A deeper understanding of the parameters driving response and resistance to immunotherapy is needed to improve the low response rates observed in breast cancer patients. Research into immunotherapy... Show moreA deeper understanding of the parameters driving response and resistance to immunotherapy is needed to improve the low response rates observed in breast cancer patients. Research into immunotherapy response has predominantly focused on T cells, however effective immune responses require tightly regulated crosstalk between innate and adaptive immune cells. By combining profiling of blood and tumors from metastatic breast cancer patients with mechanistic studies in mouse models, we uncovered the critical role of eosinophils in immunotherapy response, and we provide proof-of-principle for eosinophil engagement to enhance immunotherapy efficacy. Focusing on resistance mechanisms to immunotherapy, we demonstrate that neoadjuvant immunotherapy triggers persistent and systemic regulatory T cell activation which blunts therapeutic efficacy against metastatic spread of breast tumors. In addition, we demonstrate that neutrophils in the tumor microenvironment pose a barrier to immunotherapy response through T cell suppression. Lastly, we demonstrate that combining the immunomodulatory agent PD1-IL2v with cisplatin is a powerful approach to induce a broad activation of systemic and intratumoral adaptive and innate immunity, resulting in effective immunotherapy responses. Overall, this work identifies several key players and their interconnectivities in anti-tumor immunity and tumor-induced immunosuppression that may be therapeutically exploited to improve immunotherapy responses for breast cancer patients. 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
Pathogenic variants in PALB2 and CHEK2 are associated with an increased risk of breast cancer. By contrast, for missense variants of uncertain significance (VUS) in these genes, the associated... Show morePathogenic variants in PALB2 and CHEK2 are associated with an increased risk of breast cancer. By contrast, for missense variants of uncertain significance (VUS) in these genes, the associated breast cancer risk is often unclear. To aid in their clinical classification, functional assays that determine the impact of missense VUS on PALB2 and CHK2 protein function have been performed in this thesis. By means of these functional analyses, numerous missense VUS (in both PALB2 and CHEK2) have been identified that are, from a functional viewpoint, just as damaging as known pathogenic (i.e., truncating) variants. In agreement, we observe that the level of impaired protein function correlates with the degree of increased breast cancer risk. Overall, these findings suggest that damaging PALB2 and CHEK2 missense VUS are associated with a risk of breast cancer similar to that of protein-truncating variants in these genes. This indicates the urgency of expanding the functional characterization of missense VUS in both PALB2 and CHEK2 to further understand the associated cancer risk. Show less
Breast cancer has a high mortality in women worldwide. Tumor cells experience hypoxia, which is accompanied by alterations in cell metabolism and can drive metastasis by triggering an epithelial... Show moreBreast cancer has a high mortality in women worldwide. Tumor cells experience hypoxia, which is accompanied by alterations in cell metabolism and can drive metastasis by triggering an epithelial–mesenchymal transition (EMT) in the tumor cells. Yes-associated protein (YAP) and a transcriptional co-activator with PDZ-binding (TAZ) are two transcriptional co-activators involved in growth, metabolism, and metastasis in cancer. Breast cancer can be divided into different subtypes. One criterium underlying such subtypes is based on the levels of Human Epidermal growth factor Receptor 2 (HER-2), Estrogen Receptor (ER) and Progesterone Receptor (PR). The subtypes include luminal-like (luminal A and luminal B), HER-2 enriched and basal-like (often “triple negative”). Triple negative breast cancer (TNBC) has a lower survival rate due to the lack of therapeutic targets. Fundamental research exploring the molecular mechanisms at work in cancer cells and their response to a hypoxic environment may contribute to insights for future clinical treatment. This thesis focused on profiling breast cancer cells belonging to distinct subtypes under acute and chronic hypoxia, investigating the crosstalk between hypoxia regulated pathways and YAP/TAZ signaling in luminal breast cancer versus TNBC cells, and identification of the potential targets of TAZ in breast cancer cells. Show less
The immune system plays a dual role in cancer development. Besides the potential to eliminate cancer cells, immunoregulatory mechanisms exist that counteract anti-tumor immunity.Research in this... Show moreThe immune system plays a dual role in cancer development. Besides the potential to eliminate cancer cells, immunoregulatory mechanisms exist that counteract anti-tumor immunity.Research in this thesis focusses on the role of regulatory T cells (Tregs), a type of adaptive immune cell that plays a major role in tumor-associated immunosuppression. Specifically, the role of Tregs was investigated during the development of primary- and metastatic breast cancer, and in the context of novel immunotherapeutics. This was done by using advanced genetically engineered mouse models that recapitulate human breast cancer.The results in this thesis describe that breast tumors are, already early in their development, able to mobilise Tregs in the tumor-draining lymph nodes, thereby creating a local immunosuppressive niche leading to increased lymph node metastasis. In addition, it was found that the immunotherapeutic treatments anti-PD1 and anti-CTLA4 inadvertently activate Tregs, resulting in a diminished efficacy of this treatment in mice bearing breast tumors. Finally, we describe a mechanism by which intratumoral macrophages are critical promote the intratumoral accumulation of Tregs in breast tumors.Insights from this thesis may eventually contribute to the development of therapeutic applications that are aimed at overcoming immunoregulatory mechanisms in breast cancer. Show less
Due to a shorter remaining life expectancy, the risk of recurrence in older patients with low risk breast cancer is often reduced and the benefit of treatments may be very limited, especially with... Show moreDue to a shorter remaining life expectancy, the risk of recurrence in older patients with low risk breast cancer is often reduced and the benefit of treatments may be very limited, especially with adjuvant treatments. In the first part of this thesis, we studied the interplay between breast cancer mortality and other-cause mortality. In the second part of this thesis, we investigated the effect of surgery and radiotherapy in subsets of the older population of patients with breast cancer in which the actual treatment effect is questionable. Show less
This thesis describes the steps necessary for the addition of the tumor-stroma ratio (TSR) into clinical practice as high-risk factor besides the TNM classification. The route from laboratory... Show moreThis thesis describes the steps necessary for the addition of the tumor-stroma ratio (TSR) into clinical practice as high-risk factor besides the TNM classification. The route from laboratory biomarker development to clinical implementation is followed. During this process, the relationship of the TSR to other available biomarkers for prognostic information for breast and colon cancer patients is investigated. Additionally, the prognostic value of the TSR in lung cancer is studied. Show less
Major achievements in the field of immune oncology have demonstrated the ability of the immune system to induce a response against cancer. The prognostic impact of pre-existing immunity in several... Show moreMajor achievements in the field of immune oncology have demonstrated the ability of the immune system to induce a response against cancer. The prognostic impact of pre-existing immunity in several cancer types, including breast and colon cancer, demonstrates the influence of the immune system on disease progression. At the same time, immunotherapeutic approaches that aim to enhance antitumor immune reactions have significantly improved the clinical outcome for a subset of patients. However, a large proportion of patients (60-80%) do not respond to immunotherapeutic treatments. To extend the benefit of immunotherapeutic strategies to a larger number of patients, it is imperative to understand the mechanisms associated with immune responsiveness. Different variables have been described to influence the development of antitumor immunity in cancer patients, including the tumor’s genetic program, the genetic makeup of the patients, and environmental factors such as the microbiome. These factors likely act in concert to modulate antitumor immune responses. This thesis aimed to dissect the molecular determinants of cancer immune responsiveness in human tumors. A systems biology approach was used to define underlying factors that shape the tumor microenvironment and reveal potential mechanisms of immune escape. Show less
The main objective of work presented in this thesis was to explore the clinical utility of the Polygenic Risk Score (PRS) based on breast cancer associated common low risk variants, which explain ... Show moreThe main objective of work presented in this thesis was to explore the clinical utility of the Polygenic Risk Score (PRS) based on breast cancer associated common low risk variants, which explain ~18% of the familial relative risk, for individual breast cancer risk prediction. It did so by generating knowledge about the PRS in the Dutch general population and in clinic-based breast cancer families, as well as in a large international population of BRCA1/2 pathogenic variant carriers. We have validated the association of the PRS with breast cancer for women in both the Dutch population and breast cancer families and showed a better risk-discrimination by adding the PRS to family-based risk prediction. Secondly, we have shown that addition of the PRS to family-based risk prediction has an impact on screening recommendations for many non-carriers and carriers of a pathogenic variant in a moderate breast cancer gene. The results will support implementation of comprehensive risk prediction in the clinic, and may help women to make more informed choices about their optimal clinical management. Show less
The clinical potential of applying synthetic lethality to cancer treatment is famously demonstrated by the BRCA1/PARP1 paradigm: a tumor specific defect in BRCA1 – a component of the DNA double... Show moreThe clinical potential of applying synthetic lethality to cancer treatment is famously demonstrated by the BRCA1/PARP1 paradigm: a tumor specific defect in BRCA1 – a component of the DNA double-strand break (DSB) repair pathway homologous recombination (HR) – results in a remarkable sensitivity to PARP1 inhibition (PARPi). Despite spectacular initial responses in patients, resistance to PARPi treatment may develop and must be overcome to maximally exploit this interaction in the clinic. Genetically engineered (mouse) model systems have shown that PARPi resistance may arise through inactivation of the 53BP1 pathway. The 53BP1 pathway normally protects DSB ends from resection and the removal of this “brake” restores HR in the absence of BRCA1. However, how the 53BP1 pathway protects DSB ends from resection has remained elusive. In this thesis, advances in 3D tumor organoid culture protocols and CRISPR/Cas9 (screening) technology were applied to identify and validate new components of the 53BP1 pathway that render BRCA1 deficient cells resistant to PARPi upon their loss. Furthermore, a new acquired vulnerability that can be therapeutically exploited to deplete such PARPi resistant cells is described. Together, this thesis provides mechanistic insight in DSB repair and illustrates how such fundamental knowledge may stand at the basis to combat resistance. Show less
The studies in this thesis contribute to more accurate risk assessment and prognosis prediction for DCIS and to better response evaluation of IBC treatment.For the Ductal Carcinoma In Situ (DCIS)... Show moreThe studies in this thesis contribute to more accurate risk assessment and prognosis prediction for DCIS and to better response evaluation of IBC treatment.For the Ductal Carcinoma In Situ (DCIS) studies, unbiased cohorts were used within the international Grand Challenge PRECISION consortium, funded by Cancer Research UK and KWF Dutch Cancer Society. DCIS is graded as low-, intermediate-, or high-grade depending on how abnormal the DCIS-cells look like. However, we showed that pathologists often disagree on grade. To overcome this limitation, we found that almost all DCIS scored as non-high-grade by the majority of pathologists express the estrogen receptor (ER) and are negative for the growth factor receptor HER2, whereas high-grade DCIS is mixed in expression for ER and HER2. We also provided insights in the recurrence risks of DCIS after treatment. See also https://cancergrandchallenges.org/teams/precision.The studies on Invasive Breast Cancer (IBC) were performed on a hospital-based cohort. We found for example substantial variation in tumour response evaluation for HER2-positive IBC after pre-operative chemotherapy due to different guidelines used. For accurate outcome analysis, reducing such variation is mandatory. Therefore, we are working on reaching international consensus of response evaluation. Show less
This thesis describes the effects of shortterm fasting on chemotherapy outcome in patients with breast cancer and the IGF-1 and insulin pathway as a target for cancer therapy and as a biomarker for... Show moreThis thesis describes the effects of shortterm fasting on chemotherapy outcome in patients with breast cancer and the IGF-1 and insulin pathway as a target for cancer therapy and as a biomarker for chemotherapy outcome.Preclinical research is evaluated, which shows that short-term fasting during chemotherapy is effective. The effects of short-term fasting in humans is not evident yet. Although the first small clinical studies of short-term fasting as adjunct to chemotherapy are promising in terms of decreased toxicity and enhanced efficacy, the exact mechanism and effects are not established yet. More studies and a longer follow-up are needed to prove this.Insulin-like growth factor 1 (IGF-1) and insulin are members of the IGF-1 pathway, which is involved in cell growth and proliferation. The effects of the IGF-1 pathway on chemotherapy outcome and the pathway itself as target for cancer therapy are described. The disappointing results of clinical studies of IGF-1R inhibitors may be caused by the complexity of the IGF-1R pathway. Lowering both insulin and IGF-1, perhaps with a short-term fasting intervention, serves as a possible target in cancer therapy. Show less
Invasive lobular carcinoma (ILC) is the second most common type of breast cancer. Hallmarks of ILC include disruption of adherens junctions and hyperactivation of phosphoinositide 3-kinase (PI3K)... Show moreInvasive lobular carcinoma (ILC) is the second most common type of breast cancer. Hallmarks of ILC include disruption of adherens junctions and hyperactivation of phosphoinositide 3-kinase (PI3K)-mTOR signaling. The tumor suppressor PTEN regulates PI3K signaling. We present a preclinical mouse model of ILC metastasis, based on inactivation of the adherens junction protein E-cadherin and the tumor suppressor p53 and surgical excision of primary tumors. In this model, pharmacological mTOR inhibition blocks growth of primary tumors as well as metastatic disease, and this response is partially dependent on the adaptive immune system. Loss of E-cadherin mouse mammary epithelium leads to apoptosis, and PTEN activation alone results in squamous metaplastic mammary tumors, but a combination of these events leads to ILC formation, indicating a causal role of PI3K signaling together with E-cadherin loss in ILC. Combined somatic loss of the adherens junction molecule p120 and p53 in the mouse mammary gland leads to metaplastic mammary tumors, and loss of p120 in breast cancer cell lines promotes anoikis resistance through hypersensitization of growth factor receptor (GFR) signaling. Combined inactivation of E-cadherin, p120 and p53 induces basal-like tumors, with an epithelial-to- mesenchymal-transition (EMT) phenotype, and no ILC formation. Show less
Since cancer survival rates are rising, there is growing attention for longterm side effects of cancer and its treatment. A common side effect is the negative impact of treatment on sexuality of... Show moreSince cancer survival rates are rising, there is growing attention for longterm side effects of cancer and its treatment. A common side effect is the negative impact of treatment on sexuality of patients and their partners. Patient and partners as well as healthcare professionals experience several barriers to discuss this topic, like lack of time and lack of knowlegde. Two-thirds of the cancer patients reported to be in need of information regarding sexual health; especially those who were younger, who reported a negative impact of cancer on sexuality and those who were diagnosed less than two years ago. Patients and partners reported to prefer to discuss sexual health with nurse practitioners throughout the treatment proces. Besides, satisfaction with sexual life after treatment is related to satisfaction before treatment, not only with current sexual function.Widely available information and defining responsibility within the oncology treatment team would be helpful to improve communication around sexual health in cancer care. Additionally, specialized clinics would tackle soms frequently reported barriers of discussing sexuality. More reseach is needed on the implementation of sexual healthcare in oncology practice to deliver continuum of care, which will ultimately improve patient care. Show less
Tumors are complex ecosystems containing not just cancer cells, but a large variety of cell types, including immune cells. Moreover, tumors have a systemic influence: they can signal long distances... Show moreTumors are complex ecosystems containing not just cancer cells, but a large variety of cell types, including immune cells. Moreover, tumors have a systemic influence: they can signal long distances using soluble molecules and hijack non-neoplastic cells (such as immune cells) in distant organs for their own benefit, thus maximising their metastatic potential. The phenotype of immune cells in tumors and in systemic environments is therefore a key determinant of cancer progression and response to therapy.This thesis aims to understand what governs the tumor-immune ecosystem. We argue that cancer-intrinsic genetic aberrations have a dominant role in determining the tumor immune contexture, as well as systemic inflammatory activation. Understanding the intricate connection between the genetics of breast cancer and anti-tumor immune responses will help develop personalised immune intervention strategies for cancer, tailored to the genetic makeup of a patient’s tumor. Furthermore, we examine in detail the role of neutrophils in cancer-induced systemic inflammation, and how they influence the progression and spread of breast cancer. While tumors can be highly heterogeneous in nature, we show that neutrophils themselves also have a tremendous phenotypic diversity. Mapping this heterogeneity in neutrophil phenotypes may help to utilise these cells in cancer immunotherapy. Show less
Over a century ago, the systematic evaluation of medical treatment outcomes was first described by Ernest Codman. At that time a too progressive thought, while the value of measuring the quality of... Show moreOver a century ago, the systematic evaluation of medical treatment outcomes was first described by Ernest Codman. At that time a too progressive thought, while the value of measuring the quality of care is seen nowadays.The NABON Breast Cancer Audit (NBCA) provides insight into the various care processes that are part of the complex multidisciplinary treatment of patients diagnosed with breast cancer. Results of the NBCA show a good and still improving quality of care for patients diagnosed with breast cancer. In addition, this thesis provides insight into a sub-area where variation between hospitals is found; the performance of an immediate breast reconstruction in patients undergoing mastectomy. Many factors influence this hospital variation, such as patient and tumour factors, hospital factors and differences in indications for immediate breast reconstruction between surgeons and plastic surgeons. Physicians should inform patients about the various breast cancer treatment options. Informing a patient about immediate breast reconstruction results in a 14 times higher chance of actually undergoing a breast reconstruction after mastectomy.These factors should to be evaluated and improved as quality of life is higher for patients with immediate breast reconstruction than with mastectomy alone. By continuously measuring, providing feedback and acting on the data found at both a national level and in the consultation room, Ernest Codman's heritage will certainly be realized. Show less
The objective of this study was to investigate the expression and function of GRHL2 in different breast cancer subtypes. In Chapter 2, we focused on the expression of GRHL2 in human breast cancer... Show moreThe objective of this study was to investigate the expression and function of GRHL2 in different breast cancer subtypes. In Chapter 2, we focused on the expression of GRHL2 in human breast cancer and the distinct effects of GRHL2 knockout on aspects of growth versus migration in basal A and luminal-like subtypes. In Chapter 3, ChIP-seq was used to explore the genomic landscape of GRHL2 binding sites in basal A and luminal-like subtypes of breast cancer and this data was used to predict shared and distinct GRHL2 target genes. In Chapter 4, based on a conditional GRHL2 knockout system, we determined the dynamic changes in genome-wide DNA transcription triggered by loss of GRHL2 in luminal-like breast cancer cells and used the data to predict affected pathways. In Chapter 5, ChIP-seq and BrU-seq data were integrated to identify genes whose transcription is controlled by GRHL2 and establish gene expression networks regulated by GRHL2 in luminal-like breast cancer. 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
Ductal carcinoma in situ (DCIS) is considered to be a non-obligate precursor of invasive breast cancer (IBC). Optimal clinical management of DCIS remains controversial, as we are unable to... Show moreDuctal carcinoma in situ (DCIS) is considered to be a non-obligate precursor of invasive breast cancer (IBC). Optimal clinical management of DCIS remains controversial, as we are unable to identify those DCIS lesions with invasive potential. As a result, current treatment guidelines for DCIS dictate that all women diagnosed with DCIS should undergo treatment to prevent the development of IBC. This makes that many women, who have a low-risk to develop subsequent IBC, are being harmed by this intensive treatment without any benefit. Furthermore, definite proof of DCIS progression to IBC is still lacking.The objectives of this thesis were to identify prognostic markers predictive of the development of subsequent ipsilateral IBC after DCIS and to explore the clonal relatedness of patient-matched DCIS and subsequent ipsilateral IBC. To achieve this, histopathological analysis and molecular profiling were performed using a patient group which was part of a nation-wide population-based cohort including all women diagnosed with and treated for DCIS with breast conserving surgery alone in the Netherlands between 1989 and 2005. The results presented in this thesis will help to stratify a woman’s individual risk of subsequent invasive breast cancer development and will help to avoid overtreatment. Show less
In this thesis, we aimed to better understand the underlying mechanisms involved in TNBC progression and metastasis formation and discover new targets to reduce breast cancer related deaths. We... Show moreIn this thesis, we aimed to better understand the underlying mechanisms involved in TNBC progression and metastasis formation and discover new targets to reduce breast cancer related deaths. We performed an imaging-based RNAi phenotypic cell migration screen in two highly motile TNBC cancer cell lines to provide a repository of signaling determinants that functionally drive TNBC cell motility. Interestingly, two modulators essential for cancer cell migration were known to be involved in RNA splicing, making us decide to focus on the role of RNA splicing in breast cancer progression. We next summarized the current knowledge about splicing factors in breast cancer development and progression and identified co-regulated splicing factors that were associated with aggressive breast cancer phenotypes and metastasis formation that was not only restricted to breast cancer, increasing the global understanding of the role of the spliceosome in cancer development and progression. Moreover, the role of splicing factors in two major processes in cancer progression, cell migration and proliferation, was examined. Finally, using RNA sequencing, we systematically compared the transcriptomes of 14 breast cancer cell lines cultured both in 2D and 3D conditions to unravel the reprogramming that is associated with the invasive phenotype of basal B TNBC. Show less
