Characterization, stability, and in vivo efficacy studies of recombinant human CNTF and its permeation into the neural retina in ex vivo organotypic retinal explant culture models

Jaakko Itkonen (Corresponding Author), Ada Annala, Shirin Tavakoli, Blanca Arango-Gonzalez, Marius Ueffing, Elisa Toropainen, Marika Ruponen, Marco G. Casteleijn, Arto Urtti (Corresponding Author)

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    Ciliary neurotrophic factor (CNTF) is one of the most studied neuroprotective agents with acknowledged potential in treating diseases of the posterior eye segment. Although its efficacy and mechanisms of action in the retina have been studied extensively, it is still not comprehensively understood which retinal cells mediate the therapeutic effects of CNTF. As with therapeutic proteins in general, it is poorly elucidated whether exogenous CNTF administered into the vitreous can enter and distribute into the retina and hence reach potentially responsive target cells. Here, we have characterized our purified recombinant human CNTF (rhCNTF), studied the protein’s in vitro bioactivity in a cell-based assay, and evaluated the thermodynamic and oligomeric status of the protein during storage. Biological activity of rhCNTF was further evaluated in vivo in an animal model of retinal degeneration. The retinal penetration and distribution of rhCNTF after 24 h was studied utilizing two ex vivo retina models. Based on our characterization findings, our rhCNTF is correctly folded and biologically active. Moreover, based on initial screening and subsequent follow-up, we identified two buffers in which rhCNTF retains its stability during storage. Whereas rhCNTF did not show photoreceptor preservative effect or improve the function of photoreceptors in vivo, this could possibly be due to the used disease model or the short duration of action with a single intravitreal injection of rhCNTF. On the other hand, the lack of in vivo efficacy was shown to not be due to distribution limitations; permeation into the retina was observed in both retinal explant models as in 24 h rhCNTF penetrated the inner limiting membrane, and being mostly observed in the ganglion cell layer, distributed to different layers of the neural retina. As rhCNTF can reach deeper retinal layers, in general, having direct effects on resident CNTF-responsive target cells is plausible.
    Original languageEnglish
    Article number611
    Pages (from-to)1-31
    Number of pages31
    Issue number7
    Publication statusPublished - 30 Jun 2020
    MoE publication typeA1 Journal article-refereed


    J.I. was funded by The Finnish Cultural Foundation, The Paulo Foundation, Evald and Hilda Nissi Foundation, and Päivikki and Sakari Sohlberg Foundation. S.T. was funded by the European Union’s Horizon 2020 research and innovation programme Marie Skłodowska-Curie ITN (NANOMED, grant 676137). B.A.-G. was funded by FFB Grant PPA-0717-0719-RAD, Kerstan Foundation and ProRetina Foundation. M.U. was funded by Land Baden-Württemberg and Kerstan Foundation. A.A., E.T. and M.R. were funded by The University of Eastern Finland. M.C. was funded by the Academy of Finland Key-project (303884). A.U. was funded by Academy of Finland (project 311122).


    • CNTF
    • Intravitreal delivery
    • Neuroprotection
    • Protein aggregation
    • Retinal penetration
    • Stability


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