Therapeutic vaccines differ from prophylactic vaccines in their ability to induce a strong and specific cytotoxic CD8+ T cell (CTL) response rather than an antibody-mediated reaction. CTLs are activated by presenting T cell receptors with short peptides associated with major histocompatibility complex (MHC) class I molecules. These peptide–MHC class I complexes are present on the surface of antigen-presenting cells (APCs), which can also provide the co-stimulatory signals required for optimal CTL activation. Dendritic cells (DCs) are the most potent APCs capable of initiating a primary immune response, and when activated and mature, they serve as effective stimulators of T cell activation.
Rationale for the mechanistic action of EDA
Toll-Like Receptors (TLRs) play an important role in DC activation, allowing for the recognition of molecules that are broadly shared by pathogens. In the case of TLR-4, activation can also be mediated by a variant of fibronectin (FN), a large endogenous extracellular matrix glycoprotein. The Extra Domain A (EDA) of FN is produced by alternative splicing in response to tissue injury. Although its biological functions are not well understood, after binding to TLR-4, EDA induces the expression of pro-inflammatory cytokines. Moreover, it was recently shown that EDA activates DCs through TLR-4, suggesting that EDA might recruit DCs to sites of injury and trigger their maturation.
These findings led the group of Dr. Juan José Lasarte at CIMA to investigate vaccines based on EDA–antigen fusion proteins as a strategy to target viral or tumour antigens to DCs in vivo. Fusion of EDA to disease-specific antigens not only can ensure efficient antigen targeting to DCs, but might also simultaneously participate in the upregulation of co-stimulatory signals as well as DC maturation and migration to draining lymph nodes. In this regard, vaccines based on the EDA platform have been shown to enhance antiviral or antitumoural T cell responses when tested in experimental models.
Formune is currently conducting the pre-clinical development of its first EDA-based vaccine candidate EDA-HPVE7 for the treatment of Human Papilloma Virus (HPV)-associated malignancies, such as cervical cancer.
Cervical carcinoma is the second most common cancer in women worldwide, with an estimated 0.5 million new cases and 275,000 deaths occurring annually. The development of cervical cancer is strongly associated with chronic infection of the genital tract with high-risk Human Papilloma Virus (HPV), most commonly HPV16 and HPV18, which are linked to approximately 70% of all cervical cancer cases.
Although most HPV infections resolve spontaneously, in a small percentage of cases, high-risk HPV genotypes can exert alterations in cell cycle control of keratinocytes. These modifications can be evidenced morphologically through cytological changes called low-grade squamous intraepithelial lesions (LSILs) (also known as CIN1). Approximately 20% of CIN1 will progress to CIN2, with 30% of these lesions ultimately becoming cervical carcinoma.
High-risk HPVs have been associated not only with ano-genital carcinomas (mainly cancers of the uterine cervix), but also with other cancers (e.g., oropharyngeal carcinomas and cutaneous malignancies [non-melanoma skin cancers]).
Current cervical cancer treatment involves surgery, radiotherapy and/or chemotherapy. Although surgery can be used to remove the uterus, cervix or other nearby affected organs, this approach is generally only useful for early and medium stage cancer. On the other hand, radiation therapy is mainly utilized to treat medium or late stage tumours, whereas chemotherapy employs a variety of drugs to eradicate cancer (e.g., carboplatin, cisplatin, paclitaxel, fluororacil and cyclophosphamide) and is usually administered concurrently with radiotherapy.
Recently developed prophylactic cervical cancer HPV vaccines, Gardasil (Merck) and Cervarix (GlaxoSmithKline), have been shown to prevent infections with strains HPV16 and HPV18. While these vaccines were proven to be efficacious for the prevention of disease when given to girls at early ages (11–12 years), they have no effect on established infections. Thus, no therapeutic vaccines are currently available, and since HPV-associated cervical cancers may arise decades after the initial infection, such vaccines are urgently required.
Two viral proteins, E6 and E7, which are consistently expressed in cervical tumours, are required for both the induction and maintenance of the transformed phenotype. Moreover, since cell immunity to E7 is associated with clinical and cytological resolution of HPV-induced lesions, E7 protein constitutes an important immunological target for tumour regression therapy. Thus, development of a therapeutic HPVE7 vaccine is expected to significantly improve the treatment of precancerous lesions and cervical cancer.
During the last years, the group of Dr. Juan José Lasarte at CIMA, alone or in collaboration with other research groups, such as the Institute Pasteur or “Instituto de Agrobiotecnología” (CSIC-UPNA-Gobierno de Navarra), has published several scientific publications in internationally recognized scientific journals demonstrating the efficacy of the EDA vaccine platform.
1. "The extra domain A from fibronectin targets antigens to toll like receptor expressing cells and induces cytotoxic T cell responses in vivo". Juan J. Lasarte, Noelia Casares, Marta Gorraiz, Sandra Hervás-Stubbs, Laura Arribillaga, Cristina Mansilla, Maika Durantez, Diana Llopiz, Pablo Sarobe, Francisco Borrás-Cuesta, Jesús Prieto and Claude Leclerc. The Journal of Immunology 2007, 178:748-756.
2. “Immunization against Hepatitis C Virus with a fusion protein containing the extra domain A from fibronectin and the Hepatitis C Virus NS3 protein”. Cristina Mansilla, Marta Gorraiz, Marta Martinez, Noelia Casares, Laura Arribillaga, Francesc Rudilla, Iciar Echeverria, José Ignacio Riezu-Boj, Pablo Sarobe, Francisco Borrás-Cuesta, Jesús Prieto and Juan José Lasarte. Journal of Hepatology-2009-00486.R1.
3. “The vaccine adjuvant extra domain A from fibronectin retains its proinflammatory properties when expressed in tobacco chloroplasts”. Inmaculada Farran, Iva McCarthy-Suárez, Francisco Río-Manterola, Cristina Mansilla, Juan José Lasarte, Ángel M. Mingo-Castel. Planta. 2010 Mar;231(4):977-90.
4. “Adjuvant combination and antigen targeting as a strategy to induce polyfunctional and high-avidity T-cell responses against poorly immunogenic tumors”. Aranda F, Llopiz D, Díaz-Valdés N, Riezu-Boj JI, Bezunartea J, Ruiz M, Martínez M, Durantez M, Mansilla C, Prieto J, Lasarte JJ, Borrás-Cuesta F, Sarobe P. Cancer Res 2011;71:3214-3224. Published OnlineFirst
5. “Eradication of large tumors expressing human papillomavirus E7 protein by therapeutic vaccination with E7 fused to the extra domain A from fibronectin”. Mansilla C, Berraondo P, Durantez M, Martínez M, Casares N, Arribillaga L, Rudilla F, Fioravanti J, Lozano T, Villanueva L, Sarobe P, Borrás F, Leclerc C, Prieto J, Lasarte JJ. Int J Cancer. 2011 Sep 2. doi: 10.1002/ijc.26412.
6. “The extradomain A of fibronectin (EDA) combined with poly(I:C) enhances the immune response to HIV-1 p24 protein and the protection against recombinant Listeria monocytogenes-Gag infection in the mouse model”. San Román B, De Andrés X, Muñoz PM, Obregón P, Asensio AC, Garrido V, Mansilla C, Arribillaga L, Lasarte JJ, De Andrés D, Amorena B, Grilló MJ. Vaccine. 2012 Mar 28;30(15):2564-9.
7. “The extradomain A of fibronectin enhances the efficacy of lipopolysaccharide defective Salmonella bacterins as vaccines in mice”. Román BS, Garrido V, Muñoz PM, Arribillaga L, García B, De Andrés X, Zabaleta V, Mansilla C, Farrán I, Lasa I, De Andrés D, Amorena B, Lasarte JJ, Grilló MJ. Vet Res. 2012 Apr 19;43:31.
8. “A fusion protein between streptavidin and the endogenous TLR4 ligand EDA targets biotinylated antigens to dendritic cells and induces T cell responses in vivo”. Arribillaga L, Durantez M, Lozano T, Rudilla F, Rehberger F, Casares N, Villanueva L, Martinez M, Gorraiz M, Borrás-Cuesta F, Sarobe P, Prieto J, Lasarte JJ. Biomed Res Int. 2013;2013:864720
Project 1: Development of a therapeutic vaccine against cervical cancer based on a fusion protein between Fibronectin EDA and HPV E7 antigen.
“El consorcio liderado por Formune y compuesto por la Fundación para la Investigación Médica Aplicada (FIMA), el Centro Nacional de Biotecnología (CNB) y la Universidad de Navarra (UNAV) ejecuta el proyecto DESARROLLO DE UNA VACUNA TERAPÉUTICA FRENTE AL CÁNCER DE CÉRVIX BASADA EN LA PROTEÍNA DE FUSIÓN ENTRE EL DOMINIO EXTRA A DE LA FIBRONECTINA Y EL ANTÍGENO E7 DEL HPV financiado por el subprograma Retos Colaboración, enmarcado en el Programa Estatal de Investigación, Desarrollo e Innovación Orientada a los Retos de la Sociedad, en el marco del Plan Estatal de Investigación Científica y Técnica y de Innovación 2013-2016, con número de expediente RTC-2014-1615-1”.
Formune has been granted with two “RETOS DE COLABORACIÓN” projects financed by the Ministry of Economy and Competitiveness of the Spanish Government for 2014-2016
Project 2: Validation of Annexin 2 as a predictor of recurrent endometrial cancer and design of new therapeutic strategies.
“El consorcio liderado por Althia y compuesto por Formune, Institut de Recerca de Vall d’Hebron e IRBLleida ejecuta el proyecto VALIDACIÓN DE ANEXINA 2 COMO PREDICTOR DE RECURRENCIA EN EL CÁNCER DE ENDOMETRIO. DISEÑO DE NUEVAS ESTRATEGIAS TERAPÉUTICAS (ANXARECU), financiado por el subprograma Retos Colaboración, enmarcado en el Programa Estatal de Investigación, Desarrollo e Innovación Orientada a los Retos de la Sociedad, en el marco del Plan Estatal de Investigación Científica y Técnica y de Innovación 2013-2016, con número de expediente RTC-2014-3110-1”.
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