TB) remains a disease of global health concern and a leading cause of mortality arising from an infectious agent. Protective immunity to TB remains unclear. Suppressor of cytokine signaling-3 (SOCS3) and signal transduction and activator of transcription-3 (STAT3) genes have shown potential to influence innate immunity. We, therefore, explored the expression of SOCS3 and STAT3 and their implications on the innate immunity in TB patients and their healthy close contacts.MethodsWe recruited 72 TB patients and 62 healthy contacts from a high TB and HIV endemic setting (Lusaka, Zambia). We used RT-PCRT and flow cytometry to quantify the expression of SOCS, STAT3 and cytokines respectively. Data was analysed Stata version 14.0 and figures were developed in GraphPad prism version 9.1.0 (221). Assessment for associations for categorical and continuous variables was analysed using the Chi-square test and Mann-Whitney test respectively. Spearman’s rank correlation was used to evaluate the relationship between SOCS3 and IL-6. A p-value < 0.05 was considered statistically significant.ResultsHealthy contacts markedly expressed SOCS3 in both unstimulated and stimulated whole blood in comparison to TB patients (p <0.0001). STAT3 was elevated in TB patients in TB patients in stimulated blood only. IL-6 (P = < 0.0001) and IL-10 (P = <0.0001), were significantly expressed in Healthy contacts in comparison to TB patients. TNF-α (p = 0.044) were markedly elevated in TB patients in comparison to healthy contacts. IL-6 and SOCS3 correlated significantly in healthy contacts only (r = 0.429, p = 0.02).ConclusionsBoth SOCS3 and STAT3 are genes of importance in mounting protective innate immunity against TB. We propose that SOCS3 stimulation and inhibition of STAT3 as possible approaches in gene therapy and vaccine development for TB.

Emergence of immune escape at dominant SARS-CoV-2 killer T cell epitope

SummaryWe studied the prevalent cytotoxic CD8 T cell response mounted against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Spike glycoprotein269-277 epitope (sequence YLQPRTFLL) via the most frequent human leukocyte antigen (HLA) class I worldwide, HLA A∗02. The Spike P272L mutation that has arisen in at least 112 different SARS-CoV-2 lineages to date, including in lineages classified as “variants of concern,” was not recognized by the large CD8 T cell response seen across cohorts of HLA A∗02+ convalescent patients and individuals vaccinated against SARS-CoV-2, despite these responses comprising of over 175 different individual T cell receptors. Viral escape at prevalent T cell epitopes restricted by high frequency HLAs may be particularly problematic when vaccine immunity is focused on a single protein such as SARS-CoV-2 Spike, providing a strong argument for inclusion of multiple viral proteins in next generation vaccines and highlighting the need for monitoring T cell escape in new SARS-CoV-2 variants.

“Mutation blacklist” and “mutation whitelist” of SARS-CoV-2

Over the past two years, scientists throughout the world have completed more than 6 million SARS-CoV-2 genome sequences. Today, the number of SARS-CoV-2 genomes exceeds the total number of all other viral genomes. These genomes are a record of the evolution of SARS-CoV-2 in the human host, and provide information on the emergence of mutations. In this study, analysis of these sequenced genomes identified 296,728 de novo mutations (DNMs), and found that six types of base substitutions reached saturation in the sequenced genome population. Based on this analysis, a “mutation blacklist” of SARS-CoV-2 was compiled. The loci on the “mutation blacklist” are highly conserved, and these mutations likely have detrimental effects on virus survival, replication, and transmission. This information is valuable for SARS-CoV-2 research on gene function, vaccine design, and drug development. Through association analysis of DNMs and viral transmission rates, we identified 185 DNMs that positively correlated with the SARS-CoV-2 transmission rate, and these DNMs where classified as the “mutation whitelist” of SARS-CoV-2. The mutations on the “mutation whitelist” are beneficial for SARS-CoV-2 transmission and could therefore be used to evaluate the transmissibility of new variants. The occurrence of mutations and the evolution of viruses are dynamic processes. To more effectively monitor the mutations and variants of SARS-CoV-2, we built a SARS-CoV-2 mutation and variantmonitoring and pre-warning system (MVMPS), which can monitor the occurrence and development of mutations and variants of SARS-CoV-2, as well as provide pre-warning for the prevention and control of SARS-CoV-2 (https://www.omicx.cn/). Additionally, this system could be used in real-time to update the “mutation whitelist” and “mutation blacklist” of SARS-CoV-2.

The attenuated ASFV strains MK-200 and FK-32/135 as possible models for investigation of protective immunity by ASFV infection.

African swine fever (ASF) is an infectious disease of domestic and wild pigs of all breeds and ages, with the acute form of the disease being characterized by high fever, hemorrhages in the reticuloendothelial system and a high mortality rate. Registered safe and efficacious ASF vaccines are not available. The development of experimental ASF vaccines, particularly live attenuated, have considerably intensified in the last years. There is much variability in experimental approaches undertaken by laboratories attempting to develop first generation vaccines, rendering it difficult to interpret and make comparisons across trials. ASF virus (ASFV) genotyping does not fully correlate with available cross-protection data and may be of limited value in predicting cross-protective vaccine efficacy. Recently, ASFV strains were assigned to a respective nine groups by seroimmunotype (from I to IX): in vivo the grouping is based on results of cross protection of pigs survived after their infection with a virulent strain (bioassay), while in vitro this grouping is based on hemadsorption inhibition assay (HADIA) data. Here we demonstrate the antigenic and protective properties of two attenuated ASFV strains MK200 and FK-32/135. Pronounced differences in the HADIA and in immunological test in animals allow us to consider them and the corresponding reference virulent strains of the ASFV of Mozambique-78 (seroimmunotype III, genotype V) and France-32 (seroimmunotype IV, genotype I) as useful models for studying the mechanisms of protective immunity and evaluation of the candidate vaccines.

Germinal center activity and B cell maturation are associated with protective antibody responses against Plasmodium pre-erythrocytic infection.

Blocking Plasmodium, the causative agent of malaria, at the asymptomatic pre-erythrocytic stage would abrogate disease pathology and prevent transmission. However, the lack of well-defined features within vaccine-elicited antibody responses that correlate with protection represents a major roadblock to improving on current generation vaccines. We vaccinated mice (BALB/cJ and C57BL/6J) with Py circumsporozoite protein (CSP), the major surface antigen on the sporozoite, and evaluated vaccine-elicited humoral immunity and identified immunological factors associated with protection after mosquito bite challenge. Vaccination achieved 60% sterile protection and otherwise delayed blood stage patency in BALB/cJ mice. In contrast, all C57BL/6J mice were infected similar to controls. Protection was mediated by antibodies and could be passively transferred from immunized BALB/cJ mice into naïve C57BL/6J. Dissection of the underlying immunological features of protection revealed early deficits in antibody titers and polyclonal avidity in C57BL/6J mice. Additionally, PyCSP-vaccination in BALB/cJ induced a significantly higher proportion of antigen-specific B-cells and class-switched memory B-cell (MBCs) populations than in C57BL/6J mice. Strikingly, C57BL/6J mice also had markedly fewer CSP-specific germinal center experienced B cells and class-switched MBCs compared to BALB/cJ mice. Analysis of the IgG γ chain repertoires by next generation sequencing in PyCSP-specific memory B-cell repertoires also revealed higher somatic hypermutation rates in BALB/cJ mice than in C57BL/6J mice. These findings indicate that the development of protective antibody responses in BALB/cJ mice in response to vaccination with PyCSP was associated with increased germinal center activity and somatic mutation compared to C57BL/6J mice, highlighting the key role B cell maturation may have in the development of vaccine-elicited protective antibodies against CSP.

Review of Diagnostic and Vaccination Approaches of Infectious Bursal Disease of Poultry

Infectious bursal disease, also known as Gumboro disease, is a highly contagious and acute viral disease of poultry characterised by the destruction of lymphoid cells. Diagnosis of infectious bursal disease involves consideration of the flocks’ history, clinical signs, and lesions. The objectives of this paper are to highlight various commonly used diagnostic methods for infectious bursal disease and to review advances made in diagnostic methods and vaccination strategies for infectious bursal disease, with special emphasis on the strengths and weaknesses of each of those techniques. Isolation of infectious bursal disease virus followed by its serological assay and histopathological examination of the bursa is regarded as the gold standard method of infectious bursal disease diagnosis. Serological tests such as agar gel, immune diffusion, enzyme-linked immuno sorbent assay, and viral neutralisation tests are commonly used laboratory assays in diagnosing infectious bursal disease viruses. Recently, the most accurate and relatively fast diagnostic method, the molecular technique, is widely used. The molecular diagnostic technique is the simplest and most sensitive of the diagnostic techniques reviewed. The virus causes immunosuppression, so the infected chicken recovers from the acute disease but becomes more susceptible to infections by other pathogens. Therefore, prevention is important and vaccination has become the principal control measure of infectious bursal disease virus infection in chickens. Conventional attenuated live and killed vaccines are the most commonly used vaccines. With the advancement of knowledge and technology, new generation or genetically-engineered vaccines like deoxyribonucleic acid and subunit vaccines have been used. Various vaccination strategies, such as in ovo, at hatch, and post hatch vaccination, are used. Hatchery vaccination is becoming a common practice. Based on this review paper, more affordable and effective infectious bursal disease vaccines that are affordable and readily available must be identified with further cost-benefit analysis.

Recommendations on qPCR/ddPCR assay validation by GCC.

Gene therapy, cell therapy and vaccine research have led to an increased use of qPCR/ddPCR in bioanalytical laboratories. CROs are progressively undertaking the development and validation of qPCR and ddPCR assays. Currently, however, there is limited regulatory guidance for the use of qPCR and a complete lack of any regulatory guidelines for the use of the newer ddPCR to support regulated bioanalysis. Hence, the Global CRO Council in Bioanalysis (GCC) has issued this White Paper to provide; 1) a consensus on the different validation parameters required to support qPCR/ddPCR assays; 2) a harmonized approach to their validation and 3) a consistent development of standard operating procedures (SOPs) for all the bioanalytical laboratories using these techniques.

Severe Acute Respiratory Syndrome Coronavirus 2 Infection Versus Vaccination in Pregnancy: Implications for Maternal and Infant Immunity.

BackgroundSARS-CoV-2 infection has been associated with adverse maternal and neonatal outcomes, yet uptake of SARS-CoV-2 vaccines during pregnancy and lactation has been slow. As a result, millions of pregnant and lactating women and their infants remain susceptible to the virus.MethodsWe measured Spike-specific immunoglobulin G (anti-S IgG) and A (anti-S IgA) in serum and breastmilk (BM) samples from 3 prospective mother-infant cohorts recruited in two academic medical centers. The primary aim was to determine the impact of maternal SARS-CoV-2 immunization vs infection and their timing on systemic and mucosal immunity.ResultsThe study included 28 mothers infected with SARS-CoV-2 in late pregnancy (INF), 11 uninfected mothers who received 2 doses of the BNT162b2 vaccine in the latter half of pregnancy (VAX-P) and 12 uninfected mothers who received 2 doses of BNT162b2 during lactation (VAX-L). VAX dyads had significantly higher serum anti-S IgG compared to INF dyads (p < .0001), while INF mothers had higher BM: serum anti-S IgA ratios compared to VAX mothers (p = .0001). Median IgG placental transfer ratios were significantly higher in VAX-P compared to INF mothers (p < 0.0001). There was a significant positive correlation between maternal and neonatal serum anti-S IgG after vaccination (r = 0.68, p = 0.013), but not infection.ConclusionsBNT161b2 vaccination in late pregnancy or lactation enhances systemic immunity through serum anti-S Ig, while SARS-CoV-2 infection induces mucosal over systemic immunity more efficiently through BM Ig production. Next generation vaccines boosting mucosal immunity could provide additional protection to the mother-infant dyad. Future studies should focus on identifying the optimal timing of primary and/or booster maternal vaccination for maximal benefit.

2021 White Paper on Recent Issues in Bioanalysis: Mass Spec of Proteins, Extracellular Vesicles, CRISPR, Chiral Assays, Oligos; Nanomedicines Bioanalysis; ICH M10 Section 7.1; Non-Liquid & Rare Matrices; Regulatory Inputs(Part 1A - Recommendations on Endogenous Compounds, Small Molecules, Complex Methods, Regulated Mass Spec of Large Molecules, Small Molecule, PoC &Part 1B - Regulatory Agencies' Inputs on Bioanalysis, Biomarkers, Immunogenicity, Gene

The 15th edition of the Workshop on Recent Issues in Bioanalysis (15th WRIB) was held on 27 September to 1 October 2021. Even with a last-minute move from in-person to virtual, an overwhelmingly high number of nearly 900 professionals representing pharma and biotech companies, contract research organizations (CROs), and multiple regulatory agencies still eagerly convened to actively discuss the most current topics of interest in bioanalysis. The 15th WRIB included 3 Main Workshops and 7 Specialized Workshops that together spanned 1week in order to allow exhaustive and thorough coverage of all major issues in bioanalysis, biomarkers, immunogenicity, gene therapy, cell therapy and vaccines. Moreover, in-depth workshops on biomarker assay development and validation (BAV) (focused on clarifying the confusion created by the increased use of the term "Context of Use - COU"); mass spectrometry of proteins (therapeutic, biomarker and transgene); state-of-the-art cytometry innovation and validation; and, critical reagent and positive control generation were the special features of the 15th edition. This 2021 White Paper encompasses recommendations emerging from the extensive discussions held during the workshop, and is aimed to provide the bioanalytical community with key information and practical solutions on topics and issues addressed, in an effort to enable advances in scientific excellence, improved quality and better regulatory compliance. Due to its length, the 2021 edition of this comprehensive White Paper has been divided into three parts for editorial reasons. This publication (Part 1A) covers the recommendations on Endogenous Compounds, Small Molecules, Complex Methods, Regulated Mass Spec of Large Molecules, Small Molecule, PoC. Part 1B covers the Regulatory Agencies' Inputs on Bioanalysis, Biomarkers, Immunogenicity, Gene & Cell Therapy and Vaccine. Part 2 (ISR for Biomarkers, Liquid Biopsies, Spectral Cytometry, Inhalation/Oral & Multispecific Biotherapeutics, Accuracy/LLOQ for Flow Cytometry) and Part 3 (TAb/NAb, Viral Vector CDx, Shedding Assays; CRISPR/Cas9 & CAR-T Immunogenicity; PCR & Vaccine Assay Performance; ADA Assay Comparabil ity & Cut Point Appropriateness) are published in volume 14 of Bioanalysis, issues 10 and 11 (2022), respectively.