Athanasios Mantalaris

A framework for the design, modeling and optimization of biomedical systems

Abstract We present an overview of the key building blocks of a design framework for modeling and optimization of biomedical systems with main focus on leukemia, that we have been developing in the Biological Systems Engineering Laboratory and the Centre for Process Systems Engineering at Imperial College. The framework features the following areas: (i) a three-dimensional, biomimetic, in vitro platform for culturing both healthy and diseased blood; (ii) a novel, hollow fiber bioreactor that upgrades this in vitro platform to enable expansion and continuous harvesting of healthy and diseased blood; (iii) a global optimization-based approach for the design and operation of the aforementioned bioreactor; (iv) a pharmacokinetic / pharmacodynamic model representing patient response to Acute Myeloid Leukemia treatment; (v) an experimental framework for cell cycle modeling and quantitative analysis of environmental stress. This manuscript recapitulates the progress made in the different areas as well as the way in which these areas are connected, finally leading to a hybrid in vitro/in silico platform which allows the optimization of the ex vivo expansion of healthy and diseased blood.

Cyclin and DNA Distributed Cell Cycle Model for GS-NS0 Cells

Mammalian cell cultures are intrinsically heterogeneous at different scales (molecular to bioreactor). The cell cycle is at the centre of capturing heterogeneity since it plays a critical role in the growth, death, and productivity of mammalian cell cultures. Current cell cycle models use biological variables (mass/volume/age) that are non-mechanistic, and difficult to experimentally determine, to describe cell cycle transition and capture culture heterogeneity. To address this problem, cyclins—key molecules that regulate cell cycle transition—have been utilized. Herein, a novel integrated experimental-modelling platform is presented whereby experimental quantification of key cell cycle metrics (cell cycle timings, cell cycle fractions, and cyclin expression determined by flow cytometry) is used to develop a cyclin and DNA distributed model for the industrially relevant cell line, GS-NS0. Cyclins/DNA synthesis rates were linked to stimulatory/inhibitory factors in the culture medium, which ultimately affect cell growth. Cell antibody productivity was characterized using cell cycle-specific production rates. The solution method delivered fast computational time that renders the model’s use suitable for model-based applications. Model structure was studied by global sensitivity analysis (GSA), which identified parameters with a significant effect on the model output, followed by re-estimation of its significant parameters from a control set of batch experiments. A good model fit to the experimental data, both at the cell cycle and viable cell density levels, was observed. The cell population heterogeneity of disturbed (after cell arrest) and undisturbed cell growth was captured proving the versatility of the modelling approach. Cell cycle models able to capture population heterogeneity facilitate in depth understanding of these complex systems and enable systematic formulation of culture strategies to improve growth and productivity. It is envisaged that this modelling approach will pave the model-based development of industrial cell lines and clinical studies.

Bone and cartilage regeneration with the use of umbilical cord mesenchymal stem cells.

Introduction: The production of functional alternatives to bone autografts and the development new treatment strategies for cartilage defects are great challenges that could be addressed by the field of tissue engineering. Umbilical cord mesenchymal stem cells (MSCs) can be used to produce cost-effective, atraumatic and possibly autologous bone and cartilage grafts. Areas covered: MSCs can be isolated from umbilical cord Wharton’s jelly, perivascular tissue and blood using various techniques. Those cells have been characterized and phenotypic similarities with bone marrow-derived MSCs (BM-MSCs) and embryonic stem cells have been found. Findings on their differentiation into the osteogenic and chondrogenic lineage differ between studies and are not as consistent as for BM-MSCs. Expert opinion: MSCs from umbilical cords have to be more extensively studied and the mechanisms underlying their differentiation have to be clarified. To date, they seem to be an attractive alternative to BM-MSCs. However, further research with suitable scaffolds and growth factors as well as with novel scaffold fabrication and culture technology should be conducted before they are introduced to clinical practice and replace BM-MSCs.

A mathematical model of subpopulation kinetics for the deconvolution of leukaemia heterogeneity

Acute myeloid leukaemia is characterized by marked inter- and intra-patient heterogeneity, the identification of which is critical for the design of personalized treatments. Heterogeneity of leukaemic cells is determined by mutations which ultimately affect the cell cycle. We have developed and validated a biologically relevant, mathematical model of the cell cycle based on unique cell-cycle signatures, defined by duration of cell-cycle phases and cyclin profiles as determined by flow cytometry, for three leukaemia cell lines. The model was discretized for the different phases in their respective progress variables (cyclins and DNA), resulting in a set of time-dependent ordinary differential equations. Cell-cycle phase distribution and cyclin concentration profiles were validated against population chase experiments. Heterogeneity was simulated in culture by combining the three cell lines in a blinded experimental set-up. Based on individual kinetics, the model was capable of identifying and quantifying cellular heterogeneity. When supplying the initial conditions only, the model predicted future cell population dynamics and estimated the previous heterogeneous composition of cells. Identification of heterogeneous leukaemia clones at diagnosis and post-treatment using such a mathematical platform has the potential to predict multiple future outcomes in response to induction and consolidation chemotherapy as well as relapse kinetics.

Current clinical evidence for the use of mesenchymal stem cells in articular cartilage repair

ABSTRACT Introduction: Articular cartilage is renowned for its poor intrinsic capacity for repair. Current treatments for osteoarthritis are limited in their ability to reliably restore the native articular cartilage structure and function. Mesenchymal stem cells (MSCs) present an attractive treatment option for articular cartilage repair, with a recent expansion of clinical trials investigating their use in patients. Areas covered: This paper provides a current overview of the clinical evidence on the use of MSCs in articular cartilage repair. Expert opinion: The article demonstrates robust clinical evidence that MSCs have significant potential for the regeneration of hyaline articular cartilage in patients. The majority of clinical trials to date have yielded significantly positive results with minimal adverse effects. However the clinical research is still in its infancy. The optimum MSC source, cell concentrations, implantation technique, scaffold, growth factors and rehabilitation protocol for clinical use are yet to be identified. A larger number of randomised control trials are required to objectively compare the clinical efficacy and long-term safety of the various techniques. As the clinical research continues to evolve and address these challenges, it is likely that MSCs may become integrated into routine clinical practice in the near future.

On the model‐based optimization of secreting mammalian cell (GS‐NS0) cultures

The global bio‐manufacturing industry requires improved process efficiency to satisfy the increasing demands for biochemicals, biofuels, and biologics. The use of model‐based techniques can facilitate the reduction of unnecessary experimentation and reduce labor and operating costs by identifying the most informative experiments and providing strategies to optimize the bioprocess at hand. Herein, we investigate the potential of a research methodology that combines model development, parameter estimation, global sensitivity analysis, and selection of optimal feeding policies via dynamic optimization methods to improve the efficiency of an industrially relevant bioprocess. Data from a set of batch experiments was used to estimate values for the parameters of an unstructured model describing monoclonal antibody (mAb) production in GS‐NS0 cell cultures. Global Sensitivity Analysis (GSA) highlighted parameters with a strong effect on the model output and data from a fed‐batch experiment were used to refine their estimated values. Model‐based optimization was used to identify a feeding regime that maximized final mAb titer. An independent fed‐batch experiment was conducted to validate both the results of the optimization and the predictive capabilities of the developed model. The successful integration of wet‐lab experimentation and mathematical model development, analysis, and optimization represents a unique, novel, and interdisciplinary approach that addresses the complicated research and industrial problem of model‐based optimization of cell based processes. Biotechnol. Bioeng. 2015;112: 536–548.

Investigational drugs for fracture healing: preclinical & clinical data.

ABSTRACT Introduction: The need for fracture healing enhancement for the management of fracture complications such as non-union and for the achievement of early function in fracture patients is constantly increasing. Therefore, the development and evaluation of novel pharmaceutical agents is mandatory in order to accelerate the process and increase bone union rates. Areas covered: This review summarizes the most recent knowledge on the pharmacological enhancement of fracture repair. It provides a synopsis of the most important preclinical and clinical studies published over the past five years on long bone fracture healing. Expert opinion: To date, limited drugs seem to have the potential for clinical use in fracture healing enhancement and the field is progressing very slowly. Among anti-osteoporotic drugs, only PTH and anti-sclerostin antibodies have such a potential but further research is needed before clinical use. The same applies also to BMPs, the use of which still carries major drawbacks that should be overcome before their widespread clinical utilization. Other drugs and growth factors, such as statins, VEGF, FGF, EPO, could be future key players in fracture healing but evidence is still lacking. Further in depth understanding of the healing process is essential in order to identify novel effective pharmacological agents.

In Silico Closed-Loop Control Validation Studies for Optimal Insulin Delivery in Type 1 Diabetes

This study presents a general closed-loop control strategy for optimal insulin delivery in type 1 Diabetes Mellitus (T1DM). The proposed control strategy aims toward an individualized optimal insulin delivery that consists of a patient-specific model predictive controller, a state estimator, a personalized scheduling level, and an open-loop optimization problem subjected to patient-specific process model and constraints. This control strategy can be also modified to address the case of limited patient data availability resulting in an “approximation” control strategy. Both strategies are validated in silico in the presence of predefined and unknown meal disturbances using both a novel mathematical model of glucose-insulin interactions and the UVa/Padova Simulator model as a virtual patient. The robustness of the control performance is evaluated under several conditions such as skipped meals, variability in the meal time, and metabolic uncertainty. The simulation results of the closed-loop validation studies indicate that the proposed control strategies can potentially achieve improved glycaemic control.

Fibronectin stimulates the osteogenic differentiation of murine embryonic stem cells.

Conditioned medium from human hepatocarcinoma cells (HepG2‐CM) has been shown to stimulate the osteogenic/chondrogenic differentiation of murine embryonic stem cells (mESCs). HepG2‐CM is considered to contain visceral endoderm (VE)‐like signals and attempts have recently been made to characterize it, using proteomic profiling, with fibronectin being identified as one promising candidate. Herein, we investigated whether fibronectin is able to mimic the activities of HepG2‐CM during the osteogenic differentiation of mESCs. Specifically, the addition of RGD peptides and heparin in HepG2‐CM significantly reduced the growth‐ and adhesion‐promoting effects of HepG2‐CM, in addition to suppressing its osteogenic‐inductive activity. Furthermore, direct addition of fibronectin to basal medium was able to reproduce, at least partially, the function of HepG2‐CM. In particular, fibronectin induced the early onset of osteogenic differentiation in mESCs, as confirmed by gene expression of osteogenic markers, and resulted in the three‐fold higher calcium deposition at day 11 of osteogenic culture compared to the control group. These data clearly suggest that fibronectin contributes to the biological activities of HepG2‐CM and plays a stimulatory role during the process of osteogenesis in mESCs. Copyright

Transcriptional kinetics of the cross-talk between the ortho-cleavage and TOL pathways of toluene biodegradation in Pseudomonas putida mt-2

The TOL plasmid promoters are activated by toluene leading to gene expression responsible for the degradation of the environmental signal. Benzoate is formed as an intermediate, activating the BenR protein of the chromosomal ortho-cleavage pathway that up-regulates the chromosomal PbenA promoter and the TOL Pm promoter resulting in cross-talk between the two networks. Herein, the transcriptional kinetics of the PbenR and PbenA promoters in conjunction with TOL promoters was monitored by real-time PCR during toluene biodegradation of different concentrations in batch cultures. The cross-talk between the two pathways was indicated by the simultaneous maximal expression of the Pm and PbenR promoters, as well as the transcriptional activation from PbenA occurring prior to PbenR, which indicates the potential up-regulation of PbenA by the TOL XylS protein. The repressory effect of toluene on Pr was evident for concentrations higher than 0.3mM suggesting a threshold value for restoring the promoters activity, while all the other promoters followed a specific expression pattern, regardless of the initial inducer concentration. Induction of the system with higher toluene concentrations revealed an oscillatory behaviour of Pm, the expression of which remained at high levels until the late exponential phase, demonstrating a novel function of this network.