Generation and characterization of dromedary Tenascin-C and Tenascin-W specific antibodies

Dhaouadi, Sayda; Murdamoothoo, Devadarssen; Tounsi, A.; Erne, William; Benabderrazek, Rahma; Benlasfar, Zakaria; Hendaoui, L.; Chiquet‐Ehrismann, Ruth; Boubaker, Samir; Orend, Gertraud; Hendaoui, Ismaïl; Bouhaouala‐Zahar, Balkiss

doi:10.1016/j.bbrc.2020.05.077

Cited by 7 publications

(7 citation statements)

References 28 publications

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“…Therefore, the BotI-like toxin molecule is immunogenic and elicited a humoral response. These results are in line with our previous studies using scorpion toxin ( 36 , 37 ), metallo-beta-lactamase enzyme ( 55 ), or Tenacin protein ( 56 ) to raise specific immune responses in dromedary. We then constructed an immune VHH library of 3.0×10 6 individual clones, 87% of which carried a phagemid with an insert of the correct size of a VHH.…”

Section: Discussionsupporting

confidence: 91%

Neutralizing Dromedary-Derived Nanobodies Against BotI-Like Toxin From the Most Hazardous Scorpion Venom in the Middle East and North Africa Region

et al. 2022

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Scorpion envenoming is a severe health problem in many regions causing significant clinical toxic effects and fatalities. In the Middle East/North Africa (MENA) region, Buthidae scorpion stings are responsible for devastating toxic outcomes in human. The only available specific immunotherapeutic treatment is based on IgG fragments of animal origin. To overcome the limitations of classical immunotherapy, we have demonstrated the in vivo efficacy of NbF12-10 bispecific nanobody at preclinical level. Nanobodies were developed against BotI analogues belonging to a distinct structural and antigenic group of scorpion toxins, occurring in the MENA region. From Buthus occitanus tunetanus venom, BotI-like toxin was purified. The 41 N-terminal amino acid residues were sequenced, and the LD50 was estimated at 40 ng/mouse. The BotI-like toxin was used for dromedary immunization. An immune VHH library was constructed, and after screening, two nanobodies were selected with nanomolar and sub-nanomolar affinity and recognizing an overlapping epitope. NbBotI-01 was able to neutralize 50% of the lethal effect of 13 LD50 BotI-like toxins in mice when injected by i.c.v route, whereas NbBotI-17 neutralized 50% of the lethal effect of 7 LD50. Interestingly, NbBotI-01 completely reduced the lethal effect of the 2 LD50 of BotG50 when injected at 1:4 molar ratio excess. More interestingly, an equimolar mixture of NbBotI-01 with NbF12-10 neutralized completely the lethal effect of 7 and 5 LD50 of BotG50 or AahG50, at 1:4 and 1:2 molar ratio, respectively. Hence, NbBotI-01 and NbF12-10 display synergic effects, leading to a novel therapeutic candidate for treating Buthus occitanus scorpion stings in the MENA region.

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Section: Discussionsupporting

confidence: 91%

Neutralizing Dromedary-Derived Nanobodies Against BotI-Like Toxin From the Most Hazardous Scorpion Venom in the Middle East and North Africa Region

et al. 2022

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“…We purified recombinant TNC of human origin and used this molecule to elicit a potent immune response in the dromedary. A substantial proportion of polyclonal heavy chain IgG subclasses bound to TNC and recognized TNC by IF staining in tumor tissue (51). This encouraged us to generate nanobodies.…”

Section: Discussionmentioning

confidence: 99%

“…The anti-hTNC nanobody phage-display VHH-library was constructed as previously described with slight modifications (39,49,50). Briefly, 3 days after the last boost of antigen injection, 150 mL of anti-coagulated blood sample was collected from the jugular vein of the immunized dromedary as recently detailed (51). Peripheral blood mononuclear cells (PBMCs) were extracted by density gradient centrifugation using Lymphoprep (catalog number 17-829 LONZA, Basel, Switzerland).…”

Section: Generation Of the Phage-display Vhh-librarymentioning

confidence: 99%

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Novel Human Tenascin-C Function-Blocking Camel Single Domain Nanobodies

et al. 2021

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The extracellular matrix (ECM) molecule Tenascin-C (TNC) is well-known to promote tumor progression by multiple mechanisms. However, reliable TNC detection in tissues of tumor banks remains limited. Therefore, we generated dromedary single-domain nanobodies Nb3 and Nb4 highly specific for human TNC (hTNC) and characterized the interaction with TNC by several approaches including ELISA, western blot, isothermal fluorescence titration and negative electron microscopic imaging. Our results revealed binding of both nanobodies to distinct sequences within fibronectin type III repeats of hTNC. By immunofluroescence and immunohistochemical imaging we observed that both nanobodies detected TNC expression in PFA and paraffin embedded human tissue from ulcerative colitis, solid tumors and liver metastasis. As TNC impairs cell adhesion to fibronectin we determined whether the nanobodies abolished this TNC function. Indeed, Nb3 and Nb4 restored adhesion of tumor and mesangial cells on a fibronectin/TNC substratum. We recently showed that TNC orchestrates the immune-suppressive tumor microenvironment involving chemoretention, causing tethering of CD11c+ myeloid/dendritic cells in the stroma. Here, we document that immobilization of DC2.4 dendritic cells by a CCL21 adsorbed TNC substratum was blocked by both nanobodies. Altogether, our novel TNC specific nanobodies could offer valuable tools for detection of TNC in the clinical practice and may be useful to inhibit the immune-suppressive and other functions of TNC in cancer and other diseases.

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“…These results promote the newly developed TN-C-specific Nbs as novel molecular tools for detecting TN-C in tumors (diagnostic biomarker) as well as for their potential use as anti-cancer treatment options by blocking the oncogenic functions of TN-C (Table 1). Although TN-W-specific single-domain antibodies have been developed in the dromedary as well (188), so far they only have been tested for the specific detection and imaging of TN-W and not for blocking its functions.…”

Section: Function Blocking Antibodiesmentioning

confidence: 99%

Revisiting the Tenascins: Exploitable as Cancer Targets?

Tucker

Degen

2022

Front. Oncol.

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For their full manifestation, tumors require support from the surrounding tumor microenvironment (TME), which includes a specific extracellular matrix (ECM), vasculature, and a variety of non-malignant host cells. Together, these components form a tumor-permissive niche that significantly differs from physiological conditions. While the TME helps to promote tumor progression, its special composition also provides potential targets for anti-cancer therapy. Targeting tumor-specific ECM molecules and stromal cells or disrupting aberrant mesenchyme-cancer communications might normalize the TME and improve cancer treatment outcome. The tenascins are a family of large, multifunctional extracellular glycoproteins consisting of four members. Although each have been described to be expressed in the ECM surrounding cancer cells, tenascin-C and tenascin-W are currently the most promising candidates for exploitability and clinical use as they are highly expressed in various tumor stroma with relatively low abundance in healthy tissues. Here, we review what is known about expression of all four tenascin family members in tumors, followed by a more thorough discussion on tenascin-C and tenascin-W focusing on their oncogenic functions and their potential as diagnostic and/or targetable molecules for anti-cancer treatment purposes.

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Generation and characterization of dromedary Tenascin-C and Tenascin-W specific antibodies

Cited by 7 publications

References 28 publications

Neutralizing Dromedary-Derived Nanobodies Against BotI-Like Toxin From the Most Hazardous Scorpion Venom in the Middle East and North Africa Region

Neutralizing Dromedary-Derived Nanobodies Against BotI-Like Toxin From the Most Hazardous Scorpion Venom in the Middle East and North Africa Region

Novel Human Tenascin-C Function-Blocking Camel Single Domain Nanobodies

Revisiting the Tenascins: Exploitable as Cancer Targets?

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