Biologics

Background: Introducing unnatural base pairs into a natural, double-stranded DNA construct is a powerful tool within synthetic biology. Accordingly, the ability to detect these unnatural base pairs has many applications, including the study and detection of semisynthetic organisms. Objective and Methods: The work described here aimed to select human antibodies for the specific recognition of Hirao’s base pair dDs–dPn in various natural DNA contexts by using a combination of phage and yeast display technologies. Results: We selected a total of six antibodies in yeast-displayed scFv format, and further tested three of these antibodies in soluble form as minibodies and IgGs. We also describe an assay that can be used to detect plasmids containing dDs–dPn pair. Conclusions: Our antibodies did not afford the desired specificity or sensitivity for detection of a single unnatural base pair among thousands of natural. However, our data indicate not only that such detection is possible but also that these antibodies may be candidates for further affinity and specificity maturation. Full article

Ustekinumab is a fully human IgG1k monoclonal antibody that binds with high affinity and specificity to the p40 subunit of interleukins (IL-) 12 and 23, inhibiting their activity by preventing binding to their receptors. The European extension of the patent (Supplementary Protection Certificate) of ustekinumab expired on 20 July 2024. Biosimilar alternatives to ustekinumab are now an additional option for treating patients. The efficacy data for this drug in moderate-to-severe psoriasis obtained both from clinical trials and indirect comparisons through meta-analyses, are superior to those of etanercept and adalimumab, and its safety profile is more favorable than that of tumor necrosis factor (TNF) inhibitors. Several ustekinumab biosimilars have already been approved by regulatory agencies: between October 2023 and October 2024, Wezlana^® (Amgen ABP 654), Uzpruvo^® (Alvotech AVT04) and Pyzchiva^® (Samsung/Bioepis SB17) have been approved by both the US Food and Drug Administration (FDA) and the European Medicines Agency (EMA). SteQeyma^® (Celltrion Healthcare CT-P43) was approved by the EMA in August 2024. Otulfi^® (Fresenius Kabi/Formycon) was approved by the FDA in October 2024. Several other potential biosimilar candidates are under development, including BAT2206 (Bio-Thera), DMB-3115 (Dong-A ST), QX001S (Qyuns Therapeutic), BFI-751 (BioFactura), NeuLara (Neuclone), ONS3040 (Oncobiologics), and BOW090 (Epirus Biopharmaceuticals). In most cases, these monoclonal antibodies are expressed in cell lines (e.g., Chinese Hamster Ovary, CHO) different from those used for the originator (Sp2/0 spleen cell murine myeloma); of note, the cell line of origin is not a requirement for biosimilarity in the totality-of-evidence comparison exercise and may facilitate the production and reduce the immunogenicity of biosimilars originated in CHO cultures. This narrative review summarizes the available data on characteristics of the full comparability exercises and comparative clinical trials of these drugs. Full article

Antiretroviral therapy (ART) has significantly extended the lifespan of people living with Human Immunodeficiency Virus (HIV) or Acquired Immunodeficiency Syndrome (AIDS), thereby transforming the disease into a manageable chronic condition. However, this increased longevity has led to a higher incidence of non-AIDS-defining cancers (NADCs) among this population. In this holistic review, we explore the complex interactions between HIV, ART, and cancer development, focusing on how ART influences tumor initiation and progression in people living with HIV/AIDS (PLWHA). Our findings from this reveal several critical aspects of cancer risk in PLWHA. Firstly, while ART restores immune function, it does not fully normalize it. Chronic immune activation and persistent inflammation continue to be prevalent, creating a conducive environment for oncogenesis. Additionally, PLWHA are more susceptible to persistent infections with oncogenic viruses such as human papillomavirus (HPV) and Epstein–Barr virus (EBV), further increasing cancer risk. Some ART drugs have been implicated in genotoxicity and mitochondrial dysfunction, potentially promoting tumorigenesis. ART-induced metabolic changes, including insulin resistance and dyslipidemia, are also associated with heightened cancer risk. Common NADCs in PLWHA include lung cancer, liver cancer, anal cancer, and Hodgkin lymphoma, each with distinct etiologies linked to both HIV-related and ART-related factors. The interplay between HIV infection, chronic inflammation, immune restoration via ART, and the direct effects of ART drugs creates a unique cancer risk profile in PLWHA. Although ART reduces the incidence of AIDS-defining cancers, it does not confer the same protective effect against NADCs. Persistent HIV-related inflammation and immune activation, despite viral suppression, are key factors in cancer development. Additionally, long-term exposure to ART may introduce new oncogenic risks. These insights highlight the need for integrated cancer screening and prevention strategies tailored to PLWHA. Future research is needed to focus on identifying biomarkers for early cancer detection and developing ART regimens with lower oncogenic potential. Healthcare providers should be vigilant in monitoring PLWHA for cancer and adopt comprehensive screening protocols to mitigate the increased cancer risk associated with ART. Full article

Protein aggregation may lead to detrimental changes in brain and several other tissues. Amyloids or large protein aggregates are formed in different brain areas under multiple diseases classified as proteinopathies. However, our understanding of the initiation, elongation, and spread of amyloid aggregates is limited. Our current knowledge about these diseases is generic and we lack specific mechanisms for several diseases affecting memory, movement, and behavior. Multiple studies have indicated the involvement of vesicular transport in the spread of aggregates formed inside the brain. For example, the trafficking of amyloid precursor protein (APP) occurs from Golgi to Endosome using an adapter protein complex. Amyloids, once formed, may also affect cholesterol (an important membrane constituent), homeostasis, and overall membranous transport. A disruption of vesicular transport could be deleterious for synaptic neurotransmission. Alterations caused by amyloid proteins in vesicular transport may form a feedback loop and thus contribute further to the pathogenesis of Alzheimer’s disease (AD) and many others. In this review, we are providing recent updates on this crisscross puzzle and exploring an evolving correlation between amyloid formation and vesicular transport. Full article

The integration of biologics in endovascularly treated intracranial aneurysms is a significant area of focus in an evolving field. By presenting the clinical relevance, pathogenesis, management (historical and current), and emerging biologics themselves, this work provides a broad overview of the current landscape of the biologics under current investigation. Growth factors, cytokines, and biologic-coated coils are compared and described as modalities to increase healing, aneurysm occlusion, and long-term recovery. These emerging biologics may increase the efficacy and durability of less invasive endovascular methods and potentially change standard practice with continued exploration. Full article

This study investigated the potential of chicken byproduct hydrolysates (CBH) characterized by a mixture of low-molecular-weight peptides (<1.35 kDa) and larger peptides (<17.5 kDa) as a treatment for metabolic syndrome (MS), from a histological and histopathological point of view. This study aimed to evaluate the effects of CBH obtained using plant proteases (BP: B. pinguin, BK: B. karatas, BRO: bromelain) on the histological and histopathological analysis of the liver and kidney in an MS-induced murine model. Methods: Thirty adult male Wistar rats were randomly assigned to six groups (n = 5): (1) standard diet (STD); (2) MS with a hypercaloric diet (MS + HC); (3) CBH-BP (200 mg/kg of body weight); (4) CBH-BK (200 mg/kg of body weight); (5) CBH-BRO (200 mg/kg of body weight); (6) carnosine (CAR) 50 mg/kg of body weight. Liver and kidney samples were processed by conventional hematoxylin and eosin (H&E) histological techniques, Masson’s trichrome stain (MTS), and the periodic acid–Schiff (PAS) histochemical method. A scoring scale was used for the histopathological evaluation with scores ranging from 0 (normal tissue) to 4 (severe damage). Results: CBHs demonstrated a significant therapeutic effect (p < 0.05) on hepatic and renal morphological alterations induced by MS. Hepatic scores for lipid inclusions, vascular congestion, and cellular alteration were all reduced to below two. Similarly, renal scores for tubular degeneration, vascular congestion, and dilation of Bowman’s space were also decreased to less than two. The therapeutic efficacy of CBHs was comparable to that of the positive control, CAR (β-alanyl-L-histidine). Conclusions: CBH-BP, CBH-BK, and CBH-BRO treatments reduced morphological alterations observed in liver and kidney tissues, which is relevant since from a histological and histopathological point of view, it allows us to understand at the cellular and tissue level the effects that these treatments can have on a living organism, indicating a potential to improve organ health in people with MS. Full article

Helminths have developed intricate mechanisms to survive and evade the host’s immune responses. Hence, understanding the excretory-secretory products (ESPs) by helminths is crucial for developing control tools, including drug targets, vaccines, and potential therapies for inflammatory and metabolic disorders caused by them. Proteomics, the large-scale analysis of proteins, offers a powerful approach to unravel the complex proteomes of helminths and gain insights into their biology. Proteomics, as a science that delves into the functions of proteins, has the potential to revolutionize clinical therapies against parasitic infections that have developed anthelminthic resistance. Proteomic technologies lay a framework for accompanying genomic, reverse genetics, and pharmacokinetic approaches to provide more profound or broader coverage of the cellular mechanisms that underlie the response to anthelmintics. With the development of vaccines against helminth infections, proteomics has brought a major change to parasitology. The proteome of helminths can be analyzed comprehensively, revealing the complex network of proteins that enable parasite survival and pathogenicity. Furthermore, it reveals how parasites interact with hosts’ immune systems. The current article reviews the latest advancements in helminth proteomics and highlights their valuable contributions to the search for anthelminthic vaccines. Full article

During early development of biopharmaceuticals, suboptimal producing clones and production conditions can result in limited quantities of high-purity products. Here we describe a systematic approach, which requires minimal amounts of protein (~10 mg) to assess critical quality attributes of a monoclonal antibody (mAb). A commercial anti-PCSK9 IgG2 (evolocumab, Repatha^®) and an early-stage biosimilar candidate were compared head-to-head using a range of high-throughput physicochemical and in-vitro binding analytical methods. Overall, both mAbs were shown to be highly pure and primarily monomeric, to share an identical primary structure, and to have similar higher-order structural integrity, apparent solubility, aggregation propensity, and physical stability profiles under temperature and pH stress conditions. Low levels of dimers were detected for the innovator (1.2%) and the biosimilar candidate mAb (0.3%), which also presented fragments (1.2%). Regarding charge heterogeneity, the amount of the main charge isoform was 53.6% for the innovator and 61.6% for the biosimilar candidate mAb. Acidic species were 38% for the innovator and 30% for the biosimilar candidate. Variations in the relative content of a few N-glycan species were found. The in-vitro binding affinity to PCSK9 was monitored, and no differences were detected. The mathematical approach called “error spectral difference” (ESD), proposed herein, enabled a quantitative comparison of the biophysical datasets. The workflow used in the present work to characterize CQAs at early stages is helpful in supporting the development of biosimilar mAb candidates. Full article

Inflammatory bowel disease (IBD), which encompasses both ulcerative colitis (UC) and Crohn’s disease (CD), is a major health burden worldwide. There are increasing concerns surrounding the impacts of this disease due to significant rises in the prevalence rates of IBD across the world. In consideration of the complexities of managing IBD along with this marked rise in prevalence and incidence, developing new forms of treatment for this condition has become a major priority. In recent years, a potential new form of treatment for IBD has emerged in the form of biologic therapies. While there is a high level of optimism due to the development of these therapies, there is also a clear need to evaluate their effectiveness, and their overall safety profiles. For this review, we have evaluated three specific biologics used for the treatment IBD. More precisely, the focus of this review is to analyze and critically appraise the literature for vedolizumab, ustekinumab, and golimumab, and determine their roles in the management of UC and CD, respectively. After doing so, we have also briefly synthesized important new findings regarding the role of dietary and nutritional approaches. In doing so, we have aimed to contextualize the findings regarding biologics, and, in order to evaluate potential new treatment approaches for the future to augment biologic therapies, we have discussed the potential for combined approaches that incorporate the usage of both biologics and nutritional interventions for patients. Full article

The solute carrier family 35 (SLC35) comprises multiple members of transporters, including a group of proteins known as nucleotide sugar transporters (NSTs), an adenosine triphosphate (ATP) transporter, 3′-phosphoadenosine 5′-phosphosulfate (PAPS) transporters, and transporters of unknown function. To date, seven subfamilies (A to G) and 32 members have been classified into this large SLC35 family. Since the majority of glycosylation reactions occur within the lumen of the endoplasmic reticulum (ER) and Golgi apparatus, the functions of NSTs are indispensable for the delivery of substrates for glycosylation. Recent studies have revealed the diverse functions of this family of proteins in the regulation of numerous biological processes, including development, differentiation, proliferation, and disease progression. Furthermore, several congenital disorders of glycosylation (CDGs) resulting from variations in the SLC35 family member genes have been identified. To elucidate the pathology of these diseases, a variety of knockout mice harboring mutations in the family member genes have been generated and employed as animal models for CDGs. This review presents a historical overview of the SLC35 family, with a particular focus on recent advances in research on the functions of this family and their relationship to human diseases. Full article

The measurement of rabies-neutralizing antibody is important for monitoring the response to rabies vaccination. For all the purposes of measurement, such as routine monitoring of vaccine response in humans and animals, serosurveys, and biologics qualification, accurate and precise results are necessary. The risks associated with sample handling variation, which may impact the test results, can be overlooked within a laboratory. To determine the robustness of rabies-neutralizing antibodies in human and animal serum, samples were treated to mimic various possible deviations in the sample handling protocols. Potential deviations were designed to investigate common client inquiries and possible sample conditions experienced during shipping, storage, and laboratory processes. The treatments included the duration that sera were kept at a temperature greater than that of a refrigerator (room temperature, zero hours to two weeks), the number and duration of heat inactivation treatments (i.e., heat inactivation directly from freezer storage, etc.), the number of freeze–thaw cycles (zero, four, or six cycles), and the storage duration of sample dilutions in chamber slides before the addition of virus (zero hours to overnight). The results provided evidence for the robustness of rabies antibodies and the antibodies’ neutralizing function in uncontaminated, clear human and animal serum. In addition, prolonged heat exposure was identified as exerting the greatest impact on the measurement of rabies antibodies. Full article

Purpose: Knee osteoarthritis (KOA) is a very common cartilage disorder affecting millions of people globally and is characterized by pain, stiffness, swelling, loss of articular cartilage, and osteophyte formation, resulting in disability. The presently available treatments for KOA are palliative. Hence, there is a need to explore a non-surgical treatment portfolio. Bone marrow aspirate concentrate (BMAC) is one of the predominant attention-drawing managements/treatments for KOA in recent times due to its potential advantages of disease-modifying and regeneration capacities. Principle: This study aimed to evaluate the role of single-injection autologous BMAC as a therapeutic option in the treatment of KOA and evaluate the functional and clinical outcomes of KOA patients. In this study, 132 patients with KOA (Kellgren and Lawrence (KL) grade II and III) were included as per the inclusion criteria. Autologous bone marrow was aspirated and separated, and concentrated bone marrow aspirate was administered into the knee joint of the affected individual. Results: At the end of the 12th month (end of the follow-up period), 95% of patients showed complete pain relief and improvement in joint function, which shows that the results were promising and encouraging. Unpaired t-test results also indicated that the two-tailed p-value is less than 0.0001, and the difference is extremely statistically significant. No adverse effects were observed in the study patients. Conclusions: BMAC therapy has potential, with satisfactory, efficient, and durable results in KL grades II and III in KOA patients. This can be a safe alternative therapy in the treatment of KOA, especially in the early grades of OA. In summary, to the best of our knowledge, this is the first study from India that evaluated BMAC efficacy both subjectively and objectively in KOA (KL-II and KL-III) patients. Full article

Hyperpigmentation skin disorders are marked by an abnormal accumulation or export of melanin pigment synthesized within melanocytes and pose a significant aesthetic concern. The search for novel natural compounds that exhibit pharmacological potential for treating pigmentation disorders is growing. In this study, kahweol (KW) and cafestol (CFS), two structural analogs of coffee diterpenes, were evaluated and compared for their effects on melanogenesis using B16F10 mouse melanoma cells and primary human melanocytes derived from Asian and African American skin. To the best of our knowledge, there are no reports of the effects of KW and CFS on melanogenesis yet. We first screened nontoxic concentrations of both compounds using an MTS assay after 72 h incubations and subsequently tested their effects on melanin synthesis and export. Cellular tyrosinase activity and cell-free mushroom tyrosinase activity were assayed to study the mechanisms of melanogenesis suppression. Human melanocytes from a moderately pigmented donor (HEMn-MP cells) and from a darkly pigmented donor (HEMn-DP cells) were next examined, and effects on cellular viability, melanin content, cellular tyrosinase activity, and melanin export (quantitated via dendricity) were similarly examined for both compounds. Our results show that KW and CFS did not significantly affect intracellular melanin content but suppressed extracellular melanin in B16F10 cells and dendritic parameters in human melanocytes, indicating their unique capacity to target extracellular melanogenesis and melanin export. Although KW showed a greater extracellular melanogenesis inhibitory capacity in B16F10 cells, in both primary melanocyte cells, CFS emerged as a potent inhibitor of melanin export compared to KW. Together, these results reveal novel modes of action of both compounds and indicate a promise to use CFS as a novel candidate for treating hyperpigmentation disorders of the human skin for clinical and cosmetic use. Additional research is necessary to shed light on the molecular pathways and the efficacy of melanogenesis inhibition by CFS in 3D human skin equivalents and in vivo studies. Full article

Viral vectors are agents enabling gene transfer and genome editing and have widespread utility across the healthcare and biotechnology sectors. In January 2023, the International Bureau for Weights and Measures’ Consultative Committee for Amount of Substance (CCQM) held a workshop on Metrology for Viral systems as molecular tools. The workshop brought together international leaders from across regulatory, industry, government science, and metrology sectors to better understand key challenges for the community: Exploring current limitations in the measurement of virus-derived, virus-based, and virus-like systems in terms of quantification and characterisation; surveying the state-of-the-art in analytical methods and reference material provision for these entities; and initiating a dialog for the strategic development and implementation of suitable standardisation approaches for this sector. This article presents the workshop background and rationale, presentation summaries, conclusions, and recommendations. Full article

Background: Janus kinase (JAK) inhibitors represent a novel class of oral therapies showing efficacy in treating ulcerative colitis (UC) and Crohn’s disease (CD), challenging conventional treatment paradigms. Summary: This review provides an overview of the potential novel uses of JAK inhibitors, focusing on their current approved indications and exploring possibilities beyond these indications. Tofacitinib and filgotinib are approved for UC, while upadacitinib is approved for both UC and CD. Additionally, their potential in acute severe UC, as steroid alternatives, and in managing fistulizing CD or extraintestinal manifestations are discussed. Key Message: JAK inhibitors play an important role in IBD (inflammatory bowel disease) treatment; however, clinicians must balance their promising efficacy with safety concerns. Individualized care and vigilance are essential for optimizing therapeutic benefits while mitigating potential adverse effects. Further research is necessary to clarify their efficacy, safety, and potential applications. Full article

Traumatic Brain Injury (TBI) is a debilitating condition that poses a significant public health concern. Historically linked to motor vehicle accidents, the epidemiology of TBI has evolved. Falls now emerge as the predominant cause, particularly among older adults. Sport-related TBIs have also garnered increased attention due to concerns regarding long-term neurological sequelae. To date, therapeutic interventions remain limited and have yet to yield substantial clinical benefits. Cell-based therapies offer promising avenues for neural repair and regeneration: endogenous stem cell therapies capitalize on endogenous pools that can be triggered by the injury and further enhanced by therapeutic approaches. In contrast, exogenous cell therapies provide an exogenous source of cells. However, challenges such as age-related decline in neurogenesis, age-related inflammation, and the heterogeneity of TBI present significant hurdles to overcome. Moreover, translating stem cell research from the laboratory to clinical applications necessitates the adherence to good manufacturing practice standards, which presents distinct obstacles. Addressing these challenges requires a multifaceted approach, including careful patient selection in clinical trials, appropriate experimental models, and the optimization of therapeutic techniques. Ultimately, a combination of strategies is likely to yield the most promising outcomes in the pursuit of effective TBI therapies. Full article

Cardiovascular disease remains the main cause of mortality in the 21st century. Hypertension, vessel atherosclerosis, and familial hypercholesterolemia (FH) are responsible for increased mortality and morbidity in patients. Therapies for cardiovascular disease are based on drug treatment options, but in the era of precision medicine, personalized treatments are being developed. Studies have shown that these conditions have a strong genetic background, creating an opportunity for the implementation of gene therapy for these diseases. Currently, gene therapy is not widely used in clinical practice. Recent advances in this research field are making gene therapy a very promising preventive and therapeutic tool for cardiovascular disease. Essential hypertension’s (EH) pathophysiology is mostly based on the activation of both the sympathetic nervous system and the renin angiotensin aldosterone system (RAAS), natriuretic peptide production, and endothelial dysfunction. Plasmid DNA and viral vectors can be used, targeting the main mechanisms in the pathogenesis of EH. Many preclinical studies have been developed across the years, presenting a significant decrease in blood pressure. Nevertheless, no clinical studies have been developed studying the implementation of gene therapy in EH. Atherosclerotic damage is caused by monogenic diseases or is deteriorated by the activation of inflammation in the vessel wall. Gene therapy studies have been developed in the pre- and clinical phases targeting the lipoprotein and cholesterol metabolism and the inflammation of the vessels. FH is a common inherited metabolic disease associated with high levels of cholesterol in the blood. Clinical trials of gene therapy have been developed and presented optimistic results. In this review, the challenges of gene therapy for cardiovascular disease are outlined. Nevertheless, more clinical trials are needed to be performed for the development of convenient and safe drug schemes for our patients. Full article

Tick-borne diseases account for a large proportion of vector-borne illnesses. They include, for example, a variety of infections caused by bacteria, spirochetes, viruses, rickettsiae, and protozoa. We aim to present a review that demonstrates the connection between the diagnosis, treatment, prevention, and the significance of certain emergency tick-borne diseases in humans and their clinical–epidemiological features. This review covers three diseases: anaplasmosis, ehrlichiosis, and babesiosis. The emergence of ehrlichiosis and anaplasmosis is become more frequently diagnosed as the cause of human infections, as animal reservoirs and tick vectors have increased in numbers and humans have inhabited areas where reservoir and tick populations are high. They belong to the order Rickettsiales and the family Anaplasmataceae, and the clinical manifestations typically coexist. Furthermore, prompt diagnosis and appropriate treatment are critical to the patient’s recovery. Similar to malaria, babesiosis causes hemolysis. It is spread by intraerythrocytic protozoa, and the parasitemia dictates how severe it can get. Left untreated, some patients might have a fatal outcome. The correct diagnosis can be difficult sometimes; that is why an in-depth knowledge of the diseases is required. Prevention, prompt diagnosis, and treatment of these tick-borne diseases depend on the understanding of their clinical, epidemiological, and laboratory features. Full article

Heparan sulfate proteoglycans are highly glycosylated proteins in which heparan sulfate, a glycosaminoglycan sugar chain, is an acidic sugar chain consisting of a repeating disaccharide structure of glucuronic acid and N-acetylglucosamine is locally sulfated. Syndecan, one of the transmembrane HSPGs, functions as a receptor that transmits signals from the extracellular microenvironment to the inside of the cell. In the vascular system, heparan sulfate proteoglycans, a major component of the glycocalyx, enable the binding of various plasma-derived molecules due to their diversity, epimerization of glycosaminoglycans chains, long chains, and sulfation. Heparan sulfate proteoglycans present in the extracellular matrix serve as a reservoir for bioactive molecules such as chemokines, cytokines, and growth factors. Aberrant expression of heparan sulfate proteoglycans, heparanase, and sulfatase is observed in many pathological conditions. Therefore, it can be applied to therapeutic strategies for a wide range of fields including Alzheimer’s disease, heart failure, cancer, organ transplants, diabetes, chronic inflammation, aging, and autoimmune diseases. Full article