Using GEMMs to investigate novel therapeutic strategies for colorectal cancer

Despite recent advances in the clinic to integrate novel targeted therapeutic agents into standard therapy, colorectal cancer (CRC) remains a significant cause of mortality. The high attrition rate of novel compounds at phase III clinical trials for CRC, has been attributed to limited information from pre-clinical strategies, in particular, the use of inadequate xenograft models. In response, this thesis aimed to utilise robust and relevant genetically engineered mouse models of CRC to evaluate a number of novel therapeutic strategies. Whilst mutations in the tumour suppressor APC are crucial for initiation of CRC, mutations which lead to activation of the PI3K and MAPK signalling pathways, such as through loss of the tumour suppressor protein PTEN and activation of oncogenic KRAS, have been implicated in promoting progression of CRC. As such, combinations of these genetic alterations within the murine intestine, using the Cre-LoxP system, lead to differing mouse models of invasive intestinal adenocarcinoma. This thesis reports therapeutic targeting of the PI3K and MAPK signalling pathways using the dual PI3K and mTOR inhibitor NVP-BEZ235, and the MEK inhibitor MEK162, respectively. For this, compounds were initially evaluated for pharmacodynamic and anti-tumour effects through short term exposure experiments. These analyses yielded a range of effects, some of which appeared predictive of long term efficacy, others which were contradictory and some which revealed novel feedback mechanisms. Furthermore, these agents were assessed in a long term therapeutic setting to evaluate the effect of continuous treatment on longevity and tumour burden of tumour models. Here, whilst dual PI3K/mTOR inhibition significantly increased longevity of all mouse models, MEK inhibition was only effective in the Apc and Apc Kras mutant settings, identifying Pten loss as a marker of non-response to MEK inhibition, independently and also in the Kras mutant setting. Furthermore, analysis of the combination therapy in short term settings identified scheduling of these agents to be key to achieve concomitant pathway inhibition, particularly in the Apc Pten deficient tumour setting. Ultimately, when evaluated in a long term setting, although the combination therapy displayed no further benefit in the Pten deficient setting, this had additive benefits in the Kras mutant setting and synergistic benefits in the Pten Kras mutant setting. Nevertheless despite the promise of targeted therapy, standard chemotherapeutic agents such as 5-flurouracil (5-FU) remain the backbone of therapy for CRC, regardless of only moderate benefits of 10-15% in advanced tumour settings. Furthermore, resistance to 5-FU predominantly through upregulation of the enzyme thymidylate synthase (TS) frequently 2 occurs in human tumours. Investigations reported here aimed to target tumours with upregulated TS using novel analogues of the anti-viral agent Brivudin (BVDU), metabolites of which are converted to anti-cancer metabolites by TS. Initially, In vitro characterisation of a small library of compounds reported here identified a number of potent compounds. Following further in vitro characterisation and short term evaluation in the Apc Pten tumour model of invasive adenocarcinoma, two lead compounds: CPF472 and CPF3172 were taken forward for long term experiments. Subsequently, this study evaluated and identified compounds which showed increased efficacy in the TS upregulated setting. Taken together, the investigations presented in this thesis highlight the utility of appropriate mouse models in evaluating novel therapeutic strategies and generating clinically relevant hypotheses

Dynamic Performance Improvement Of Induction Motor Using Radr Controller

The primary concern of researchers is the control of induction motor. Proportional-integral- derivative (PID) control and direct torque control (DTC) have been proposed for induction motors [1-6]. Due to the following issues control of induction motor is challenging: 1) the dynamic system of an induction motor is highly nonlinear; 2) the rotor resistance varies because of heating. In this paper, RADR control with reduced order extended state observer (RESO) is applied for induction motor control. For controlling a slip ring induction motor the RADAR control is employed. All the three phases are assumed to be identical and symmetrically located with respect to each other. In this paper Induction Motor Model Responses with RADR and DTC control are compared in  MATLAB/SIMULINK

Physiological and Hematological Profiles among the Highlanders of Mount Kinabalu

Previous studies show that people residing at the highlands had the potential in physical endurance. The main objective of this study was to obtain the physiological and haematological profile of the highlanders residing at Mount Kinabalu.  27 male and four female porters and guides with age ranged from 17 to 51 years old working at the Sabah National Park voluntarily participated in the study. Anthropometric measurement, physiological attributes and hematological components were measured. Even though the participants had high haemoglobin levels, their VO2max were below the average level. Follow-up studies should consider assessment of other physiological factors at higher altitudes.Keywords: highlanders; endurance; physiological attributes; hematological componentseISSN 2398-4279 © 2018. The Authors. Published for AMER ABRA cE-Bs by e-International Publishing House, Ltd., UK. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/). Peer–review under responsibility of AMER (Association of Malaysian Environment-Behaviour Researchers), ABRA (Association of Behavioural Researchers on Asians) and cE-Bs (Centre for Environment-Behaviour Studies), Faculty of Architecture, Planning & Surveying, Universiti Teknologi MARA, Malaysia.

Human resources and models of mental healthcare integration into primary and community care in India: Case studies of 72 programmes

Background: Given the scarcity of specialist mental healthcare in India, diverse community mental healthcare models have evolved. This study explores and compares Indian models of mental healthcare delivered by primary-level workers (PHW), and health workers’ roles within these. We aim to describe current service delivery to identify feasible and acceptable models with potential for scaling up. Methods: Seventy two programmes (governmental and non-governmental) across 12 states were visited. 246 PHWs, coordinators, leaders, specialists and other staff were interviewed to understand the programme structure, the model of mental health delivery and health workers’ roles. Data were analysed using framework analysis. Results: Programmes were categorised using an existing framework of collaborative and non-collaborative models of primary mental healthcare. A new model was identified: the specialist community model, whereby PHWs are trained within specialist programmes to provide community support and treatment for those with severe mental disorders. Most collaborative and specialist community models used lay health workers rather than doctors. Both these models used care managers. PHWs and care managers received support often through multiple specialist and non-specialist organisations from voluntary and government sectors. Many projects still use a simple yet ineffective model of training without supervision (training and identification/referral models). Discussion and conclusion Indian models differ significantly to those in high-income countries—there are less professional PHWs used across all models. There is also intensive specialist involvement particularly in the community outreach and collaborative care models. Excessive reliance on specialists inhibits their scalability, though they may be useful in targeted interventions for severe mental disorders. We propose a revised framework of models based on our findings. The current priorities are to evaluate the comparative effectiveness, cost-effectiveness and scalability of these models in resource-limited settings both in India and in other low- and middle- income countries

Assessment of the In Vivo

The objective of tailoring medicines for cancer patients according to the molecular profile of their disease holds great promise for the improvement of cancer therapy. Nevertheless, this approach has been limited, in part, due to the lack of predictive and informative preclinical studies. Herein, we describe an assessment of the therapeutic potential of targeting PI3K/mTOR and MAPK signaling in genetically defined mouse models of colorectal cancer mirroring disease subtypes targeted for novel therapy in the FOCUS4 trial. Our studies demonstrate that dual PI3K/mTOR inhibition is highly effective in invasive adenocarcinoma models characterized by combinatorial mutations in Apc and Pten; Apc and Kras; and Apc, Pten and Kras. MEK inhibition was effective in the combinatorial Apc and Kras setting, but had no impact in either Apc Pten mutants or in Apc Pten Kras triple mutants. Furthermore, we describe the importance of scheduling for combination studies and show that although no additional benefit is gained in Apc Pten mice, combination of PI3K/mTOR and MAPK inhibition leads to an additive benefit in survival in Apc Kras mice and a synergistic increase in survival in Apc Pten Kras mice. This is the first study using robust colorectal cancer genetically engineered mouse models to support the validity of PI3K/mTOR and MEK inhibitors as tailored therapies for colorectal cancer and highlight the potential importance of drug scheduling in the clinic

Colorectal tumors require NUAK1 for protection from oxidative stress

The authors wish to thank the staff of the CRUK Beatson Institute Biological Services Unit for animal husbandry and assistance with in vivo experiments; the staff of the CRUK BI Histology core facility and William Clark of the NGS core facility; David McGarry, Rene Jackstadt, Jiska Van der Reest, Justin Bower and Heather McKinnon for many helpful discussions, and countless colleagues at the CRUK BI and Glasgow Institute of Cancer Sciences for support; Prem Premsrirut & Mirimus Inc. for design and generation of dox-inducible Nuak1 shRNA expressing mice Nathanael Gray for initial provision of NUAK1 inhibitors. Funding was provided by the University of Glasgow and the CRUK Beaton Institute. J.P. was supported by European Commission Marie Curie actions C.I.G. 618448 “SERPLUC” to D.J.M.; N.M. was supported through Worldwide Cancer (formerly AICR) grant 15-0279 to O.J.S. & D.J.M.; B.K. was funded through EC Marie Curie actions mobility award 705190 “NuSiCC”; T.M. was funded through British Lung Foundation grant APHD13-5. The laboratories of S.R.Z. (A12935), O.J.S. (A21139) and M.D. (A17096) are funded by Cancer Research UK. O.J.S. was additionally supported by European Research Council grant 311301 “ColoCan”.Peer reviewedPostprin

Targeted Genomic Sequencing of TSC1 and TSC2 Reveals Causal Variants in Individuals for Whom Previous Genetic Testing for Tuberous Sclerosis Complex Was Normal

Tuberous sclerosis complex (TSC) is caused by inactivating variants in TSC1 and TSC2. Somatic mosaicism, as well as the size and complexity of the TSC1 and TSC2 loci, makes variant identification challenging. Indeed, in some individuals with a clinical diagnosis of TSC, diagnostic testing fails to identify an inactivating variant. To improve TSC1 and TSC2 variant detection, we screened the TSC1 and TSC2 genomic regions using targeted HaloPlex custom capture and next-generation sequencing (NGS) in genomic DNA isolated from peripheral blood of individuals with definite, possible or suspected TSC in whom no disease-associated variant had been identified by previous diagnostic genetic testing. We obtained &gt;95% target region coverage at a read depth of 20 and &gt;50% coverage at a read depth of 300 and identified inactivating TSC1 or TSC2 variants in 83/155 individuals (54%); 65/113 (58%) with clinically definite TSC and 18/42 (43%) with possible or suspected TSC. These included 19 individuals with deep intronic variants and 54 likely cases of mosaicism (variant allele frequency 1-28%; median 7%). In 13 cases (8%), we identified a variant of uncertain significance (VUS). Targeted genomic NGS of TSC1 and TSC2 increases the yield of inactivating variants found in individuals with suspected TSC.</p

Genomic investigations of unexplained acute hepatitis in children

Since its first identification in Scotland, over 1,000 cases of unexplained paediatric hepatitis in children have been reported worldwide, including 278 cases in the UK1. Here we report an investigation of 38 cases, 66 age-matched immunocompetent controls and 21 immunocompromised comparator participants, using a combination of genomic, transcriptomic, proteomic and immunohistochemical methods. We detected high levels of adeno-associated virus 2 (AAV2) DNA in the liver, blood, plasma or stool from 27 of 28 cases. We found low levels of adenovirus (HAdV) and human herpesvirus 6B (HHV-6B) in 23 of 31 and 16 of 23, respectively, of the cases tested. By contrast, AAV2 was infrequently detected and at low titre in the blood or the liver from control children with HAdV, even when profoundly immunosuppressed. AAV2, HAdV and HHV-6 phylogeny excluded the emergence of novel strains in cases. Histological analyses of explanted livers showed enrichment for T cells and B lineage cells. Proteomic comparison of liver tissue from cases and healthy controls identified increased expression of HLA class 2, immunoglobulin variable regions and complement proteins. HAdV and AAV2 proteins were not detected in the livers. Instead, we identified AAV2 DNA complexes reflecting both HAdV-mediated and HHV-6B-mediated replication. We hypothesize that high levels of abnormal AAV2 replication products aided by HAdV and, in severe cases, HHV-6B may have triggered immune-mediated hepatic disease in genetically and immunologically predisposed children

Quantico and Intersectionality

Quantico aired September 2015 and has garnered a steady following of viewers, curious to see how the show will progress. The show follows Alex Parrish (Priyanka Chopra), a half Indian and half Caucasian, recruit for the FBI being framed for a terrorist attack in New York City. Along with Parrish there are about a half dozen other characters portrayed in a very intersectional way through their race, gender, or status on the show. Chopra’s character is particularly fascinating because she is the lead of the show but as a half Indian she is not portrayed as other Indians are in the media. In various scenes Parrish mentions some of the stereotypes or perceptions that she is faced with because of her identities and roles. I will explore how the concept of the mythical norm is portrayed on the show Quantico along with the idea of the traffic intersection as explained by Kimberle Crenshaw. Both are clearly visible throughout the show not just with the lead character, Alex Parrish, but also other leads. I will evaluate how the basement theory (also by Crenshaw but explained by Anna Carastathis) relates to the lead characters of Quantico, particularly Alex Parrish, and explore how the basement theory is also shown in various ways on the show. Finally I will review the benefits of having a show like Quantico that portrays more diversity and backgrounds for a greater variety and range of characters