Background: The targeted delivery of bioactive molecules with antibodies specific to tumorassociated antigens represents a promising strategy for improving the efficacy of tumor therapy. The large isoform of tenascin-C, an abundant glycoprotein of the tumor extracellular matrix, is strongly overexpressed in adult tissue undergoing tissue remodeling, including wound healing and neoplasia, and has been implicated in a variety of different cancers while being virtually undetectable in most normal adult tissues. Experimental Design: We have used antibody phage technology to generate good-quality human recombinant antibodies (F16 and P12) specific to the alternatively spliced domains A1 and D of the large isoform of tenascin-C. The tumor-targeting properties of F16 and P12 were assessed by biodistribution studies in tumor xenografts using the antibodies in small immunoprotein (SIP) format. Results: SIP(F16) selectively accumulated at the tumor site with 4.5%ID/g at 24 hours in the U87 glioblastoma model but was rapidly cleared from other organs (tumor-to-organ ratios, f10:1). The accumulation of SIP(P12) in the tumor was lower compared with SIP(F16) and persistent levels of radioactivity were observed in the intestine. Conclusions:These data suggest that the F16 antibody, specific to domain A1of tenascin-C, is a promising building block for the development of antibody-based pharmaceuticals in view of its excellent tumor-targeting performance and the strong expression of the antigen in a variety of primary and metastatic tumors.A promising avenue toward the development of more selective anticancer therapies consists in the targeted delivery of bioactive molecules (antibody constant regions, cytokines, drugs, radionuclides, photosensitizers, procoagulant factors, etc.) to the tumor environment by means of ligands specific to good-quality tumor-associated antigens and endowed with suitable pharmacokinetic properties (1 -7).Antigens that are preferentially expressed in the modified tumor extracellular matrix are, in many respects, ideal targets for tumor-targeting applications (2, 8). Extracellular matrix components are often more abundant and more stable than antigens located on the surface of tumor cells. Furthermore, they typically exhibit a low shedding profile and are well accessible to agents (such as antibody derivatives) coming from the bloodstream. In collaboration with the Zardi group (Genova, Italy), our group has extensively shown the tumortargeting potential of antibodies directed against components of the tumor extracellular matrix using, as an example, the L19 human monoclonal antibody (9), specific to the extradomain B of fibronectin (EDB), a marker of angiogenesis (10 -12). The L19 antibody was shown to efficiently target tumor neovasculature and stromal structures in animal models of cancer (13) The EDB domain of fibronectin is a good-quality marker of angiogenesis, which is overexpressed in a variety of solid tumors (renal cell carcinoma, colorectal carcinoma, hepatocellular carcinoma, high-g...

We had previously reported that splice isoforms of tenascin-C containing the extra-domain C are virtually absent in normal adult tissues but are highly abundant in high-grade astrocytomas, with a prominent peri-vascular pattern of expression. We now report that the extra-domain C of tenascin-C is strongly expressed in the majority of lung cancers, with a vascular and stromal pattern of expression. Using antibody phage technology, we have generated a human monoclonal antibody (G11), with a dissociation constant K(D) = 4.2 nM for the human domain C. The G11 antibody, expressed in scFv and in mini-antibody (SIP) format, as well as a scFv-interleukin-2 fusion protein, was then characterized in quantitative biodistribution studies using mice grafted subcutaneously with U87 gliomas, revealing a selective tumor uptake, with tumor/blood ratios up to 11.8:1 at 24 h. A radioiodinated preparation of SIP(G11) was also investigated in a double tracer study using an orthotopic rat glioma model, confirming the antibody's ability to preferentially localize at the tumor site, with tumor/brain ratios superior to the ones observed with (18)F-fluorodeoxyglucose. These tumor-targeting properties, together with the strong immunohistochemical staining of human tumor sections, indicate that the G11 antibody may be used as a portable targeting moiety for the selective delivery of imaging and therapeutic agents to gliomas and lung tumors.

Biologic treatment options such as tumor necrosis factor (TNF) inhibitors have revolutionized the treatment of inflammatory diseases, including rheumatoid arthritis. Recent data suggest, however, that full and long-lasting responses to TNF inhibitors are limited because of the activation of the pro-inflammatory TH17/interleukin (IL)-17 pathway in patients. Therefore, dual TNF/IL-17A inhibition is an attractive avenue to achieve superior efficacy levels in such diseases. Based on the marketed anti-TNF antibody adalimumab, we generated the bispecific TNF/IL-17A-binding FynomAb COVA322. FynomAbs are fusion proteins of an antibody and a Fyn SH3-derived binding protein. COVA322 was characterized in detail and showed a remarkable ability to inhibit TNF and IL-17A in vitro and in vivo. Through its unique mode-of-action of inhibiting simultaneously TNF and the IL-17A homodimer, COVA322 represents a promising drug candidate for the treatment of inflammatory diseases. COVA322 is currently being tested in a Phase 1b/2a study in psoriasis ( Identifier: NCT02243787).

Background: IL-17A is a proinflammatory cytokine involved in various inflammatory diseases.Results: Potent IL-17A inhibitors were generated by fusing human Fyn SH3 domain derivatives (Fynomers) to the Fc part of an antibody.Conclusion: The linker length between the Fc part and the binding domain influences significantly the potency of Fc fusion proteins.Significance: Fynomers represent a promising class of therapeutic proteins.

The antibody-mediated targeted delivery of cytokines to sites of disease is a promising avenue for cancer therapy, but it is largely unexplored for the treatment of chronic inflammatory conditions. Using both radioactive and fluorescent techniques, the human monoclonal antibodies L19 and G11 (specific to two markers of angiogenesis that are virtually undetectable in normal adult tissues) were found to selectively localize at arthritic sites in the murine collagen-induced model of rheumatoid arthritis following intravenous (i.v.) administration. The same animal model was used to study the therapeutic action of the L19 antibody fused to the cytokines IL-2, tumour necrosis factor (TNF) and IL-10.