Aprovechamiento energético del agua de las minas

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There is no doubt that the development of Imatinib has been a major triumph for the treatment of CML, continually demonstrating its significance in the clinic by inducing complete haematological response (CHR) as well as cytogenetic response (CCyR) in the vast majority of patients, while lowering the risk and rate of

progression into Blast phase (CML-BP) (Druker et al., 2006; Hochhaus et al.,2009;

Deininger et al., 2009). What’s more, since its clinical implementation many CML patients now can expect to have a life expectancy similar to that of the general population (Gambacorti-Passerini et al., 2011). Unfortunately, even with this high success rate there are still many patients for whom Imatinib treatment is inadequate or produces adverse effects, while resistant to Imatinib treatment through Bcr-Abl- dependent or -independent means has demonstrated a significant problem for treatment (Deininger et al., 2009). A cyclic trend appears to be emerging of new Bcr-Abl tyrosine kinase inhibitors (TKIs) being developed to tackle this problem, only to induce a novel mechanism of resistance themselves, requiring the need to develop more drugs further perpetuating this trend (Quintás-Cardama et al., 2007; Bixby and Talpaz, 2011). At the stem of this problem is the necessity for continual, lifelong treatment with TKIs in order to prevent disease reoccurrence due to residual disease, thereby increasing the potential for this resistance to develop. It is evident that Bcr-Abl TKI treatment alone will only ever conceal or prolong the course of this disease, with the real solution for a complete treatment of CML requiring the

removal of residual disease. This has presented a large obstacle for CML treatment, which has mounted much concern, prompting a re-evaluation of therapeutic

approach and has demonstrated the need for a greater understanding of this disease in order to development alternative strategies for treatment.

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Bcr-Abl expression is one of the key requirements for oncogenesis in CML (McLaughlin et al., 1987; Lugo et al., 1990), achieving oncogenesis in many ways by mimicking the effects of growth factor stimulation in haematopoietic stem and progenitor cells thereby inducing disease phenotype. A number of studies have shown that induced or constitutive expression of Bcr-Abl increases the intracellular levels of Reactive Oxygen Species (ROS) (Sattler et al., 2000; Kim et al., 2005; Naughton et al., 2009; Reddy et al., 2011), which are traditionally seen as the key factors for genomic instability in CML cells, inducing the development of drug resistance as well as influencing disease progression (Nowicki et al., 2004; Koptyra

et al., 2006; Rassool et al., 2007; Sallmyr et al., 2008; Nieborowska-Skorska et al.,

2012). However, these particular studies have also identified ROS production as a prerequisite for complete signalling activity downstream of Bcr-Abl, identifying them as intracellular signalling molecules, a concept which has now become widely accepted in all cell types (Rhee et al., 2005a; Rhee et al., 2005b; Toledano et al., 2010; Bae et al., 2011).

Naughton et al. (2009) demonstrated a link between this ROS production downstream of Bcr-Abl signalling and the professional ROS generators, the NADPH Oxidases (Nox). Considering Bcr-Abl activity in a way mimics growth factor

stimulation it is therefore reasonable to expect it to activate Nox proteins, which are known to be heavily influenced by signalling cascades induced downstream of growth factor stimulation (Woolley et al., 2013). What is of significance is that Nox- derived ROS have been demonstrated to be involved in a host of cellular activities in leukaemias, driving disease phenotype by increasing survival, migration,

proliferation and even differentiation (Kim et al., 2005; Naughton et al., 2009; Sardina et al., 2010; Hole et al., 2011; Reddy et al., 2011). Although a link between

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Bcr-Abl activity and Nox-mediated ROS generation was established by Naughton et

al. (2009), it was unclear how exactly Bcr-Abl was influencing this process.

Throughout this study the K562 cell line, which constitutively express Bcr- Abl, were used as a CML model. In the first part of this study these cells were used to elucidate a possible novel mechanism of regulation of Nox-dependent ROS production downstream of Bcr-Abl signalling. This work established that specific inhibition of Bcr-Abl activity reduced ROS generation which coincided with the degradation of p22phox, a membrane-bound protein essential for full activity of Nox proteins 1, 2, 3 and 4 (Ambasta et al., 2004; Ueno et al., 2005). Consequently, p22phox was demonstrated to be an important mediator of ROS production

downstream of Bcr-Abl signalling. Inhibition of GSK-3 downstream of both the

PI3K/Akt and Raf/MEK/ERK signalling pathways, both of which are activated by Bcr-Abl, was identified to be pivotal for p22phox protein maintenance. These studies thereby established a link between Nox-derived ROS and Bcr-Abl activity through the maintenance of p22phox protein levels (Landry et al., 2013). Interestingly, such control of Nox activity downstream of tyrosine kinase activity is not unique and this method of p22phox degradation and has also been demonstrated on the

reintroduction of von Hippel-Lindau tumour suppressor gene (VHL) into VHL- deficient carcinoma cells (Block et al., 2007; Block et al., 2010) and upon inhibition of the FLT3-ITD oncogene in the Acute Myeloid Leukaemia (AML) MV-411 cell line (Woolley et al., 2012).

Further examination in K562 cells demonstrated a significant role for p22phox-mediated ROS production in cell proliferation. Rather significantly enhanced proliferation is a major contributory factor to CML disease phenotype demonstrating the importance of p22phox and Nox proteins in CML pathogenesis.

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Numerous studies have also demonstrated such a role for Nox-derived ROS in cell proliferation with some studies drawing a direct link to an importance in p22phox expression and further highlighting the significance of these results in CML (Jeong

et al., 2004; Sturrock et al., 2006; Petry et al., 2006; Reddy et al., 2011). Here

p22phox-mediated ROS production was shown to influence cell proliferation by having a positive effect on the G1/S transition of the cell cycle, a role for which Nox-derived ROS has been previously highlighted (Venkatachalam et al., 2008). It was suggested that p22phox function mediated this effect by indirectly inhibiting pRb activity through oxidative inactivation of phosphatases known to be important for pRb activation, Protein Phosphatase 1 (PP1) and Protein Phosphatase 2A (PP2A).This was believed to influence Cyclin E proteins levels and therefore cell cycle progression (Rao and Clayton 2002; O’Loghlen et al., 2003; Kolupaeva and Janssens, 2013). Significantly, work by Naughton et al. (2009) has also implemented

Nox-derived ROS in the inhibition of PP1 activity downstream of Bcr-Abl

signalling.

In addition to having a positive role in cell proliferation, p22phox-mediated Nox-derived ROS was also identified in this study to be important in the overall viability of K562 cells with its removal demonstrating minor increases in the level of apoptotic cell death. Treatments utilising Bcr-Abl inhibition via Imatinib in

combination with chemotherapeutic agents have been studied as an approach to overcome the known obstacles currently facing CML treatment (Gu et al., 2005; Tseng et al., 2005; Giallongo et al., 2011; Bonifacio et al., 2012). Similarly rather than using broad spectrum drugs much research is now focused on the simultaneous targeting of Bcr-Abl in combination with pathways identified as important in CML maintenance and progression (Helgason et al., 2011; O’Hare et al., 2012). Although

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only demonstrating a minor role in the overall survival of these cells this study still identified the importance of p22phox expression in CML pathogenesis. It was evident however that p22phox inhibition alone did not demonstrate a significant enough effect to suggest it as sole treatment for CML. Indeed, combining p22phox removal with Imatinib treatment significantly enhanced the effect of Imatinib treatment. It was concluded from this work that p22phox removal in a way weakens cells, priming Bcr-Abl inhibition to produce a substantially greater effect on cell death demonstrating the clinical potential of targeting Nox proteins in combination with Bcr-Abl inhibition. This potential was further established after DPI was used in combination with Imatinib and Nilotinib, demonstrating a substantial and synergistic increase in cell death through augmentation of apoptosis. Unfortunately, the off target effects of DPI make it a less than ideal compound for therapeutic use,

highlighting the need for the development of more specific and better characterised Nox inhibitors. Regardless of this, these studies still provided the basis and

reasoning for future analysis of Nox inhibition as a potential therapeutic in CML treatment.

Nox-derived ROS production can lead to increased genomic instability (Weyemi et al., 2012a; Weyemi et al., 2012b), a phenomena which can influence progression into Blast phase (CML-BP) as well as development of TKI resistance (Nowicki et al., 2004; Koptyra et al., 2006; Rassool et al., 2007; Sallmyr et al.,

2008; Slupianek et al., 2011;Chakraborty et al., 2012; Nieborowska-Skorska et al.,

2012). Therefore, considering the effectiveness of DPI treatment in combination with Bcr-Abl inhibition there is a potential for Nox inhibitors in reducing the