Modeling infection and antiviral therapy of enteric viruses using primary intestinal organoids

In this thesis, we first highlighted important clinical complications associated with rotavirus infection in setting of orthotopic organ transplantation, I then analyzed the incidence of rotavirus infection, its diagnosis, its pathogenesis, how to rationally use of immunosuppressive agents in patients at risk for rotavirus infection, especially by studying the interactions of rotavirus with specific immunosuppressants, and also how to manage rotavirus infection in organ recipients. I also found 3% incidence of rotavirus infections among 6176 transplantations. I conclude that rotavirus infections occurring in transplantation patients remain clinically largely not diagnosed, and thus more attention should be paid to this pathogen. The situation might be improved through the development of new superior model systems allowing new directions of research. I demonstrated that primary intestinal organoids can support infection with both laboratory rotavirus strains and with patient-derived rotavirus isolates. Thus, the organoid model might become exceedingly useful for obtaining new scientific insights but may also become important for individualized assessment of the efficacy of different antivirals in a particular patient, next to their potential for developing new and effective medicines against rotavirus. In support for this notion I found that the broadly used antivirals including interferon a and ribavirin inhibit rotavirus replication utilizing the organoid model. I thus propose that organoids provide a promising novel avenue for investigating rotavirus-host interactions and the evaluation of medicines against rotavirus. I profiled common used immunosuppressive medicines on enteric rotavirus and norovirus infections. I found that CsA moderately inhibits rotavirus and norovirus infections, and that MPA is very in this respect for both viruses and also with high barrier towards the development of d.rug resistance. Mechanistically, the antiviral effects of MPA are mediated through inhibition of its canonical cellular target IMPDH and depend on the resulting guanosine nucleotide depletion. I also found that 6-TG potently inhibits rotavirus via blocking the formation of the active form of Rael {GTP-Racl), again with high barrier towards the development of drug resistance. I hope my findings will become an important reference for clinicians to design optimal immunosuppressive therapy for rotavirus infected transplantation patients and aid the development of novel antiviral medicines against rotavirus. I also demonstrated that Pl3K-Akt-mTOR signaling pathway sustains rotavirus infection and that the clinically used mTOR inhibitor rapamycin significantly inhibits rotavirus infection. Rapamycin induces autophagy via 4E-BP1, and induction of autophagy exerted antiviral effect on rotavirus, suggesting that this is the mechanistic explanation of this finding. Although I perceive that the findings presented in this thesis contributed to a better understanding of the complex nature of enteric viruses including rotavirus and norovirus, I also have to acknowledge that more work is needed to reduce the burden of rotavirus and norovirus infection and hence there is plenty of opportunities for my successors either at the Erasmus MC or elsewhere to further contribute to such efforts.