TUMOR MICROENVIRONMENT IN EXPERIMENTAL PRECLINICAL MOUSE MODELS OF HUMAN CANCER: MORPHOLOGICAL APPROACH

One of the recent advancements in oncological research has been the recognition of the tumor microenvironment (TME) as a relevant participant during all stages of the evolution of a neoplastic process. Indeed, over the past decades, tumors have been considered through a changing perspective: no longer as a growth of homogeneous neoplastic cells, but as an actual organ composed of different cell populations and structures: the parenchyma being the neoplastic population and the stroma, including the vascular network and infiltrating cells. The tumor microenvironment has a dual role in tumor biology, both promoting and antagonizing tumor development, growth, and local or distant invasiveness. According to its leading role in influencing tumor biology each component of the TME could be considered as a potential pharmacological target to be enhanced or antagonized, in order to influence tumor behavior. Accordingly, the study of the TME could provide new insights in the tumor biology and offers numerous potential targets for the development of novel therapeutic strategies. In this context, morphological techniques represent useful tools for the investigation of the TME, allowing the evaluation of the spatial distribution of the different elements, and provide useful complementary information to clinical and other data obtained in experimental in vivo studies. In this thesis, the three main classes of the TME components -tumor-associated vasculature, immune-inflammatory cells and tumor stroma- are illustrated in three different chapters and relevant experimental studies described. However, it should be considered that the various aspects of TME are not separate entities but are all involved in a dynamic system with complex structural and functional interactions. Chapter 1 – Tumor-associated vasculature Tumor angiogenesis has been identified as a hallmark of cancer, due to its central role in supporting tumoral growth, providing nutrient supply, removing catabolites and enabling tumoral metastatic dissemination. Most of the solid tumors are characterized by an “angiogenetic switch” in which an imbalance between pro- and anti-angiogenic factors sustains a dysregulated angiogenetic process, leading to the formation of an altered vascular network composed of structurally and functionally abnormal blood vessels. Drugs targeting tumor vasculature has been extensively studied as a mean to interfere with tumoral growth as well as to promote the delivery and/or effect of co-administered compounds to the tumor. In the first study of this chapter, we demonstrated the therapeutic efficacy and the antiangiogenic effect of a novel compound developed by binding sunitinib (a well-known antiangiogenic drug) to a selective binder of αVβ3 integrin thus promoting its delivery to the target site (tumors expressing αVβ3 integrin). The other studies of this chapter investigated the relation between tumor vasculature and tumor hypoxia. In particular, this relation was investigated to uncover the potential mechanism underlying the synergistic effect of the administration of an antiangiogenic compound (cediranib) with a poly-ADP ribose polymerase (PARP) inhibitor (olaparib) in a panel of patient-derived xenografts of ovarian carcinoma. Chapter 2 – Tumor immune microenvironment In most cancers, both innate and acquired immunity have a driving role during all stages of tumor development and progression. Depending on the cell population and/or molecular stimuli received, they can act in a dual way, antagonizing or promoting tumor growth. Three selected studies were described in chapter 2 and investigated: 1. The role of NK cells in hindering metastasis engraftment in a metastatic model of synovial sarcoma. After the combined administration of an heparanase-inhibitor with a tyrosine kinase inhibitor a significant reduction of lung metastases was observed and immunohistochemical analyses demonstrated the role of NK cells in this phenomenon. 2. The macrophage polarization status in a panel of xenotransplanted thyroid carcinoma tumors. The mononuclear-phagocyte populations infiltrating the tumors were evaluated by immunohistochemistry. 3. The role of inflammation in the development of colorectal cancer was evaluated in mice (wild type and EMILIN1-mutant), undergoing administration of AOM-SS (chemical carcinogenesis model). EMILIN1 mutant mice developed more numerous and more severe tumoral lesions compared to wild type, as well as increased inflammatory infiltrate was observed, unveiling a potential contribution of Emilin 1 in the pathogenesis of colorectal adenocarcinoma. Chapter 3 – Tumor stroma Tumor stroma represents not only the scaffold in which tumors growth, but also an intricate network of molecules and signals influencing tumor biology. The first study of this chapter investigated stroma-derived circulating molecules as a potential tool for the early diagnosis of pancreatic ductal adenocarcinoma (PDAC). Selected molecules (MMP-7, TIMP-1 and Throbospondin-2) were tested in KC genetically engineered mice (modeling the early stages of PDAC development) and patient-derived xenografts (modeling tumor progression), by serum ELISA and by immunohistochemistry. The second study evaluated the potential improvement in the biodistribution of chemotherapeutic drugs derived from the combined treatment with hyaluronidase. Tumor-bearing mice (ovarian carcinoma and pancreatic carcinoma models) were treated with chemotherapy alone (paclitaxel) or combined with hyaluronidase. Hyaluronidase treatment reduced the amount of stromal hyaluronic acid (as demonstrated by Alcian blue stain) and improved intratumor distribution of paclitaxel (as analyzed by mass spectrometry).