The regulation of cancer cell invasive behaviour by Chloride Interacellular Channel 3 (CLIC3) and Transglutaminasae 2

A study into the role of secreted CLIC3 in tumour cell invasion. The initiation and progression of cancers is thought to be linked to their relationship with a population of activated fibroblasts, which are associated with tumours. I have used an organotypic approach, in which plugs of collagen I are preconditioned with fibroblastic cells, to characterise the mechanisms through which carcinoma-associated fibroblasts (CAFs) influence the invasive behaviour of tumour cells. I have found that immortalised cancer-associated fibroblasts (iCAFs) support increased invasiveness of cancer cells, and that this is associated with the ability of CAFs to increase the fibrillar collagen content of the extracellular matrix (ECM). To gain mechanistic insight into this phenomenon, an in-depth SILAC-based mass proteomic analysis was conducted, which allowed quantitative comparison of the proteomes of iCAFs and immortalised normal fibroblast (iNFs) controls. Chloride Intracellular Channel Protein 3 (CLIC3) was one of the most significantly upregulated components of the iCAF proteome. Knockdown of CLIC3 in iCAFs reduced the ability of these cells to remodel the ECM and to support tumour cell invasion through organotypic plugs. A series of experiments, including proteomic analysis of cell culture medium that had been preconditioned by iCAFs, indicated that CLIC3 itself was a component of the iCAF secretome that was responsible for the ability of iCAFs to drive tumour cell invasiveness. Moreover, addition of soluble recombinant CLIC3 (rCLIC3) was sufficient to drive the extension of invasive pseudopods in cancer cell lines, and to promote disruption of the basement membrane in a 3D in vitro model of the ductal carcinoma in situ (DCIS) to invasive ductal carcinoma (IDC) transition. My investigation into the mechanism through which extracellular CLIC3 drives tumour cell invasiveness led me to focus on the relationship between CLIC3 and the ECM modifying enzyme, transglutaminase-2 (TG2). Through this, I have found that TG2 physically associates with CLIC3 and that TG2 is necessary for CLIC3 to drive tumour cell invasiveness. These data identifying CLIC3 as a key pro-invasive factor, which is secreted by CAFs, provides an unprecedented mechanism through which the stroma may drive cancer progression.

Autonomous formation flying: unified control and collision avoidance methods for close manoeuvring spacecraft

The idea of spacecraft formations, flying in tight configurations with maximum baselines of a few hundred meters in low-Earth orbits, has generated widespread interest over the last several years. Nevertheless, controlling the movement of spacecraft in formation poses difficulties, such as in-orbit high-computing demand and collision avoidance capabilities, which escalate as the number of units in the formation is increased and complicated nonlinear effects are imposed to the dynamics, together with uncertainty which may arise from the lack of knowledge of system parameters. These requirements have led to the need of reliable linear and nonlinear controllers in terms of relative and absolute dynamics. The objective of this thesis is, therefore, to introduce new control methods to allow spacecraft in formation, with circular/elliptical reference orbits, to efficiently execute safe autonomous manoeuvres. These controllers distinguish from the bulk of literature in that they merge guidance laws never applied before to spacecraft formation flying and collision avoidance capacities into a single control strategy. For this purpose, three control schemes are presented: linear optimal regulation, linear optimal estimation and adaptive nonlinear control. In general terms, the proposed control approaches command the dynamical performance of one or several followers with respect to a leader to asymptotically track a time-varying nominal trajectory (TVNT), while the threat of collision between the followers is reduced by repelling accelerations obtained from the collision avoidance scheme during the periods of closest proximity. Linear optimal regulation is achieved through a Riccati-based tracking controller. Within this control strategy, the controller provides guidance and tracking toward a desired TVNT, optimizing fuel consumption by Riccati procedure using a non-infinite cost function defined in terms of the desired TVNT, while repelling accelerations generated from the CAS will ensure evasive actions between the elements of the formation. The relative dynamics model, suitable for circular and eccentric low-Earth reference orbits, is based on the Tschauner and Hempel equations, and includes a control input and a nonlinear term corresponding to the CAS repelling accelerations. Linear optimal estimation is built on the forward-in-time separation principle. This controller encompasses two stages: regulation and estimation. The first stage requires the design of a full state feedback controller using the state vector reconstructed by means of the estimator. The second stage requires the design of an additional dynamical system, the estimator, to obtain the states which cannot be measured in order to approximately reconstruct the full state vector. Then, the separation principle states that an observer built for a known input can also be used to estimate the state of the system and to generate the control input. This allows the design of the observer and the feedback independently, by exploiting the advantages of linear quadratic regulator theory, in order to estimate the states of a dynamical system with model and sensor uncertainty. The relative dynamics is described with the linear system used in the previous controller, with a control input and nonlinearities entering via the repelling accelerations from the CAS during collision avoidance events. Moreover, sensor uncertainty is added to the control process by considering carrier-phase differential GPS (CDGPS) velocity measurement error. An adaptive control law capable of delivering superior closed-loop performance when compared to the certainty-equivalence (CE) adaptive controllers is finally presented. A novel noncertainty-equivalence controller based on the Immersion and Invariance paradigm for close-manoeuvring spacecraft formation flying in both circular and elliptical low-Earth reference orbits is introduced. The proposed control scheme achieves stabilization by immersing the plant dynamics into a target dynamical system (or manifold) that captures the desired dynamical behaviour. They key feature of this methodology is the addition of a new term to the classical certainty-equivalence control approach that, in conjunction with the parameter update law, is designed to achieve adaptive stabilization. This parameter has the ultimate task of shaping the manifold into which the adaptive system is immersed. The performance of the controller is proven stable via a Lyapunov-based analysis and Barbalat’s lemma. In order to evaluate the design of the controllers, test cases based on the physical and orbital features of the Prototype Research Instruments and Space Mission Technology Advancement (PRISMA) are implemented, extending the number of elements in the formation into scenarios with reconfigurations and on-orbit position switching in elliptical low-Earth reference orbits. An extensive analysis and comparison of the performance of the controllers in terms of total Δv and fuel consumption, with and without the effects of the CAS, is presented. These results show that the three proposed controllers allow the followers to asymptotically track the desired nominal trajectory and, additionally, those simulations including CAS show an effective decrease of collision risk during the performance of the manoeuvre.