WO2025190901A1

WO2025190901A1 - Use of myeloperoxidase (mpo) inhibitors for promoting cd8+ t cell responses

Info

Publication number: WO2025190901A1
Application number: PCT/EP2025/056536
Authority: WO
Inventors: Sophie UGOLINI; Anais ROGER
Original assignee: Aix Marseille Universite; Centre National de la Recherche Scientifique CNRS; Institut National de la Sante et de la Recherche Medicale INSERM
Current assignee: Aix Marseille Universite; Centre National de la Recherche Scientifique CNRS; Institut National de la Sante et de la Recherche Medicale INSERM
Priority date: 2024-03-13
Filing date: 2025-03-11
Publication date: 2025-09-18
Anticipated expiration: 2026-09-13

Abstract

Host protection against infectious diseases involves complex cross-regulation between the immune system and the nervous system. However, the molecular mechanisms behind this regulation remain poorly understood, particularly in the context of viral infections. Using a mouse model of herpes simplex virus 1 (HSV-1) infection, the inventors demonstrate a role of primary sensory neurons in regulating the antiviral innate and adaptive immune response in the skin and the skin draining lymph node. The inventors show that an excess of neutrophil myeloperoxidase (MPO) activity in the skin inhibits the dendritic cell response, thereby limiting the priming of HSV-1-specific CD8⁺ T cells in the skin draining lymph node. This study reveals novel site-specific neuroimmune regulatory mechanisms controlling antiviral host defense, opening up novel therapeutic perspectives. Thus, the present invention relates to a method of promoting CD8⁺ T-cell responses in a subject in need thereof comprising administering to the patient a therapeutically effective amount of a myeloperoxidase inhibitor.

Description

USE OF MYELOPEROXIDASE (MPO) INHIBITORS FOR PROMOTING CD8+ T

CELL RESPONSES

FIELD OF THE INVENTION:

The present invention is in the field of medicine, in particular virology and immunology.

BACKGROUND OF THE INVENTION:

Host protection against infection is mainly mediated by the immune system. However, recent studies have revealed that the nervous system plays an important role in controlling inflammatory processes during infectious diseases. As a barrier organ, the skin is one of the first lines of defence against pathogens. At steady state, the skin-resident immune system includes macrophages, dendritic cells (DC), mast cells and T cell subsets¹. These cells act as sentinels, reacting to physical injuries or pathogens by mounting a robust inflammatory response inducing the recruitment of additional immune cells. Neutrophils are among the first cells to migrate from the blood to the site of assault. They can kill pathogens and produce cytokines, chemokines, and proteolytic enzymes promoting the recruitment and activation of other cells, including monocytes² . Skin DC take-up foreign antigens and present them to naive T cells in the skin-draining lymph nodes (dLN), initiating an adaptive T cell response³'⁴. The early events occurring at the site of infection are therefore crucial for the development of protective immunity.

The skin is also innervated by a dense meshwork of low- and high-threshold sensory nerves⁵'⁶. These nerves include the nociceptive sensory neurons (also called nociceptors), which specialise in detecting noxious stimuli and eliciting pain perception. During inflammatory processes and infections, pathogen-derived molecules, lipids and immune cell-derived mediators act on the peripheral nerve terminals of nociceptive sensory neurons⁷. These mediators include ATP, prostaglandins and leukotrienes, bradykinin, histamine, growth factors and cytokines⁸. These mediators bind to their receptors on the neurons, modifying neuron excitability and increasing action potential generation. The resulting signals are transduced to the spinal cord and relayed to the brain for processing, leading to the perception of pain. Neuronal responses to pathogens and injury are induced over a scale of milliseconds, whereas immune cell response induction takes hours or days. The early response of neurons may therefore make an important contribution to host defence, the elimination of threats, and tissue repair processes. Upon activation, nociceptive sensory neurons release a number of mediators locally that can modulate the activity of immune cells present in the damaged skin⁸. However, depending on the experimental model considered, these neurons may have either pro- or antiinflammatory activities, suggesting that their regulatory role depends on the pathological context⁹'¹¹

Nociception has long been known to be modulated during infections caused by members of the herpesvirus family¹²'¹³ . One of these viruses, herpes simplex virus type 1 (HSV-1), is a neurotropic virus highly prevalent in the human population. HSV-1 is transmitted principally by oral contact, and causes infections in or around the mouth, which may manifest as painful blisters or ulcers. After the initial mucocutaneous disease, the virus infects the peripheral nervous system. Infected individuals often experience a tingling, itching or burning sensation around the mouth, before the appearance of skin/mucosal lesions, indicating an early activation of nociceptive pathways. Viral replication occurs in the DRGs and the virions then travel by anterograde transport to the skin, where a secondary growth phase occurs, leading to a zosteriform skin lesion across the entire dermatome. The initiation of the first phase of infection triggers a strong immune response involving both innate and adaptive immune cells, which control viral replication and limit tissue damage¹⁴. HSV-1 then enters latency, but reactivation may occur periodically in neurons—, causing recurrent symptoms.

We have shown that primary sensory neurons expressing the Navi.s sodium channel, including most nociceptive sensory neurons, play a major role in regulating host responses to HSV-1 infection¹². These neurons are required to control neutrophil infiltration in the skin, thereby limiting the severity of tissue damage¹⁶. This downregulation of neutrophil influx by Navi.s⁺ sensory neurons is also required to trigger virus-specific CD8⁺ T-cell priming by dendritic cells (DCs) in the skin-draining lymph nodes (dLNs)¹⁶. However, the molecular mechanisms underlying this neuroimmune regulation are unknown.

SUMMARY OF THE INVENTION:

The present invention is defined by the claims. In particular, the present invention relates to the use of myeloperoxidase (MPO) inhibitors for promoting CD8+ T cell responses.

DETAILED DESCRIPTION OF THE INVENTION:

Host protection against infectious diseases involves complex cross-regulation between the immune system and the nervous system. However, the molecular mechanisms behind this regulation remain poorly understood, particularly in the context of viral infections. Using a mouse model of herpes simplex virus 1 (HSV-1) infection, the inventors demonstrate a role of primary sensory neurons in regulating the antiviral immune response in the skin and in the skin draining lymph nodes. They investigated the mechanisms underlying this regulation. They found that sensory neurons are required to limit the influx of neutrophils in the skin. Neutrophils produce high levels of myeloperoxidase (MPO). The inventors show that an excess of MPO activity in the skin inhibits the dendritic cell response, thereby limiting the priming of HSV-l-specific CD8⁺ T cells in the skin draining lymph node. This study reveals novel regulatory mechanisms controlling antiviral host defense, opening up novel therapeutic perspectives.

Accordingly, the first object of the present invention relates to a method of promoting CD8+ T-cell responses in a subject in need thereof comprising administering to the patient a therapeutically effective amount of a myeloperoxidase inhibitor.

As used herein, the term “subject” is interchangeable with the term “individual” or “patient”, and may refer to a subject to be treated by the methods disclosed herein. In particular embodiment, the patient is affected or likely to suffer from an infection, more particularly a skin infection. In particular embodiment, the patient is affected or likely to suffer from a viral infection, more particularly a skin viral infection. In particular embodiment, the patient is affected or likely to suffer from a cancer, more particularly a skin cancer. In some embodiments, the patient is a mammal. Non-limiting examples of mammals include rodents (e.g., mice and rats), primates (e.g., lemurs, bushbabies, monkeys, apes, and humans), rabbits, dogs (e.g., companion dogs, service dogs, or work dogs such as police dogs, military dogs, race dogs, or show dogs), horses (such as race horses and work horses), cats (e.g., domesticated cats), livestock (such as pigs, bovines, donkeys, mules, bison, goats, camels, and sheep), and deer. In some embodiments, the mammal is a human. In some embodiments, the patient is a human infant. In some embodiments, the patient is a human child. In some embodiments, the patient is a human adult.

As used herein “CD8+ T cell” has its general meaning in the art and refers to a subset of T cells, which express CD8 on their surface. They are MHC class I-restricted, and function as cytotoxic T cells. “CD8+ T cells” are also called “cytotoxic T lymphocytes” (CTL), “T-killer cell”, or “cytolytic T cells”. CD8 antigens are members of the immunoglobulin superfamily and are associative recognition elements in major histocompatibility complex class I-restricted interactions.

As used herein, the term “CD8+ T cell response” refers to the specific proliferation and activation of CD8+ T cell effector functions induced by an antigen. In particular, CD8+ T cell effector functions may include, but are not limited to, lysis of antigen-pulsed, antigen-precursor pulsed, or naturally antigen-presenting target cells; secretion of cytokines, such as IFN-y, TNF- a or IL-2 induced by a antigen; and secretion of effector molecules such as granzymes or perforins induced by antigen or degranulation.

As used herein, the expression “promoting CD8+ T cell responses” means activating, enhancing, inducing, initiating, or stimulating CD8+ T cell responses. In particular, the method of the present invention is particularly suitable for promoting CD8+ T cell responses in skin.

As used herein, the term “skin” denotes the skin of the face, of the body and the scalp. The term “skin” includes, without limitation, the lips, skin of the face, hands, arms, neck, scalp, and chest. The skin of a mammal is composed of an epidermis layer, a dermis layer, and a subcutaneous layer. The epidermis is the outer layer of the skin. The dermis, which is the middle layer of the skin, contains nerve endings, sweat glands and oil (sebaceous) glands, hair follicles, and blood vessels. The subcutaneous layer is made up of fat and connective tissue that houses larger blood vessels and nerves.

As used herein the term "antigen" refers to a molecule capable of being specifically bound by the T cell receptor (TCR) of CD8+ T cells if processed and presented by MHC class I molecules. An antigen is thus capable of being recognized by the immune system and/or being capable of inducing a cellular immune response leading to the activation of CD8+ T cells.

In some embodiments, the antigen may be a protein derived from cancer. The cancers, include, but are not limited to, Acute lymphoblastic leukemia; Acute myeloid leukemia; Adrenocortical carcinoma; AIDS-related cancers; AIDS-related lymphoma; Anal cancer; Appendix cancer; Astrocytoma, childhood cerebellar or cerebral; Basal cell carcinoma; Bile duct cancer, extrahepatic; Bladder cancer; Bone cancer, Osteosarcoma/Malignant fibrous histiocytoma; Brainstem glioma; Brain tumor; Brain tumor, cerebellar astrocytoma; Brain tumor, cerebral astrocytoma/malignant glioma; Brain tumor, ependymoma; Brain tumor, medulloblastoma; Brain tumor, supratentorial primitive neuroectodermal tumors; Brain tumor, visual pathway and hypothalamic glioma; Breast cancer; Bronchial adenomas/carcinoids; Burkitt lymphoma; Carcinoid tumor, childhood; Carcinoid tumor, gastrointestinal; Carcinoma of unknown primary; Central nervous system lymphoma, primary; Cerebellar astrocytoma, childhood; Cerebral astrocytoma/Malignant glioma, childhood; Cervical cancer; Childhood cancers; Chronic lymphocytic leukemia; Chronic myelogenous leukemia; Chronic myeloproliferative disorders; Colon Cancer; Cutaneous T-cell lymphoma; Desmoplastic small round cell tumor; Endometrial cancer; Ependymoma; Esophageal cancer; Ewing's sarcoma in the Ewing family of tumors; Extracranial germ cell tumor, Childhood; Extragonadal Germ cell tumor; Extrahepatic bile duct cancer; Eye Cancer, Intraocular melanoma; Eye Cancer, Retinoblastoma; Gallbladder cancer; Gastric (Stomach) cancer; Gastrointestinal Carcinoid Tumor; Gastrointestinal stromal tumor (GIST); Germ cell tumor: extracranial, extragonadal, or ovarian; Gestational trophoblastic tumor; Glioma of the brain stem; Glioma, Childhood Cerebral Astrocytoma; Glioma, Childhood Visual Pathway and Hypothalamic; Gastric carcinoid; Hairy cell leukemia; Head and neck cancer; Heart cancer; Hepatocellular (liver) cancer; Hodgkin lymphoma; Hypopharyngeal cancer; Hypothalamic and visual pathway glioma, childhood; Intraocular Melanoma; Islet Cell Carcinoma (Endocrine Pancreas); Kaposi sarcoma; Kidney cancer (renal cell cancer); Laryngeal Cancer; Leukemias; Leukemia, acute lymphoblastic (also called acute lymphocytic leukemia); Leukemia, acute myeloid (also called acute myelogenous leukemia); Leukemia, chronic lymphocytic (also called chronic lymphocytic leukemia); Leukemia, chronic myelogenous (also called chronic myeloid leukemia); Leukemia, hairy cell; Lip and Oral Cavity Cancer; Liver Cancer (Primary); Lung Cancer, Non-Small Cell; Lung Cancer, Small Cell; Lymphomas; Lymphoma, AIDS-related; Lymphoma, Burkitt; Lymphoma, cutaneous T-Cell; Lymphoma, Hodgkin; Lymphomas, Non-Hodgkin (an old classification of all lymphomas except Hodgkin's); Lymphoma, Primary Central Nervous System; Marcus Whittle, Deadly Disease; Macroglobulinemia, Waldenstrom; Malignant Fibrous Histiocytoma of Bone/Osteosarcoma; Medulloblastoma, Childhood; Melanoma; Melanoma, Intraocular (Eye); Merkel Cell Carcinoma; Mesothelioma, Adult Malignant; Mesothelioma, Childhood; Metastatic Squamous Neck Cancer with Occult Primary; Mouth Cancer; Multiple Endocrine Neoplasia Syndrome, Childhood; Multiple Myeloma/Plasma Cell Neoplasm; Mycosis Fungoides; Myelodysplastic Syndromes; Myelodysplastic/Myeloproliferative Diseases; Myelogenous Leukemia, Chronic; Myeloid Leukemia, Adult Acute; Myeloid Leukemia, Childhood Acute; Myeloma, Multiple (Cancer of the Bone-Marrow); Myeloproliferative Disorders, Chronic; Nasal cavity and paranasal sinus cancer; Nasopharyngeal carcinoma; Neuroblastoma; Non-Hodgkin lymphoma; Non-small cell lung cancer; Oral Cancer; Oropharyngeal cancer; Osteosarcoma/malignant fibrous histiocytoma of bone; Ovarian cancer; Ovarian epithelial cancer (Surface epithelial-stromal tumor); Ovarian germ cell tumor; Ovarian low malignant potential tumor; Pancreatic cancer; Pancreatic cancer, islet cell; Paranasal sinus and nasal cavity cancer; Parathyroid cancer; Penile cancer; Pharyngeal cancer; Pheochromocytoma; Pineal astrocytoma; Pineal germinoma; Pineoblastoma and supratentorial primitive neuroectodermal tumors, childhood; Pituitary adenoma; Plasma cell neoplasia/Multiple myeloma; Pleuropulmonary blastoma; Primary central nervous system lymphoma; Prostate cancer; Rectal cancer; Renal cell carcinoma (kidney cancer); Renal pelvis and ureter, transitional cell cancer; Retinoblastoma; Rhabdomyosarcoma, childhood; Salivary gland cancer; Sarcoma, Ewing family of tumors; Sarcoma, Kaposi; Sarcoma, soft tissue; Sarcoma, uterine; Sezary syndrome; Skin cancer (nonmelanoma); Skin cancer (melanoma); Skin carcinoma, Merkel cell; Small cell lung cancer; Small intestine cancer; Soft tissue sarcoma; Squamous cell carcinoma — see Skin cancer (nonmelanoma); Squamous neck cancer with occult primary, metastatic; Stomach cancer; Supratentorial primitive neuroectodermal tumor, childhood; T-Cell lymphoma, cutaneous (Mycosis Fungoides and Sezary syndrome); Testicular cancer; Throat cancer; Thymoma, childhood; Thymoma and Thymic carcinoma; Thyroid cancer; Thyroid cancer, childhood; Transitional cell cancer of the renal pelvis and ureter; Trophoblastic tumor, gestational; Unknown primary site, carcinoma of, adult; Unknown primary site, cancer of, childhood; Ureter and renal pelvis, transitional cell cancer; Urethral cancer; Uterine cancer, endometrial; Uterine sarcoma; Vaginal cancer; Visual pathway and hypothalamic glioma, childhood; Vulvar cancer; Waldenstrom macroglobulinemia and Wilms tumor (kidney cancer).

Thus, the method of the present invention is particularly suitable for the treatment of cancer, in particular skin cancer.

In other words, the invention refers to a method for treating cancer, in particular skin cancer, in a subject in need thereof comprising administering to the subject a therapeutically effective amount of a myeloperoxidase inhibitor, wherein said myeloperoxidase inhibitor promotes CD8+ T response.

As used herein, the term “skin cancer” is intended to include non-melanoma skin cancer, malignant melanoma, Merkel cell carcinoma, squamous cell carcinoma or basal cell carcinoma. Basal cell carcinomas include superficial basal cell carcinoma as well as nodular basal cell carcinoma.

In some embodiments, the antigen may be derived from a pathogen.

In some embodiments, the pathogen may be a bacterial pathogen and the antigen may be a protein derived from the bacterial pathogen. The pathogenic bacteria include, but are not limited to, Bordetella pertussis, Borrelia burgdorferi, Brucella abortus, Brucella canis, Brucella melitensis, Brucella suis, Campylobacter jejuni, Chlamydia pneumoniae, Chlamydia trachomatis, Chlamydophila psittaci, Clostridium botulinum, Clostridium difficile, Clostridium perfringens, Clostridium tetani, Corynebacterium diphtheriae, Enterococcus faecalis, Enterococcus faecium, Escherichia coli, Francisella tularensis, Haemophilus influenzae, Helicobacter pylori, Legionella pneumophila, Leptospira interrogans, Listeria monocytogenes, Mycobacterium leprae, Mycobacterium tuberculosis, Mycobacterium ulcerans, Mycoplasma pneumoniae, Neisseria gonorrhoeae, Neisseria meningitidis, Pseudomonas aeruginosa, Rickettsia rickettsii, Salmonella typhi, Salmonella typhimurium, Shigella sonnei, Staphylococcus aureus, Staphylococcus epidermidis, Staphylococcus saprophyticus, Streptococcus agalactiae, Streptococcus pneumoniae, Streptococcus pyogenes, Treponema pallidum, Vibrio cholera and Yersinia pestis.

In some embodiments, the pathogen may be a parasite and the antigen may be a protein derived from the parasite pathogen. The parasite may be a protozoan organism or disease caused by a protozoan organism such as, but not limited to, Acanthamoeba, Babesiosis, Balantidiasis, Blastocystosis, Coccidia, Dientamoebiasis, Amoebiasis, Giardia, Isosporiasis, Leishmaniasis, Primary amoebic meningoencephalitis (PAM), Malaria, Rhinosporidiosis, Toxoplasmosis — Parasitic pneumonia, Trichomoniasis, Sleeping sickness and Chagas disease. The parasite may be a helminth organism or worm or a disease caused by a helminth organism such as, but not limted to, Ancylostomiasis/Hookworm, Anisakiasis, Roundworm — Parasitic pneumonia, Roundworm — Baylisascariasis, Tapeworm — Tapeworm infection, Clonorchiasis,

Dioctophyme renalis infection, Diphyllobothriasis — tapeworm, Guinea worm — Dracunculiasis, Echinococcosis — tapeworm, Pinworm — Enterobiasis, Liver fluke — Fasciolosis, Fasciolopsiasis — intestinal fluke, Gnathostomiasis, Hymenolepiasis, Loa boa filariasis, Calabar swellings, Mansonelliasis, Filariasis, Metagonimiasis — intestinal fluke, River blindness, Chinese Liver Fluke, Paragonimiasis, Lung Fluke, Schistosomiasis — bilharzia, bilharziosis or snail fever (all types), intestinal schistosomiasis, urinary schistosomiasis, Schistosomiasis by Schistosoma japoni cum, Asian intestinal schistosomiasis, Sparganosis, Strongyloidiasis — Parasitic pneumonia, Beef tapeworm, Pork tapeworm, Toxocariasis, Trichinosis, Swimmer's itch, Whipworm and Elephantiasis Lymphatic filariasis. The parasite may be an organism or disease caused by an organism such as, but not limited to, parasitic worm, Halzoun Syndrome, Myiasis, Chigoe flea, Human Botfly and Candiru. The parasite may be an ectoparasite or disease caused by an ectoparasite such as, but not limited to, Bedbug, Head louse — Pediculosis, Body louse — Pediculosis, Crab louse — Pediculosis, Demodex — Demodicosis, Scabies, Screwworm and Cochliomyia.

In some embodiments, the pathogen may be a viral pathogen and the antigen may be a protein derived from the viral pathogen. Viruses include, but are not limited to viruses belonging to Retroviridae (i.e. Lentivirinae), like HIV (human immunodeficiency virus); Flaviviridae, which comprises (i) the Flaviviruses like Yellow fever virus (YFV) and Dengue virus, the Hepaciviruses like HCV (hepatitis C virus) and (iii) the Pestiviruses like Bovine viral diarrhea virus (BVDV); Herpesviridae, like Herpes simplex virus type 1 (HSV-1) or type 2 (HSV-2), Varicella-zoster virus (VZV), Cytomegalovirus (CMV) or Human Herpes virus type 6 (HHV- 6); Poxyiridcte. like Vaccinia; Hepadnaviridae, like HBV (hepatitis B virus); Coronaviridae, like SARS-CoV-2; Orthomyxoviridae, like influenza virus A, B and C; Togctviridcie Arenaviridae, like Arenavirus; Bunyaviridae, like Punta Toro; Paramyxoviridae, like Respiratory syncytial virus (RSV) or Parainfluenza-3 virus; and Rhabdoviridae . In particular, the virus is a herpes virus. As used herein, the term “herpes virus” includes any virus of the herpes virus family, e.g., herpes simplex virus 1 (HSV-1), herpes simplex virus 2 (HSV-2), varicella zoster virus (VZV), epstein-barr virus (EBV), and human cytomegalovirus (HCMV).

Thus in some embodiments, the method of the present invention is particularly suitable for the treatment of infections. In particular, the method of the present invention is particularly suitable for the treatment of skin infections. In particular, the method of the present invention is suitable for promoting antiviral CD8+ T cell responses. In particular, the method of the present invention is suitable for promoting antiviral CD8+ T cell responses in skin. In particular, the method of the present invention is suitable for promoting CD8+ T cell responses against herpes viruses. In particular, the method of the present invention is suitable for promoting CD8+ T cell responses against herpes viruses in skin. In particular, the method of the present invention is suitable for promoting CD8+ T cell responses against HSV-1. In particular, the method of the present invention is suitable for promoting CD8+ T cell responses against HSV-1 in skin.

As used herein, the term “skin infection” herein refers to any infection of the skin that presents with one or more occurring or reoccurring symptoms such as erythema, warmth, swelling, tenderness, pain, ulcers, lesion(s), nodules, fever, scaling, plaques, papules, pustules, cysts, and the like. Non-limiting examples of skin infections include cellulitis; erysipelas; impetigo; folliculitis; furuncles; carbuncles; secondarily infected dermatoses such as atopic dermatitis, allergic contact dermatitis and psoriasis; secondarily infected traumatic lesions; acne; and other skin disorders associated with infectious pathogens.

In some embodiments, the invention includes a method of treating a viral-induced skin lesion comprising administering to the subject a therapeutically effective amount of a myeloperoxidase inhibitor.

The lesion that results from the virus will vary according to the virus and the clinical presentation that results from the infection. A typical clinical presentation of virally-induced skin lesions are warts and benign tumors. The method is suitable for treatment of numerous types of warts, including but not limited to verrucae warts, plantar warts, flat warts, and genital warts in both men and women. Treatment of skin lesion caused by Molluscum contagiosum, a member of the poxvirus group, by the method of the invention is also contemplated. Another member of the poxviridae family is Varicella-Zoster, which presents in its recurrent form as lesions commonly known as shingles and in an acute form commonly known as chicken pox.

In some embodiments, the invention relates to the use of a MPO inhibitor for the treatment of verrucal virus-based diseases of the skin. The verrucal virus-based diseases of the skin are preferably caused by papillomaviruses, in particular human papillomaviruses (HPV) or molluscipox virus. These are especially preferably verrucal virus-based disease of the skin selected from the group consisting of verrucae vulgares (common warts), verrucae planae juvenilis (juvenile (plane) warts), verrucae plantares (plantar warts), condylomata acuminata (venereal warts), verrucae planae (flat warts), verrucae filiformes (bristle warts) or molluscum contagiosum (dimple warts).

In some embodiments, the invention relates to the use of a MPO inhibitor for the treatment of herpesvirus-based diseases of the skin. The herpesvirus-based diseases of the skin are preferably caused by herpes simplex virus type 1, herpes simplex virus type 2 or varicella zoster virus. These are especially preferably herpesvirus-based diseases of the skin selected from the group comprising herpes labiales, herpes nasalis, keratoconjunctivitis herpetica, stomatitis herpetica, herpes facialis, herpes buccalis, herpes genitales, herpes perianalis and herpes glutealis and herpes zoster.

As used herein, the term "treatment" or "treat" refer to both prophylactic or preventive treatment as well as curative or disease modifying treatment, including treatment of patient at risk of contracting the disease or suspected to have contracted the disease as well as patients who are ill or have been diagnosed as suffering from a disease or medical condition, and includes suppression of clinical relapse. The treatment may be administered to a patient having a medical disorder or who ultimately may acquire the disorder, in order to prevent, cure, delay the onset of, reduce the severity of, or ameliorate one or more symptoms of a disorder or recurring disorder, or in order to prolong the survival of a patient beyond that expected in the absence of such treatment. By "therapeutic regimen" is meant the pattern of treatment of an illness, e.g., the pattern of dosing used during therapy. A therapeutic regimen may include an induction regimen and a maintenance regimen. The phrase "induction regimen" or "induction period" refers to a therapeutic regimen (or the portion of a therapeutic regimen) that is used for the initial treatment of a disease. The general goal of an induction regimen is to provide a high level of drug to a patient during the initial period of a treatment regimen. An induction regimen may employ (in part or in whole) a "loading regimen", which may include administering a greater dose of the drug than a physician would employ during a maintenance regimen, administering a drug more frequently than a physician would administer the drug during a maintenance regimen, or both. The phrase "maintenance regimen" or "maintenance period" refers to a therapeutic regimen (or the portion of a therapeutic regimen) that is used for the maintenance of a patient during treatment of an illness, e.g., to keep the patient in remission for long periods of time (months or years). A maintenance regimen may employ continuous therapy (e.g., administering a drug at regular intervals, e.g., weekly, monthly, yearly, etc.) or intermittent therapy (e.g., interrupted treatment, intermittent treatment, treatment at relapse, or treatment upon achievement of a particular predetermined criteria [e.g., disease manifestation, etc.]). In some embodiments, the method of the present invention is also suitable for promoting CD8+ T cells responses during vaccination. In particular, the method of the present invention is particularly suitable for enhancing the potency of a vaccine.

Accordingly, the present invention relates to a method for enhancing the potency of a vaccine administered to a subject comprising: administering to the subject a pharmaceutically effective amount of a MPO inhibitor in combination with the vaccine.

A further object of the present invention relates to a method of vaccinating a subject in need thereof comprising administering to the subject a therapeutically effective combination of a vaccine with a MPO inhibitor, wherein administration of the combination results in enhanced vaccine efficacy relative to the administration of the vaccine alone.

As used herein, the term “vaccine” refers to an antigenic composition usually comprising an infectious factor or a portion of an infectious factor, such as an antigen, optionally in combination with an immune adjuvant, administered into the body to elicit an immune response (e.g. CD8+ T cell response). The antigenic portion may be a microorganism such as a virus or bacterium; a natural product purified from a microorganism; or a synthetic or genetically engineered protein, peptide, polysaccharide, or similar product. In some embodiments, the antigenic portion of the vaccine of the present invention is comprised of a T cell epitope.

As used herein, the term “vaccination” or “vaccinating” means, but is not limited to, a process to elicit an immune response in a subject against a particular antigen.

As used herein, the term "vaccine composition" is intended to mean a composition which can be administered to humans or to animals in order to induce an immune system response; this immune system response can result in the activation of certain cells, in particular CD8+ T cells.

As used herein, “improving vaccine efficacy” or “improving the efficacy of a vaccine” or the like refers to any change or alteration in the immune response of a subject that is capable of rendering the vaccine of the invention more effective in terms of protection. In some embodiments, this may involve accelerating the appearance of an immune response (e.g. CD8+ T cell responses) and/or improving the persistence or strength of an immune response to the vaccine. In particular, the method of the present invention is particularly suitable for improving the efficacy of a vaccine that is topically, subcutaneously or intradermally administrated to the subject.

As used herein, the term "intradermal administration" refers to the delivery of the vaccine into the dermis layer of the skin of the subject. In contrast in intradermal administration, "subcutaneous administration" refers to the administration of a substance into the subcutaneous layer and "topical administration" refers to the administration of a substance onto the surface of the skin.

Thus, in some embodiments, the MPO inhibitor of the present invention can be used as an adjuvant. As used herein, the term “adjuvant” refers to a compound that can induce and/or enhance the immune response against an antigen when administered to a subject or an animal. It is also intended to mean a substance that acts generally to accelerate, prolong, or enhance the quality of specific immune responses to a specific antigen.

As used herein the term “myeloperoxidase” or “MPO” has its general meaning in the art and refers to a heme-containing enzyme. The enzyme uses hydrogen peroxide to oxidize chloride to hypochlorous acid. Other halides and pseudohalides (like thiocyanate) are also physiological substrates to MPO.

As used herein, the term “MPO inhibitor” refers to any compound natural or not which is capable of inhibiting the activity of MPO. MPO inhibitors are well known in the art. The term encompasses any MPO inhibitor that is currently known in the art or that will be identified in the future, and includes any chemical entity that, upon administration to a patient, results in inhibition or down-regulation of a biological activity associated with activation of the MPO. The term also encompasses inhibitor of expression. The MPO inhibition of the compounds may be determined using various methods well known in the art.

In particular embodiment, the myeloperoxidase inhibitor is an inhibitor of myeloperoxidase expression or an inhibitor of myeloperoxidase activity.

By "biological activity" of a myeloperoxidase is meant inhibiting the dendritic cell response and inhibiting CD8+ T cell response. Tests for determining the capacity of a compound to be an inhibitor of myeloperoxidase are well known to the person skilled in the art, as disclosed in WO 02/090575.

Examples of compounds that can be used as MPO-inhibitors are compounds described in WO 2006/062465, WO 2006/062465, WO 2003/089430, WO 2003/089430, or WO 2003/089430. In particular, W02003/089430, W02005/037835, W02007/120097, W02007/120098 and WO2007/142576 disclose thioxantine derivatives and the use thereof as MPO inhibitors in therapy. W02006/062465 and WO2007/142577 disclose 2-thioxo-l,2,3,4-tetrahydro- pyrrolo[3,2-d]pyrimidin-4-one derivatives claimed to be inhibitors of MPO. W02009/025618 discloses thioxantine and 2-thioxo-l,2,3,4-tetrahydro-pyrrolo[3,2-d]pyrimidin-4-one derivatives and the use thereof as MPO inhibitors for the treatment of multiple system atrophy (MSA) and Huntington's disease (HD) and for neuroprotection. MPO inhibitors are also disclosed in Soubhye J, Van Antwerpen P, Dufrasne F. A patent review of myeloperoxidase inhibitors for treating chronic inflammatory syndromes (focus on cardiovascular diseases, 2013-2019). Expert Opin Ther Pat. 2020 Aug;30(8):595-608. doi:

10.1080/13543776.2020.1780210. Epub 2020 Jun 18. Erratum in: Expert Opin Ther Pat. 2020 Sep 14; : 1. PMID: 32510253 and in Chaikijurajai T, Tang WHW. Myeloperoxidase: a potential therapeutic target for coronary artery disease. Expert Opin Ther Targets. 2020 Jul;24(7):695- 705. doi: 10.1080/14728222.2020.1762177. Epub 2020 May 7. PMID: 32336171; PMCID: PMC7387188.

In some embodiment, the MPO inhibitor is AZD5904 which has the formula of:

In some embodiments, the MPO inhibitor is l-[[2-[(lR)-l-aminoethyl]-4- chlorophenyl]methyl]-2-sulfanylidene-5H-pyrrolo[3,2-d]pyrimidin-4-one (Mitiperstat or AZD4831).

In some embodiments, the MPO inhibitor is an inhibitor of MPO expression. An “inhibitor of expression” refers to a natural or synthetic compound that has a biological effect to inhibit the expression of a gene. In a preferred embodiment of the invention, said inhibitor of gene expression is a siRNA, an antisense oligonucleotide or a ribozyme. For example, anti-sense oligonucleotides, including anti-sense RNA molecules and anti-sense DNA molecules, would act to directly block the translation of MPO mRNA by binding thereto and thus preventing protein translation or increasing mRNA degradation, thus decreasing the level of MPO, and thus activity, in a cell. For example, antisense oligonucleotides of at least about 15 bases and complementary to unique regions of the mRNA transcript sequence encoding MPO can be synthesized, e.g., by conventional phosphodiester techniques. Methods for using antisense techniques for specifically inhibiting gene expression of genes whose sequence is known are well known in the art (e.g. see U.S. Pat. Nos. 6,566,135; 6,566,131; 6,365,354; 6,410,323; 6,107,091; 6,046,321; and 5,981,732). Small inhibitory RNAs (siRNAs) can also function as inhibitors of expression for use in the present invention. MPO gene expression can be reduced by contacting a subject or cell with a small double stranded RNA (dsRNA), or a vector or construct causing the production of a small double stranded RNA, such that MPO gene expression is specifically inhibited (i.e. RNA interference or RNAi).

Inhibitors of gene expression according to the present invention may be based nuclease therapy (like Talen or Crispr).

The term “nuclease” or “endonuclease” means synthetic nucleases consisting of a DNA binding site, a linker, and a cleavage module derived from a restriction endonuclease which are used for gene targeting efforts. The synthetic nucleases according to the invention exhibit increased preference and specificity to bipartite or tripartite DNA target sites comprising DNA binding (i.e. TALEN or CRISPR recognition site(s)) and restriction endonuclease target site while cleaving at off-target sites comprising only the restriction endonuclease target site is prevented. The guide RNA (gRNA) sequences direct the nuclease (i.e. Cas9 protein) to induce a sitespecific double strand break (DSB) in the genomic DNA in the target sequence.

Restriction endonucleases (also called restriction enzymes) as referred to herein in accordance with the present invention are capable of recognizing and cleaving a DNA molecule at a specific DNA cleavage site between predefined nucleotides. In contrast, some endonucleases such as for example Fokl comprise a cleavage domain that cleaves the DNA unspecifically at a certain position regardless of the nucleotides present at this position. Therefore, preferably the specific DNA cleavage site and the DNA recognition site of the restriction endonuclease are identical. Moreover, also preferably the cleavage domain of the chimeric nuclease is derived from a restriction endonuclease with reduced DNA binding and/or reduced catalytic activity when compared to the wildtype restriction endonuclease. According to the knowledge that restriction endonucleases, particularly type II restriction endonucleases, bind as a homodimer to DNA regularly, the chimeric nucleases as referred to herein may be related to homodimerization of two restriction endonuclease subunits. Preferably, in accordance with the present invention the cleavage modules referred to herein have a reduced capability of forming homodimers in the absence of the DNA recognition site, thereby preventing unspecific DNA binding. Therefore, a functional homodimer is only formed upon recruitment of chimeric nucleases monomers to the specific DNA recognition sites. Preferably, the restriction endonuclease from which the cleavage module of the chimeric nuclease is derived is a type IIP restriction endonuclease. The preferably palindromic DNA recognition sites of these restriction endonucleases consist of at least four or up to eight contiguous nucleotides. Preferably, the type IIP restriction endonucleases cleave the DNA within the recognition site which occurs rather frequently in the genome, or immediately adjacent thereto, and have no or a reduced star activity. The type IIP restriction endonucleases as referred to herein are preferably selected from the group consisting of: Pvull, EcoRV, BamHl, Bcnl, BfaSORF1835P, Bfil, Bgll, Bglll, BpuJl, Bse6341, BsoBl, BspD6I, BstYl, CfrlOl, Ecll8kl, EcoO1091, EcoRl, EcoRll, EcoRV, EcoR1241, EcoR12411, HinPl l, Hindi, Hindlll, Hpy991, Hpyl881, Mspl, Muni, Mval, Nael, NgoMIV, Notl, OkrAl, Pabl, Pad, PspGl, Sau3 Al, Sdal, Sfil, SgrAl, Thai, VvuYORF266P, Ddel, Eco571, Haelll, Hhall, Hindll, and Ndel. Antisense oligonucleotides, siRNAs, shRNAs and ribozymes of the invention may be delivered in vivo alone or in association with a vector. In its broadest sense, a "vector" is any vehicle capable of facilitating the transfer of the antisense oligonucleotide, siRNA, shRNA or ribozyme nucleic acid to the cells and typically cells expressing MPO. Typically, the vector transports the nucleic acid to cells with reduced degradation relative to the extent of degradation that would result in the absence of the vector. In general, the vectors useful in the invention include, but are not limited to, plasmids, phagemids, viruses, other vehicles derived from viral or bacterial sources that have been manipulated by the insertion or incorporation of the antisense oligonucleotide, siRNA, shRNA or ribozyme nucleic acid sequences. Viral vectors are a preferred type of vector and include, but are not limited to nucleic acid sequences from the following viruses: retrovirus, such as moloney murine leukemia virus, harvey murine sarcoma virus, murine mammary tumor virus, and rous sarcoma virus; adenovirus, adeno-associated virus; SV40-type viruses; polyoma viruses; Epstein-Barr viruses; papilloma viruses; herpes virus; vaccinia virus; polio virus; and RNA virus such as a retrovirus. One can readily employ other vectors not named but known to the art. As used herein, the term "therapeutically effective amount" of the MPO inhibitor as above described is meant a sufficient amount to provide a therapeutic effect. It will be understood, however, that the total daily usage of the compounds and compositions of the present invention will be decided by the attending physician within the scope of sound medical judgment. The specific therapeutically effective dose level for any particular subject will depend upon a variety of factors including the disorder being treated and the severity of the disorder; activity of the specific compound employed; the specific composition employed, the age, body weight, general health, sex and diet of the subject; the time of administration, route of administration, and rate of excretion of the specific compound employed; the duration of the treatment; drugs used in combination or coincidental with the specific polypeptide employed; and like factors well known in the medical arts. For example, it is well within the skill of the art to start doses of the compound at levels lower than those required to achieve the desired therapeutic effect and to gradually increase the dosage until the desired effect is achieved. However, the daily dosage of the products may be varied over a wide range from 0.01 to 1,000 mg per adult per day. Typically, the compositions contain 0.01, 0.05, 0.1, 0.5, 1.0, 2.5, 5.0, 10.0, 15.0, 25.0, 50.0, 100, 250 and 500 mg of the active ingredient for the symptomatic adjustment of the dosage to the subject to be treated. A medicament typically contains from about 0.01 mg to about 500 mg of the active ingredient, preferably from 1 mg to about 100 mg of the active ingredient. An effective amount of the drug is ordinarily supplied at a dosage level from 0.0002 mg/kg to about 20 mg/kg of body weight per day, especially from about 0.001 mg/kg to 7 mg/kg of body weight per day.

Typically, the inhibitor of the present invention is combined with pharmaceutically acceptable excipients, and optionally sustained-release matrices, such as biodegradable polymers, to form pharmaceutical compositions. "Pharmaceutically" or "pharmaceutically acceptable" refer to molecular entities and compositions that do not produce an adverse, allergic or other untoward reaction when administered to a mammal, especially a human, as appropriate. A pharmaceutically acceptable carrier or excipient refers to a non-toxic solid, semi-solid or liquid filler, diluent, encapsulating material or formulation auxiliary of any type. Typically, the pharmaceutical compositions contain vehicles, which are pharmaceutically acceptable for a formulation capable of being injected. These may be in particular isotonic, sterile, saline solutions (monosodium or disodium phosphate, sodium, potassium, calcium or magnesium chloride and the like or mixtures of such salts), or dry, especially freeze-dried compositions which upon addition, depending on the case, of sterilized water or physiological saline, permit the constitution of injectable solutions. The pharmaceutical forms suitable for injectable use include sterile aqueous solutions or dispersions; formulations including sesame oil, peanut oil or aqueous propylene glycol; and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersions. It must be stable under the conditions of manufacture and storage and must be preserved against the contaminating action of microorganisms, such as bacteria and fungi. Sterile injectable solutions are prepared by incorporating the inhibitor at the required amount in the appropriate solvent with several of the other ingredients enumerated above, as required, followed by filtered sterilization. Generally, dispersions are prepared by incorporating the various sterilized active ingredients into a sterile vehicle which contains the basic dispersion medium and the required other ingredients from those enumerated above. In the case of sterile powders for the preparation of sterile injectable solutions, the preferred methods of preparation are vacuum-drying and freeze-drying techniques which yield a powder of the active ingredient plus any additional desired ingredient from a previously sterile-filtered solution thereof.

The invention will be further illustrated by the following figures and examples. However, these examples and figures should not be interpreted in any way as limiting the scope of the present invention.

FIGURES:

Figure 1: Increased neutrophil counts and MPO levels in mice lacking TRPV1⁺ neurons after HSV-1 infection. (A) Experimental procedure used to deplete TRPV1⁺ neurons in C57BL/6 mice with resiniferatoxin (RTX) treatment. (B) Representative image of dorsal root ganglia of PBS-treated mice (left panel) and RTX-treated mice (right panel) showing depletion of TRPV1⁺ neurons. (C) Experiment design used to monitor immune cells recruitment after HSV-l-TK' skin infection. (D) Absolute number of neutrophils in the skin at day 5 post-infection (p.i) and type 2 conventional dendritic cells (cDC2), Langerhans cells and virus-specific CD8⁺ T cells in the skin dLN at day 8 p.i. (n= 13-14 mice per group). (E, F) At day 5 p.i, the skin was collected, and proteins were extracted to quantify MPO concentration by ELISA. (n=5 mice per group). PBS corresponds to control mice injected with PBS (control vehicle) and RTX corresponds to RTX-treated mice injected with RTX. The data are presented as mean ± SEM and are representative of at least two independent experiments. P values (* p < 0.05, ** p < 0.01, **** p < 0.0001) were obtained using a Mann-Whitney test (two-tailed) for d and f. Figure 2: MPO mediates antiviral CD8⁺ T-cell responses by controlling neutrophil recruitment and myeloperoxidase levels in the skin. (A) Experimental protocol used to monitor immune responses in skin and skin dLN of infected PBS- or RTX-treated mice injected with 4-ABAH inhibitor or with the vehicle only. (B) After treatment, at day 5 p.i, the skin was collected, and proteins were extracted to quantify MPO concentration by ELISA. (n= 5 mice per group). (C) Absolute number of neutrophils in the skin at day 8 post-infection (p.i) with HSV-l-TK' of infected PBS- or RTX- mice treated id with 4-ABAH or the vehicle. (n= 10 mice per group). (D, E) Absolute number of indicated cells in the skin dLN at day 8 post-infection (p.i) with HSV-l-TK' of infected PBS- or RTX- mice treated id with 4-ABAH or the vehicle. (n=5 to 10 mice per group). PBS corresponds to control mice injected with PBS (control vehicle) and RTX corresponds to RTX-treated mice injected with RTX. The data are presented as mean ± SEM and are representative of at least two independent experiments. P values (* p < 0.05, ** p < 0.01, **** p < 0.0001) were obtained using a One-way ANOVA test followed by a Dunn’s multiple comparisons test.

EXAMPLE:

Methods:

Drug treatment

Resiniferatoxin (Coger, AG-CN2-054-MC05) (RTX)-treated mice were obtained by injecting subcutaneously into the flank 4-week-old C57BL/6 with three escalating doses (30 pg/kg, 70 pg/kg and 100 pg/kg) of RTX on consecutive days to allow specific ablation of TRPV1⁺ sensory neurons. Control littermates were injected with vehicle only (2% DMSO in IX PBS). Deletion of TRPV1⁺ sensory neurons was confirmed by immunofluorescence in DRG and flicktail assays. The tails of the mice were immersed in a water bath maintained at 52°C, and the latency to tail flick was recorded, with a maximum of approximately 10 seconds. For myeloperoxidase (MPO) inhibition, mice were injected intradermally (id) with 13 pg/g of 4- ABAH in 25 pl (Merck, 141909) from day 0 to day 4 post-infection with HSV-l-TK'.

Viral infection

Mice were inoculated with HSV-l-TK' and HSV-1-TK⁺ after flank scarification, as previously described¹⁷. Briefly, female mice of 6-12-week-old were anaesthetized by intraperitoneal injection (ip) of ketamine (2%)/Rompun (5%) solution in saline buffer (10 pl/g). The left flank of each mouse was clipped and depilated with Veet hair removal cream. A small area of skin near the top of the spleen was abraded with a MultiPro power tool (Dremel, Racine, WI) composed of a grindstone attachment (3.2 mm), held on the skin for 20 seconds to create a small abrasion. Then, a 10 pl of viral suspension (1.10⁶ plaque forming units (PFU)) was applied to the abraded skin. To contain the virus during the initial infection, a small piece of OpSite Flexigrid (Smith & Nephew, Hull, UK) was placed over the inoculation site. To prevent removal of the OpSite Flexigrid and disruption of the viral infection, the flank of the mouse was wrapped with Micropore tape followed by Transpore tape (3M Health Care, St. Paul, MN). The tape and Flexigrid were removed 48 hours after infection. To generate the UV-killed HSV- 1 model, an aliquot of HSV-OVA-TK⁺ virus suspension was irradiated with type-C ultraviolet (UV, X = 254 nm; voltage 8 W; source less than 5 cm from the target) for 30 minutes. All virus strains were grown on confluent monolayers of Vero cells (CSL) by PFU assay (see below) in a medium containing 10% FBS, 50 pM 2-mercaptoethanol, 2 mM L-glutamine, 100 U/ml penicillin and 100 pg/ml streptomycin and titrated by Plaque-forming unit (PFU) assay (see below).

Myeloperoxidase (MPO) quantification

Skin samples were homogenized using the Minute™ Total Protein Extraction Kit for Skin Tissue (Invent Biotechnologies) containing a protease inhibitor cocktail (Halt Protease Inhibitor 100X, Thermofisher) according to manufacturer’s instructions, and stored at -80°C until use. MPO quantification from skin samples was performed using the Myeloperoxidase Mouse ELISA kit (Thermofisher, 18064022) according manufacturer’s instructions.

Skin and draining lymph nodes (dLN) cell isolation

Skin-cell isolation. Skin samples (12 x 12-mm punch biopsy full-thickness pieces) were cut into small fragments and incubated in a collagenase/dispase/DNase solution (0.2 mg/ml collagenase type IV (Sigma), 0.2 mg/ml dispase (GIBCO) and 1 mg/ml DNase (Roche) in RPMI 1640 complete medium for 1 hour at 37°C with agitation. Then, the tissue was dissociated using a 2.5 ml syringes with 18G needles, and the resulting suspension was filtered on a 100pm cell strainer (Startedt). Cells were washed with FACS buffer (5 mM EDTA in IX PBS) to obtain a homogeneous cell suspension. dLN-cell isolation. Lymph nodes were crushed in 20 % FCS in IX PBS on a Falcon™ Round- Bottom polystyrene tube with a cell strainer snap cap (Falcon). Cells were washed with FACS buffer (5 mM EDTA in IX PBS) to obtain a homogeneous cell suspension. Quantification and statistical analysis

Normalization of absolute number by flow cytometry

For each mouse, a 12x12mm of skin punch biopsy was collected and a brachial draining lymph node (dLN). Each organ was digested to obtain a single cell suspension. An equal amount of Sphere™ Blank Calibration Particles (556296, BD Biosciences) was added to each sample to normalize them so that the same number of beads were acquired in each sample.

Statistical analysis

Statistical analyses were performed using GraphPad Prim software (version 9). Data is represented as mean ± standard error mean (SEM). When two groups are compared, Mann- Whitney t test was used. For multiples comparison analysis, normality and lognormality tests (Shapiro-Wilk test or Kolmogorov-Smirnov test) was used. If datas passed normality test, ordinary one-way ANOVA test followed by Dunnett’s, Sidak’s or Tukey’s multiple comparisons test was used. If datas not passed normality test, Kruskal-Wallis test followed by Dunn’s multiple comparisons test was used. Multiple comparisons of two parameters were analyze using a Two-way ANOVA followed by a Sidak’s multiple comparisons.

Results:

TRPV1⁺ neurons limit skin inflammation and induce a robust virus-specific CD8⁺ T-cell response to HSV-1

We then investigated the molecular and cellular mechanisms underlying the inflammatory phenotype observed in nociceptor-deficient mice¹⁶, by focusing on the role of TRPV1⁺ neurons, which were also activated by HSV-1 infection (data not shown). Most TRPV1 -expressing DRG neurons are peptidergic neurons. We investigated the function of TRPV1⁺ neurons by chemically ablating these neurons with resiniferatoxin (RTX), as previously described²⁰ (Figure 1A). The ablation of TRPV1⁺ neurons in RTX-treated mice was confirmed by immunofluorescence staining in DRGs (Figure IB) It resulted in a loss of heat sensation and white patches of hair, as expected (data not shown). Skin scarification without infection resulted in similar small lesions were in RTX-treated and control mice (data not shown). By contrast, HSV-l-TK' infection in mice lacking TRPV1⁺ neurons resulted in significantly more severe inflammatory lesions than in control mice (data not shown). The larger skin lesions observed in RTX-treated mice were associated with a greater influx of neutrophils into the skin at day 5 pi (Figure 1C and ID). Moreover, RTX-treated mice had a defect of DC subsets, with smaller numbers of cDC2 and Langerhans cells in the skin dLNs (Figure ID). This impaired DC response was associated with a defect of virus-specific CD8⁺ T-cell expansion in the skin dLNs of these mice at day 8 pi, as previously observed in mice lacking the entire population of Navi.8⁺ neurons (data not shown)¹⁶. TRPV1⁺ neurons are, therefore, required to limit neutrophil infiltration into the skin and to promote virus-specific CD8⁺ T-cell priming in cutaneous dLN after HSV-1 infection.

Neutrophil depletion and myeloperoxidase inhibition in the skin promotes antiviral CD8⁺ T-cell responses

We then investigated the mechanisms which promoted antiviral CD8⁺ T-cell responses in the skin dLNs of HSV-1 -infected mice. We previously showed that DCs isolated from the dLNs of infected mice lacking NaviU sensory neurons were less efficient at priming CD8⁺ T cells ex iv ⁶. Moreover, neutrophil depletion was sufficient to restore a robust virus-specific CD8⁺ T- cell response in mice lacking NaviU sensory neurons, suggesting that excessive neutrophil influx into the skin during HSV-1 infection has direct effects on DC and T-cell responses. We therefore hypothesized that mediators produced by neutrophils may be involved in modulating the adaptive immune response. Upon activation, neutrophils produce several mediators, including neutrophil elastase and MPO, often in association with neutrophil extracellular traps (NETs). We therefore investigated whether excessive MPO levels due to the high influx of neutrophils in the skin could underlie the modulation of the adaptive immune response to HSV- 1. MPO levels were higher in the skin of RTX- treated mice compared to PBS-treated mice after HSV-1 infection (Figure IE and IF) showing that in the absence of TRPVU sensory fibers, there is more MPO in the skin. RTX- or PBS-treated mice received id injections of the MPO inhibitor 4-aminobenzoic acid hydrazide (4-ABAH) from day 0 to day 4 pi (Figure 2A). This treatment effectively reduces MPO concentration in the skin of infected mice (Figure 2B) and had no effect on neutrophil infiltration, which remained excessive in the skin of RTX- treated mice (Figure 2C). However, MPO inhibition was sufficient to restore the number of DCs in the dLNs to normal levels and to enhance the virus-specific CD8⁺ T-cell response in the dLNs of RTX-treated mice at day 8 pi (Figures 2D and 2E).

Overall, these data support a model in which HSV-1 infection activates TRPVU sensory neurons, limiting the influx of neutrophils and, consequently, MPO levels in the skin of HSV- 1 -infected mice. Importantly, blockade of MPO activity in infected mice lacking TRPVU neurons is sufficient to restore a robust T cell response. This neuroimmune regulation plays a crucial role in limiting HSV-l-induced tissue damage and promotes efficient DC and CD8⁺ T- cell responses. Discussion:

The initiation and resolution of inflammatory processes are two essential sequential steps in host resistance to infectious diseases. Indeed, inflammatory processes play a key role in pathogen elimination, but must be tightly controlled to avoid excessive tissue damage once the pathogens have been eliminated. The sensory nervous system may have proinflammatory or anti-inflammatory properties, depending on the type of invading pathogen and the immune responses triggered^21,22. These different and sometimes opposing functions are probably related to the great heterogeneity of the sensory neurons that innervate tissues and the ways in which they are activated. The neuroimmune mechanisms involved in the control of viral infections are poorly understood.

We previously showed that Navi.C peripheral sensory neurons play an important role in regulating the immune response to HSV-1, and in particular the local recruitment of neutrophils in the skin¹⁶. Neutrophils release several mediators upon activation, including MPO, a local mediator of tissue damage. In the context of HSV-1 infection, we found that MPO inhibition restored the DC response and the priming of virus-specific CD8⁺ T cells in mice lacking TRPV1⁺ sensory neurons.

Overall, these data suggest that excessive neutrophil infiltration into the skin of nociceptordeficient mice increases the local production of MPO, with deleterious effects on the antiviral adaptive immune response.

Finally, this study opens up interesting therapeutic perspectives. It suggests that specific and selective inhibition of MPO may be of potential interest for increasing the efficacy of vaccines or antiviral treatments.

REFERENCES:

Throughout this application, various references describe the state of the art to which this invention pertains. The disclosures of these references are hereby incorporated by reference into the present disclosure.

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Claims

CLAIMS:

1. A method of promoting CD8⁺ T-cell responses in a subject in need thereof comprising administering to the subject a therapeutically effective amount of a myeloperoxidase inhibitor.

2. The method of claim 1, wherein the CD8⁺ T cell responses is promoted in the skin

3. The method of claim 1 or 2, for treating cancer in a subject in need thereof.

4. The method of claim 3, wherein the cancer is skin cancer.

5. The method of claim 1 or 2 for treating infection in a subject in need thereof.

6. The method of claim 5, wherein the infection is a skin infection

7. The method of claim 5 or 6, wherein the infection is induced by a bacterial pathogen, a parasite or a virus.

8. The method of claim 1 for treating a viral -induced skin lesion.

9. The method of claim 7 and 8, wherein the virus is a herpes virus.

10. The method of claim 1 for treating verruca virus-based diseases of the skin or herpesvirusbased diseases of the skin.

11. A method for enhancing the potency of a vaccine administered to a subject comprising administering to the subject a pharmaceutically effective amount of a myeloperoxidase inhibitor in combination with the vaccine.

12. A method of vaccinating a subject in need thereof comprising administering to the subject a therapeutically effective combination of a vaccine with a myeloperoxidase inhibitor, wherein administration of the combination results in enhanced vaccine efficacy relative to the administration of the vaccine alone.

13. The method of anyone claim 1 to 12, wherein the myeloperoxidase inhibitor is an inhibitor of myeloperoxidase expression or an inhibitor of myeloperoxidase activity.