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41.
42.
Is there an approach for optimizing the specific binding of antibodies to target cells?
Ario De Marco, invited lecture at foreign university

Found in: osebi
Keywords: Nanobodies, avidity, modeling, off-target accumulation
Published: 31.01.2017; Views: 1320; Downloads: 0
.pdf Fulltext (81,48 KB)

43.
Nanobody technology: principles & applications
Ario De Marco, invited lecture at foreign university

Abstract: Antibodies possess unattainable capacity to bind selectively and at high affinity their cognates. For this reason they have been largely used in applications which rely on specific molecular recognition. Biosensors, nanoparticles, and even cells can be functionalized with antibodies for improving sensitivity and target specificity. However, conventional antibodies (IgGs) are large molecules (150 kDa) that are difficult to engineer. In the last years, antibody fragments have become more and more popular as an effective alternative and specifically nanobodies raised enthusiasm because of their minimal mass (14 kDa), high stability, relative similarity to human sequences, and simplified mutagenesis. Pre-immune nanobody libraries have the further advantage of enabling blind selection for antigens that can be used to discriminate between subpopulations of cells and vesicles. The panning can be performed directly on intact cells and the resulting binders are specific for the native antigen conformation
Found in: osebi
Keywords: Nanobodies, in vitro panning, cancer biomarkers, recombinant antibody engineering
Published: 03.05.2017; Views: 1852; Downloads: 0
.pdf Fulltext (37,92 KB)

44.
Biotechnological applications of nanobody pre-immune libraries
Ario De Marco, invited lecture at foreign university

Found in: osebi
Keywords: nanobodies, pre-immune libraries, in vitro selection, biomarkers, recombinant proteins
Published: 16.05.2017; Views: 1146; Downloads: 0
.pdf Fulltext (331,92 KB)

45.
Whole-cell biopanning with a synthetic phage display library of nanobodies enabled the recovery of follicle-stimulating hormone receptor inhibitors
Ronan Crepin, Gianluca Veggiani, Selma Djender, Anne Beugnet, Francois Planeix, Christophe Pichon, Sandrine Moutel, Sebastian Amigorena, Franck Pérez, Nicolae Ghinea, Ario De Marco, 2017, original scientific article

Abstract: Antibodies are essential reagents that are increasingly used in diagnostics and therapy. Their specificity and capacity to recognize their native antigen are critical characteristics for their in vivo application. Follicle-stimulating hormone receptor is a GPCR protein regulating ovarian follicular maturation and spermatogenesis. Recently, its potentiality as a cancer biomarker has been demonstrated but no antibody suitable for in vivo tumor targeting and treatment has been characterized so far. In this paper we describe the first successful attempt to recover recombinant antibodies against the FSHR and that: i) are directly panned from a pre-immune library using whole cells expressing the target receptor at their surface; ii) show inhibitory activity towards the FSH-induced cAMP accumulation; iii) do not share the same epitope with the natural binder FSH; iv) can be produced inexpensively as mono- or bivalent functional molecules in the bacterial cytoplasm. We expect that the proposed biopanning strategy will be profitable to identify useful functional antibodies for further members of the GPCR class.
Found in: osebi
Keywords: nanobodies, GPCR proteins, FSHR, panning, pre-immune antibody library
Published: 19.10.2017; Views: 1034; Downloads: 0
.pdf Fulltext (982,69 KB)

46.
Engineered cross-reacting nanobodies simplify comparative oncology between humans and dogs
Elisa Mazzega, Ario De Marco, 2017, original scientific article

Found in: osebi
Keywords: HER2, nanobodies, inter-species cross-reactivity, thermophoresis
Published: 19.10.2017; Views: 1254; Downloads: 0
.pdf Fulltext (1,58 MB)

47.
48.
Nanobodies: towards rational design of immune-reagents
Ario De Marco, 2017, published scientific conference contribution abstract (invited lecture)

Abstract: Antibodies are irreplaceable reagents in both research and clinical practice. Despite their relevance, the structural complexity of conventional mono- and polyclonal antibodies (IgG) has always been a limit for their engineering towards reagents optimized for specific applications, such as in vivo diagnostics and therapy. Furthermore, their isolation is time consuming, their production expensive, and their functionalization results often in heterogeneous macromolecule populations. These drawbacks promoted the search for both innovative antibody isolation strategies and alternative scaffolds. In vitro panning of pre-immune collections of recombinant antibody fragments allows for the simple and fast recovery of binders. Since they did not undergo somatic maturation, their affinity for targets can be insufficient but on the other hand they can be rapidly mutated by standard molecular biology techniques to generate second-generation antibodies among which to identify clones with improved characteristics. Both stochastic and rational methods have been proposed for the optimization process. Random mutagenesis followed by panning at stringent conditions has been successful used to select binders with improved physical characteristics. Rational methods try to identify in silico key residues involved in the regulation of specific antibody features, such as stability or binding affinity. The accuracy of these methods usually depends on the calculation resources. In this perspective, smaller molecules can be analyzed “better” than larger because of their restricted number of residues. Nanobodies small dimensions have been long appreciated since enable better tissue penetration, shorter clearance time, higher yields. Now it becomes evident that this characteristic makes them also optimal objects for modeling.
Found in: osebi
Keywords: recombinant antibody modeling, nanobody engineering, molecular dynamics and docking
Published: 21.03.2018; Views: 1278; Downloads: 0
.pdf Fulltext (638,20 KB)

49.
In vitro isolation of anti Alexandrium minutum nanobodies from a pre-immune library
Ario De Marco, Marina Cabrini, Alfred Beran, Elisa Mazzega, 2017, published scientific conference contribution abstract

Found in: osebi
Keywords: nanobodies, toxic microalgae, biopanning
Published: 04.12.2017; Views: 777; Downloads: 8
.pdf Fulltext (61,76 KB)
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50.
Naked protein nanoparticles
Ario De Marco, 2017, published scientific conference contribution abstract (invited lecture)

Found in: osebi
Keywords: Protein nanoparticles, targeted delivery, cell uptake
Published: 15.12.2017; Views: 823; Downloads: 140
.pdf Fulltext (2,09 MB)

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