ItemInhibition of the endosomal recycling pathway to overcome resistance to cancer therapies(University College Cork, 2022) Fletcher, Kelsey; Lindsay, AndrewCancer is a major public health threat with incidence and mortality rates continuing to rise every year. Breast and prostate cancer are the most diagnosed cancers in women and men, respectively. Both cancers account for roughly 30% of all cancers diagnosed in each sex. Despite the continuous development of new therapies, drug resistance is a growing problem and is a major cause of cancer treatment failure, accounting for many cancer recurrences and deaths. Therefore, new drugs and treatment regimens are urgently required to overcome this resistance. Recent findings from our own lab and others have found that inhibition of the endosomal recycling pathway may be a promising strategy to downregulate clinically relevant cell surface proteins and to overcome drug resistance. This thesis focuses on two clinically relevant hormone receptors that are strongly linked to the development and progression of breast and prostate cancer linked to disease development and progression, the estrogen receptor alpha and the androgen receptor. The aim of this project was to confirm data obtained from a reverse-phase protein array (RPPA) study performed by our lab that found that the endosomal recycling inhibitor primaquine downregulates ER-α and AR expression. We used Western blot, quantitative RT-PCR, and immunofluorescence microscopy to confirm the RPPA results. We found that another endosomal recycling inhibitor, monensin, also potently downregulates these hormone receptors and that both inhibitors synergise with tamoxifen and enzalutamide, standard-of-care therapies for breast and prostate cancer. Keywords Cancer, prostate, breast, androgen receptor, estrogen receptor, endosomal recycling pathway, primaquine, monensin, drug resistance, tamoxifen, lapatinib, enzalutamide, synergy. ItemDevelopment of patient resources for oncology patients with dysphagia and disease-related malnutrition(University College Cork, 2021-12-21) Hanna, Michelle; Ryan, Aoife; Power, Derek; Breakthrough Cancer Research; Oesophageal Cancer FundCancer is a leading cause of both morbidity and mortality globally . Additionally, its incidence is continuing to rise year on year. Thus the development of treatments and supports to tackle and reduce cancer morbidity and mortality is of crucial importance. Current research highlights that the loss of body weight, and significantly lean body mass, contributes to increased risk of mortality in those with cancer [2, 3]. Cancer itself and its treatments can cause a variety of debilitating side-effects. These can in turn affect a person’s appetite and their ability to chew and swallow normally. Those suffering from cancers of the head and neck and upper gastrointestinal tract are particularly susceptible to dysphagia (difficulty swallowing), often related to the location of the tumour and its treatment . These negative effects on appetite and normal eating lead to reduced food intake and quality of life, significant weight loss and malnutrition. A supportive, multifaceted, evidence-based approach from an early stage is required to attenuate the development of malnutrition and thus minimise its detrimental impact on treatment outcomes and quality of life. Involuntary weight loss affects between 50-80% of those with cancer and is associated with reduced quality of life, psychological implications, increased treatment-related complications and poorer survival . Currently, no known cure exists for cancer-induced weight loss. The European Society for Clinical Nutrition and Metabolism (ESPEN) guidelines for cancer patients recommend nutrition counselling as the first line of nutrition therapy, and state that the best way to increase energy and protein intake is with normal food: a food-first approach . Prior to the publication of ‘Good Nutrition for Cancer Recovery’ by University College Cork in 2014 no evidence-based resource existed that provided patients with understandable and reliable nutritional information and nourishing recipes to help attenuate cancer-induced weight loss through a food-first approach. A 130-page, evidence-based resource containing 52 high protein, high calorie recipes was created but has been out of print since 2018. To address this, as part of this thesis, a second expanded edition was created: a 250+ page resource containing information, advice, and a bank of 116 nourishing recipes. This is described in chapter 2. All recipes are fortified with high-protein, high-calorie additions and portion sizes were adjusted to ensure meals were small in volume for those with a reduced appetite. The revision of the original recipes also allowed for an enhanced protein to fat ratio and all recipes comply with the energy and protein goals set by the British Dietetic Association (BDA) for nutritionally vulnerable patients . Patients with solid tumours of their upper gastrointestinal tract are especially vulnerable to cancer-induced weight loss, exacerbated by dysphagia . Difficulty chewing and swallowing reduce dietary intake and puts this population at an increased risk of malnutrition. In 2016, a novel written resource, ‘Eating Well with Swallowing Difficulties in Cancer’ was developed by University College Cork, aiming to help attenuate malnutrition and sarcopenia in cancer patients experiencing dysphagia. It consisted of 59 nourishing, texture-modified recipes and evidence-based nutrition advice and information based on the ‘Irish Consistency Descriptors for Modified Fluids and Food (2009)’ [APPENDIX 4]. Ireland’s changeover in 2019 to the International Dysphagia Diet Standardisation Initiative (IDDSI) led to changes in framework structure, terminology used and testing methods, thus all dysphagia resources required updating for compliance. IDDSI is a global initiative providing universal terminology describing food texture and fluid thickness, as used in dysphagia diets . Chapter 3 of this thesis discusses the development of the written resources aimed to help optimise oral intake and attenuate weight loss in head and neck and upper gastrointestinal cancer patients experiencing dysphagia, according to the IDDSI framework. It outlines the creation of appealing recipes that are nourishing and texture-modified for IDDSI levels 3 and 4. These resources aim to help those with cancer in achieving nutrition requirements with all recipes developed to comply fully with the appropriate IDDSI level. It is hoped that these cookbooks with help both patients and their families cope with the challenges of a texture- modified diet and that they will become a useful resource for dietitians and other health care professionals in providing advice on IDDSI meals. Recipes were fortified with high protein, high calorie additions; portion sizes were adjusted to low-volume for easier eating, and texture was altered to meet modified texture guidelines. Thousands of copies of these resource will be printed and distributed to oncology centres nationwide in order to disseminate this knowledge, free of charge, to cancer patients. Chapter 4 of this thesis describes the development of an evidence-based cancer information and support website. With so much unfiltered information available, there is often a lot of confusion around what to believe in relation to diet and cancer. This can leave people extremely vulnerable to misleading information at what is already a very difficult time in their lives. To address this, a one-stop-shop evidence-based website was created. The aim of this website was to dispel the most common myths and misinformation on diet and cancer and provide simplified evidence-based information for people throughout their cancer journey. The main sections covered included problems which may affect eating, cancer prevention, cancer survivors, dysphagia and cancer myths and misconceptions. The overall conclusions are summarised in Chapter 5. Malnutrition is a condition with a multifactorial aetiology and thus must be treated with an effective multimodal approach. However, this is outside the scope of this thesis which deals with its nutritional aspect. Dietary intake may be optimised by providing patients with a resource of high-calorie high-protein, nourishing recipes to improve oral intake and, potentially, attenuate loss of weight and muscle mass – such as the cookbook ‘Good Nutrition for Cancer Recovery’. For those with cancer experiencing swallowing difficulties in addition to malnutrition, separate resources with specific tailored advice, based on IDDSI guidelines, were published. Prior to these booklets being published, no resource existed to tackle the challenging task of creating high protein high calorie recipes which were also texture-modified according to the IDDSI guidelines. To support both projects and patients throughout their cancer journeys, a comprehensive website with easily accessible evidence-based information was created. ItemSensitising pancreatic cancer cell lines to electrochemotherapy by modulation of the cell cycle(University College Cork, 2021-03) Cooke, Katie Deniese; Forde, Patrick; Collins, Dearbhaile; Breakthrough Cancer ResearchWe chose to research pancreatic cancer as pancreatic adenocarcinoma (PAC) is quite the aggressive disease, which even with the vast improvements in cancer treatments over recent years still has limited treatment options. Monotherapy treatment of pancreatic cancer with chemotherapy drugs initially show hopeful results, decreased disease symptoms and can lead to a slight improvement in survival but unfortunately does often ends in recurrence (1–3). Surgery has shown to be successful for respectable tumours followed by chemotherapy. Chemotherapy alone has proven to be associated with chemoresistance and can be quite intense, often patients are not able to continue with strong treatment regimes. FOLFIRINOX is classed as the gold standard for treating PAC, however, patient fitness ultimately determines suitability. Electrochemotherapy (ECT) is a new and emerging cancer therapy which allows for decreased doses of chemotherapy drugs to be used. We have evaluated pancreatic cancer cell lines response to low dose chemotherapy and Electrochemotherapy. The human pancreatic cell lines BxPC-3, PANC-1, and MIAPaCa-2 were used for all experiments in this thesis. Based on previous studies we predicted that PANC-1 cells would be most resistant to drug treatment followed by BxPC-3 and MIAPaCa-2 cells. All cell lines survived treatment with Gemcitabine (GEM) and Nab-paclitaxel (Nab), however, they had decreased survival when treated with 5’Fluorouracil (5’FU). When Electrochemotherapy (ECT) was applied to the cell line using GEM, Nab and 5’FU, cells survived and recovered but again had decreased survival and recovery with 5’FU ECT. Cells were also treated with Oxaliplatin (Ox) ECT, this had more of an impact to cell survival than with GEM/Nab/5’FU ECT. Bleomycin and cisplatin are the conventional drugs used with ECT due to their effectiveness (4,5), but we chose Ox as it is part of the FOLFIRONOX cocktail which is the gold standard for treating patients with PAC. Electrochemotherapy (ECT) has emerged as a promising treatment strategy for patients with pancreatic cancer (PC). ECT is safe for patients and has reduced side effects of systemic chemotherapy as drugs are concentrated to treatment site(6). ECT has shown to be very effective in treating melanoma (7). ECT has recently been used to treat locally advanced PC (8). Development of preclinical and clinical agents that inhibit cell cycle progression have proven effective in the treatment of cancer. Targeting cyclin dependent kinases (CDKs) and cell cycle checkpoint proteins can induce cell cycle arrest and apoptosis in cancer cells. Cell cycle deregulation is regularly seen in pancreatic cancer. Cells grow out of control and can lead to mortality. Controlling the deregulation with cell cycle inhibitors has shown to be beneficial for survival. Our study aimed to investigate CDK4/6 inhibitors and their ability to inhibit cell cycle progression. We specifically chose CDK4/6 inhibitors as they have proven to be quite effective in previous studies of breast cancer(9). Our hypothesis was that using a cell cycle inhibitor as a pre-treatment to ECT would make cells more susceptible to treatment. We used Palbociclib (PAL), a CDK4 inhibitor which causes cell cycle arrest at G1 and prevents cells progressing to S phase. We carried out experiments to investigate CDK4 levels in cells treated with PAL and cell cycle profiles of these cells post treatment. PAL prevents cell from passing to S phase of the cell cycle. We could not finish our planned work due to time, ceased funding and the COVID-19 global pandemic. We theorise that this combination treatment could enhance the efficiency of ECT and could potentially be translated into a clinical study as a new treatment option and improve patient quality of life.