Prostate cancer is the second leading cause of cancer death in men in Europe and the US. In the context of previous preclinical experiments and clinical studies there are certain assumptions predicating successful application of immunotherapy in the treatment of patients with prostate cancer. Promising results have been achieved by a combination of different treatment modalities which provide a synergistic antitumor effect. One of these combinatorial options is the use of antitumor vaccines and adoptive T cell transfer. The topic of this thesis is to provide a fresh insight into the past and current trends following the long-term candidate's department program in the field of anti-tumor immunotherapy. The experimental part of this thesis revolves around our own results published in this field. The introductory chapter delivers a basic overview of cellular mechanisms of anti-tumor immunity and the role of individual immune components in these processes. Following chapters are dedicated to current immunotherapeutic approaches with emphasis on the adoptive T cell transfer and implication of this technology in the treatment of prostate cancer. The results section describes the establishment of our protocol for adoptive T cell transfer as well as the protocol for ex vivo enrichment of human T cell...

The aim of the study was to evaluate the expression of immunosuppressive (FOXP3, NKG2A), and cytotoxic (perforin) or cytotoxic / immunomodulating (IFNγ) T-cell properties representing biomarkers in the peripheral blood CD8h T-cell population of patients with advanced renal cell carcinoma (RCC) or high risk cutaneous melanoma and healthy controls by multicolour flow cytometry. Determination of the percentage of functionally competing T-cell subsets (especially immunosuppressive) in the CD8hCD57+ T-cell subpopulation in future may serve as one of parameters enabling to assess the overall status of antitumor immune response and select cancer patients most suitable for antitumor immunotherapy while dismissing those to whom it would be ineffective or even harmful. / Darbo tikslas buvo įvertinti imunosupresines (FOXP3, NKG2A), citotoksines (perforin) bei citotoksines / imunomoduliuojančias (IFNγ) savybes atspindinčių žymenų raiškos skirtumus išplitusiu inkstų vėžiu ar didelės rizikos odos melanoma sergančių pacientų periferinio kraujo CD8h T limfocitų populiacijoje, lyginant su kontroline grupe. Skirtingas T limfocitų savybes atspindinčių žymenų raiška buvo tiriama tėkmės citometrijos būdu. Nustatyta, kad inkstų vėžiu ar odos melanoma sergančių pacientų periferiniame kraujyje Įvairių subpopuliacijų (ypač imunosupresinės) nuošimčio nustatymas CD8hCD57+ T limfocitų populiacijoje ateityje gali būti naudingas klinikinėje praktikoje, individualizuojant priešnavikinę imunoterapiją ir selektyviai parenkant tik tuos pacientus, kuriems imuninės sistemos aktyvinimas sukeltų navikinių ląstelių naikinimą, o ne dar labiau gilintų imunosupresiją.

Darbo tikslas buvo įvertinti imunosupresines (FOXP3, NKG2A), citotoksines (perforin) bei citotoksines / imunomoduliuojančias (IFNγ) savybes atspindinčių žymenų raiškos skirtumus išplitusiu inkstų vėžiu ar didelės rizikos odos melanoma sergančių pacientų periferinio kraujo CD8h T limfocitų populiacijoje, lyginant su kontroline grupe. Skirtingas T limfocitų savybes atspindinčių žymenų raiška buvo tiriama tėkmės citometrijos būdu. Nustatyta, kad inkstų vėžiu ar odos melanoma sergančių pacientų periferiniame kraujyje Įvairių subpopuliacijų (ypač imunosupresinės) nuošimčio nustatymas CD8hCD57+ T limfocitų populiacijoje ateityje gali būti naudingas klinikinėje praktikoje, individualizuojant priešnavikinę imunoterapiją ir selektyviai parenkant tik tuos pacientus, kuriems imuninės sistemos aktyvinimas sukeltų navikinių ląstelių naikinimą, o ne dar labiau gilintų imunosupresiją. / The aim of the study was to evaluate the expression of immunosuppressive (FOXP3, NKG2A), and cytotoxic (perforin) or cytotoxic / immunomodulating (IFNγ) T-cell properties representing biomarkers in the peripheral blood CD8h T-cell population of patients with advanced renal cell carcinoma (RCC) or high risk cutaneous melanoma and healthy controls by multicolour flow cytometry. Determination of the percentage of functionally competing T-cell subsets (especially immunosuppressive) in the CD8hCD57+ T-cell subpopulation in future may serve as one of parameters enabling to assess the overall status of antitumor immune response and select cancer patients most suitable for antitumor immunotherapy while dismissing those to whom it would be ineffective or even harmful.

L’immunothérapie basée sur le transfert de lymphocytes T (LT) spécifiques de la tumeur est une approche prometteuse contre le cancer. Pour activer et amplifier de tels LT, principale étape limitante de cette approche, des cellules présentatrices d’antigène artificielles (CPAA) ont été développées au laboratoire. Ces CPAA ont été construites à partir de fibroblastes murins NIH/3T3 transduits à l’aide de vecteurs gammarétroviraux afin d’exprimer les principaux éléments nécessaires à l’activation de LT humains. Ces CPAA nous permettent d’obtenir des LT mémoires souches (TSCM : CD95+CD45RA+CD62L+CCR7+), LT très peu différenciés récemment identifiés chez l’homme. Ces TSCM ont été décrits comme étant du plus grand intérêt pour l’immunothérapie en raison de leur capacité d’auto-renouvellement et de leur faculté à se différencier en LT effecteursefficaces. Pour optimiser l’amplification de TSCM spécifiques, nous avons notamment étudié les effets sur les LT de l’expression de différentes molécules de costimulation par nos CPAA (CD80, CD70 et 4-1BBL). Les protéines MART-1 et MELOE-1, surexprimées dans les mélanomes, ont été utilisées comme antigènes modèles pour ces travaux. Les CPAA CD80+CD70+ et CD80+CD70+4-1BBL+ sont les plus prometteuses pour maintenir le phénotype des TSCM. Une étude exhaustive des CPAA CD80+CD70+ a montré que nous pouvions obtenir un plus grand nombre de TSCM fonctionnels spécifiques de MART-1 et de MELOE-1 de manière reproductible avec ces CPAA. Dans une seconde étude, nous avons pu montrer que les CPAA CD80+CD70+4-1BBL+ permettaient d’obtenir le plus grand nombre de LT spécifiques fonctionnels et très peu différenciés après purification et restimulation de LT spécifiques stimulés une première fois par les CPAA CD80+CD70+. Ces travaux devraient nous permettre, après le développement d’un modèle murin, de proposer de nouvelles stratégies d’immunothérapie basées sur l’obtention grâce à nos CPAA optimisées de LT spécifiques anti-tumoraux capables d’assurer une protection à long terme aux patients. / Immunotherapy based on the transfer of tumor-specific T lymphocytes (TLs) is a promising approach against cancer. To activate and amplify such TLs, main limiting step of this approach, artificial antigen presenting cells (AAPCs) have been developed in the laboratory. These AAPCs have been constructed from NIH/3T3 murine fibroblasts transduced with gammaretroviral vectors to express the principal elements required to activate human TLs. With these AAPCs, we can obtain anti-tumor stem cell memory TLs (TSCM: CD95+CD45RA+CD62L+CCR7+), which are very limitedly differentiated TLs recently identified in humans. These TLs have been recently described as cells of great interest for immunotherapy because of their self-renewal capacity and their ability to differentiate into effective effector TLs. To improve the amplification of specific TSCM, we notably studied the effects on TLs of the expression of different co-stimulatory molecules by our AAPCs (CD80, CD70 and 4-1BBL). MART-1 and MELOE-1, proteins that are overexpressed in melanoma, were used as model antigens in this work. CD80+CD70+ and CD80+CD70+4-1BBL+ AAPCs appear to be the most promising ones for maintaining a TSCM phenotype. An exhaustive study of CD80+CD70+ AAPCs showed that we could reproducibly get greater numbers of MART-1- and MELOE-1-specific functional TSCM with these AAPCs. In another study, we have shown that CD80+CD70+4-1BBL+ AAPCs enabled us to get the greatest number of functional and very limitedly differentiated specific TLs after purification and restimulation of specific TLs stimulated first with CD80+CD70+ AAPCs. This work should allow us, after the development of a murine model, to propose new immunotherapy strategies based on the possibility of obtaining with our optimized AAPCs anti-tumor specific TLs capable of ensuring patient long term protection.