While immune checkpoint blockade (ICB) (CTLA4/ PD1) therapy has resulted in durable responses in patients with advanced stage cutaneous melanoma, some patients do not benefit from this, due to... Show moreWhile immune checkpoint blockade (ICB) (CTLA4/ PD1) therapy has resulted in durable responses in patients with advanced stage cutaneous melanoma, some patients do not benefit from this, due to resistance mechanisms. This thesis aims to improve response of a subset of melanoma patients to immunotherapy. Firstly, mouse melanoma tumor models are developed that closely resemble some of the resistance mechanisms occurring in patient tumors, which are useful for research. Thereafter, a prognostic biomarker is used to identify the subset of melanoma patients that are unlikely to benefit from ICB therapy. It is then evaluated whether this subset of patients would benefit from combining histone deacetylase inhibitor with ICB. Despite encouraging pre-clinical results showing enhanced anti-tumor immune responses using this combination, this did not result in improved responses in the clinic. Furthermore, this thesis focuses on an immune cell subset, namely regulatory T cells (Tregs), which are known to suppress anti-tumor immune responses. Thus, their presence in the tumor microenvironment (TME) is detrimental to tumor cell killing. This thesis provides an increased understanding of the metabolic adaptation of Tregs to conditions in the TME and propose that targeting these adaptations might overcome the suppression by Tregs and enhance responses to immunotherapy. Show less
The aim of this thesis was to investigate if a text-mining tool is suitable for collecting real-world data from electronic health records to evaluate cancer treatments in clinical practice. By... Show moreThe aim of this thesis was to investigate if a text-mining tool is suitable for collecting real-world data from electronic health records to evaluate cancer treatments in clinical practice. By investigating a range of use cases including treatments of patients with renal cell carcinoma, hepatocellular carcinoma, melanoma, breast cancer, and COVID-19, it showed that the text-mining tool is a suitable method of data needed for the evaluation of treatment patterns, effectiveness, safety, prognostic factors, and guideline adherence. The discussion showed that enhancing the data quality and actively using real-world data for treatment evaluation regarding treatment policies are some of the next steps. Show less
Communication between cells is essential for the proper function of tissues in the human body. In cancer, this communication is disrupted. The signaling pathway initiated by Transforming Growth... Show moreCommunication between cells is essential for the proper function of tissues in the human body. In cancer, this communication is disrupted. The signaling pathway initiated by Transforming Growth Factor beta (TGF-beta) signaling molecules controls many processes in cells, including cell division, cell differentiation and cell migration. Dysregulation of TGF-beta signaling contributes to the formation and progression of many cancers. The inhibition of the TGF-beta signaling pathway is therefore an attractive anti-cancer therapy. Unfortunately, these therapies result in many side effects. The research described in this thesis aims to better understand TGF-beta signaling in different types of cancer. Through the development of new fluorescent tools, the activity and dynamics of TGF-beta signaling can be better studied in vitro and in vivo. In melanoma, the tumor microenvironment appeared to influence the effect of TGF-beta. In cervical cancer, TGF-beta activity levels could distinguish two subtypes of cervical cancer. In bladder cancer, a E3 ubiquitin ligase mediated a balance of BMP signaling, affecting bladder cancer progression. These studies all highlight the complexity of TGF-beta signaling and the need to fully understand TGF-beta signaling in different contexts to improve the use TGF-beta inhibition based therapies in the clinic. Show less
This thesis aims to improve the treatment of patients with stage III melanoma. The first part describes different aspects of treatment with Talimogene Laherparepvec (T-VEC), a genetically modified... Show moreThis thesis aims to improve the treatment of patients with stage III melanoma. The first part describes different aspects of treatment with Talimogene Laherparepvec (T-VEC), a genetically modified herpes virus, which is used as oncolytic immunotherapy for skin and lymph node metastases in melanoma patients. We show that patients with a low tumor burden have the best outcomes, suggesting T-VEC should be used earlier on in the course of the disease. We present a prediction model, allowing a more accurate selection of patients for T-VEC monotherapy. Two studies focused on the use of T-VEC in clinical practice and the results allowed us to make recommendations on the use of PET/CT and dermoscopy during T-VEC treatment. Part two focuses on the value of surveillance and screening imaging in high-risk melanoma patients. We show that FDG-PET/CT is a valuable imaging tool to detect recurrence after complete resection of stage III disease, even shortly after surgery (before starting adjuvant therapy). Finally, we conclude that nodal staging with US as adjunct to SLNB is useful in the work- up of stage IIB/C melanoma, as it can lead to alterations in treatment and prevent unnecessary surgery. Show less
Death in all types of melanomas is generally caused by metastasis. Uveal melanoma (UM) is the most common intraocular melanoma, there are currently no (patient-derived) animal models that... Show moreDeath in all types of melanomas is generally caused by metastasis. Uveal melanoma (UM) is the most common intraocular melanoma, there are currently no (patient-derived) animal models that faithfully recapitulate metastatic dissemination of UM. Here we generate embryonic zebrafish models for both the primary and disseminated stage of ocular melanoma. In doing so we can recapitulate the etiology of cancer in its totality. Subsequently, we developed a patient-derived zebrafish xenograft (zf-PDX) model, using spheroid cultures generated from metastatic and primary UM tissues. Harnessing this versatile model, we reveal high sensitivity of circulating UM cells to ferroptosis induction in vivo by Erastin and RSL3, implicating ferroptosis as a new potential therapy in metastatic UM.Increased melanin levels in cutaneous melanoma are associated with decreased patient survival. Melanin levels in primary uveal melanoma patient cells positively correlate with their metastatic potential in zebrafish. Modulation of melanin levels of pan-melanoma cells results in enhanced/reduced metastatic potential upon increased or decreased melanin levels, respectively. Melanin depletion sensitizes melanoma cells to ferroptosis inducers in zebrafish leading to a decreased metastatic burden. Collectively, our data identify melanin biosynthetic enzymes as potential future target to treat melanoma and show that melanin protects metastasizing melanoma cells from ferroptosis. Show less
Ocular melanoma is a rare disease that originates from melanocytes in the eye. It is the most prevalent primary ocular malignancy in adults, and has a high metastatic rate. Two important questions... Show moreOcular melanoma is a rare disease that originates from melanocytes in the eye. It is the most prevalent primary ocular malignancy in adults, and has a high metastatic rate. Two important questions for good patient care are: 1) How to differentiate between (benign) nevi, and (malignant) melanoma?, and 2) How to treat this tumor best, particularly in cases with metastases?This thesis addresses two types of ocular melanoma: melanoma of the internal parts of the eye (uveal melanoma) and melanoma of the mucous membrane covering the eye (conjunctival melanoma). This thesis combines patient-related projects with projects from the lab.With new imaging techniques we demonstrate that oxygen values differ in eyes with melanoma compared to other eyes including those with a nevus. We use OCT-angiography to depict tumour vessels non-invasively in conjunctival and iris lesions. These two techniques may be used in the future to differentiate lesions, and to monitor patients after treatment.With studies in the lab we show that new drugs (immunotherapy) that are recently used in cutaneous melanoma, can also be used to treat conjunctival melanoma. We show that vascular growth in uveal melanoma is related to other (genetic and immunologic) characteristics, providing new clues for therapy. Show less
Although clinical aspects of melanoma have been extensively studied, the literature largely concerns relatively healthy 20-70 years old patients. Special populations, such as the elderly, children,... Show moreAlthough clinical aspects of melanoma have been extensively studied, the literature largely concerns relatively healthy 20-70 years old patients. Special populations, such as the elderly, children, patients with multiple primary melanoma and those with familial melanoma, are frequently excluded from clinical studies. The studies presented in this thesis were aimed to assess prognostic factors and management of patients with clinically localized melanoma, in particular among the aforementioned special populations. Show less
A type I immune response is crucial for adequate tumor eradication by the immune system. However, tumors often gain evasion mechanisms that create barriers to the generation or effectiveness of a... Show moreA type I immune response is crucial for adequate tumor eradication by the immune system. However, tumors often gain evasion mechanisms that create barriers to the generation or effectiveness of a type I immune response. Among these barriers is the suppression of effective T cell priming and the inhibition of proper T cell infiltration and function in tumors. At present, the only therapies to target these barriers are focused on direct inhibition of T cell function by the tumor, through checkpoint molecules. These therapies are thus dependent on an existing type I response, and are generally not successful when tumors have insufficient T cells primed or infiltrated. This thesis has revealed ways to improve T cell priming and the infiltration of T cells in tumors. Show less
Melanoma is the most aggressive and lethal type of skin cancer since it has the ability to spread to other organs in the body making it harder to control the disease.In this thesis, we aim to... Show moreMelanoma is the most aggressive and lethal type of skin cancer since it has the ability to spread to other organs in the body making it harder to control the disease.In this thesis, we aim to explore the degree to which epigenetics play a role in melanoma, namely, inherited and acquired epigenetic alterations in melanoma susceptibility and development. Show less
In summary, this thesis focused on the understanding the underlying mechanisms driving TNBC metastatic progression. We established DUB activity profiling methods and identified UCHL1 as a candidate... Show moreIn summary, this thesis focused on the understanding the underlying mechanisms driving TNBC metastatic progression. We established DUB activity profiling methods and identified UCHL1 as a candidate oncoprotein that promotes TGFβ-induced breast cancer metastasis. Importantly, we found UCHL1 activity inhibitor as a potential drug for TNBC therapy and developed UCHL1 activity-based probe. For vemurafenib-resistance melanoma, we provided insights that targeting TGFβ signaling may help to overcome drug resistant phenotype. Show less
The aim of the research described in this thesis was to gain a better understanding of the role of the different immune cells and the different FcR on their cell surface, in antibody therapy and... Show moreThe aim of the research described in this thesis was to gain a better understanding of the role of the different immune cells and the different FcR on their cell surface, in antibody therapy and to investigate whether the effectiveness of the tumor-killing mechanisms, activated by the antibodies, can be improved. First, we investigated the role of FcR in various immune responses using a genetically modified mouse, in which all FcR were missing. After defining that role, we studied antibody therapy in a mouse model for melanoma in two different ways: on the one hand, after vaccination using a viral vector that expressed a melanoma antigen, on the other hand, by injecting a melanoma-specific antibody in combination with other substances that activate the immune system. Show less
The focus of this thesis is uveal melanoma (UM) which, once metastasized, is lethal due to lack of effective treatment options. To repress p53 activity approximately 65% of UM tumors express high... Show moreThe focus of this thesis is uveal melanoma (UM) which, once metastasized, is lethal due to lack of effective treatment options. To repress p53 activity approximately 65% of UM tumors express high levels of the p53 inhibitory proteins MDMX or MDM2. The aim of this thesis is to unravel the oncogenic function of MDMX and provide new treatment options for patients with metastasized UM. Chapter 2 describes the regulation of the transcriptome by MDMX in UM and proposes novel p53-independent effects of MDMX, i.e. FOXO inhibition. In chapter 3 the opportunities of a combined targeting of two common signaling pathways as therapeutic intervention for metastasized UM patients is investigated. Genetic interference with either MDMX or PKC δ expression or activity showed that beneficial effects can already be achieved by a more specific targeting, which is presumable less toxic to the patient. In chapter 4 it is described, opposed to what has been reported before, that enhancer of zeste homolog 2 (EZH2) inhibition poses a valuable novel therapeutic invention for UM. In chapter 5 it is shown that combining two clinically approved drugs, Quisinostat and Flavopiridol, could serve as an effective therapeutic intervention for UM patients. Show less
This thesis focused on different aspects of melanoma treatment with immunotherapy and targeted therapy. In chapter 2 we search for biomarkers that could be associated with overall survival in... Show moreThis thesis focused on different aspects of melanoma treatment with immunotherapy and targeted therapy. In chapter 2 we search for biomarkers that could be associated with overall survival in patients treated with ipilimumab. In chapter 3 we describe diarrea, a commonly seen side effect of immunotherapy. Here we show that there is no significant correlation between grade of diarrhea and severity of colitis as seen during endoscopy. Chapters 4 and 5 describe patients with brain metastases and/or leptomeningeal metastases. In chapter 4 we show the difference in overall survival in patients treated with vemurafenib, dabrafenib or the combination of dabrafenib + trametinib. Chapter 5 focusses on the treatment of leptomeningeal metastases. Here a significant difference in overall survival was noted between treated and untreated patients. Furthermore LDH was a predictive biomarker for overall survival. In chapter 6 we show that treating patients with vemurafenib beyond progression of disease has a significant impact on overall survival. Lastly in chapter 7 we review the past, present and future of treating patients with different kinds of cancer with tumor-infiltrating lymphocytes. Show less
Being a member of a melanoma family is a major risk factor for cutaneous malignant melanoma. In this thesis clinical characteristics and management of melanoma families are discussed. In the first... Show moreBeing a member of a melanoma family is a major risk factor for cutaneous malignant melanoma. In this thesis clinical characteristics and management of melanoma families are discussed. In the first part of the thesis clinical and histological characteristics of melanoma (patients) from families with a (p16-Leiden) mutation in the high penetrance melanoma susceptibility gene CDKN2A were compared with the general population. Significant differences with respect to several characteristics are reported. In the second part of the thesis the yield, effectiveness, and causes for failure of surveillance of melanoma families are discussed. We report that surveillance is associated with a more favorable tumor stage. Several aspects of surveillance, including interval melanomas, surveillance interval, noncompliance, and overdiagnosis are discussed. Based on analyses of melanoma detection rates in families with different family and genetic characteristics, we propose a risk stratification for members of melanoma families. In the third part of the thesis we investigate the impact of dermoscopy on management decisions. It is demonstrated that dermoscopy by dermoscopy experts in the setting of melanoma family surveillance resulted in a considerable reduction of unnecessary excisions. This effect was considerably less in the setting of dermoscopy non-experts examining patients in general dermatology clinics. Show less
Melanoma is a malignancy that arises from melanocytes, the pigment-producing cells that can be predominantly found in the eye or the epidermal basal layer of the skin. Mainly due to increased UV... Show moreMelanoma is a malignancy that arises from melanocytes, the pigment-producing cells that can be predominantly found in the eye or the epidermal basal layer of the skin. Mainly due to increased UV exposure, the incidence of melanoma has doubled worldwide over the past three decades (200.000 new cases in 2008). Primary melanomas can be easily treated by surgical resection, leading to a good prognosis for stage I patients. However, metastasized melanoma is almost completely resistant to therapeutic modalities such as radio- and chemotherapy, resulting in a median overall survival of less than one year for this patient group. Despite considerable efforts, for over 20 years there was no melanoma treatment developed that could improve survival of stage IV patients. However, the treatment of unresectable metastasized melanoma has progressed markedly in recent years due to the development of both immunotherapies that stimulate anti-tumor immunity and targeted therapies that block oncogenic proteins. This thesis will focus on pre-clinical work concerning the optimization of melanoma treatment. In detail, it will address for both targeted therapies and immunotherapies factors that play a role in the identification of response-predictive biomarkers, the toxicity of treatments, and the potential efficacy of combination treatments. Show less
Cutaneous melanoma is the most aggressive form of skin cancer and its incidence among Caucasian populations has increased whereas mortality rates are stabilizing or decreasing. The total burden of... Show moreCutaneous melanoma is the most aggressive form of skin cancer and its incidence among Caucasian populations has increased whereas mortality rates are stabilizing or decreasing. The total burden of melanoma is expected to be increasing. As effective treatment options for advanced melanoma are lacking, melanoma prevention may be the key issue in melanoma disease control. Although sun protection programs have increased awareness, they have not resulted in a decreased melanoma incidence. In addition, most melanoma risk factors are not amenable. Alternative approaches such as cancer chemoprevention are, therefore, important research topics. Several agents, such as statins, non-steroidal anti-inflammatory drugs, and angiotensin-converting enzyme inhibitors, have been claimed to have chemopreventive properties. However, it is unknown which of these have the best potential to be useful. This thesis presents: - epidemiologic cancer registry-based studies from The Netherlands on the epidemiology of extracutaneous melanoma and on the burden of disease due to cutaneous melanoma - a qualitative review discussing candidate drugs for melanoma chemoprevention, their possible mechanisms of action, and evidence for their chemopreventive efficacy, safety and tolerability - pharmacoepidemiological studies testing hypotheses on chemopreventive activityof several drugs on melanoma - pharmacoepidemiological studies on the association between estrogen use and melanoma. Show less
Cutaneous and uveal melanoma are malignant tumours with no treatment available once the metastases occur. Despite both melanomas are highly immunogenic, and often despite the presence of potent... Show moreCutaneous and uveal melanoma are malignant tumours with no treatment available once the metastases occur. Despite both melanomas are highly immunogenic, and often despite the presence of potent anti-tumour immune cells in patients__ blood, in more than 95% of patients, tumour growth remains unaffected. Hereby we investigate the mechanisms that help melanomas to escape from the spontaneous or activated by vaccination cytotoxicity of T lymphocytes and discuss the impact of local microenvironment created by melanoma, focusing on the role of immunomodulatory dendritic cells. Show less
For a tumor cell to propagate, it must survive extremely stressful conditions that would normally trigger the cell to die. Cancer cells however survive, probably due to evasion of the apoptotic... Show moreFor a tumor cell to propagate, it must survive extremely stressful conditions that would normally trigger the cell to die. Cancer cells however survive, probably due to evasion of the apoptotic cell death pathway. It follows that a detailed understanding of the regulation of the apoptotic pathways in cancer cells can improve the anti-cancer treatments. Part 1 of this thesis describes our in vitro studies regarding the regulation of apoptosis in melanoma cells, since melanoma is a form of cancer that is highly resistant to anti-cancer therapies. c-Myc enhances the apoptosis sensitivity of the cells. The protein Apaf-1 is not involved in this sensitivity. A yet unidentified serine protease plays an important role in the initiation of apoptosis upon DNA damage. Part 2 of this thesis describes our studies regarding both the regulation of apoptosis in rectal carcinoma and its prognostic value for rectal cancer patients. To evaluate the impact of (radiation-induced) tumor cell apoptosis on clinical outcome of cancer patients, the level of apoptosis have been determined in non-irradiated and irradiated rectal carcinoma samples. The level of tumor cell apoptosis is scored by immunohistochemical stainings of the carcinoma samples, and by measuring caspase-3 activity. Both studies show that high levels of apoptosis is associated with a low local recurrence risk. A genetic approach is used to identify factors that play a role in the regulation of apoptosis in rectal carcinoma in vivo. After evaluation two microarray procedures, the most convenient procedure is used to compare the gene expression profiles of tumors with high levels of apoptosis with low-apoptotic tumors. The difference in expression of several of the identified genes are confirmed on protein expression level by immunohistochemistry, and show two subsets of high-apoptotic tumors. These data suggest two different regulations of apoptosis in vivo. The prognostic value of one of the identified proteins, HLA-DR, has been studied in more detail and epithelial HLA-DR expression is significantly associated with lower recurrences and better survival for rectal cancer patients. 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
This thesis describes several topics on angiogenesis and screening in uveal melanoma, the most frequent eye tumour in adults. The expression of vascular endothelial growth factor and other... Show moreThis thesis describes several topics on angiogenesis and screening in uveal melanoma, the most frequent eye tumour in adults. The expression of vascular endothelial growth factor and other angiostimulating factors in this tumour are described. In a second part, the use of different screening tests in screening for metastases of uveal melanoma is investigated. Show less
