WO2010056148A1 - Vaccin par voie muqueuse pour l’immunothérapie de maladies liés à des virus du papillome humain - Google Patents

Vaccin par voie muqueuse pour l’immunothérapie de maladies liés à des virus du papillome humain Download PDF

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WO2010056148A1
WO2010056148A1 PCT/RU2009/000422 RU2009000422W WO2010056148A1 WO 2010056148 A1 WO2010056148 A1 WO 2010056148A1 RU 2009000422 W RU2009000422 W RU 2009000422W WO 2010056148 A1 WO2010056148 A1 WO 2010056148A1
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hsp70
treatment
protein
hpv
human papillomavirus
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PCT/RU2009/000422
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English (en)
Russian (ru)
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Всеволод Иванович КИСЕЛЕВ
Петр Георгиевич СВЕШНИКОВ
Михаил Александрович ПАЛЬЦЕВ
Original Assignee
Kiselev Vsevolod Ivanovich
Sveshnikov Petr Georgievich
Paltsev Mikhail Alexandrovich
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Application filed by Kiselev Vsevolod Ivanovich, Sveshnikov Petr Georgievich, Paltsev Mikhail Alexandrovich filed Critical Kiselev Vsevolod Ivanovich
Priority to UAA201103142A priority Critical patent/UA98254C2/ru
Publication of WO2010056148A1 publication Critical patent/WO2010056148A1/fr

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K39/12—Viral antigens
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/12—Antivirals
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/60—Medicinal preparations containing antigens or antibodies characteristics by the carrier linked to the antigen
    • A61K2039/6031—Proteins
    • A61K2039/6043—Heat shock proteins
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2710/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA dsDNA viruses
    • C12N2710/00011—Details
    • C12N2710/20011—Papillomaviridae
    • C12N2710/20034—Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein

Definitions

  • the invention relates to the field of genetic engineering, specifically it relates to a composition of recombinant proteins based on the oncoprotein E7 of human papillomavirus fused to the heat shock protein of mycobacteria Hsp70 in combination with chitosan and pharmacologically acceptable additives for the manufacture of suppositories.
  • the invention also relates to vaccine therapy for diseases caused by human papilloma viruses.
  • HPV human papillomavirus
  • HPV DNA is detected on average in 98.9% and 94.7% of genital warts and cervical cancer samples, respectively.
  • HPV is found in 35% of cases, with 6, 11, lb and 18 types - only in 21.6%.
  • the majority of HPV 16 DNA in genital warts cells is in episomal form, and in cervical cancer samples it is integrated into the cellular genome.
  • several large-scale epidemiological studies have been carried out on the geographical distribution of various types of HPV in cervical samples from patients with invasive cervical cancer (Cervical cancer).
  • HPV types 16/18 are detected, however, further ranking of HPV types has pronounced geographical features. For example, in China, the next most important HPV types are 58 and 52, in Africa - HPV type 45, in Central / South America - HPV type 31. (Wao Y, Li N., Smith J.
  • HPV carriage is not life-long is important: according to the WHO (2001), in the absence of aggravating factors for 3 years, low-severity squamous cell epithelial lesions containing HPV are regressed in 50 - 62% of cases . According to scientists at the University of California, in 70% of young HPV-infected women, HPV DNA ceases to be detected for 24 months. Speed elimination is significantly reduced when infected with several types of HPV, as well as in the presence of a history of genital warts. At the same time, among women with at least 3 times positive results of HPV determination, the risk of severe oncological lesions of the epithelium increased by 14 times.
  • the expression of the E6 and E7 genes is regulated by the product of the E2 gene, which is a repressor of the transcription of these genes. That is why, while the virus is in an episomal state, benign processes of growth of infected tissues are observed.
  • a key event in the malignancy of cells is the integration of the virus into the cell genome, which is accompanied by a deletion of the E2 gene. This event has two important consequences:
  • any antiviral drugs are powerless to stop the process of tumor transformation, since infected cells do not contain the virus in the traditional sense, and all therapeutic measures should be aimed at eliminating cells with an integrated form of the HPV genome.
  • Cell cycle control and cell differentiation is carried out by E6 and E7 proteins through their interaction and inactivation of such “key” proteins-regulators of proliferative activity and apoptosis of cells as p53 and retinoblastoma protein (pRB). Uncontrolled proliferation of infected cells leads to the accumulation of genetic damage and, ultimately account, to malignancy.
  • Oncoprotein E7 uses several pathways to regulate the cell cycle.
  • E7 is able to form a stable complex with the pRB protein, causing its degradation, which leads to the release of the transcription factor E2F, which stimulates the transcription of genes necessary for DNA replication and S-phase of the cell cycle.
  • E7 also affects the activity of a number of cell cycle proteins, such as A and E cyclins, cdk2 kinase and inhibitors of cyclin-dependent kinase p21 and p27 (Kiselev B.I., Kiselev O.I., Etiological role of human papillomavirus in the development of cervical cancer uterus: genetic and pathogenetic mechanisms, Cytokines and inflammations, 2003, 2, 4, 31-38; ⁇ Helrs WC, Varpes J.A. apd Loba DC, Humaparillotavigas: molotargetgeps aprpresult forpolitelaprilapriltapl, 1999, 7, 4-8).
  • VLP virus-like particles
  • a model of virus-like particles (vigas-likelihoods-VLP), self-assembled from the main HPV-Ll capsid protein, is widely used to study the protective role of virus-neutralizing antibodies.
  • Immunization of animals with VLP protects them against experimental infection homologous viruses.
  • the passive transfer of serum from mice immunized with VLP 5 to other experimental animals also has a protective effect, which confirms the protective role of virus-neutralizing antibodies (Type RW, Impeve evaciop, Hpap parillomavis-asserceper, Neper, 2002, 2; Zhou J., Liu WJ., Repg SW et al.
  • HPV - associated disease is substantially higher among patients with transplant clinics (Rark JS et al Ipastivatiop ° F ipterferop regulatoru factor-1 tumor surrressor rroteip bu HPV E7 opsorroteip Imrlisatiop for the E7- medated immupe evasiop meshapism Ip servisal sarsipogepesis J. Viol... Chet 2000, 275, 6764-6769) and HIV-infected, i.e. in cases with deep impaired cellular immunity (Frazer L, et al.
  • T-helpers T-helpers
  • CDs + cytotoxic T-cells
  • the skin lesions associated with HPV in patients receiving immunosuppressive therapy often disappear when the treatment course is canceled Le. SJ. Et at. E6 apd E7 optorotepiofs of humaprilomavigas 16 iphibit IL-18-ipfpu humid rerherall blodopopuslear-apd NK sells. Jtipol, 2001, 167, 497-504).
  • the target for immunotherapy of tumors is normal or mutant cell proteins, while E6 and E7 are completely foreign to the host’s immune system, and therefore should be very immunogenic (Przeriorka D. apd. Srivastava R., Non Shock Protei - Rertide comlexes as immüütäraru for humap sapser. Molesilartisipipe Todau, 1998, 11, 478-484).
  • HPV protein E7 type 16 is capable of inducing cytotoxic immunity, and injection of this protein in combination with various adjuvants enhances the cytotoxic reaction.
  • E7 is synthesized in small quantities during the reproductive phase of the HPV life cycle.
  • its synthesis increases significantly after the integration of viral DNA into the cellular genome, since this process, as a rule, is accompanied by the loss of the E2 gene encoding the repressor of the E7 gene.
  • the induction of specific cellular immunity against the E7 protein is not observed in the body.
  • Some experimental models shed light on the amazing ability of Oncoprotein E7 to induce tolerance.
  • a skin transplant from a transgenic mouse synthesizing E7 in epithelial cells is transplanted with a genetically related mouse, then the expected transplant rejection does not occur. If the mice are introduced stimulating formation of anti-inflammatory cytokines (killed bacteria or endotoxins), then the graft quickly rejects. However, transplant rejection against the background of the introduction of cytokine inducers occurs only during the graft engraftment, and a second transplant of the same mouse transplant results in rejection without additional induction of cytokines.
  • E6 or E7 The constant synthesis of oncoproteins is necessary to maintain the tumor phenotype of cells. Obviously, the appearance in the body of cells carrying foreign antigens (E6 or E7) should induce appropriate immune responses. However, patients with cervical carcinomas have a very low T-cell immunity against E6 and E7.
  • Kiselev O.I. Kiselev V.I.
  • Sveshnikov P.G. Recombinant protein composition, method for preparing such a composition, pharmaceutical kit for immunotherapy and prophylactic vaccination of tumor diseases of the anogenital sphere, method and mmunotherapy and preventive vaccination based on it).
  • the company Nvepta is developing an injectable form of a vaccine preparation containing the E7 protein, "fused" to the heat shock protein M. tuberculosis with a molecular weight of 65 IdDa.
  • N ° 2229307 describes the development of a vaccine based on the hybrid proteins E7 and Hp 70 M.tibersilosis.
  • the authors describe an injectable dosage form with which it is possible to achieve regression of tumors induced by the virus in experimental models with transplantable tumors.
  • the demonstrated vaccination efficacy in these models confirms the correct selection of the E7 protein as the molecular target against which immune responses are induced.
  • cervical dysplasias developing in women due to chronic papillomavirus infection of the cervix uteri the induction of systemic immunity cannot guarantee the regression of malignancy processes, since the cervix has a certain isolation from systemic immunity in the regulation of local immunological reactions.
  • mucosal vaccines are preferential induction of local mucosal immunity, which is required for pathologies that affect the mucous membranes, as in the case of papillomavirus infection.
  • suppositories or “sprays” can be considered, which allow the vaccine preparation to be applied to the mucosa in the form of lyophilized powder
  • the route of administration should include stimulation of local adaptive and innate immunity, namely: in the case of PIIIM, the route of administration is vaginal; in case of colorectal cancer, the rectal route of administration.
  • the recombinant protein which is the active substance of the vaccine preparation, should contain amino acid sequences of proteins that have an immunomodulatory effect on the formation of cytokines and improve the presentation of the E7 antigen.
  • the dosage form of the vaccine should contain ingredients that increase adhesion on the mucous membranes and the affinity of the protein components of the vaccine for antigen-presenting cells.
  • E7 type 16 or type 18 fusion proteins As the main antigens, fused to M.tibercylsis Hsp70.
  • a hybrid protein is a single polypeptide chain consisting of the first protein - HPV type E7, type 16 or 18.
  • E7 oncoprotein is the main inducer of malignancy of epithelial cells infected with human papilloma viruses (Yip E.K. , Theole O HPV E6 and E7 ESPs are supported by the HPV-Associated Test Server, Sapser Res. Therap., 2005, 37, 6, 319-324)
  • the E7 protein has pronounced immune properties.
  • the E7 protein that transformed cells manage to "escape" from the body's immune responses (Kapoda S., Faheu L.M. apd Kast WM, Meshapism used bu rillomaviruses so espare teh host immresie reserpse, Serepteregres, 2007, 7, 2007, 89).
  • the E7 protein is an optimal target for immunotherapy.
  • Another part of this protein is represented by the heat shock protein from M.tibercylosis, which has a high affinity for antigen-presenting cells and is a powerful inducer of cytokinrv synthesis (Prozero D.
  • type 6 E7 protein as an antigen, which is more common than other types of HPV when the rectal mucosa is damaged.
  • chitosan in the ratio of protein: chitosan 1-1 to 1-10 is included in the composition of vaccines as an adhesive agent that promotes the penetration of antigens through the mucous membranes and a strong stimulator of nonspecific immune reactions.
  • the optimal ratio of protein to chitosan in the dosage form was studied experimentally . It was found that the maximum effect in the induction of immunological reactions against antigenic determinants of proteins was observed when using a ratio in the range of 1-1 to 1-10. With a decrease or increase in the concentration of chitosan in relation to a fixed dosage of protein, a decrease in the efficiency of the induction of immunological reactions was observed.
  • Vitepsol cocoa butter or type A solid fat as a lipophilic base
  • polyethylene glycols of various molecular weights as a hydrophilic base
  • nipagin as a hydrophilic base
  • nipagin as a hydrophilic base
  • nipagin as a hydrophilic base
  • nipagin as a hydrophilic base
  • nipagin as a hydrophilic base
  • nipagin nipagin
  • nipazole or aminoglycosides as antimicrobial agents
  • succinic acid and sodium benzoate - as stabilizers or preservatives
  • thiomersal and thiomersal as stabilizers and antimicrobial agents
  • patent RU 2224542 C2 A61 K39 / 116, 9/02, A 61P 13/00
  • Patent RU 2150268 A61K9 / 02, A61K35
  • the present invention also relates to methods of treating diseases associated with human papillomavirus, comprising topical application of a mucosal vaccine.
  • a hybrid protein E7 type 16-Hsp70 or E7 type 18-Hsp70 in combination with chitosan in a ratio of 1: 0.1-10 and pharmacologically acceptable additives, is used to treat cervical dysplasia caused by human papillomavirus.
  • the prepared suppository is administered intravaginally.
  • suppositories are prescribed 2-4 times with an interval of a week with mild cervical dysplasia and 4-8 times with an interval of one week with dysplasia of the 2nd and 3rd degree.
  • an E7 type 16-Hsp70 or E7type 6-Hsp70 hybrid protein is used, in combination with chitosan in a ratio of 1: 0.1-10 and pharmacologically acceptable additives.
  • the prepared suppository is administered rectally. Suppositories are prescribed for the night with a frequency of once a week. The maximum clinical effect is achieved with the appointment of 4-8 suppositories.
  • E7 Hsp70 fusion proteins in the vaginal suppository 1.
  • the proposed invention can significantly increase the effectiveness of the treatment of diseases associated with human papillomavirus.
  • the obtained effect is achieved due to the preferential induction of local immunological reactions due to mucosal application of recombinant proteins in complex with chitosan.
  • Using the proposed mucosal vaccine can significantly reduce the cost of treatment compared with the known methods of treating cervical cancer and PKK.
  • the proposed vaccine is intended for local use, which eliminates the unwanted and extremely life-threatening side effects of injectable drugs, for example, anaphylactic shock.
  • the proposed treatment method significantly simplifies the treatment process - the patient can undergo treatment outside the clinic, self-administering the drug.
  • FIG. 1 The dynamics of the serum IgG immune response in different methods of immunization of RSL mice E76-Hsp70,
  • Figure 3 The level of secretion of IFN-gamma (l) and IL-4 (2) by lymphocytes from inguinal lymph nodes (A) and spleen (B), stimulated in culture by a mixture of recombinant proteins E716 and E718.
  • Figure 4 The level of secretion of IFN-gamma (l) and IL-4 (2) lymphocytes from inguinal lymph nodes, stimulated in culture by a mixture of recombinant proteins E716 and E76.
  • Recombinant fusion proteins E711-Hsp70, E716-Hsp70, E718-Hsp70, E76-Hsp70 (RSL) were purified from inclusion bodies by solubilization in 8M urea, followed by metal chelate and ion exchange chromatography to a homogeneous state. The degree of protein purity was monitored by electrophoresis on a 10% polyacrylamide gel under denaturing conditions, the protein concentration was determined by the Lowry method using bovine serum albumin as a standard.
  • the recombinant fusion protein (RSL) was dialyzed to remove preservatives and stabilizers against phosphate-buffered saline (PBS, pH7.2), diluted or concentrated to the required concentration, and sterilized filtration was performed. After these procedures, solutions of recombinant proteins were mixed with a solution of chitosan in various proportions and used to make suppositories.
  • PBS phosphate-buffered saline
  • mice Used adult female mice (6-8 weeks old) of the Balb / s line (Nursery RAMS “Stolbovaya”) in the amount of 5 pieces per group. Immunization was performed three times at intervals of 4 weeks (0-28-56 days). During intranasal immunization, mice were injected with 20 ⁇ l of the RSL drug with chitosan (10 ⁇ l into each nostril) using a micropipette under halotan anesthesia. For vaginal and rectal immunization using a micropipette, 10 ⁇ g of protein in complex with chitosan was introduced in a volume of 20 ⁇ l.
  • Subcutaneous immunization was carried out at the withers of mice by injection of 200 ⁇ l of a suspension of RSL with AlhudroGeC.
  • the RSL preparations were adsorbed on 0.5% (w / w) aluminum hydroxide gel (Alhudroel., Bepostag), overnight.
  • the dose of RSL for each immunization was 10 ⁇ g of protein per mouse.
  • Control groups of mice were immunized in a similar way using 0.5% (w / v) chitosan solution and 0.5% (w / w) Alhudrogel solution for the intranasal and subcutaneous route of administration, respectively.
  • Nasopharyngeal swabs for the determination of specific IgA were obtained 14 days after the third immunization.
  • the mice were sacrificed by cervical dislocation, decapitated, and the nasopharynx was washed from the side of the trachea with 0.5 ml PBS.
  • Nasopharyngeal swabs were stored at -20 0 C until ELISA.
  • mice were sacrificed by cervical dislocation 14 days after the third immunization and the lungs, spleen and mediastinal lymph nodes were aseptically removed.
  • Lymphocyte cultures were obtained by careful mechanical grinding of tissues, lysis of red blood cells, and three-time washing of cells by low-speed centrifugation in RPMI 1640 medium with the addition of 10% fetal serum. 20mM L-Glutamine and antibiotics.
  • Example N ° 3
  • Figure 1 shows the dynamics of the serum IgG immune response in different ways of immunizing RSL mice E76-Hsp70: Group 1 - E76-Hsp70 + chitosan, intranasally; Group 2 (control) - chitosan, intranasally; Group 3 (control) - aluminum hydroxide, subcutaneously; Group 4 - E76-Hsp70 + aluminum hydroxide, subcutaneous.
  • the ordinate shows the logarithm of the geometric mean titer of IgG antibodies.
  • FIG. 1 shows the magnitude of the mucosal normalized IgA antibody response during intranasal and subcutaneous immunization of RS7 E711-Hsp70 mice.
  • Example Ns4 Comparison of a specific T-cell response during intravaginal and subcutaneous immunization with a mixture of RSH E716-Hsp70 and E718-Hsp70
  • mice Used adult female mice (age 8-10 weeks) line Balb / s in the amount of 5 pieces per group. Immunization was carried out 3 times with an interval of 10 days according to the scheme: 0-10-20 days.
  • intravaginal immunization i.vag.
  • the mice were injected with 20 ⁇ g of a mixture of RSL 10 ⁇ g of E716-Hsp70 and E718-Hsp70, respectively, in 16 ⁇ l of a 0.5% (w / v) chitosan solution (Chitosap, Celechemia AG, Switzerland) using a micropipette into the vagina.
  • Subcutaneous immunization was carried out at the withers of mice by injection of 200 ⁇ l of a suspension of a mixture of RSL (20 ⁇ g) with 0.5% (weight / weight) aluminum hydroxide gel (Alhudrogel, Vreptag Biocestor, Denmark). Control groups of mice were immunized similarly, using a 0.5% (w / v) chitosan solution and a 0.5% (w / w) Alhdrogel solution for the intravaginal and subcutaneous route of administration, respectively. 14 days after the third immunization, the mice were sacrificed and the spleen and inguinal lymph nodes were aseptically removed.
  • Lymphocyte cultures were obtained by careful mechanical grinding of tissues, erythrocyte lysis and three-time washing of cells by low-speed centrifugation in RPMI 1640 medium with the addition of 10% fetal serum, 20 mM L-glutamine and antibiotics. Lymphocyte cultures obtained from the spleen and inguinal lymph nodes of mice immunized with a mixture of RSH E716-Hsp70 and E718-Hsp70 and control groups were examined for secretion of interferon gamma (IFN-gamma) and interleukin 4 (IL-4) after stimulation with recombinant proteins E716- Hsp70 and E718-Hsp70 using ELISPOT kits (BD BIOSECIEPS, Oxford, UK).
  • IFN-gamma interferon gamma
  • IL-4 interleukin 4
  • Figure 3 shows the level of secretion of IFN-gamma (l) and IL-4 (2) by lymphocytes from inguinal lymph nodes (A) and spleen (B), stimulated in culture with a mixture of recombinant proteins E716 and E718. Mice were immunized:
  • mice Used adult female mice (age 8-10 weeks) line Balb / s in the amount of 5 pieces per group. Immunization was carried out 3 times with an interval of 10 days according to the scheme: 0-10-20 days.
  • rectal immunization the mice were injected with 20 ⁇ g of a mixture of RSL 10 ⁇ g of E716-Hsp70 and E76-Hsp70, respectively, in 16 ⁇ l of a 0.5% (weight / volume) solution of chitosan (Chitosap, Celechem AG, Switzerland) using a micropipette in the rectum.
  • Subcutaneous immunization was carried out at the withers of mice by injection of 200 ⁇ l of a suspension of a mixture of RSL (20 ⁇ g) with 0.5% (w / w) aluminum hydroxide gel (Alhudrogel, Vreptag Biocestor, Denmark).
  • the control groups of mice were immunized in a similar way using 0.5% (w / v) chitosan solution and 0.5% (w / w) Alhudrogel solution for rectal and subcutaneous administration, respectively.
  • mice 14 days after the third immunization, mice were sacrificed and inguinal lymph nodes were aseptically removed. Lymphocyte cultures were obtained by careful mechanical grinding of tissues, erythrocyte lysis and three-time washing of cells by low-speed centrifugation in RPMI 1640 medium with the addition of 10% fetal serum, 20 mM L-glutamine and antibiotics.
  • Lymphocyte cultures obtained from inguinal lymph nodes of mice immunized with a mixture of RSH E716-Hsp70 and E76-Hsp70 and control groups were examined for the secretion of interferon gamma (IFN-gamma) and interleukin 4 (IL-4) after stimulation with recombinant proteins E716-Hsp70 and E76-Hsp70 using ELISPOT kits (BD BIOSECIPSES, Oxford, UK). The data are shown in figure 4.
  • IFN-gamma interferon gamma
  • IL-4 interleukin 4
  • Figure 4 reflects the level of secretion of IFN-gamma (l) and IL-4 (2) by lymphocytes from inguinal lymph nodes stimulated in culture with a mixture of recombinant proteins E716 and E76.
  • Mice were immunized: Gray columns - E716-HSP70 + E76-HSP70 + chitosan, rectally; Black columns - chitosan, rectally; White columns - aluminum hydroxide, subcutaneously; Shaded columns - E716-HSP70 + E76-HSP-70 + aluminum hydroxide, subcutaneous.
  • the ordinate shows the number of cytokine-secreting lymphocytes per 1 million cells.
  • MB example shows the number of cytokine-secreting lymphocytes per 1 million cells.
  • Hsp70 induces the production of pro-inflammatory cytokines in the DC: IL-I beta, IL-6, IL-12, and TNF-alpha, which indicates a pronounced adjuvant effect of Hsp70.
  • the study of changes in the phenotype of DC during Hsp70 stimulation showed that the proportion of cells expressing markers of mature DC (CD38, CD40, CD83, MHC2 molecules) increases. Of the whole spectrum of markers of mature DCs does not increase the expression of co-stimulatory molecules CD86 and CDlIc. The change in the level of TLR2 and TLR4, which are more typical for immature DCs, remains unclear.
  • Vitepsol HW 6 TRIS of the company MP Biomedisals Ips. cat. JChb 195605. Composition per 1 suppository:
  • Suppositories were prepared by pouring. Calculation of the components was carried out per 100 suppositories of 3.0 g each.
  • a solution of recombinant proteins E7 of types 16 and 18 (c 1 mg / ml) in 0.0 IM TRIS 50 ml

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Abstract

L’invention se rapporte au domaine du génie génétique et peut être utilisée en thérapie contre des maladies associées au virus du papillome humain. Le vaccin par voie muqueuse comprend une quantité efficace de protéine hybride se composant d’oncoprotéine E7 du virus du papillome humain combinée à une protéine de choc thermique de mycobactéries Hsp70, du chitosane dans une proportion de 1:0,1-10 par rapport à la protéine hybride, ainsi que des additifs pharmaceutiquement acceptables pour la préparation de suppositoires. La présente invention permet d’augmenter fortement l’efficacité de traitement de maladies associées au virus du papillome humain, d’abaisser considérablement le coût du traitement par rapport aux méthodologies connues de traitement du cancer du col de l’utérus et cancer du colon, d’éliminer les effets secondaires indésirables pouvant mettre en danger la vie du patient et résultant de préparations à injecter, comme un choc anaphylactique du fait d’une application locale, et de simplifier le processus thérapeutique, le patient pouvant suivre le traitement hors clinique en s’administrant lui-même la préparation.
PCT/RU2009/000422 2008-11-13 2009-08-20 Vaccin par voie muqueuse pour l’immunothérapie de maladies liés à des virus du papillome humain WO2010056148A1 (fr)

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RU2489481C1 (ru) * 2012-01-16 2013-08-10 Федеральное государственное унитарное предприятие "Государственный научно-исследовательский институт генетики и селекции промышленных микроорганизмов" (ФГУП "ГосНИИгенетика") СПОСОБ ПОЛУЧЕНИЯ БЕЛКА E7-HSP70 И ШТАММ ДРОЖЖЕЙ Saccharomyces cerevisiae ДЛЯ ЕГО ОСУЩЕСТВЛЕНИЯ
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RU2229307C1 (ru) * 2002-10-22 2004-05-27 ООО "Фирма"БиоМедИнвест" Композиция рекомбинантных белков, способ получения такой композиции, фармацевтический набор реагентов для иммунотерапии и профилактической вакцинации опухолевых заболеваний аногенитальной сферы, способ иммунотерапии и профилактической вакцинации на его основе
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