AAV8-Mediated In Vivo Overexpression of miR-155 Enhances the Protective Capacity of Genetically Attenuated Malarial Parasites

Abstract

Malaria, caused by protozoan Plasmodium parasites, remains a prevalent infectious human disease due to the lack of an efficient and safe vaccine. This is directly related to the persisting gaps in our understanding of the parasite's interactions with the infected host, especially during the clinically silent yet essential liver stage of Plasmodium development. Previously, we and others showed that genetically attenuated parasites (GAP) that arrest in the liver induce sterile immunity, but only upon multiple administrations. Here, we comprehensively studied hepatic gene and miRNA expression in GAP-injected mice, and found both a broad activation of IFNγ-associated pathways and a significant increase of murine microRNA-155 (miR-155), that was especially pronounced in non-parenchymal cells including liver-resident macrophages (Kupffer cells). Remarkably, ectopic upregulation of this miRNA in the liver of mice using robust hepatotropic adeno-associated virus 8 (AAV8) vectors enhanced GAP's protective capacity substantially. In turn, this AAV8-mediated miR-155 expression permitted a reduction of GAP injections needed to achieve complete protection against infectious parasite challenge from previously three to only one. Our study highlights a crucial role of mammalian miRNAs in Plasmodium liver infection in vivo and concurrently implies their great potential as future immune-augmenting agents in improved vaccination regimes against malaria and other diseases.

Plasmodium meets AAV—the (un)likely marriage of parasitology and virology, and how to make the match

“…In contrast, other, nonparenchymal liver cell types that are relevant and interesting in the malaria liver‐stage context, that is, Kupffer cells, stellate cells, and LSECs, are largely refractory to transduction with wild‐type AAV capsids. In our previous work, we did observe AAV8 activity in Kupffer cells in mouse livers following systemic vector administration, but the efficiency was clearly below that in hepatocytes in the same animals . We have recently made similar findings for another wild‐type serotype, AAV6, and stellate cells in mouse liver (D. Grimm & H. Willenbring, unpublished results).…”

“…Lastly, our own two groups have previously harnessed AAV8 vectors to dysregulate miR-155 in mouse livers prior to vaccination with genetically attenuated P. berghei parasites (GAP, [3,4]), following our discovery that this immunoregulatory host miRNA is highly elevated upon Plasmodium liverstage infection [50]. Our concomitant result that GAP infusion stimulates expression of TNFa (tumor necrosis factor alpha) and IFNc shed light on the underlying molecular mechanism, considering that these two cytokines are known upstream regulators of miR-155 expression.…”

“…Next to proteins, an additional class of important host factors are microRNA (miRNA), roughly 22nucleotide-long RNA that are master regulators of at least 60% of all mammalian genes and as such play central roles in pathological processes, including cancer and pathogen infections [49]. Recently, we dissected the effects of Plasmodium sporozoite infection on host miRNA expression under physiologically relevant conditions in livers of adult mice [50]. Combined microarray and PCR screens using total liver RNA obtained after infection with either wild-type or attenuated Plasmodium strains revealed not only an up-regulation of IFN-associated genes (confirming and extending previous work, see above and [2,3]) but also a dysregulation of 11 of 698 studied miRNA.…”

MicroRNAs and Malaria – A Dynamic Interaction Still Incompletely Understood

“…Previously, it had been shown that GAP arrested in the liver, induce sterile immunity, but only after several administrations. It was found that GAP-injected mice displayed a broad activation of IFNγ-associated pathways and a significant increase in miR-155 within macrophages in the liver [50]. This upregulation enhanced the protective capacity of GAP substantially, highlighting the crucial role of mammalian miRNAs in Plasmodium liver infection.…”

Roles of microRNAs in T cell immunity: Implications for strategy development against infectious diseases

“…A recent study revealed that the inhibition of miR‐21 using a recombinant AAV8 could inhibit pathogenesis in mouse liver following infections with Schistosoma japonicum . Likewise, another study highlighted the role of the mammalian miR‐155 in immune regulation and showed that an AAV8‐based delivery strategy protects mice from infections with malaria . Recently, the U.S. Food and Drug Administration (FDA) approved Kymriah, a cell‐based gene therapy for certain pediatric and young adult patients with a form of acute lymphoblastic leukemia.…”