<p>  </p> <p>Cancer is well known as a disease involving genetic disorders, which make them distinguishable from normal tissue by the altered molecular signatures. Theoretically, malignant cells can be recognized and attacked by innate and adaptive immune system as “non-self” species, and the idea to take advantage of host immunity to treat cancer has been discussed for over a century. Through the multi-disciplinary research efforts from immunology, cancer biology, cell engineering <em>etc.</em>, cancer immunotherapy has been successfully translated from benchside to bedside. While the clinical application of immunotherapeutic regimens has achieved extraordinary success including the unprecedented long-term survival of metastatic melanoma patients, we must take it seriously that only a small proportion (about 20% on average) of patients benefit from immunotherapy, and many develop secondary progression after the initial response. Advancements have been made in biomarker development to identify the group the patients who may benefit from immunotherapy, yet the accuracy and adaptability remain to be improved. In general, the performance of immunotherapy is hardly satisfactory as the current situation.</p> <p><br></p> <p>The effect out of T cell-mediated immune response is mediated by plenty pairs of receptor-ligand interactions in the immune synapse between T cells and target cells. Despite the T cell receptor-mediated first signal and CD28-mediated second signal, a huge collection of co-signals molecules serves unneglectable roles to keep the T-cell immune response fine-tuned under appropriate threshold. Inadequate co-signaling transduction result in with immune deficiency or autoimmunity depending on the type of signal (stimulatory or inhibitory). 4-1BB is a significant co-receptor which is mainly expressed on T cells and delivers activation signal to drive T cell proliferation and cytotoxicity. 4-1BB is targetable for cancer treatment and can be used as a tumor-reactive T cell marker as well. Hence, it is of substantial importance to understand how co-signaling molecules, such as 4-1BB, are regulated under specific physiological or pathological conditions.</p> <p><br></p> <p>Proteins are regulated at multiple levels, including transcription, translation, localization, and interaction with other biomolecules (covalently or non-covalently). Post-translational modification (PTM) constitutes a critical type of mechanism that elicit multidimensional effects to the biophysical properties of target proteins. Herein, I sought to elaborate how 4-1BB, an TNFRSF family co-stimulatory receptor, is possessed and regulated by PTMs, particularly ubiquitination and <em>N</em>-glycosylation. In the first part of this study, I confirmed that 4-1BB is degraded through ubiquitination-proteasome pathway and identified FBXL20 to be the E3 ligase subunit mediating 4-1BB polyubiquitination. While conducting the first section, I noticed that 4-1BB is heavily <em>N</em>-glycosylated and thereby dissected the biological significance of this modification which made up the second part of this study. I experimentally characterized that 4-1BB necessitates its <em>N</em>-glycans to be efficaciously transported to cell membrane through the secretory pathway. Plus, the glycosylated 4-1BB has short half-life. Without the spatial hindrance established by <em>N</em>-linked carbohydrate moieties, the exposed C121 residue of 4-1BB can be used to forms stable multimer which intracellular retention and stabilization of 4-1BB.</p> <p><br></p> <p>This study uncovered the post-translational mechanisms of action of 4-1BB regulation for the first time. More fundamentally, we provided a blueprint to study the post-translational regulation network of immune receptors which may be applied for future investigations in other targets. Our ultimate hope is to be able to grasp the dynamic of key immune regulators in the context of microenvironment and based on which pair the right therapeutics with the correct populations.</p>

Bakgrund: Varje år drabbas cirka 1200 personer av hjärntumör. En hjärntumör påverkarpatienter fysiskt- och psykiskt. Det kan innebära allvarliga symtom, biverkningar avbehandling samt en förändrad livssituation. För att underlätta vården behöver patienterförklara deras upplevelser. Syfte: Att sammanställa och belysa forskning som beskriverpatienters upplevelser av vård vid hjärntumör. Metod: Litteraturöversikt av tolvvetenskapliga artiklar som analyserades enligt analysgranskning för litteraturöversikt.Resultat: Resultatet framkom tre teman: 1. Behov av kontinuerligt stöd och information 2.Mötet som är avgörande för ett meningsfullt liv 3. Uppleva utanförskap. Subteman var: 1.Att uppleva behov av att erhålla information; Att uppleva behov av stöd 2. Att få en godvårdrelation och kommunikation; Osäkerhet och rädsla i vårdmöten 3. Uppleva bristandedelaktighet; Att förlora sin autonomi Konklusion: Stöd och en god, trygg vårdrelation krävsför att patienter ska uppnå gott välbefinnande. God och individualiserad information behövsnär de fysiska- och psykiska förändringarna sker för patienten. Känslor av utanförskap ochförlorad autonomi drabbar patienterna. Kliniska implikationer: Studien belyser patientersupplevelser av vård vid hjärntumör. Sjuksköterskor får i denna studie ökad kunskap ochförståelse för hur dessa patienter bör vårdas i syfte att ge en god omvårdnad med respekt förpatientens rätt till självbestämmande och välbefinnande. / Background: Every year, around 1,200 people suffer from brain tumors. A brain tumor havephysical as well as psychological affects on these patients. It can cause serious symptoms,side effects of treatment and a changed life situation. To facilitate care, patients need toexplain their experiences. Aim: To compile and illustrate research that describes patientsexperiences of care when suffering from a brain tumour. Method: Literature review oftwelve scientific articles that were analyzed according to the method of analysis for literaturereview. Result: The result revealed three themes: 1. The need for continuous support andinformation. 2. The meeting that is crucial for a meaningful life. 3. To experience exclusion.The subthemes were: 1. To experience the need of obtaining information; To experienceneeds of support. 2. To acquire a good caring relationship and communication; Insecuritiesand fear of care meetings. 3. To experience lack of participation; To lose their autonomy.Conclusion: Support and comforting, safe caring relationship is required in order forpatients to experience appropriate well-being. Kind and individualised information arerequired when the physical and psychological changes takes place for the patients. Feelingsof exclusion and lack of autonomy affects patients. Clinical implications: This studyillustrates the patients experiences of care when suffering from a brain tumour. In this studynurses achieve increased knowledge and understanding about how these patients should becared for in order to provide good care with respect for the patients right of self determinationand well-being.

Sarcoid-like tumours have been reported in Cape mountain zebra (Equus zebra zebra) in two South African game parks recently. These tumours caused severe distress to the animals and also made them unsightly for tourists visiting the parks. The aim of this investigation was to identify and characterize the infectious agent considered to be involved in the aetiology of sarcoid in the Cape mountain zebra. Bovine papillomaviruses (BPV) -1 and -2 deoxyribonucleic acid (DNA) were detected by the polymerase chain reaction (PCR) in sarcoid tumour tissue, but not from blood specimens or unaffected skin. Differentiation between BPV-1 and -2 was made by using the restriction endonuclease BstXI on PCR products of the E5 open reading frame (ORF). A hybridization probe real-time assay was developed for the specific and sensitive detection and differentiation of BPV-1 and -2 DNA in blood, skin and sarcoid tumour samples. For the specific detection of BPV-1, an increase in fluorescence was detected at 640 nm and of BPV-2 at 705 nm. The test is extremely sensitive and able to detect 1.5 genome copies/reaction. The presence of BPV-1 and -2 DNA could be demonstrated in the blood of sarcoid-affected and -unaffected zebras even in the blood of zebras from parks where sarcoids have never been observed. The phylogenetic relationships of the papillomaviruses detected in tumours in the Cape mountain zebra in comparison with a broad selection of papillomavirus sequences available in GenBank were compiled. The papillomavirus sequences retrieved from the zebras were identified as variants of either BPV-1 or BPV-2. The age of the most recent common ancestor for BPV-1 variants is estimated to be 1.40 million years (Mya) and for BPV-2 variants, 0.55 Mya. The age of the most recent common ancestor of BPV-1 and BPV-2 is estimated to be 5.34 Mya. Certain major histocompatibility (MHC) haplotypes are associated with increased risk of sarcoid tumours in horses. The zebras in these parks may have become inbred for the MHC region with increased prevalence for a haplotype, conferring increased risk for sarcoid tumours. Therefore typing system was developed to determine whether or not a high prevalence of sarcoids among zebras is associated with a MHC haplotype. Single strand conformational polymorphism was used to assess the genetic variation in MHC class II genes. The use of DQB and DRB genes demonstrated that genetic variation and sarcoids in the zebras could not be attributed to a specific haplotype. The developed real-time PCR technique was also applied in the detection of cutaneous papillomavirus in two giraffes (Giraffa camelopardalis) which were manifesting cutaneous papillomatosis, in the Kruger National Park and in a fibropapilloma in a sable antelope (Hippotragus niger), on a game farm in the Kimberley district, South Africa. In conclusion, this was the first study to confirm the presence of BPV-1 and -2 DNA in the sarcoid tumours, healthy skin and blood of sarcoid-affected and healthy free-roaming zebras from sarcoidaffected parks. The presence of BPV-1 and -2 DNA in the blood of zebras from parks where sarcoids have not been previously observed was a significant finding. / Thesis (PhD)--University of Pretoria, 2010. / Veterinary Tropical Diseases / unrestricted

TCTP est une protéine de 20 kDa que l’on retrouve souvent sous forme de dimère. Elle est fortement conservée dans la phylogénie et on la trouve dans les levures, les plantes, les invertébrés et les mammifères. Elle est localisée dans tous les compartiments de la cellule : noyau, cytoplasme, et membranes. Il s’agit d’une protéine très abondante dans des cellules souches ainsi que des cellules en croissance exponentielle, y compris les cellules tumorales. Sa fonction principale est celle d’une « protéine de survie ». TCTP a été décrite comme interagissant avec de multiples protéines dont p53, MDM2, Bcl-xL et TSAP6. Le but de mon travail est de permettre de mieux caractériser ces fonctions de TCTP et pour cela, nous avons étudié ses interactions in vitro et surtout, in vivo, dans différents modèles génétiques chez la souris. / TCTP is a 20 kDa protein frequently encountered as a dimer. It is highly conserved through phylogeny and is present inn yeast, plants, invertebrates and mammals. It is localized in all compartments of the cell: nucleus, cytoplasm, membranes. This protein is highly abundant in stem cells and during the exponential growth, including in cancer cells. It mainly functions as a survivor factor. TCTP has been described as interacting with multiple proteins, including p53, MDM2, Bcl-xL and TSAP6. The purpose of my work is to better characterize these functions of TCTP; we therefore studied its interactions in vitro, but mostly in vivo, using different murine genetic models.

Dans la première partie de cette thèse, nous étudierons l’impact de l’hétérogénéité tumorale sur les phénomènes d’invasion collective des cellules cancéreuses et de dissémination métastatique.L’hétérogénéité des populations cellulaires tumorales est observée dans la plupart des lésions cancéreuses solides. Cependant, son impact sur le phénomène de métastase – élément prépondérant dans l’établissement du pronostic vital du patient – demeure à ce jour mal compris. En utilisant un modèle numérique minimal de tumeur, nous avons cherché à déterminer quel était l’impact de l’hétérogénéité des propriétés mécaniques des cellules cancéreuses sur leur invasion dans les tissus sains entourant la tumeur. Nous nous sommes particulièrement intéressés aux différences de mobilité cellulaire au sein des diverses populations cellulaires composant une tumeur. Nos travaux établissent un lien de causalité entre l’hétérogénéité tumorale et la dissémination métastatique. De plus, ils permettent de reproduire un certain nombre de morphologie d’invasion cancéreuse telles que des protrusions pluricellulaires en forme de « doigts » ou d’agrégats. Nos expériences in silico démontrent que deux mécanismes complémentaires sont à l’œuvre au sein des tumeurs hétérogènes. Une faible proportion de cellules leaders, possédant une force mobile plus élevée, est capable d’initier et de diriger l’invasion cancéreuse, alors que les effets de mouvements collectifs au sein de la tumeur fournissent la coordination mécanique nécessaire à un phénomène d’invasion collectif continu. Ces résultats suggèrent que la dynamique d’invasion collective observée durant le processus de métastase est un phénomène universel. Celui-ci est propre aux populations de cellules aux propriétés mécaniques hétérogènes, et peut être décrit en se fondant sur un nombre limité d’hypothèses physiques, et ce malgré l’importante variabilité génétique et phénotypique qui caractérise les pathologies cancéreuses.Dans la seconde partie de cette thèse, nous continuerons à étudier l’impact de l’hétérogénéité des propriétés cellulaires, cette fois à l’échelle d’un organisme pluricellulaire et non pas seulement d’un tissu. Nous nous intéresserons au développement de l’amibe sociale Dictyostelium discoideum. Lorsque les amibes sont privées de nourriture, elles forment des agrégats pluricellulaires nommés slugs,dans lesquels les cellules initialement identiques se différencient et se ségrèguent en deux populations distinctes : les cellules prespores, à l’arrière, et les cellules prestalks, à l’avant. La formation de ce motif spatial est caractérisé par une homéostasie des proportions des types cellulaires, qui demeurent quasi constants malgré les variations importantes du nombre de cellules au sein des agrégats. Si différents modèles ont été proposés pour expliquer l’origine de ce phénomène, il demeurait nécessaire de mettre en place des expériences quantitatives afin de confirmer ou d’infirmer ces modèles. Dans ce but, nous avons développé et caractérisé une nouvelle souche cellulaire de Dictyostelium, AX2-PYR, utilisant des sondes fluorescentes génétiquement encodées permettant de distinguer les différents types cellulaires au sein des slugs. Nos résultats démontrent l’invariance du motif prespore/prestalk avec la taille des slugs sur quatre ordres de grandeur, et mettent en évidence l’existence d’un mécanisme actif de régulation des proportions reposant sur les communications intercellulaires. / In the first part of this thesis, we study the impact of tumour heterogeneity on cancer collective invasion and metastatic dissemination. Heterogeneity within tumour cell populations is commonly observed in most solid tumours, but its impact on metastasis, one of the primary determinants of the disease prognosis, remains poorly understood.Working with a simplified numerical model of tumour spheroids, weinvestigate the impact of mechanical heterogeneity of tumour cells on the onset of tumour invasion into surrounding tissues, focusing more particularly on the influence of differences in cell motility. Ourwork establishes a positive link between tumour heterogeneity and metastatic dissemination, and recapitulates a number of invasion patterns identified in vivo, such as multicellular finger-like protrusionsor tumour cell clusters. In our in silico experiments, we demonstrate that two complementary mechanisms are at play in heterogeneous tumours: a small proportion of stronger cells with a higher motile force are able to initiate and lead the escape from the tumour, while collective effects in the bulk of the tumour provide the coordination required to sustain the invasive process through multicellular streaming. This suggests that the multicellular dynamics observed during metastasis is a generic feature of mechanically heterogeneous cell populations and might rely on a limited and generic set of physical assumptions shared by most tumours in spite of the genetic and phenotypic variability amongst patients and pathologies.In the second part of our work, we continue to explore the impact of heterogeneity on population scale behaviours of multicellular systems, focusing on the development of the social amoeba Dictyosteliumdiscoideum. Under starvation Dictyostelium cells form multicellular aggregates named slugs where amoeba cells differentiate and segregate into two distinct spatial zones, the prespore (rear) and prestalk (front) cells regions. This developmental pattern is characterized by an homeostasis of cell-type proportions with respect to slug size and external perturbations. Different models have been proposed to explain theorigin and regulation of this pattern, but quantitative experiments were still needed to decipher between the proposed mechanisms. To quantitatively investigate cell differentiation and spatial patterning in live multicellular aggregates, we developed and characterized a new stable cell line, AX2-PYR, using genetically encoded fluorescent reporters of cell differentiation into prespore and prestalk cells. Our results demonstrate the scaling of the prespore/prestalk pattern over more than three orders of magnitude in slug size, and show the existence of a proportion regulation mechanism which might rely on cell-cell communications.

Les sarcomes à « génétique complexe » sont des tumeurs rares du tissu mésenchymateux. Ces tumeurs sont caractérisées par un nombre important de réarrangements chromosomiques et possèdent un génome complexe dans lequel aucune altération n’avait été retrouvée de manière spécifique et récurrente. Le but de ma thèse était donc d’identifier les conséquences de cette complexité chromosomique et de comprendre comment celle-ci pouvait être générée. Dans un premier temps, le transcriptome de 112 sarcomes à « génétique complexe » a été analysé. La recherche et la validation de transcrits chimériques a conduit l’identification de réarrangements fréquents au niveau du gènes TRIO. Ces transcrits permettent la formation soit d’une protéine tronquée et seraient des évènements issus du réarrangement global du génome de ces tumeurs. Dans un deuxième temps, nous avons alors recherché l’origine de la formation de ces altérations, en s’intéressant particulièrement à la fusion cellulaire comme mécanisme initiateur. Ce processus physiologique est observé dans des cellules mésenchymateuses comme les macrophages et les myoblastes et peut être détourné afin de permettre le développement et l’évolution tumorale. J’ai alors étudié les conséquences génomiques et phénotypiques de la fusion de fibroblastes à différents stades d’immortalisation ou à différentes phases du cycle cellulaire. Mes travaux ont alors permis de montrer que des mécanismes de fusion cellulaire conduisent à la formation d’altérations génétiques similaires à celles des sarcomes à « génétique complexe » et contribueraient à l’initiation et à la progression de ces tumeurs. / Sarcomas with a complex genetics are rare tumours from mesenchymal tissue. They are characterized by massive chromosomal without any recurrent and specific alteration. The objective of my thesis was to identify the consequences of this chromosomal complexity and mechanisms explaining how this could be generated. First, transcriptome of 112 sarcomas with a complex genetics have been analysed. Chimeric transcripts detection and validation permitted the identification of frequent rearrangements in TRIO gene. These transcripts lead to the formation of a truncated protein and they would originate from a global rearrangement of the tumour genomes. Second, we have sought the origin of these alterations, with a particular interest for the cell fusion as an initiator mechanism. This physiological process is observed in mesenchymal cells like macrophages and myoblasts and it can be hijacked to drive tumour inception and evolution. I consequently studied both genomic and phenotypic consequences of hybrids from fibroblasts at different immortalization steps or in the different cell cycle phases. This work permitted to demonstrate that cell fusion mechanism leads to the initiation of genetic alterations that mimics the ones in sarcomas with complex genetics and would contribute to their tumour initiation and progression.

Premature cellular senescence is the process of permanent cell cycle arrest in response to various inducers, such as DNA damage, oxidative stress, chemotherapy agents, and irradiation. Senescent cells produce and secrete numbers of cytokines, chemokines, growth factors, which compose specific senescence-associated secretory phenotype (SASP). Senescence is considered to be an important barrier against tumor progression. On the other hand, senescent cells can also exert protumorigenic effects in their microenvironment. Based on this concept, the major aim of this thesis was to determine tumor cells senescence in terms of different inducers, namely chemotherapeutic agent docetaxel (DTX) and cytokines IFNγ and TNFα, and to demonstrate the role of immunotherapy in senescent cells elimination. Our results show that DTX-induced senescent cells can exert a tumor-promoting effect when co-injected with proliferating cells in mice. Importantly, we demonstrate that IL-12-based immunotherapy suppresses senescence-accelerated tumor growth. These results suggest that IL-12-based immunotherapy can be effectively used in anti-tumor therapy mainly in a case when the microenvironment is altered by the presence of tumor senescent cells. On the other hand, the data we obtained in vitro show that bystander or...

The pediatric population is highly impacted by brain tumours, as they are the most common type of solid tumour affecting children. Medical advances have improved the survival rate of children with brain tumours, but many survivors still experience late effects. In adulthood, 18% of pediatric brain tumour (PBT) survivors have reported cardiovascular issues such as strokes, blood clots, and angina, but little is known about the cardiovascular health of these survivors during childhood. The primary objective of this thesis was to measure the proportion of PBT survivors with values of body mass index (BMI), waist circumference (WC), and aerobic fitness meeting the cut-offs associated with unfavourable cardiovascular health (BMI≥+2 SDs, WC≥90th percentile, and % peak oxygen uptake (VO2peak) predicted<85%). The secondary objective was to observe the effects of 12 weeks of exercise training on the BMI, WC, blood pressure (BP), and aerobic fitness of PBT survivors and describe the feasibility of an exercise training program for this population. In this thesis, 32 PBT survivors who had all received cranial radiation were included (age=12.3±3.4 years, age at diagnosis=7.0±2.5 years, time since treatment completion=4.5±2.8 years, 21/32 male). While WC was measured for 13/32 participants, BMI and aerobic fitness were measured for all. Of the participants, 5/32 children completed a 12-week pilot exercise program consisting of two group and two in-home exercise sessions per week. A control group (n=2, age=14.8±3.6 years, 1/2 male) and an intervention group (n=5, age=15.0±2.3, 3/5 male) had BMI, WC, BP, and aerobic fitness measurements taken pre- and post-training. Of the participants, 15.6% (5/32) had a BMI≥+2 SDs, 30.7% had a WC≥90th percentile, and 86.7% had a %VO2peak predicted<85%. In total, 81% (26/32) of the participants had at least one identified biomarker reflecting unfavourable cardiovascular health. A training effect was only observed in BP (change of +9%). The exercise program was feasible with an adherence rate of 88% (21/24) to the group and in-home sessions. All (5/5) participants completed the program without injuries or adverse events during the training program. Findings from this thesis indicate that 81% brain tumour survivors have at least one biomarker indicating unfavourable cardiovascular health in childhood. This thesis also provides novel information to be considered before implementing exercise as a therapy for improving the cardiovascular health of survivors. Future additional research is required to determine the appropriate duration, frequency, and intensity of aerobic exercise to stimulate a training effect on these cardiovascular biomarkers. / Thesis / Master of Science (MSc)

Purpose: Tumour hypoxia is the result of uncontrolled growth of the tumour and its vasculature and is often found in solid tumours. It has been known for some time that tumour hypoxia is associated with increased radio resistance and poorer treatment outcomes. While there are several techniques to image the tumour’s oxygenation, no metric or guideline exists that helps in automatically delineating those hypoxic cells into target volumes. Even though several hypoxic biomarkers have been developed and tested to detect visualise and localise hypoxic areas, most of these delineated areas show volumes that are not immediately suitable for dose planning (i.e. a speckled hypoxia distribution). This work deals with 18 cases of tumour hypoxia in patients with non-small cell lung cancer (NSCLC) and presents a method that gives guidance on how to construct hypoxic target volumes feasible for dose planning. Materials and Methods: PET-CT scans have been taken with the hypoxic biomarker 18F-HX4. Hypoxic volumes have been extracted using a threshold of 10mmHg. A region growing algorithm was used to develop the HTV delineation method. Individually calculated doses based on the pO2-distribution within the hypoxic target volume have been used for the construction of dose plans with 24 fractions. Results: Treatment plans that boost the hypoxic target volume whilst sparing surrounding organs at risk were possible to construct for those tumours lying outside the mediastinum. Tumours which volumes were partially or fully overlapping with the mediastinum showed conflicts with delivering the dose necessary for a tumour control probability (TCP) of at least 95% and not exceeding the dose constraints set for the mediastinum.

<p>Breast cancer remains a major cause of cancer-related deaths among women despite several advances in the field due to metastasis with a 5-year survival rate of less than 30% for metastatic breast cancer. Dissemination of tumor cells to metastatic sites begins as early as the primary tumor is diagnosed at just 4mm in size. These cells remain dormant for extended periods of time evading immune surveillance and later turn into therapy resistant metastases resulting in the poor prognosis in breast cancer patients. Hence, there is a <b>critical need </b>to improve our understanding of the metastatic programs in breast cancer and its contributors to develop better therapy options.</p><p>One such contributor is alcohol which is listed as a carcinogen by the National Toxicology Program. Alcohol consumption is a risk factor for several cancers and increases the risk of breast cancer incidence in a dose dependent manner. We have observed in preliminary studies, that alcohol consumption causes increased neutrophil extracellular trap (NET) formation in the lungs and outgrowth of previously dormant cancer cells in mice. Further, NETs increase cancer cell seeding and play a role in metastasis. Hence, we hypothesized that alcohol consumption breaks cancer cell dormancy by activating neutrophils.</p><p>In this study, we have broken cancer cell dormancy and generated a novel cell line, Alcohol-D2.OR, by inducing outgrowth of the dormant D2.OR cells in mice through alcohol consumption. Reinjection of the Alcohol-D2.OR cells, into alcohol-naïve mice results in aggressive outgrowth of the cells suggesting these cells are modified on a genetic level. Indeed, RNA sequencing analysis of the gene expression in the cells showed that these cells have significantly modified gene expression as well as modified morphology and surface protein expression than the parental D2.OR cells. Importantly, from our analysis we have identified a tumor suppressor, SPINK5 which was significantly downregulated in the alcohol line. Further, SPINK5 expression in cancer cells suppressed neutrophil activity in-vitro. Knockdown of SPINK5 in the parental D2.OR line resulted in outgrowth of the cells in-vivo with increased lung NETs highlighting the importance of this gene for maintenance of dormancy by suppression of neutrophil activity.</p><p>Hence, we have successfully identified a gene responsible for dormancy maintenance, SPINK5 which will aid in not only therapeutic intervention but also in identification of breast cancer patients likely to progress to metastasis. Further, the newly established Alcohol-D2.OR cells provide a novel tool to study other initiators of metastasis in breast cancer.</p><p>A common side-effect of most chemotherapeutic treatments is neutropenia, reduced neutrophils in circulation increasing susceptibility to infections. Hence, GM-CSF is often administered to patients to mobilize bone marrow neutrophils. However, neutrophils have been increasingly shown to promote distant metastases. Circulating disseminated cancer cells (DCCs), which are present as early as primary diagnosis, have been shown to activate neutrophils resulting in the release of neutrophil extracellular traps (NETs). These NETs alter the lung architecture providing a suitable environment for the seeding and growth of DCCs promoting lung metastases. One key player in neutrophil activation is spleen tyrosine kinase (SYK), an intracellular non-receptor kinase which is activated by the engagement of b-integrin on the neutrophil surface.</p><p>Using a chemical genetics approach we are able to specifically inhibit SYK in the murine host. Using our transgenic model of specific SYK inhibition as well as the FDA approved SYK inhibitor, fostamatinib, we see similar results of decreased lung metastases compared to controls. We also observed decreased neutrophil viability in-vitro in the presence of fibronectin, an effect that was not seen on plastic highlighting the importance of integrin mediated activity of SYK. We also observe decreased neutrophil and macrophage infiltration into the lungs upon host-specific SYK inhibition. Overall, these findings suggest a paracrine effect of SYK in stromal cells that promotes favorable tumor microenvironment (TME) and its inhibition may be a useful therapeutic option to combat DCCs from forming metastases.</p><p>Hence, through this work we address two mechanisms of neutrophil-mediated breast cancer metastasis and that therapeutic intervention by rescuing SPINK5 expression in cancer cells or inhibition of SYK in the tumor microenvironment can suppress pulmonary metastasis in breast cancer.</p>