As part of the GBC (Global Bioanalysis Consortium), the L3 assay format team has focused on reviewing common platforms used to support ligand binding assays in the detection of biotherapeutics. The ...following review is an overview of discussions and presentations from around the globe with a group of experts from different companies to allow an international harmonization of common practices and suggestions for different platforms. Some of the major platforms include Gyrolab, Erenna, RIA, AlphaLISA, Delfia, Immuno-PCR, Luminex, BIAcore, and ELISAs. The review is meant to support bioanalysts in taking decisions between different platforms depending on the needs of the analyte with a number of recommendations to help integration of platforms into a GLP environment.
As oligonucleotides (ONs) and similar nucleic acid therapeutic modalities enter development pipelines, there is continual need to develop bioanalytical methodologies addressing unique challenges they ...pose. Novel ONs back bone chemistries, especially those enabling stereochemical control, and base modifications are being exploited to improve pharmacological properties, potency, and increase half-lives. These changes have strained established methods, oftentimes precluding development of assays sensitive and specific enough to meet the needs of preclinical programs. For stereopure ONs representing a single molecular species, nontrivial presence of chain-shortened metabolites in biological samples necessitate assays with high specificity. To meet these needs, this report presents a toolbox of novel techniques, easy to implement for existing hybridization-ligation enzyme-linked immunosorbent assay formats, which address this challenge and yield significant sensitivity and specificity enhancements. Ligation efficiency was improved up to 61-fold through addition of polyethylene glycol, betaine, or dimethylsulfoxide, mitigating major differences among sequence-matched ONs of varying stereopurity, enabling sensitivities below 0.100 ng/mL for quantitation. These improvements enabled further refinement of capture probe designs engendering sufficient specificity to discriminate N-1 chain-shortened metabolites at both the 5′ and 3′ end of the ONs. These generalizable methods advance the performance of mainstay bioanalytical assays, facilitating research and development of innovative ONs therapeutics.
Many biotherapeutics currently in development have complex mechanisms of action and contain more than one domain, each with a specific role or function. Examples include antibody–drug conjugates ...(ADC), PEGylated, fusion proteins and bi-specific antibodies. As with any biotherapeutic molecule, a multi-domain biotherapeutic (MDB) can elicit immune responses resulting in the production of specific anti-drug antibodies (ADA) when administered to patients. As it is beneficial to align industry standards for evaluating immunogenicity of MDBs, this paper highlights pertinent immunogenicity risk factors and describes steps involved in the design of a testing strategy to detect and characterize binding (non-neutralizing and neutralizing, NAb) ADAs. In a common tier based approach, samples identified as ADA screen positive are confirmed for the binding specificity of the antibodies to the drug molecule via a confirmatory assay. The confirmation of specificity is generally considered as a critical step of the tier based approach in overall ADA response evaluation. Further characterization of domain specificity of polyclonal anti-MDB ADA response may be required based on the analysis of molecule specific risk factors. A risk based approach in evaluating the presence of NAbs for MDB is discussed in this article. Analysis of domain-specific neutralizing antibody reactivity should be based on the risk assessment as well as the information learned during binding ADA evaluation. Situations where additional characterization of NAb specificity is possible and justified are discussed. Case studies demonstrating applicability of the risk factor based approach are presented. In general, the presence of a domain with high immunogenicity risk or presence of a domain with high endogenous protein homology may result in an overall high immunogenicity risk level for the entire MDB and can benefit from domain specificity characterization of immune response. For low immunogenicity risk MDBs, domain specificity characterization could be re-considered at later clinical phases based on the need to explain specific clinical observations. Inclusion of domain specificity characterization in early phase clinical studies for MDBs with limited clinical immunogenicity experience may be considered to help understand its value in later clinical development. It is beneficial and is recommended to have a well-defined plan for the characterization of ADA domain specificity and data analysis prior to the initiation of sample testing. Overall, best practices for immunogenicity evaluation of complex MDBs are discussed.
Purpose
– The purpose of this paper is to present the Andaman and Nicobar Islands (ANI) as a case to delineate contextual and often paradoxical challenges in implementing centrally sponsored schemes ...in a small island economy. Government reports cite ANI's record on health outcomes to be impressive when comparing it with other Union Territories (UTs). However, using examples of two public health and nutrition schemes, the paper argues that unless ANI's development indicators are compared over few years, public policy may fail to identity its development challenges and continue its rhetoric while in reality, health outcomes may be regressing.
Design/methodology/approach
– The paper used case study research design involving mixed method approach. Quantitative data were secondary in nature, collected from local government departments. Two of the three districts of ANI formed the sample of the study. Main tools used for qualitative analysis were observation and informal interviews with government functionaries and other stakeholders.
Findings
– The findings point to the paradox of superior public spending and infrastructure in ANI in midst of deteriorating public health and child nutrition outcomes. The paper concluded that any policy making on island's trajectory of development is not only incomplete, but also faulty unless its special context is considered as a starting point.
Research limitations/implications
– Ethnographic studies on the health-seeking behaviour of ANI communities can reveal trends in access to health services and its reasons, but that was beyond the scope of the paper.
Originality/value
– The paper is arguably the first to identify development paradoxes in implementation of government health and nutrition programmes in ANI. The paper poses further questions on unpacking the paradox of development in the islands. It urges policy makers and administrators to reconsider the efficacy of centrally sponsored schemes that are implemented without being contextualised for a small island economy like ANI.
Purpose: Normal epithelial cell specific-1 ( NES1 )/ kallikrein 10 gene is expressed in normal mammary and prostate epithelial cells, but the expression of NES1 mRNA and protein is markedly
reduced ...in established breast and prostate cancer cell lines although the NES1 gene is intact. Here, we wished to assess whether NES1 expression is down-regulated in primary breast cancers.
Experimental Design: We developed and used an in situ hybridization technique with an antisense NES1 probe to detect NES1 mRNA in sections of normal breast specimens, typical
and atypical ductal hyperplasia, ductal carcinoma in situ , and infiltrating ductal carcinoma.
Results: All of the 30 normal breast specimens showed high NES1 expression. Notably, 18 (75%) of 24 typical and atypical breast hyperplasia
specimens showed high NES1 expression, with weak-to-moderate expression in 6 (25%). Significantly, 13 (46%) of 28 ductal carcinoma
in situ specimens lacked NES1 expression, and the remaining 15 (54%) showed weak-to-moderate expression. Finally, 29 of 30 (97%)
infiltrating ductal carcinoma grades I-III samples lacked NES1 mRNA, with weak expression in the remaining one sample.
Conclusions: Our results demonstrate that NES1 mRNA is expressed in normal breast tissue and benign lesions, with loss of NES1 expression
during tumor progression. We suggest that NES1 expression may serve as a molecular tool in the study of breast cancer progression.
Studies with larger series of specimens should help assess whether NES1 expression can be a diagnostic and/or prognostic marker
in breast and other cancers.
In contemporary governance discourse, accountability is found linked to almost every conceivable aspect of ‘good governance’ – from developmental effectiveness to empowerment. However, ...accountability, both as a concept and a strategy, has been reduced to rhetoric while large public programmes especially on health and nutrition continue to record dismal child nutrition outcomes even after six decades of democracy in India. Child malnutrition remains one of the most challenging public health and development issues in India. The article argues that local accountability processes and relationships between users and providers in a community are actual markers of service delivery as opposed to dominant practices of techno-managerial checks. Local institutional design shape actions of frontline health providers in service delivery which can explain differential child nutrition outcomes. The article is one of the original attempts in studying accountability in public-health service delivery from the prism of institutions in a developing democratic country like India.
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