mRNA vaccines

Abstract

The present invention generally relates to combinations of mRNA molecules encoding functional immunostimulatory proteins with CTLA4 pathway inhibitors. In particular, the present invention relates to combinations of one or more mRNA molecules encoding at least one functional immunostimulatory protein selected from the list comprising CD40L, CD70, and caTLR4, optionally with a CTLA4 pathway inhibitor, also in the form of an mRNA molecule. The present invention further relates to vaccines comprising such combinations, and to the use of the combinations and vaccines of the invention in human or veterinary medicine, particularly in the prevention and / or treatment of cell proliferative disorders.

Description

[Technical field]

[0001] The present invention generally relates to a combination of an mRNA molecule encoding a functional immunostimulatory protein with a CTLA4 pathway inhibitor. In particular, the present invention relates to a combination of one or more mRNA molecules encoding at least one functional immunostimulatory protein selected from the list comprising CD40L, CD70 and caTLR4 with a CTLA4 pathway inhibitor, optionally also in the form of an mRNA molecule. The present invention further relates to a vaccine comprising such a combination, and to the use of the combination and vaccine of the present invention in human or veterinary medicine, in particular in the prevention and / or treatment of cell proliferative disorders. [Background technology]

[0002] Induction of a strong cytolytic CD8 T cell response capable of recognizing and killing cancer cells is the main goal of any therapeutic cancer vaccine. The ability of a vaccine to induce such a cytolytic T cell response is highly dependent on the initial interaction between the vaccine and dendritic cells (DCs), which are the most potent antigen-presenting cells and instigators of T cell immunity. In contrast to protein-based vaccines, mRNA vaccines allow expression of antigens encoded by mRNA in the cytosol of DCs, which is the natural pathway for antigen processing and presentation to CD8 T cells.

[0003] In addition, in vitro transcribed mRNA produced by viral polymerases such as T7 is partially similar to viral RNA and therefore recognized by innate immune sensors, endowing the mRNA with inherent adjuvant properties (Non-Patent Document 1, Non-Patent Document 2). Nevertheless, activation of DCs by IVT mRNA is suboptimal and can be further enhanced by co-delivery of TriMix mRNA, a mixture of three mRNAs encoding the immunostimulatory proteins CD40L, CD70 and caTLR4 (Non-Patent Document 3, Non-Patent Document 4, Non-Patent Document 5). The addition of TriMix mRNA to mRNAs encoding tumor antigens has been demonstrated to strongly enhance the magnitude of T cell responses and their antitumor efficacy in preclinical models and is currently being investigated in clinical studies.

[0004] Anti-CTLA4 antibodies can disrupt tumor immune suppression and restore the anti-tumor effect of existing effector T cells, and the anti-CTLA-4 blocking antibody ipilimumab is the first immune checkpoint inhibitor approved for the treatment of cancer patients (Non-Patent Document 6). The interaction of CTLA-4 with CD80 and CD86 present on the surface of antigen-presenting cells provides an inhibitory signal to T cells during initial priming. In addition, CTLA-4 is highly expressed on the surface of regulatory T cells.

[0005] mRNA vaccines have the ability to induce / expand antitumor T cells against tumor-associated antigens encoded by mRNA. Nevertheless, the therapeutic effect of vaccine-induced T cells is often hindered by the strong immunosuppressive microenvironment present at the tumor site. Because CTLA-4 antibodies can block co-inhibitory signals during T cell priming and interfere with the immunosuppressive function of regulatory T cells, we assessed whether combining TriMix-based mRNA vaccination with anti-CTLA4 antibodies could enhance the antitumor effect.

[0006] Surprisingly, the inventors found that while anti-CTLA4 monotherapy had no therapeutic effect, the combination of TriMix-based mRNA vaccination with anti-CTLA4 therapy showed a superior anti-tumor effect, even superior to TriMix monotherapy. [Prior art documents] [Non-patent literature]

[0007] [Non-Patent Document 1] Kariko K et. Immunity 2005; 23, 165-175. [Non-Patent Document 2] Yoneyama, M. & Fujita, T. Rev. Med. Virol. 2010; 20, 4-22 [Non-Patent Document 3] Bonehill A et al. Mol. Ther. 2008; 16:1170-80. [Non-Patent Document 4] Van Lint S. et al. Cancer Res. 2012; 1;72(7):1661-71 [Non-Patent Document 5] Van Lint S. et al. Cancer Res Immunol. 2016; 4 (2):146-156 [Non-Patent Document 6] Seidel et al. Front Oncol. 2018; 8: 86. Summary of the Invention

[0008] The invention is described by the following numbered statements.

[0009] 1. one or more mRNA molecules encoding at least one functional immunostimulatory protein selected from the list including CD40L, CD70 and caTLR4; a CTLA4 pathway inhibitor that blocks or blocks signal transduction initiated by CTLA4; A combination including:

[0010] 2. The combination according to statement 1, wherein said one or more mRNA molecules encode all of the functional immunostimulatory proteins selected from the list comprising CD40L, CD70 and caTLR4.

[0011] 3. The combination of statement 1, wherein said CTLA4 pathway inhibitor is in the form of an mRNA encoding said CTLA4 pathway inhibitor.

[0012] 4. The combination according to statement 1, wherein said CTLA4 pathway inhibitor is an antagonist antibody, nanobody or derivative thereof against CTLA4.

[0013] 5. The combination according to statement 4, wherein said antagonist antibody against CTLA4 is selected from the list comprising ipilimumab and tremelimumab.

[0014] 6. A combination according to any one of statements 1 to 5, further comprising one or more mRNA molecules encoding a tumor-specific antigen.

[0015] 7. A combination according to any one of statements 1 to 6, wherein said one or more mRNA molecules are formulated for parenteral administration, more particularly for intravenous, intratumoral, intradermal, subcutaneous, intraperitoneal, intramuscular or intranodal administration.

[0016] 8. A combination according to any one of statements 1 to 7, wherein the mRNA molecule is encapsulated in a nanoparticle.

[0017] 9. A combination according to any one of statements 1 to 8, wherein said nanoparticles are selected from the list comprising lipid nanoparticles and polymeric nanoparticles.

[0018] 10. A combination according to any one of statements 1 to 7, wherein said mRNA molecule is formulated for intranodal or intratumoral administration and is in the form of a naked mRNA molecule in a suitable injection buffer such as Ringer Lactate buffer.

[0019] 11. A combination according to any one of statements 1 to 10, wherein said CTLA4 pathway inhibitor is formulated for parenteral administration, more particularly for intravenous, intratumoral, intradermal, subcutaneous, intraperitoneal, intramuscular or intranodal administration.

[0020] 12. A vaccine comprising a combination according to any one of statements 1 to 11.

[0021] 13. A combination according to any one of statements 1 to 11, or a vaccine according to statement 12, for use in human or veterinary medicine.

[0022] 14. A combination according to any one of statements 1 to 11, or a vaccine according to statement 12, for use in the prevention and / or treatment of a cell proliferative disorder.

[0023] 15. A combination according to any one of statements 1 to 11 or a vaccine according to statement 12 for use in inducing an immune response against a tumor in a subject.

[0024] Thus, in a first aspect, the present invention provides a method for producing a method for treating a cancer cell comprising the steps of: one or more mRNA molecules encoding at least one functional immunostimulatory protein selected from the list comprising CD40L, CD70 and caTLR4; a CTLA4 pathway inhibitor that blocks or blocks signal transduction initiated by CTLA4; The present invention provides a combination comprising:

[0025] In a particular embodiment of the invention, said one or more mRNA molecules encode all of the functional immunostimulatory proteins selected from the list comprising CD40L, CD70 and caTLR4.

[0026] In yet further embodiments of the invention, the CTLA4 pathway inhibitor is in the form of an mRNA encoding the CTLA4 pathway inhibitor. Alternatively, the CTLA4 pathway inhibitor is an antagonistic antibody, nanobody or derivative thereof against CTLA4. More specifically, the antagonistic antibody against CTLA4 may be selected from the list comprising ipilimumab and tremelimumab.

[0027] In certain embodiments, the combination of the present invention may further comprise one or more mRNA molecules encoding a tumor antigen.

[0028] In a further embodiment of the invention, the one or more mRNA molecules are formulated for parenteral administration, more particularly for intravenous, intratumoral, intradermal, subcutaneous, intraperitoneal, intramuscular or intranodal administration.

[0029] In a particularly preferred embodiment, the above-mentioned combinations or mRNA molecules of the invention are formulated in nanoparticles, such as lipid nanoparticles or polymeric nanoparticles.

[0030] In yet a further specific embodiment, the mRNA molecule is formulated for intranodal or intratumoral administration and is in the form of naked mRNA molecule in a suitable injection buffer, such as Ringer's lactate buffer.

[0031] The present invention also provides a combination as described herein, wherein said CTLA4 pathway inhibitor is formulated for parenteral administration, more particularly for intravenous, intratumoral, intradermal, subcutaneous, intraperitoneal, intramuscular or intranodal administration.

[0032] In a further aspect, the present invention provides a vaccine comprising a combination as described herein.

[0033] In a particular embodiment, the present invention relates to a combination or vaccine as described herein for use in human or veterinary medicine, more particularly for the prevention and / or treatment of a cell proliferative disorder, e.g. for inducing an immune response against a tumor in a subject.

[0034] With specific reference now to the drawings, it is emphasized that the description shown is by way of example and is intended only as an explanatory discussion of various embodiments of the present invention. These drawings are presented to provide what is believed to be the most useful and simplest explanation of the principles and conceptual aspects of the present invention. In this regard, no attempt has been made to show structural details of the invention in more detail than is necessary for a fundamental understanding of the invention. This description, taken together with the drawings, will make apparent to those skilled in the art how some forms of the present invention may be actually embodied. [Brief description of the drawings]

[0035] [Figure 1] Tumor growth curves of individual mice after treatment of TC-1 bearing mice with PBS, anti-CTLA4, E7-TriMix mRNA, or a combination of E7-TriMix mRNA and anti-CTLA-4. CR = complete responder, i.e., the number of tumor-free mice 90 days after tumor inoculation. [Diagram 2] Average tumor growth curves of TC-1-inoculated mice treated with PBS, anti-CTLA4, E7+TriMix mRNA, and E7+TriMix mRNA+anti-CTLA4.**p=0.0012, 2-way ANOVA with Bonferroni's multiple comparison test. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0036] As already detailed herein above, the present invention relates to a method for producing a medicament for the treatment of a pulmonary arthritis, comprising: one or more mRNA molecules encoding at least one functional immunostimulatory protein selected from the list comprising CD40L, CD70 and caTLR4; a CTLA4 pathway inhibitor that blocks or blocks signal transduction initiated by CTLA4; The present invention relates to a combination comprising:

[0037] Throughout this invention, the term "TriMix" refers to a mixture of mRNA molecules encoding CD40L, CD70 and caTLR4 immunostimulatory proteins. Use of the combination of CD40L and caTLR4 results in mature cytokine / chemokine secreting DCs, as shown for ligation of CD40 and TLR4 by addition of soluble CD40L and LPS. Introduction of CD70 into DCs enhances CD27 by inhibiting activated T cell apoptosis and supporting T cell proliferation. + It provides costimulatory signals to naive T cells. As an alternative to caTLR4, other Toll-like receptors (TLRs) can be used. Constitutively active forms of each TLR are known, and it is possible that they can be introduced into DCs to induce host immune responses. However, in our opinion, caTLR4 is the most potent activating molecule, and therefore is preferred.

[0038] The mRNA or DNA used or referred to herein may be naked mRNA or DNA or protected mRNA or DNA. Protection of DNA or mRNA increases its stability while maintaining the ability of the mRNA or DNA to be used for vaccination purposes. Non-limiting examples of protection of both mRNA and DNA may be liposomal encapsulation, protamine protection, (cationic) lipid lipoplexation, cationic or polycationic compositions of lipids, mannosylated lipoplexation, bubble liposomation, polyethyleneimine (PEI) protection, liposome-loaded microbubble protection, etc. In certain embodiments, the mRNA or DNA molecules defined herein are isolated mRNA or DNA molecules, in particular, they are preferably not part of a cell (such as a dendritic cell). The present invention particularly contemplates in vivo applications involving the direct use of isolated mRNA or DNA molecules, as opposed to ex vivo approaches involving the use of dendritic cells transfected with such mRNA or DNA molecules.

[0039] The present invention is particularly suitable for use in conjunction with tumor-specific antigens, but may also be suitably used in conjunction with other types of target-specific antigens.

[0040] The term "target" used throughout the specification is not limited to the specific examples that may be described herein. Any infectious agent, such as a virus, a bacterium or a fungus, may be targeted. In addition, any tumor or cancer cell may be targeted. The term "target-specific antigen" used throughout the specification is not limited to the specific examples that may be described herein. It is clear to the skilled artisan that the present invention relates to the induction of immune activation in APCs, regardless of the target-specific antigen presented. The antigen presented depends on the type of target intended to induce an immune response in the subject. Typical examples of target-specific antigens are markers expressed or secreted specific to tumors, bacteria and fungal cells, or specific viral proteins or structures. Without wishing to limit the scope of protection of the present invention, some examples of possible markers are given below.

[0041] The terms "neoplasm", "cancer" and / or "tumor" used throughout the specification are not intended to be limited to the types of cancer or tumor that may be exemplified. Thus, the terms encompass all proliferative disorders, such as neoplasms, dysplasias, premalignant or precancerous lesions, abnormal cell proliferation, benign tumors, malignant tumors, cancers or metastases, and the cancers may be selected from the group of leukemia, non-small cell lung cancer, small cell lung cancer, CNS cancer, melanoma, ovarian cancer, renal cancer, prostate cancer, breast cancer, glioma, colon cancer, bladder cancer, sarcoma, pancreatic cancer, colorectal cancer, head and neck cancer, liver cancer, bone cancer, bone marrow cancer, gastric cancer, duodenal cancer, esophageal cancer, thyroid cancer, blood cancer, and lymphoma. Cancer-specific antigens may be, for example, MelanA / MART1, cancer-germ cell antigen, gp100, tyrosinase, CEA, PSA, Her-2 / neu, survivin, telomerase.

[0042] In a preferred embodiment of the vaccine of the invention, the mRNA or DNA molecule(s) encode CD40L and CD70 immunostimulatory proteins. In a particularly preferred embodiment of the vaccine of the invention, the mRNA or DNA molecule(s) encode CD40L, CD70 and caTLR4 immunostimulatory proteins.

[0043] Said mRNA or DNA molecules encoding immunostimulatory proteins may be part of a single mRNA or DNA molecule. Preferably, said single mRNA or DNA molecule is capable of expressing two or more proteins simultaneously. In one embodiment, said mRNA or DNA molecules encoding immunostimulatory proteins are separated in the single mRNA or DNA molecule by a sequence encoding an internal ribosome entry site (IRES) or a self-cleaving 2a peptide.

[0044] In certain embodiments, one or more of the above mRNA molecules of the invention may further contain a translation enhancer and / or a nuclear retention element. Suitable translation enhancers and nuclear retention elements are those described in WO2015071295.

[0045] Cytotoxic T-lymphocyte antigen-4 (CTLA-4) is expressed primarily in the intracellular compartment of T cells. Upon activation of naive T cells, CTLA-4 is transported to the cell surface and enriched at the immune synapse, where it competes with CD28 for CD80 / CD86 and downmodulates TCR signaling.

[0046] The term "CTLA4 pathway inhibitor" as used herein includes any compound that blocks or blocks signal transduction initiated by CTLA4. Thus, CTLA4 pathway inhibitors may directly or indirectly affect the regulation of CTLA4, for example, by decreasing the expression of the CTLA4 receptor (i.e., transcription and / or translation) or its natural ligands B7-1 (CD80) and B7-2 (CD86). Such inhibitors include, but are not limited to, siRNAs, antisense molecules, proteins, peptides, small molecules, antibodies, nanobodies, and derivatives of any of these. In certain embodiments, the CTLA4 inhibitors may also be provided in the form of mRNAs encoding the inhibitors, such as mRNAs encoding anti-CTLA4 antibodies.

[0047] Preferred anti-CTLA4 antibodies are human antibodies that specifically bind to human CTLA4. Human antibodies offer substantial advantages in the therapeutic methods of the invention, as they are expected to minimize immunogenic and allergic reactions associated with the use of non-human antibodies in human patients.

[0048] Exemplary human anti-CTLA4 antibodies are described in detail, for example, in WO 00 / 37504. Such antibodies include, but are not limited to, 3.1.1, 4.1.1, 4.8.1, 4.10.2, 4.13.1, 4.14.3, 6.1.1, ticilimumab, 11.6.1, 11.7.1, 12.3.1.1, and 12.9.1.1, as well as tremelimumab and ipilimumab. In another embodiment, the antibody is selected from antibodies having the full-length, variable region, or CDR, heavy and light chain amino acid sequences of the antibodies defined above.

[0049] In other embodiments of the invention, the antibody inhibits binding between CTLA4 and B7-1, B7-2, or both. Preferably, the antibody has an IC of about 100 nM or less, more preferably about 10 nM or less, such as about 5 nM or less, even more preferably about 2 nM or less, or even more preferably, such as about 1 nM or less. 50Similarly, the antibody can inhibit binding to B7-1 with an IC of about 100 nM or less, more preferably 10 nM or less, e.g., even more preferably about 5 nM or less, even more preferably about 2 nM or less, or even more preferably about 1 nM or less. 50 can inhibit the binding of B7-2.

[0050] While the anti-CTLA4 antibodies discussed herein above may be preferred, one of skill in the art will understand based on the disclosure provided herein that the invention encompasses a wide variety of anti-CTLA4 antibodies and is not limited to these particular antibodies. More specifically, while human antibodies are preferred, the invention is in no way limited to human antibodies; rather, the invention encompasses antibodies useful regardless of species origin, including, inter alia, chimeric, humanized, and / or primatized antibodies.

[0051] In one embodiment, the disclosure provides a method of combination therapy of an individual comprising administering to the individual one or more immunostimulatory factors and CTLA4 pathway inhibitors. The one or more immunostimulatory factors and CTLA4 pathway inhibitors may be administered simultaneously or contemporaneously in a single composition or separate compositions, or the one or more immunostimulatory factors and CTLA4 pathway inhibitors may be administered at different times.

[0052] The compositions of the invention generally include a CTLA4 pathway inhibitor in combination with one or more immunostimulatory factors.

[0053] The suitable dosage of CTLA4 pathway inhibitor depends on many factors, including, for example, the age and weight of the individual, at least one exact pathology that needs to be treated, the severity of the pathology, the nature of the composition, the route of administration, and combinations thereof.Finally, the suitable dosage can be easily determined by those skilled in the art, such as, for example, physicians, veterinarians, scientists, and other medical and research professionals.For example, those skilled in the art can start with a low dosage and increase it until it achieves the desired therapeutic outcome or result.Alternatively, those skilled in the art can start with a high dosage and decrease it until it reaches the minimum dosage required to achieve the desired therapeutic outcome or result.

[0054] The present invention also provides a combination as described herein, wherein said mRNA is packaged in a nanoparticle.

[0055] As used herein, the term "nanoparticle" refers to any particle having a diameter that makes the particle particularly suitable for systemic administration, especially intravenous administration, of nucleic acids, typically having a diameter of less than 1000 nanometers (nm).

[0056] In a particular embodiment of the invention, the nanoparticles are selected from the list comprising lipid nanoparticles and polymeric nanoparticles.

[0057] Lipid nanoparticles (LNPs) are commonly known as nano-sized particles composed of a combination of different lipids. While many different types of lipids can be included in such LNPs, the LNPs of the present invention can be composed of, for example, a combination of ionic lipids, phospholipids, sterols, and PEG lipids.

[0058] Polymeric nanoparticles can typically be nanospheres or nanocapsules. Two main strategies are used for the preparation of polymeric nanoparticles: the "top-down" approach and the "bottom-up" approach. In the top-down approach, polymeric nanoparticles are generated by dispersion of preformed polymers, whereas in the bottom-up approach, polymerization of monomers leads to the formation of polymeric nanoparticles. Both the top-down and bottom-up methods use synthetic polymers / monomers such as poly(d,l-lactide-co-glycolide), poly(ethyl cyanoacrylate), poly(butyl cyanoacrylate), poly(isobutyl cyanoacrylate), and poly(isohexyl cyanoacrylate); stabilizers such as poly(vinyl alcohol) and didecyldimethylammonium bromide; and organic solvents such as dichloromethane and ethyl acetate, benzyl alcohol, cyclohexane, acetonitrile, acetone, etc. Recently, the scientific community has been trying to find alternatives to synthetic polymers using synthetic methods that use natural polymers and less toxic solvents.

[0059] The present invention also provides the combinations and vaccines described herein for use in human or veterinary medicine, particularly for use in the treatment of cell proliferative disorders, and more particularly for use in eliciting an immune response against a tumor in a subject.

[0060] Finally, the present invention provides a method of treating a cell proliferative disorder comprising the step of administering to a subject in need of treatment a combination or vaccine of the present invention.

[0061] Such compositions may also be of value in the veterinary field, which for the purposes herein includes not only the prevention and / or treatment of diseases in animals, but also, in the case of economically important animals such as cattle, pigs, sheep, chickens, fish, etc., enhancing the growth and / or weight of the animal, and / or the quantity and / or quality of meat or other products obtained from the animal.

[0062] The invention will now be illustrated by the following synthetic and biological examples, which are not to be construed as limiting the scope of the invention in any way. EXAMPLES

[0063] Example 1: Antitumor efficacy of TriMix-based immunization as monotherapy or in combination with anti-CTLA4 antibody in TC-1 tumor-bearing mice Materials and Methods: Synthesis of in vitro transcribed mRNA: E7 mRNA and mouse caTLR4, mouse CD70 and mouse CD40L (TriMix) mRNA were synthesized from the corresponding linearized peTheRNA plasmids by in vitro transcription as previously described (EP 3068888).

[0064] mouse: C57BL / 6 mice (female, 6 weeks old) were purchased from Janvier (Genest) and maintained under SPF (OncoDesign, Dijon) conditions according to FELASA guidelines. TC-1 cells were obtained from ATCC and cultured as previously described.

[0065] antibody: Anti-CTLA4 antibody was purchased from BioXCell (reference: BE0131; clone: ​​9H10; reactivity: mouse; isotype: hamster IgG1; storage conditions: +4°C).

[0066] Tumor inoculation and treatment schedule: On day 0, mice were administered 1 × 10 6 TC-1 tumor cells were inoculated subcutaneously. Intralymph node immunization with E7 / TriMix mRNA was performed on days 3, 8, and 13 after tumor inoculation. Mice received 10 μg of E7 mRNA combined with 30 μg of TriMix mRNA dissolved in 0.8× Ringer's lactate solution (20 μl). The mRNA was injected into the inguinal lymph node. Anti-CTLA4 antibody was injected intraperitoneally (10 mg / kg / dose) on days 3, 6, 9, and 12 after tumor inoculation.

[0067] Intralymph node injection The groin of the mice was shaved before disinfection with 70% ethanol. A small incision was made in the groin to expose the inguinal lymph node. A total volume of 10 μl of mRNA solution was injected into the inguinal lymph node using a 0.3 ml 30G insulin needle (BD Biosciences, ref. 324826A). After injection, the skin was closed with 4-0 crinerce sutures. Injections were performed into the same inguinal lymph node for all three intralymph node immunizations.

[0068] result: As detailed in Figures 1 and 2, the described anti-CTLA-4 monotherapy regimen did not provide any therapeutic benefit in the TC-1 model compared to control (PBS), with no complete responders in any group. In contrast, intranodal immunization with E7 / TriMix was shown to significantly slow TC-1 tumor growth and result in a small number of complete responders (2 / 15).

[0069] Even more surprising was that the combination of anti-CTLA4 antibody and intranodal E7 / TriMix immunization demonstrated superior antitumor efficacy compared to both intranodal E7 / TriMix monotherapy (p=0.0012) and anti-CTLA4 monotherapy, resulting in a significantly higher rate of complete responders (9 / 15).

[0070] Thus, these data clearly demonstrate the potential of combining TriMix with CTLA4 pathway inhibitor molecules in tumor therapy.

Claims

1. In the manufacture of a medicament for preventing and / or treating HPV-related cancer, one or more isolated mRNA molecules encoding all of the functional immunostimulatory proteins CD40L, CD70, and caTLR4; an mRNA molecule encoding the E7 antigen, and an antagonist antibody against CTLA4 selected from ipilimumab and tremelimumab; The use of a combination comprising The combination does not include dendritic cells, and the one or more isolated mRNA molecules are formulated for intralymphatic administration.

2. The use according to claim 1, wherein the antagonist antibody against CTLA4 is ipilimumab.

3. The use according to claim 1 or 2, wherein one or more isolated mRNA molecules encoding all of the functional immunostimulatory proteins CD40L, CD70 and caTLR4 and / or an mRNA molecule encoding the E7 antigen are formulated within nanoparticles.

4. The use according to claim 3, wherein the nanoparticles are selected from lipid nanoparticles and polymeric nanoparticles.

5. The use according to claim 1 or 2, wherein one or more isolated mRNA molecules encoding all of the functional immunostimulatory proteins CD40L, CD70 and caTLR4 and / or the mRNA molecule encoding the E7 antigen are formulated for intralymphatic administration and are in the form of naked mRNA molecules in a suitable injection buffer.

6. The use according to claim 1 or 2, wherein one or more isolated mRNA molecules encoding all of the functional immunostimulatory proteins CD40L, CD70 and caTLR4 and / or the mRNA molecule encoding the E7 antigen are formulated for intralymphatic administration and are in the form of naked mRNA molecules in Ringer's lactate buffer.

7. The use according to any one of claims 1 to 6, wherein the antagonist antibody against CTLA4 is formulated for parenteral administration.

8. The use according to any one of claims 1 to 7, wherein the antagonist antibody against CTLA4 is formulated for intravenous, intratumoral, intradermal, subcutaneous, intraperitoneal, intramuscular or intralymphatic administration.

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