Interference of p53:Twist1 interaction through competing nanobodies

D'Agostino, Serena; Mazzega, Elisa; Praček, Katja; Piccinin, Sara; Pivetta, Flavia; Armellin, Michela; Fortuna, Sara; Maestro, Roberta; Marco, Ario de

doi:10.1016/j.ijbiomac.2021.11.160

Cited by 4 publications

(4 citation statements)

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“…Our work confirms that in vitro selection of nanobodies performed imposing stringent parameters represents a valuable modality to isolate binders specific for particular domains, such as antigen epitopes and Ab paratopes [ 34 , 35 ]. We obtained a large and structurally highly heterogeneous array of clones with specificity for a specific idiotype.…”

Section: Discussionsupporting

confidence: 74%

Nanobodies Selectively Binding to the Idiotype of a Dengue Virus Neutralizing Antibody Do Not Necessarily Mimic the Viral Epitope

et al. 2023

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Vaccination against dengue virus is challenged by the fact that a generic immune response can induce antibody-dependent-enhancement (ADE) in secondary infections. Only some antibodies targeting a quaternary epitope formed by the dimerization of the virus protein E possess sufficient neutralizing capacity. Therefore, the immunization with anti-idiotypic antibodies of neutralizing antibodies might represent a safe vaccination strategy. Starting from a large pre-immune library, we succeeded in isolating a wide set of anti-idiotypic nanobodies characterized by selective and strong binding to the paratope of the neutralizing antibody 1C10. However, the mice immunized with such constructs did not produce effective antibodies, despite at least some of them eliciting an immune response selective for the nanobody variable regions. The results suggest that complex conformational epitopes might be difficult to be recreated by anti-idiotypic structures. The selection process of the anti-idiotypic candidates might be optimized by applying epitope mapping and modeling approaches aimed at identifying the key residues that is necessary to bind to trigger selective immune response.

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Section: Discussionsupporting

confidence: 74%

Nanobodies Selectively Binding to the Idiotype of a Dengue Virus Neutralizing Antibody Do Not Necessarily Mimic the Viral Epitope

et al. 2023

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“…The same principle could be applied in the future for the selection of nanobodies specific for EV sub-populations characterized by the presence of exclusive biomarkers. In this perspective, we already demonstrated the possibility to isolate nanobodies from a phage display pre-immune library panning directly on EVs or on specific epitopes of soluble antigens [ 8 , 37 ].…”

Section: Resultsmentioning

confidence: 99%

Macroporous Epoxy-Based Monoliths Functionalized with Anti-CD63 Nanobodies for Effective Isolation of Extracellular Vesicles in Urine

Neumair

D’Ercole

March

et al. 2023

IJMS

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Extracellular vesicles (EVs) have enormous potential for the implementation of liquid biopsy and as effective drug delivery means, but the fulfilment of these expectations requires overcoming at least two bottlenecks relative to their purification, namely the finalization of reliable and affordable protocols for: (i) EV sub-population selective isolation and (ii) the scalability of their production/isolation from complex biological fluids. In this work, we demonstrated that these objectives can be achieved by a conceptually new affinity chromatography platform composed of a macroporous epoxy monolith matrix functionalized with anti-CD63 nanobodies with afflux of samples and buffers regulated through a pump. Such a system successfully captured and released integral EVs from urine samples and showed negligible unspecific binding for circulating proteins. Additionally, size discrimination of eluted EVs was achieved by different elution approaches (competitive versus pH-dependent). The physical characteristics of monolith material and the inexpensive production of recombinant nanobodies make scaling-up the capture unit feasible and affordable. Additionally, the availability of nanobodies for further specific EV biomarkers will allow for the preparation of monolithic affinity filters selective for different EV subclasses.

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“…Furthermore, a nanobody was recently reported to inhibit the interaction between Twist1 and p53 [ 26 ]. Twist1 facilitates the degradation of p53 by the negative regulator murine double minute 2-Protein (MDM2), which in wild-type cancer inhibits the transcriptional activity of p53 and induces its degradation.…”

Section: Intrabodies Against Cytoplasmic or Nucleus Located Taasmentioning

confidence: 99%

“…They can be selected by phage display or ribosomal display from immune, naïve or synthetic single domain antibody repertoires [ 16 , 17 , 18 , 19 ] and inactivate their targets by altering their conformation or interfere with the binding of the target protein to its corresponding binding partner. sdAbs were isolated against intracellular TAAs [ 19 ], including the recent examples of hypoxia-inducible factor-1 (HIF-1) [ 24 ], serine/threonine protein kinase AKT2 [ 25 ], p53 C-terminal region involved in the interaction with Twist1 [ 26 ] and chemokine receptor US28 [ 27 ]. In addition to targeting intracellular TAAs, intrabodies have been selected against intracellular neoantigens, for example against HRASG12V [ 28 , 29 , 30 ] HRASG12V, KRASG12D, KRASG13D, NRASQ61R, KRASG12V, KRASQ61H [ 31 ] H - and K - Ras G12V [ 32 ], p21Ras [ 33 ] and KRASG12V [ 34 ].…”

Section: Introductionmentioning

confidence: 99%

Therapeutic Potential of Intrabodies for Cancer Immunotherapy: Current Status and Future Directions

Böldicke

2022

Antibodies

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Tumor cells are characterized by overexpressed tumor-associated antigens or mutated neoantigens, which are expressed on the cell surface or intracellularly. One strategy of cancer immunotherapy is to target cell-surface-expressed tumor-associated antigens (TAAs) with therapeutic antibodies. For targeting TAAs or neoantigens, adoptive T-cell therapies with activated autologous T cells from cancer patients transduced with novel recombinant TCRs or chimeric antigen receptors have been successfully applied. Many TAAs and most neoantigens are expressed in the cytoplasm or nucleus of tumor cells. As alternative to adoptive T-cell therapy, the mRNA of intracellular tumor antigens can be depleted by RNAi, the corresponding genes or proteins deleted by CRISPR-Cas or inactivated by kinase inhibitors or by intrabodies, respectively. Intrabodies are suitable to knockdown TAAs and neoantigens without off-target effects. RNA sequencing and proteome analysis of single tumor cells combined with computational methods is bringing forward the identification of new neoantigens for the selection of anti-cancer intrabodies, which can be easily performed using phage display antibody repertoires. For specifically delivering intrabodies into tumor cells, the usage of new capsid-modified adeno-associated viruses and lipid nanoparticles coupled with specific ligands to cell surface receptors can be used and might bring cancer intrabodies into the clinic.

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Interference of p53:Twist1 interaction through competing nanobodies

Cited by 4 publications

References 47 publications

Nanobodies Selectively Binding to the Idiotype of a Dengue Virus Neutralizing Antibody Do Not Necessarily Mimic the Viral Epitope

Nanobodies Selectively Binding to the Idiotype of a Dengue Virus Neutralizing Antibody Do Not Necessarily Mimic the Viral Epitope

Macroporous Epoxy-Based Monoliths Functionalized with Anti-CD63 Nanobodies for Effective Isolation of Extracellular Vesicles in Urine

Therapeutic Potential of Intrabodies for Cancer Immunotherapy: Current Status and Future Directions

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