Clinical Relevance of Immune Cells in Pancreatic Adenocarcinoma and their Implication in Immunotherapy

This thesis summarizes the work I have done during my PhD training, focused on the immune microenvironment of pancreatic adenocarcinoma (PDAC). The aim was to delineate key features of immunologic and tumour components shaping the biology of pancreatic cancer. Immunotherapy has been largely explored as a promising anticancer approach but not convincingly introduced yet for this tumour type. The results here summarized could contribute to a better understanding of the microenvironment of human PDAC and foster the introduction of immunotherapeutic strategies into the clinical practice. The first section of the thesis summarizes the work done to investigate the occurrence of tertiary lymphoid tissue (TLT) in PDAC. TLT has been demonstrated to coordinate the recruitment and activation of adaptive immune cells in several tumour settings. This work documents the occurrence of TLT in pancreatic cancer, addresses the prognostic role of B cells according to their spatial distribution within TLT and tests the hypothesis that TLT can serve as immunological sites that contribute to efficacy of immunotherapeutic vaccination. The second section is focused on the intersection between cancer metabolism and immune function in PDAC. In order to unearth early metabolic alterations in PDAC, we have analyzed the pancreatic juice, a relatively unexplored liquid biopsy potentially enriched with tumor-derived metabolites. Our results indicate a specific metabolic signature that allows to single out PDAC from other pancreatic pathologies and reveal an important link between glucose metabolism and accumulation of PD-1+ cells in PDAC tumors

MICAL1 regulates actin cytoskeleton organization, directional cell migration and the growth of human breast cancer cells as orthotopic xenograft tumours

The Molecule Interacting with CasL 1 (MICAL1) monooxygenase has emerged as an important regulator of cytoskeleton organization via actin oxidation. Although filamentous actin (F-actin) increases MICAL1 monooxygenase activity, hydrogen peroxide (H2O2) is also generated in the absence of F-actin, suggesting that diffusible H2O2 might have additional functions. MICAL1 gene disruption by CRISPR/Cas9 in MDA MB 231 human breast cancer cells knocked out (KO) protein expression, which affected F-actin organization, cell size and motility. Transcriptomic profiling revealed that MICAL1 deletion significantly affected the expression of over 700 genes, with the majority being reduced in their expression levels. In addition, the absolute magnitudes of reduced gene expression were significantly greater than the magnitudes of increased gene expression. Gene set enrichment analysis (GSEA) identified receptor regulator activity as the most significant negatively enriched molecular function gene set. The prominent influence exerted by MICAL1 on F-actin structures was also associated with changes in the expression of several serum-response factor (SRF) regulated genes in KO cells. Moreover, MICAL1 disruption attenuated breast cancer tumour growth in vivo. Elevated MICAL1 gene expression was observed in invasive breast cancer samples from human patients relative to normal tissue, while MICAL1 amplification or point mutations were associated with reduced progression free survival. Collectively, these results demonstrate that MICAL1 gene disruption altered cytoskeleton organization, cell morphology and migration, gene expression, and impaired tumour growth in an orthotopic in vivo breast cancer model, suggesting that pharmacological MICAL1 inhibition could have therapeutic benefits for cancer patients

Psychological treatments and psychotherapies in the neurorehabilitation of pain. Evidences and recommendations from the italian consensus conference on pain in neurorehabilitation

BACKGROUND: It is increasingly recognized that treating pain is crucial for effective care within neurological rehabilitation in the setting of the neurological rehabilitation. The Italian Consensus Conference on Pain in Neurorehabilitation was constituted with the purpose identifying best practices for us in this context. Along with drug therapies and physical interventions, psychological treatments have been proven to be some of the most valuable tools that can be used within a multidisciplinary approach for fostering a reduction in pain intensity. However, there is a need to elucidate what forms of psychotherapy could be effectively matched with the specific pathologies that are typically addressed by neurorehabilitation teams. OBJECTIVES: To extensively assess the available evidence which supports the use of psychological therapies for pain reduction in neurological diseases. METHODS: A systematic review of the studies evaluating the effect of psychotherapies on pain intensity in neurological disorders was performed through an electronic search using PUBMED, EMBASE, and the Cochrane Database of Systematic Reviews. Based on the level of evidence of the included studies, recommendations were outlined separately for the different conditions. RESULTS: The literature search yielded 2352 results and the final database included 400 articles. The overall strength of the recommendations was medium/low. The different forms of psychological interventions, including Cognitive-Behavioral Therapy, cognitive or behavioral techniques, Mindfulness, hypnosis, Acceptance and Commitment Therapy (ACT), Brief Interpersonal Therapy, virtual reality interventions, various forms of biofeedback and mirror therapy were found to be effective for pain reduction in pathologies such as musculoskeletal pain, fibromyalgia, Complex Regional Pain Syndrome, Central Post-Stroke pain, Phantom Limb Pain, pain secondary to Spinal Cord Injury, multiple sclerosis and other debilitating syndromes, diabetic neuropathy, Medically Unexplained Symptoms, migraine and headache. CONCLUSIONS: Psychological interventions and psychotherapies are safe and effective treatments that can be used within an integrated approach for patients undergoing neurological rehabilitation for pain. The different interventions can be specifically selected depending on the disease being treated. A table of evidence and recommendations from the Italian Consensus Conference on Pain in Neurorehabilitation is also provided in the final part of the pape

What is the role of the placebo effect for pain relief in neurorehabilitation? Clinical implications from the Italian consensus conference on pain in neurorehabilitation

Background: It is increasingly acknowledged that the outcomes of medical treatments are influenced by the context of the clinical encounter through the mechanisms of the placebo effect. The phenomenon of placebo analgesia might be exploited to maximize the efficacy of neurorehabilitation treatments. Since its intensity varies across neurological disorders, the Italian Consensus Conference on Pain in Neurorehabilitation (ICCP) summarized the studies on this field to provide guidance on its use. Methods: A review of the existing reviews and meta-analyses was performed to assess the magnitude of the placebo effect in disorders that may undergo neurorehabilitation treatment. The search was performed on Pubmed using placebo, pain, and the names of neurological disorders as keywords. Methodological quality was assessed using a pre-existing checklist. Data about the magnitude of the placebo effect were extracted from the included reviews and were commented in a narrative form. Results: 11 articles were included in this review. Placebo treatments showed weak effects in central neuropathic pain (pain reduction from 0.44 to 0.66 on a 0-10 scale) and moderate effects in postherpetic neuralgia (1.16), in diabetic peripheral neuropathy (1.45), and in pain associated to HIV (1.82). Moderate effects were also found on pain due to fibromyalgia and migraine; only weak short-term effects were found in complex regional pain syndrome. Confounding variables might have influenced these results. Clinical implications: These estimates should be interpreted with caution, but underscore that the placebo effect can be exploited in neurorehabilitation programs. It is not necessary to conceal its use from the patient. Knowledge of placebo mechanisms can be used to shape the doctor-patient relationship, to reduce the use of analgesic drugs and to train the patient to become an active agent of the therapy

Experimental evaluation of different hydrodynamic modelling techniques applied to the ISWEC

The appropriate numerical modelling of a Wave Energy Converter behaviour is crucial for both the initial design phase (loadings estimations) and the productivity assessment of the real device, working in a certain installation site. The most difficult aspect to be modelled are the floater motions at resonance conditions, where motions amplitude are emphasised. Several techniques can be used, ranging from the linear Cummins' equation to fully viscous CFD simulations. This paper deals with the implementation of the 3 DOFs linear hydrodynamic model developed for the time-domain simulation of ISWEC (Inertial Sea Wave Energy Converter) device. The main advantage of the model here presented is its low computational cost. A first benchmarking between the numerical model and a series of tank testing experiments, carried out on a scaled ISWEC floater prototype, is described. Experimental results are used for the identification of the non-linearity due to the viscous effects. The novelty of the identification approach is the estimation of the non-linearity contribution along each DOF taken into account. A second comparison is done against the open source code WEC-Sim that allows the calculation of the floater instantaneous wetted surface, with the aim of obtaining a cost-benefit evaluation of the different modelling techniques

Defective apoptotic cell contractility provokes sterile inflammation, leading to liver damage and tumour suppression

Apoptosis is characterized by profound morphological changes, but their physiological purpose is unknown. To characterize the role of apoptotic cell contraction, ROCK1 was rendered caspase non-cleavable (ROCK1nc) by mutating aspartate 1113, which revealed that ROCK1 cleavage was necessary for forceful contraction and membrane blebbing. When homozygous ROCK1nc mice were treated with the liver-selective apoptotic stimulus of diethylnitrosamine, ROCK1nc mice had more profound liver damage with greater neutrophil infiltration than wild-type mice. Inhibition of the damage-associated molecular pattern protein HMGB1 or signalling by its cognate receptor TLR4 lowered neutrophil infiltration and reduced liver damage. ROCK1nc mice also developed fewer diethylnitrosamine-induced hepatocellular carcinoma (HCC) tumours, while HMGB1 inhibition increased HCC tumour numbers. Thus, ROCK1 activation and consequent cell contraction are required to limit sterile inflammation and damage amplification following tissue-scale cell death. Additionally, these findings reveal a previously unappreciated role for acute sterile inflammation as an efficient tumour-suppressive mechanism.  </p