Book Chapters / Κεφάλαια Βιβλίου

Recent attempts to develop nanoparticulate systems for diagnosis and/or therapy of neurodegenerative diseases, with main focus on Alzheimer's Disease (AD) are presented. Since the main obstacle to treat brain-located pathologies is the bloodbrain barrier, a brief description of its physiology, and methodologies used for studying transport of drugs across the BBB, are mentioned. All types of nanoparticulates which have been employed to-date to target AD (and deliver drugs or imaging substances to AD-related pathological features) are described, and the results accomplished so far together with advantages/disadvantages of each specifi c category of nanoparticles are mentioned. The philosophy and main physicochemical characteristic prerequisites of nanoparticles used as systems to target diseases in general, and brain-located pathologies particularly, are analyzed. Finally, the main current accomplishments and challenges for the future are summarized.

Metastasis is the cause of almost all cancer-related deaths. It is an extremely complex, multistep process defined as the spreading of cancer cells from their primary site to distant tissues. Once metastasis occurs it causes catastrophic damage to the critical organs, which is ultimately detrimental to patients. Collective efforts of many scientists have revealed the underlying molecular mechanisms of metastasis by a considerable extent, but there is still a colossal job to be undertaken by researchers to solve this life-threatening health problem. In this chapter, metastasis is explained by focusing on underlying molecular pathways. We define the steps that a cancer cell needs to climb in order to metastasize and discuss the significant molecular actors aberrantly regulated during this process. First, we outline how these molecules are deregulated in cancer cells in order to circumvent natural barriers against metastasis. Then, we give a molecular explanation on why some cancer types metastasize to certain organs. Lastly, we look into recent therapeutic trials, which involve targeting of pathways in the metastatic cascade (Figure 18.1).

The notion that breast cancer is not a single disease but many – that there is considerable heterogeneity among different tumors – is not new, having been observed by physicians for decades. But with the advent of powerful analytical and computational tools, systems biology has provided a fascinating view of the underlying mutations, biology, and networks that drive the process of breast tumorigenesis. The information derived from analyzing these networks is also proving to be clinically useful and is beginning to be incorporated into the standard clinical management of breast cancer patients – to determine prognosis and guide the choice of therapeutics, for example. However, these advances are complicated by the heterogeneity of cancer cell phenotypes that exist within a single tumor, which drives metastasis, resistance to treatment, and, eventually, recurrence. Systems biology offers a strong tool for investigating both levels of heterogeneity to comprehensively define, and ultimately attack, the aberrant molecular networks governing breast cancer cells. Epidemiology Breast cancer is a staggering public health problem. The American Cancer Society expects that over 232,000 cases of invasive breast cancer will be diagnosed in 2014, accounting for the largest number of cancer cases in women (ACS, 2014). Nearly 40,000 deaths are predicted to occur, which rank it the second leading cause of cancer deaths in women, behind only lung cancer. Fortunately, the number of newly diagnosed cases has decreased since 2000, largely as a result of a reduction in number of post-menopausal women on hormone replacement therapy, which has been strongly linked to the development of breast cancer (Rossouw et al., 2002). Death rates have also decreased in recent years and this has been attributed to improvements in screening and early detection with mammography as well as better treatment options (ACS, 2012). However, racial disparities do exist as African American women have a lower incidence rate but an increased chance of death (ACS, 2011). While this may reflect socioeconomic factors and access to health care, there also seems to be a biological difference since African American women are more likely to be diagnosed with aggressive cancers (Amend et al., 2006; Stead et al., 2009). Additionally, over 62,000 cases of carcinoma in situ are expected in 2014 (ACS, 2014). These are non-invasive neoplasms and likely represent a pre-malignant stage in the progression toward invasive breast cancer (Burstein et al., 2004).

Atomic force microscope (AFM) is a powerful and invaluable tool for imaging and probing the mechanical properties of biological samples at the nanometric scale. The importance of nano-scale characterization and nanomechanics of soft biological tissues is becoming widely appreciated, and AFM offers unique advantages in this direction. In this chapter, we describe the procedure to collect data sets (imaging and mechanical properties measurement) of collagen gels and tumor tissues. We provide step-by-step instructions throughout the procedure, from sample preparation to cantilever calibration, data acquisition, analysis, and visualization, using two commercial AFM systems (PicoPlus and Cypher ES) and software that accompanied the AFM systems and/or are freeware available (WSxM, AtomicJ). Our protocols are written specifically for these two systems and the mentioned software; however, most of the general concepts can be readily translated to other AFM systems and software.

Cancer progression is modulated by aberrant expression and secretion of cytokines at various stages. TGFβ and BMP belong to a superfamily of around 40 secreted cytokines that regulate a plethora of biological responses in normal as well as in cancer cells. Studies have shown that these molecules can regulate a large number of processes such as cell proliferation, apoptosis, senescence, differentiation, angiogenesis, immunosuppression, cell migration, and cancer metastasis. Recent studies have shed light into the molecular mechanisms and signaling networks that govern the effects of these pivotal pathways during cancer progression. Therefore it is becoming increasingly clear that unraveling the mechanistic complexity and clinical relevance of these pathways will greatly enhance our therapeutic efforts against tumor development and evolution of malignant cells. Most of the knowledge pertaining to the TGFβ superfamily of cytokines has been elucidated from studies regarding the TGFβ isoforms. There are three TGFβ isoforms, TGFβ1, TGFβ2, and TGFβ3, which are initially synthesized as inactive 75-kDa homodimeric pro-proteins, known as pro-TGFβ. These propeptides, referred to as the latency-associated proteins (LAPs), are part of the TGFβ large latent complex (LLC) which consists of LAPs and latent TGFβ binding proteins (LTBPs) assembled together by the formation of disulfide bonds between cysteine residues [1–3]. LTBPs are members of the LTBP/fibrillin protein family, which consists of fibrillin-1, 2, and 3 as well as LTBP-1, 2, 3, and 4. Out of these proteins, LTBP-1, 3, and 4 have the unique ability to bind LAP through the third of their four 8-cystein domains [4]. The remaining cysteine domains are likely to localize LTBPs to the extracellular matrix (ECM) [5]. As a part of the LLC, TGFβ remains in an inactive form. In this state, LAPs form a non-covalent, high-affinity association with TGFβ preventing the receptor–ligand interaction [6]. LLC is primarily localized at the matrix via covalent association of the N-terminal region of LTBPs with ECM proteins [7]. During the activation step, LAPs undergo conformational changes induced by thrombospondin-1 (TSP-1) [8, 9] and cleavage by furins and other convertases leading to the release of the mature 24-kDa TGFβ dimer [10, 11], which can bind to and activate TGFβ receptors resulting in the propagation of downstream signaling events.