This thesis focuses on using liposomes in two different treatment strategies; vaccination (or immunotherapy) and delivery of a small molecule, and in two different disease models; cancer and... Show moreThis thesis focuses on using liposomes in two different treatment strategies; vaccination (or immunotherapy) and delivery of a small molecule, and in two different disease models; cancer and atherosclerosis. For each of these treatment strategies, the liposomal formulation was tailored to obtain the desired therapeutic effect. Chapter 2 reviews some of the most important physicochemical properties (size, shape, and rigidity) that determine the immunological effects of liposomes and other nanoparticles. In chapter 3 we present a detailed study on the effect of the rigidity of anionic liposomes, as measured by atomic force microscopy, on antigen-specific regulatory T-cell (Treg) responses. In chapter 4, we show that our optimized anionic liposomes can induce potent antigen-specific Treg responses, and can be used to delay atherosclerosis progression in a mouse model. Chapter 5 also focuses on liposomal treatment of atherosclerosis, but here targeted liposomes were prepared to successfully deliver a small molecule to foam cells in atherosclerotic plaques. In Chapter 6, we used cationic liposomes in combination with an adjuvant for cancer immunotherapy in mice. Finally, we summarize the overall findings in chapter 7 and discuss perspectives of using liposomes for vaccination and targeted drug delivery. Show less
Clinical responses to immunotherapy can be complete and sustained; however, compared to responses to conventional cancer therapies such as chemotherapy they are often delayed. It has become clear... Show more Clinical responses to immunotherapy can be complete and sustained; however, compared to responses to conventional cancer therapies such as chemotherapy they are often delayed. It has become clear that when different cancer therapies are combined they can potentially synergize. Understanding how individual cancer therapeutics work enables the development of the most optimal combinations of cancer therapies. In this thesis, we have used interesting combinations of chemotherapy, cancer vaccines and immune checkpoint blockade and obtained insights in the mechanisms involved in these combinations. For example, we combined chemotherapy with vaccination and observed that none of the tested chemotherapeutics had a negative effect on vaccination, while only a few displayed synergy with vaccination in tumor eradication. We found that vaccine-induced T cells produce Tumor Necrosis Factor-α which could sensitize tumor cells for chemotherapy-induced cell death. Furthermore, we have shown that different cancer therapies modulate intratumoral myeloid cells in a unique manner which can critically affect the therapeutic efficacy of the therapy. Understanding how different therapies affect the local tumor environment will help to improve clinical responses and decrease toxicity in patients. Show less
Preface: The content of this thesis is based on research that was conducted at the travel and vaccination clinic at Leiden University Medical Centre (LUMC). This clinic provides pre-travel care to... Show morePreface: The content of this thesis is based on research that was conducted at the travel and vaccination clinic at Leiden University Medical Centre (LUMC). This clinic provides pre-travel care to the general population, and to special groups of travellers, such as patients who use immunosuppressants or who have chronic diseases. The clinic is closely connected to the department of Infectious Diseases at LUMC. The setting of a travel clinic within an academic medical hospital, provides unique circumstances for medical research, like an experienced team of nurses, expertise regarding immunization, a constant flux of travellers and the knowledge and infrastructure that is required for research into microbiology, virology and parasitology. Examples of research that stem from this clinic are projects on immunization against malaria, yellow fever, travellers' diarrhea, poliomyelitis and hepatitis B, vaccination of immunocompromised patients, and projects on travel related acquisition of extended spectrum ß-lactamase producing Enterobacteriacae and on the utility of post-travel screening of asymptomatic travellers for parasites. Show less
The subject of this thesis is the study of the function and behavior of CD8+ T cells that have exited the circulation and entered a tissue in order to get rid of an infection. In particular, the... Show moreThe subject of this thesis is the study of the function and behavior of CD8+ T cells that have exited the circulation and entered a tissue in order to get rid of an infection. In particular, the research focus on 1- how effector CD8+ T cells can localize infected cells within a tissue. 2-how, once the infection has been cleared, effector CD8+ T cells become memory T cells that take up permanent residence in the tissue; also, how the patrolling behavior that these cells acquire in the tissue allows quick control of local secondary infections. 3-how tissue-resident memory CD8+ T cells that have re-encountered a specific antigen can induce an increased level of local immune protection against infections. Show less
This dissertation reports on the relation between the immune system, colorectal cancer and immunotherapy. In the first part, expression of HLA class I and expression of CXCL5 in colocectal cancer... Show moreThis dissertation reports on the relation between the immune system, colorectal cancer and immunotherapy. In the first part, expression of HLA class I and expression of CXCL5 in colocectal cancer was studied. Low expression of HLA class I in rectal tumors was associated with poor survival of rectal cancer patients. Low expression of CXCL5 in cancer cells was significantly associated with poor prognosis in a population of colorectal cancer patients and correlated with presence of intra-tumoral CD8+ T-cell infiltration. In the second part of this thesis we focused on induction of tumor specific T-cells. For immunotherapeutic purposes distinction should be made between microsatellite instable (MSI-H) and microsatellite stable (MSS) colorectal tumors, as MSI-H tumors express neo-antigens __foreign__ to the immune system while immunotherapy against MSS tumors depends on tumor associated __self__-antigens. We developed a methodology predicting immunogenic behavior of frameshift-mutated antigens present in MSI-H tumors that was based on accumulation and MHC class I presentation. This method can be used to develop cancer immunotherapy of patients at risk for MSI-H tumors. In the last two chapters we described safety and immunogenicity of a p53 synthetic long peptides vaccine combined with and without Interferon-alpha. Addition of IFN-_ to the p53-SLP_ vaccine significantly improved p53-specific after vaccination. Altogether this dissertation reports on the relation between the immune system, colorectal cancer and immunotherapy. This knowledge can be used to further optimize immunotherapeutic strategies to treat cancer patients. Show less
Microneedle-based transcutaneous immunisation is an appealing alternative to the classical manner of injecting vaccines by intramuscular or subcutaneous route. Importantly, as a consequence of the... Show moreMicroneedle-based transcutaneous immunisation is an appealing alternative to the classical manner of injecting vaccines by intramuscular or subcutaneous route. Importantly, as a consequence of the fact that the skin is in direct contact with the environment and should protect the body against pathogens, it contains more antigen presenting cells, such as dendritic cells than the muscles or subcutaneous tissue and thereby offers the possibility to induce a more effective immune response. The combination of microneedles and adjuvanted subunit vaccines may offer effective vaccination whereas ensuring patient safety and vaccine application in a painless manner. The principal aim of this thesis was to design subunit vaccine formulations that can be combined with microneedles for transcutaneous immunisation. The approaches described in this thesis have generated new insights into the main requirements for transcutaneous immunisation. Microneedles definitively have the potential to be an excellent utensil for the delivery of vaccines into the skin. However, the skin is a very elastic organ and the actual conduits formed by microneedle pre-treatment will be considerably smaller than the diameter of the microneedles. Therefore, a small antigen-adjuvant entity is the preferred formulation, as it will be transported efficiently through the microneedle conduits while it retains the co-delivery of antigen and adjuvant. Show less
The research described in this thesis investigates the effect of viral infection on the risk of rejection. It was previously thought that a viral specific T-cell receptor could not crossreact... Show moreThe research described in this thesis investigates the effect of viral infection on the risk of rejection. It was previously thought that a viral specific T-cell receptor could not crossreact against allogeneic HLA molecules, however here it is proven that this is not only possible but is also common. The research performed in this thesis has important clinical implications for solid organ (kidney) and bone marrow transplantation. Viral infection commonly generates alloreactive T-cells, and may therefore be responsible for the failure of tolerance inducing regimens in primates and humans. It is shown that vaccination can also induce HLA-specific alloreactive T-cell responses and it is therefore recommended that vaccines be given to transplantation candidates at least three months prior to expected transplantation date. It is also shown that alloreactive T-cells can be tissue specific. In the final chapter of the thesis thesis it is shown that allogeneic cell therapy could conversely be useful to elicit a viral specific T-cell response in patients with infections, such as HIV. Show less
In conclusion, this thesis describes different clinical aspects on the use of anti-TNF. Initial combination treatment with infliximab and methotrexate seems very promising in early RA and may even... Show moreIn conclusion, this thesis describes different clinical aspects on the use of anti-TNF. Initial combination treatment with infliximab and methotrexate seems very promising in early RA and may even alter the course of the disease. In patients with intolerance or inadequate response to infliximab, a second TNF-blocking agent can be successful, although, the introduction of new biologicals such as rituximab and abatacept provides other therapeutic options. Further research is needed to answer the question which drugs are indicated after failing a first TNF-blocking agent, or even as initial therapy. The use of intra-articular infliximab is inferior to intra-articular depomedrol in chronic and/or recurrent gonartrtitis and this expensive treatment should not have benefit of the doubt in these patients. In contrast, the use of infliximab in rheumatoid vasculitis seems promising in patients who have contra-indication for the recommended immunosupression. Influenza- and pneumococcal vaccinations result in less and lower antibody titers during treatment with anti-TNF, however, antibodies were formed and it remains unclear whether the lower laboratory outcomes are clinically relevant. To date, most guidelines recommend vaccination in patients using immunosuppressive therapy. During the use of anti-TNF neurological symptoms comparable to those of multiple sclerosis (MS) have been described and MR changes have been detected. The use of anti-TNF should be considered as contra-indicated in patients with signs and symptoms compatible with MS. Show less
In an epoch where every generation travels more frequently and at longer distances than the previous generation, with a mean increase of 30 million travellers per year from 1995 until today,... Show moreIn an epoch where every generation travels more frequently and at longer distances than the previous generation, with a mean increase of 30 million travellers per year from 1995 until today, physicians throughout the world are confronted with new diseases. In absolute numbers, this implies that each year, roughly 4 million travellers appeal to specialised health care, either abroad or at home, because of systemic febrile illness, diarrhoea or dermatologic disorders. During the last decades, travel medicine has evolved into a distinct discipline of Infectious Diseases, eventhough transmission of infectious agents into vulnerable populations through travel has been well know for centuries. Improvement of of protection against travel-related diseases can be achieved through knowledge on the following topics; 1. Epidemiology of travel-related diseases, 2. Morbidity and mortality of these illnesses in specific groups of travellers, 3. Adherence to travel health precautions, 4. Responsivity against vaccination, and 5. Availability of preventive measures, such as vaccines. The research described in this thesis addresses these various topics. Show less
Atherosclerosis, the predominantly underlying pathology of cardiovascular events, is the consequence of lipid deposition in the arterial wall, mostly as consequence of high levels of serum... Show moreAtherosclerosis, the predominantly underlying pathology of cardiovascular events, is the consequence of lipid deposition in the arterial wall, mostly as consequence of high levels of serum cholesterol. Treatment of atherosclerosis is mainly focused at the reduction of cholesterol levels by lipid lowering medication, such as statins. Despite the use of statins and prophylactic treatments, such as a reduction in blood pressure and a reduction in risk factors to prevent atherosclerosis, cardiovascular disease is still the major cause of death in the Western world.1-3 As the struggle against atherosclerosis continues and its prevalence is increasing in the world, it is pivotal to find new targets for implementing new strategies against atherosclerosis. Inflammation is considered to be a major component in the process of atherosclerosis and is involved in initiation, progression and destabilization of the atherosclerotic lesion. Although the beneficial effects of statins on atherosclerosis may partly be ascribed to their anti-inflammatory properties, relatively little is known about the exact mechanism and contribution of different inflammatory cells and products in atherosclerosis. The aim of this thesis was to further elucidate the contribution of various components of the inflammatory response in atherosclerosis and thereby finding new intervention points to reduce the incidence and the consequences of atherosclerosis. Show less
Worldwide, more than a billion people are infected with helminths. These worm infections are chronic in nature and can lead to considerable morbidity. Immunologically these infections are... Show moreWorldwide, more than a billion people are infected with helminths. These worm infections are chronic in nature and can lead to considerable morbidity. Immunologically these infections are interesting; chronic helminth infections are characterized by skewing towards a T helper 2 type response as well as regulatory responses. The latter is thought to prevent strong immune responses against parasitic worms, allowing their long-term survival and restricting pathology. This regulatory network is thought to also temper responses to non-helminth antigens, like allergens or self antigens, possibly leading to lower prevalence of allergies and autoimmune diseases in subjects that are chronically infected with helminths. Thus, helminths may bear molecules that have potential therapeutic action against allergies and possibly other inflammatory diseases. However, on the other side of the coin, this would predict that helminth infected subjects might not respond strongly to third party antigens like vaccines. Helminth derived molecules have been identified that induce T helper 2 and regulatory responses via modulation of dendritic cells and some appear to do so via TLR signaling. New insights into these pathways could be useful to antagonize suppression and hence boost vaccine efficacy or to optimize suppression induced by helminth derived molecules and control inflammatory diseases. Show less
This thesis describes clinical and immunological aspects of immunoablative therapy followed by reinfusion of T-cell depleted autologous stem cells in patients with progressive refractory Juvenile... Show moreThis thesis describes clinical and immunological aspects of immunoablative therapy followed by reinfusion of T-cell depleted autologous stem cells in patients with progressive refractory Juvenile Idiopathic Arthritis (JIA). After an intensive immunoablative therapy in order to eradicate auto-agressive T cells, autologous hematopoietic stem cells of bone marrow, purged of potentially autoreactive mature T lymphocytes, were reinfused as rescue to reduce the aplastic phase after autologous stem cell transplantation (ASCT). Chapter 1 addresses the background and rationale leading to this study. Chapter 2 describes the safety, efficacy, complications and long term clinical outcome after ASCT in 22 patients with JIA, who were refractory to conventional medication. In chapter 3 the efficacy and safety of ASCT in 34 JIA patients transplanted in 9 different centers in Europe were evaluated. In chapter 4 the immunological effects of the conditioning on cellular infiltrates and expression of cytokines in the synovial tissue of two JIA patients were studied before and 6 months after ASCT. The subject of chapter 5 of this thesis is macrophage activation syndrome in systemic JIA patients. The actual effect of conditioning in vivo and graft manipulation ex vivo on the intended elimination of the adaptive (auto)immunological memory after ASCT was studied in 19 JIA/systemic lupus erythematodes (SLE) and 10 multiple sclerosis (MS) patients (chapter 7). In order to obtain reference values for anti-rabies specific humoral and cellular immune responses after a single and booster vaccination, we conducted a study in 18 healthy controls as described in chapter 6. Show less
This thesis introduces a novel T cell vaccination method that uses a tattoo machine to inject DNA in the skin of the vaccinee. In comparison to other experimental vaccination methods DNA tattooing... Show moreThis thesis introduces a novel T cell vaccination method that uses a tattoo machine to inject DNA in the skin of the vaccinee. In comparison to other experimental vaccination methods DNA tattooing is very strong: besides small laboratory animals also large animals mount strong T cell responses upon tattoo DNA vaccination. Show less
T cells recognize pathogen-derived antigens and are crucial for fighting pathogens such as viruses and bacteria. In addition, T cells are able to recognize and attack certain types of tumors, in... Show moreT cells recognize pathogen-derived antigens and are crucial for fighting pathogens such as viruses and bacteria. In addition, T cells are able to recognize and attack certain types of tumors, in particular virally induced tumors. In this thesis we aimed 1) to obtain more insight into antigen-specific T cell responses and 2) to study how antigen-specific T cell responses can be improved. For the first aim we generated new tools that by enabling the visualization of antigen-specific CD4+ and CD8+ T cells allow the study of the dynamics of antigen-specific T cell responses in time throughout an ongoing immune response (chapter 2). In addition, we developed a novel technique that enables the study of family relationships between different T cell populations. This technique for instance allows us to determine whether two different types of effector T cell populations arise from the same or different pool(s) of na_ve T cells (chapter 5). For the second aim, we analyzed whether antigen-specific T cell responses can be manipulated by providing increased costimulation in the form of constitutive triggering of CD27 (chapter 3) or by generating CD4+ T cells that are modified by the introduction of MHC class I restricted TCRs (chapter 4). Show less
Aderverkalking (atherosclerose) is een ziekte waarbij door verdikking van de vaatwand vernauwing van slagaderen optreedt. Door deze vernauwing is het mogelijk dat er te weinig zuurstofrijk bloed de... Show moreAderverkalking (atherosclerose) is een ziekte waarbij door verdikking van de vaatwand vernauwing van slagaderen optreedt. Door deze vernauwing is het mogelijk dat er te weinig zuurstofrijk bloed de organen bereikt. In het geval van atherosclerose in slagaderen die hart of hersen van bloed moeten verzien leidt de vernauwing niet zelden tot de dood. Het feit dat atherosclerose de belangrijkste onderliggende oorzaak is van wereldwijde sterfte onderstreept het maatschappelijk belang van ontwikkeling van een adequate therapie tegen atherosclerose. In dit proefschrift werden nieuwe vaccinatietechnieken aangewend om het proces van atherosclerose tegen te gaan. Door de tolerantie voor lichaamseigen stoffen of cellen te doorbreken met deze vaccinatietechnieken bleek het mogelijk te zijn atherosclerose in muizen sterk af te remmen. Het wegnemen van de functie van ontstekingsbevorderende eiwitten (interleukinen 12 en 17) met deze vaccinatietechieken leidde tot een 70-90% afname in atherosclerose. Ook het met vaccinatie specifiek opruimen van cellen die betrokkken zijn bij atherosclerose-geassocieerde processen, zoals nieuwvaatvorming, bleek een geschikte methode te zijn om atherosclerose tegen te gaan. Het voordeel van de nieuwe vaccinatietechnieken die bij dit onderzoek zijn gebruikt is gelegen in het feit dat met enkele injecties voor lange tijd (ten minste 24 weken) bescherming tegen atherosclerose bewerkstelligd kan worden. Andere voordelen van deze vaccinaties zijn het gebrek aan afweerreacties tegen lichaamsvreemde medicijnen en de lage productiekosten van dergelijke vaccins. De combinatie van de sterke afname in atherosclerose en de bijkomende voordelen van de vaccinatietechnieken leidde in dit proefschrift tot de conclusie dat vaccinatie een belangrijke bijdrage kan leveren aan de ontwikkeling van therapieën tegen hart- en vaatziekten in mensen. Show less
