**1. Introduction**

Mesenchymal stem cells (MSCs) are successfully used in various fields of regenerative medicine [1]. Cell therapy is based on the ability of MSCs to migrate to the sites of pathology. They are able to exert anti-inflammatory and immunomodulatory effects upon allogeneic transplantation, as well as

in autoimmune diseases [2–5]. Obtaining genetically modified MSCs expressing introduced genes significantly expands the possibilities of both cellular and genetic therapy, ensuring the delivery of therapeutic molecules to the sites of damage and inflammation [6,7]. MSCs transduced by the interferon β (IFN-β) gene have been shown to reduce the signs of inflammation and the severity of the disease and to improve the condition of the CNS in experimental multiple sclerosis [8]. Positive results with modified MSCs have been obtained in myocardial infarction [9] and in cancer therapy [10].

These results suggest that obtaining and using modified MSCs (mMSCs) that harbor viral genes could be effective for the development of antiviral vaccines. This approach has a number of advantages over traditional vaccine technologies. mMSCs can express many proteins simultaneously, thus ensuring a wide range of epitopes with the correct post-translational modifications as during natural infection. They are also capable of delivering, expressing, and presenting an antigen for a long time. Indeed, Tomchuck et al. demonstrated in an experimental model of HIV infection that cellular vaccines based on transfected MSCs could be developed [11].

One of the urgent but unresolved problems of biology and medicine is the development of an effective vaccine towards hepatitis C virus (HCV). HCV is considered as one of the main etiological agents of chronic liver disease, including terminal stages – cirrhosis and hepatocarcinoma. In up to 80% of cases, acute hepatitis C transfers into chronic disease, which may be caused by a very high heterogeneity of viral genome and the existence of quasispecies, interference of the virus with innate and adaptive immune response pathways, and the formation of "escape" variants of HCV that are not recognized by the immune system [12,13]. Anti-HCV therapy based on a combination of pegylated recombinant IFN-α and ribavirin eliminates the virus in no more than 50% of patients [14]. Modern therapy using direct-acting antivirals (DAA) that target HCV NS3, NS5A, and NS5B proteins makes it possible to cure up to 99% of patients, but the extremely high cost of DAAs makes access to the treatment limited, as exemplified by higher rates of detection of new cases compared to number of patients treated with these drugs [15] Another factor that limits access to treatment is unawareness of a majority (ca. 80%) of patients of their status. In addition, there is a lot of evidence showing that HCV can remain in the liver cells and peripheral blood mononuclear cells (PBMC) of patients after the disappearance of viral RNA in serum, thus establishing occult infection [16,17]. Such ongoing viral replication in hepatocytes can lead to continuous liver injury and may underlie the absence of improvement in clinical outcomes after a sustained virological response achieved in a majority of patients [16].

The development of anti-HCV vaccines can contribute to global efforts to eradicate the virus. Numerous attempts to develop a vaccine whose action is based on broadly neutralizing antibodies against structural proteins failed, most likely due to very high variability of E1 and E2 glycoproteins and escape of virions due to bound lipoproteins and glycans [18]. Efficient vaccines could be based on the recombinant viral proteins/peptides that contain B- and T-cell epitopes or DNA plasmids/viral vectors, ensuring their expression [19]. However, the optimal set of HCV genes or their fragments that should be present in the vaccine has not yet been determined. Literature data suggest that none of the candidate vaccines triggered a full preventive and therapeutic response against HCV [20,21]. Recently, several vaccines against HCV based on dendritic cells (DC) have been reported [22,23]. DCs are highly specialized antigen-presenting cells (APC), so DC-based vaccination based on ex vivo stimulated and matured DCs loaded with HCV specific antigens is an attractive approach to elicit sustained anti-viral response to HCV proteins. However, generation of DCs can require substantial time and expense. It was shown that induction of T cell immune responses by DC vaccination is highly dependent on efficient antigen loading of the DCs [24]. The trials showed that such vaccines did not clear HCV infection in chronic hepatitis C patients despite induction of pronounced T-cell response [25,26]. Thus, the development of MSC vaccines based on genetic and cellular technologies could be one of the most promising strategies to prevent the spread of infectious diseases.

The aim of this work was to obtain MSCs expressing HCV genes and to analyze the humoral and cellular immune responses of animals to the introduction of these modified MSCs. As a model we chose genes of nonstructural HCV proteins that form viral replicase complex, as they are more conservative than structural proteins and in total comprise two-thirds of the entire HCV proteome [27]. Several lines of evidence show that clearance of acute HCV infection in chimpanzees and humans is temporally associated with early, strong, and broadly reactive T cell responses against multiple non-structural viral proteins [12,28]. The non-structural proteins are considered as the dominant targets for CD8+ and CD4+ cells [27]. A robust HCV-specific CD8+ and CD4+ T cell responses to the non-structural proteins has the potential to restrict infection, eliminate virus-infected cells after challenge, and prevent persistent infection at the very least [29].
