Browsing by Author "Smirnova, Olga"

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    Cellular immune response induced by dna immunization of mice with drug resistant integrases of hiv-1 clade a offers partial protection against growth and metastatic activity of integrase-expressing adenocarcinoma cells
    (2021-06) Isaguliants, Maria; Krotova, Olga; Petkov, Stefan; Jansons, Juris; Bayurova, Ekaterina; Mezale, Dzeina; Fridrihsone, Ilze; Kilpelainen, Athina; Podschwadt, Philip; Agapkina, Yulia; Smirnova, Olga; Kostic, Linda; Saleem, Mina; Latyshev, Oleg; Eliseeva, Olesja; Malkova, Anastasia; Gorodnicheva, Tatiana; Wahren, Britta; Gordeychuk, Ilya; Starodubova, Elizaveta; Latanova, Anastasia; Research Department
    Therapeutic DNA-vaccination against drug-resistant HIV-1 may hinder emergence and spread of drug-resistant HIV-1, allowing for longer successful antiretroviral treatment (ART) up-to relief of ART. We designed DNA-vaccines against drug-resistant HIV-1 based on consensus clade A integrase (IN) resistant to raltegravir: IN_in_r1 (L74M/E92Q/V151I/N155H/G163R) or IN_in_r2 (E138K/G140S/Q148K) carrying D64V abrogating IN activity. INs, overexpressed in mammalian cells from synthetic genes, were assessed for stability, route of proteolytic degradation, and ability to induce oxidative stress. Both were found safe in immunotoxicity tests in mice, with no inherent carcinogenicity: their expression did not enhance tumorigenic or metastatic potential of adenocarcinoma 4T1 cells. DNA-immunization of mice with INs induced potent multicytokine T-cell response mainly against aa 209–239, and moderate IgG response cross-recognizing diverse IN variants. DNA-immunization with IN_in_r1 protected 60% of mice from challenge with 4Tlluc2 cells expressing non-mutated IN, while DNA-immunization with IN_in_r2 protected only 20% of mice, although tumor cells expressed IN matching the immunogen. Tumor size inversely correlated with IN-specific IFN-γ/IL-2 T-cell response. IN-expressing tumors displayed compromised metastatic activity restricted to lungs with reduced metastases size. Protective potential of IN immunogens relied on their immunogenicity for CD8+ T-cells, dependent on proteasomal processing and low level of oxidative stress.
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    Fusion to Flaviviral Leader Peptide Targets HIV-1 Reverse Transcriptase for Secretion and Reduces Its Enzymatic Activity and Ability to Induce Oxidative Stress but Has No Major Effects on Its Immunogenic Performance in DNA-Immunized Mice
    (2017-06-22) Latanova, Anastasia; Petkov, Stefan; Kuzmenko, Yulia; Kilpelaïnen, Athina; Ivanov, Alexander; Smirnova, Olga; Krotova, Olga; Korolev, Sergey; Hinkula, Jorma; Karpov, Vadim; Isaguliants, Maria; Starodubova, Elizaveta; Rīga Stradiņš University
    Reverse transcriptase (RT) is a key enzyme in viral replication and susceptibility to ART and a crucial target of immunotherapy against drug-resistant HIV-1. RT induces oxidative stress which undermines the attempts to make it immunogenic. We hypothesized that artificial secretion may reduce the stress and make RT more immunogenic. Inactivated multidrug-resistant RT (RT1.14opt-in) was N-terminally fused to the signal providing secretion of NS1 protein of TBEV (Ld) generating optimized inactivated Ld-carrying enzyme RT1.14oil. Promotion of secretion prohibited proteasomal degradation increasing the half-life and content of RT1.14oil in cells and cell culture medium, drastically reduced the residual polymerase activity, and downmodulated oxidative stress. BALB/c mice were DNA-immunized with RT1.14opt-in or parental RT1.14oil by intradermal injections with electroporation. Fluorospot and ELISA tests revealed that RT1.14opt-in and RT1.14oil induced IFN-γ/IL-2, RT1.14opt-in induced granzyme B, and RT1.14oil induced perforin production. Perforin secretion correlated with coproduction of IFN-γ and IL-2 (R=0,97). Both DNA immunogens induced strong anti-RT antibody response. Ldpeptide was not immunogenic. Thus, Ld-driven secretion inferred little change to RT performance in DNA immunization. Positive outcome was the abrogation of polymerase activity increasing safety of RT-based DNA vaccines. Identification of the molecular determinants of low cellular immunogenicity of RT requires further studies.
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    Genetic instability and anti-HPV immune response as drivers of infertility associated with HPV infection
    (2021-12) Isaguliants, Maria; Krasnyak, Stepan; Smirnova, Olga; Colonna, Vincenza; Apolikhin, Oleg; Buonaguro, Franco M.; Rīga Stradiņš University
    Human papillomavirus (HPV) is a sexually transmitted infection common among men and women of reproductive age worldwide. HPV viruses are associated with epithelial lesions and cancers. HPV infections have been shown to be significantly associated with many adverse effects in reproductive function. Infection with HPVs, specifically of high-oncogenic risk types (HR HPVs), affects different stages of human reproduction, resulting in a series of adverse outcomes: 1) reduction of male fertility (male infertility), characterized by qualitative and quantitative semen alterations; 2) impairment of couple fertility with increase of blastocyst apoptosis and reduction of endometrial implantation of trophoblastic cells; 3) defects of embryos and fetal development, with increase of spontaneous abortion and spontaneous preterm birth. The actual molecular mechanism(s) by which HPV infection is involved remain unclear. HPV-associated infertility as Janus, has two faces: one reflecting anti-HPV immunity, and the other, direct pathogenic effects of HPVs, specifically, of HR HPVs on the infected/HPV-replicating cells. Adverse effects observed for HR HPVs differ depending on the genotype of infecting virus, reflecting differential response of the host immune system as well as functional differences between HPVs and their individual proteins/antigens, including their ability to induce genetic instability/DNA damage. Review summarizes HPV involvement in all reproductive stages, evaluate the adverse role(s) played by HPVs, and identifies mechanisms of viral pathogenicity, common as well as specific for each stage of the reproduction process.
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    HIV-1 Reverse Transcriptase Promotes Tumor Growth and Metastasis Formation via ROS-Dependent Upregulation of Twist
    (2019-12-02) Bayurova, Ekaterina; Jansons, Juris; Skrastina, Dace; Smirnova, Olga; Mezale, Dzeina; Kostyusheva, Anastasia; Kostyushev, Dmitry; Petkov, Stefan; Podschwadt, Philip; Valuev-Elliston, Vladimir; Sasinovich, Sviataslau; Korolev, Sergey; Warholm, Per; Latanova, Anastasia; Starodubova, Elizaveta; Tukhvatulin, Amir; Latyshev, Oleg; Selimov, Renat; Metalnikov, Pavel; Komarov, Alexander; Ivanova, Olga; Gorodnicheva, Tatiana; Kochetkov, Sergey; Gottikh, Marina; Strumfa, Ilze; Ivanov, Alexander; Gordeychuk, Ilya; Isaguliants, Maria; Department of Pathology; Rīga Stradiņš University
    HIV-induced immune suppression results in the high prevalence of HIV/AIDS-associated malignancies including Kaposi sarcoma, non-Hodgkin lymphoma, and cervical cancer. HIV-infected people are also at an increased risk of "non-AIDS-defining" malignancies not directly linked to immune suppression but associated with viral infections. Their incidence is increasing despite successful antiretroviral therapy. The mechanism behind this phenomenon remains unclear. Here, we obtained daughter clones of murine mammary gland adenocarcinoma 4T1luc2 cells expressing consensus reverse transcriptase of HIV-1 subtype A FSU_A strain (RT_A) with and without primary mutations of drug resistance. In in vitro tests, mutations of resistance to nucleoside inhibitors K65R/M184V reduced the polymerase, and to nonnucleoside inhibitors K103N/G190S, the RNase H activities of RT_A. Expression of these RT_A variants in 4T1luc2 cells led to increased production of the reactive oxygen species (ROS), lipid peroxidation, enhanced cell motility in the wound healing assay, and upregulation of expression of Vimentin and Twist. These properties, particularly, the expression of Twist, correlated with the levels of expression RT_A and/or the production of ROS. When implanted into syngeneic BALB/C mice, 4T1luc2 cells expressing nonmutated RT_A demonstrated enhanced rate of tumor growth and increased metastatic activity, dependent on the level of expression of RT_A and Twist. No enhancement was observed for the clones expressing mutated RT_A variants. Plausible mechanisms are discussed involving differential interactions of mutated and nonmutated RTs with its cellular partners involved in the regulation of ROS. This study establishes links between the expression of HIV-1 RT, production of ROS, induction of EMT, and enhanced propagation of RT-expressing tumor cells. Such scenario can be proposed as one of the mechanisms of HIV-induced/enhanced carcinogenesis not associated with immune suppression.