Kinetics of hepatitis B virus replication in anti-HBc positive/HBsAg-negative people with HIV switching to tenofovir sparing therapy.

Objectives: To unravel the still unexplored HBV-replicative kinetics in anti-HBc-positive/HBsAg-negative people-with-HIV (PWH) suspending tenofovir disoproxil-fumarate/tenofovir-alafenamide (TDF/TAF).

Methods: A total of 101 anti-HBc-positive/HBsAg-negative PWH switching to TDF/TAF-sparing therapy were included. Serum HBV-DNA and HBV-RNA were quantified by droplet-digital-PCR at switching (T0), within 12 months (T1) and 12-24 months postswitch (T2).

Chagas Disease in the Non-Endemic Area of Rome, Italy: Ten Years of Experience and a Brief Overview.

Chagas disease (CD) is a parasitic infection endemic in Latin America and also affects patients in Western countries due to migration flows. This has a significant impact on health services worldwide due to its high morbidity and mortality burden. This paper aims to share our experience at the National Institute for Infectious Diseases "Lazzaro Spallanzani", IRCCS, in Rome, Italy, where to date, a total of 47 patients-mainly Bolivian women-diagnosed with CD have received treatment with benznidazole, with all but one presenting with chronic disease.

3D topographies promote macrophage M2d-Subset differentiation.

cellular models denote a crucial part of drug discovery programs as they aid in identifying successful drug candidates based on their initial efficacy and potency. While tremendous headway has been achieved in improving 2D and 3D culture techniques, there is still a need for physiologically relevant systems that can mimic or alter cellular responses without the addition of external biochemical stimuli. A way forward to alter cellular responses is using physical cues, like 3D topographical inorganic substrates, to differentiate macrophage-like cells.

Determinants of worse liver-related outcome according to HDV infection among HBsAg positive persons living with HIV: Data from the ICONA cohort.

Objectives: We aimed to study hepatitis D virus (HDV) prevalence and risk of progression to severe liver-related events (SLRE) in HBsAg positive people living with HIV (PLWH) in Italy; role of HDV-RNA copy levels, HCV coinfection and nadir CD4 counts were also investigated.

Methods: People living with HIV (PLWH) from Italian Foundation cohort Naïve antiretrovirals (ICONA) with available HBsAg and HDV Ab were enrolled. HBsAg, HDV Ab, HDV-RNA and HDV genotypes were tested.

Bulk Reformatting of Antibody Fragments Displayed on the Surface of Yeast Cells to Final IgG Format for Mammalian Production.

While yeast surface display (YSD) has gained traction for antibody hit discovery efforts with the first therapeutic YSD-isolated antibody sintilimab approved in 2018, a major drawback that remains is the time-consuming reformatting of monoclonal antibody (mAb) candidates. By using a Golden Gate cloning (GGC)-dependent workflow, the bulk transfer of genetic information can be performed from antibody fragments displayed on yeast cells to a bidirectional mammalian expression vector. Herein, we describe in-depth protocols for the reformatting of mAbs, starting from the generation of Fab fragment libraries in YSD vectors and ending up with IgG molecules in bidirectional mammalian vectors in a consolidated two-pot, two-step procedure.

Generation of a symmetrical trispecific NK cell engager based on a two-in-one antibody.

To construct a trispecific IgG-like antibody at least three different binding moieties need to be combined, which results in a complex architecture and challenging production of these molecules. Here we report for the first time the construction of trispecific natural killer cell engagers based on a previously reported two-in-one antibody combined with a novel anti-CD16a common light chain module identified by yeast surface display (YSD) screening of chicken-derived immune libraries. The resulting antibodies simultaneously target epidermal growth factor receptor (EGFR), programmed death-ligand 1 (PD-L1) and CD16a with two Fab fragments, resulting in specific cellular binding properties on EGFR/PD-L1 double positive tumor cells and a potent ADCC effect.

Parahydrogen-induced polarization allows 2000-fold signal enhancement in biologically active derivatives of the peptide-based drug octreotide.

Octreotide, a somatostatin analogue, has shown its efficacy for the diagnostics and treatment of various types of cancer, i.e., in octreotide scan, as radio-marker after labelling with a radiopharmaceutical.

Targeted Phagocytosis Induction for Cancer Immunotherapy via Bispecific MerTK-Engaging Antibodies.

The Tyro, Axl, and MerTK receptors (TAMRs) play a significant role in the clearance of apoptotic cells. In this work, the spotlight was set on MerTK, as it is one of the prominent TAMRs expressed on the surface of macrophages and dendritic cells. MerTK-specific antibodies were previously isolated from a transgenic rat-derived immune library with suitable biophysical properties.

TriTECM: A tetrafunctional T-cell engaging antibody with built-in risk mitigation of cytokine release syndrome.

Harnessing the innate power of T cells for therapeutic benefit has seen many shortcomings due to cytotoxicity in the past, but still remains a very attractive mechanism of action for immune-modulating biotherapeutics. With the intent of expanding the therapeutic window for T-cell targeting biotherapeutics, we present an attenuated trispecific T-cell engager (TCE) combined with an anti- interleukin 6 receptor (IL-6R) binding moiety in order to modulate cytokine activity (TriTECM). Overshooting cytokine release, culminating in cytokine release syndrome (CRS), is one of the severest adverse effects observed with T-cell immunotherapies, where the IL-6/IL-6R axis is known to play a pivotal role.

Streamlining the Transition From Yeast Surface Display of Antibody Fragment Immune Libraries to the Production as IgG Format in Mammalian Cells.

Yeast-surface display (YSD) is commonly applied to screen Fab immune or naïve libraries for binders of predefined target molecules. However, reformatting of isolated variants represents a time-intensive bottleneck. Herein, we present a novel approach to facilitate a lean transition from antibody screening using YSD Fab libraries to the production of full-length IgG antibodies in Expi293-F cells.

Antibody Library Screening Using Yeast Biopanning and Fluorescence-Activated Cell Sorting.

Yeast surface display (YSD) emerged as a prominent screening methodology for the isolation of monoclonal antibodies (mAbs) against various antigens. However, phage display remains the gold standard in cell panning-based screenings to isolate mAbs against difficult-to-screen targets, such as G-protein coupled receptors (GPCR) and ion channels. Herein we describe a step-by-step protocol to establish and perform the isolation of mAbs using YSD in a fluorescence-activated cell sorting (FACS)-assisted biopanning manner, yielding a variety of antibodies binding their antigen with high affinity in the natural environment of the cell.

A Generic Strategy to Generate Bifunctional Two-in-One Antibodies by Chicken Immunization.

Various formats of bispecific antibodies exist, among them Two-in-One antibodies in which each Fab arm can bind to two different antigens. Their IgG-like architecture accounts for low immunogenicity and also circumvents laborious engineering and purification steps to facilitate correct chain pairing. Here we report for the first time the identification of a Two-in-One antibody by yeast surface display (YSD) screening of chicken-derived immune libraries.

Milking the Cow: Cattle-Derived Chimeric Ultralong CDR-H3 Antibodies and Their Engineered CDR-H3-Only Knobbody Counterparts Targeting Epidermal Growth Factor Receptor Elicit Potent NK Cell-Mediated Cytotoxicity.

In this work, we have generated epidermal growth factor receptor (EGFR)-specific cattle-derived ultralong CDR-H3 antibodies by combining cattle immunization with yeast surface display. After immunization, ultralong CDR-H3 regions were specifically amplified and grafted onto an IGHV1-7 scaffold by homologous recombination to facilitate Fab display. Antigen-specific clones were readily obtained by fluorescence-activated cell sorting (FACS) and reformatted as chimeric antibodies.

Inflammation and microbial translocation measured prior to combination antiretroviral therapy (cART) and long-term probability of clinical progression in people living with HIV.

Background: Despite the effectiveness of cART, people living with HIV still experience an increased risk of serious non-AIDS events, as compared to the HIV negative population. Whether pre-cART microbial translocation (MT) and systemic inflammation might predict morbidity/mortality during suppressive cART, independently of other known risk factors, is still unclear. Thus, we aimed to investigate the role of pre-cART inflammation and MT as predictors of clinical progression in HIV+ patients enrolled in the Icona Foundation Study Cohort.

Recombinant Antibody Production Using a Dual-Promoter Single Plasmid System.

Monoclonal antibodies (mAbs) have demonstrated tremendous effects on the treatment of various disease indications and remain the fastest growing class of therapeutics. Production of recombinant antibodies is performed using mammalian expression systems to facilitate native antibody folding and post-translational modifications. Generally, mAb expression systems utilize co-transfection of heavy chain () and light chain () genes encoded on separate plasmids.

Design of a Trispecific Checkpoint Inhibitor and Natural Killer Cell Engager Based on a 2 + 1 Common Light Chain Antibody Architecture.

Natural killer cell engagers gained enormous interest in recent years due to their potent anti-tumor activity and favorable safety profile. Simultaneously, chicken-derived antibodies entered clinical studies paving the way for avian-derived therapeutics. In this study, we describe the affinity maturation of a common light chain (cLC)-based, chicken-derived antibody targeting EGFR, followed by utilization of the same light chain for the isolation of CD16a- and PD-L1-specific monoclonal antibodies.

Expeditious Generation of Biparatopic Common Light Chain Antibodies Chicken Immunization and Yeast Display Screening.

Bispecific (BsAb) and biparatopic (BpAb) antibodies emerged as promising formats for therapeutic biologics exhibiting tailor-made functional properties. Over recent years, chicken-derived antibodies have gained traction for diagnostic and therapeutic applications due to their broad epitope coverage and convenience of library generation. Here we report the first generation of a biparatopic common light chain (cLC) chicken-derived antibody by an epitope binning-based screening approach using yeast surface display.

From cell line development to the formulated drug product: The art of manufacturing therapeutic monoclonal antibodies.

Therapeutic monoclonal antibodies and related products have steadily grown to become the dominant product class within the biopharmaceutical market. Production of antibodies requires special precautions to ensure safety and efficacy of the product. In particular, minimizing antibody product heterogeneity is crucial as drug substance variants may impair the activity, efficacy, safety, and pharmacokinetic properties of an antibody, consequently resulting in the failure of a product in pre-clinical and clinical development.

Engineering therapeutic antibodies for patient safety: tackling the immunogenicity problem.

Established monoclonal antibodies (mAbs) allow treatment of cancers, autoimmune diseases and other severe illnesses. Side effects either arise due to interaction with the target protein and its biology or result from of the patient's immune system reacting to the foreign protein. This immunogenic reaction against therapeutic antibodies is dependent on various factors.

Schistosomiasis, strongyloidiasis and Chagas disease: the leading imported neglected tropical diseases in Italy.

Background: In recent years, an increasing number of individuals affected by neglected tropical diseases (NTDs) have been observed in Italy, due to migration, international travels and climate changes. Reliable data on the current NTD epidemiology in Italy and the health system preparedness on this issue are not available.

Methods: We report the results of a survey on selected NTDs (schistosomiasis, strongyloidiasis, echinococcosis, Chagas disease, leishmaniasis, cysticercosis, filariasis and scabies) in nine Italian sentinel centres, in order to investigate their occurrence throughout the country and identify which ones are a priority for public health interventions, development of protocols for case management, and training activities.

Unexpected human cases of cutaneous anthrax in Latium region, Italy, August 2017: integrated human-animal investigation of epidemiological, clinical, microbiological and ecological factors.

On 31 August, a veterinarian and a farmworker were hospitalised for skin lesions. Both had been exposed to a dead cow on 19 August on a farm near Rome, where eight further cattle died of confirmed anthrax later the same month. At admission, the first case showed a black depressed eschar and another smaller lesion on one hand.

[The biological resources and molecule archives organization: turn a need into an opportunity for the Smart Specialization Strategy of the Lazio region.].

Smart Specialization Strategy (S3) of Lazio defines smart specialization strategies to bring out the excellence of the territory with prospects of success on the global market. Chemical-pharmaceutical, biomedical and biotechnological field is one of the 7 sectors considered of greatest interest for the S3. Key engine of biotechnology development are biological materials and associated data, stored in biobanks.