Oleksii S Rukhlenko

University College Dublin | UCD · Systems Biology Ireland

Clinically used RAF inhibitors are ineffective in RAS mutant tumors because they enhance homo- and heterodimerization of RAF kinases, leading to paradoxical activation of ERK signaling. Overcoming enhanced RAF dimerization and the resulting resistance is a challenge for drug design. Combining multiple inhibitors could be more effective, but it is u...

Migrating cells need to coordinate distinct leading and trailing edge dynamics but the underlying mechanisms are unclear. Here, we combine experiments and mathematical modeling to elaborate the minimal autonomous biochemical machinery necessary and sufficient for this dynamic coordination and cell movement. RhoA activates Rac1 via DIA and inhibits...

Increasing evidence suggests that the reactivation of initially inhibited signaling pathways causes drug resistance. Here, we analyze how network topologies affect signaling responses to drug treatment. Network-dependent drug resistance is commonly attributed to negative and positive feedback loops. However, feedback loops by themselves cannot comp...

Understanding cell state transitions and purposefully controlling them is a longstanding challenge in biology. Here we present cell state transition assessment and regulation (cSTAR), an approach for mapping cell states, modelling transitions between them and predicting targeted interventions to convert cell fate decisions. cSTAR uses omics data as...

Fluid shear stress (FSS) from blood flow sensed by vascular endothelial cells (ECs) determines vessel behavior, but regulatory mechanisms are only partially understood. We used cell state transition assessment and regulation (cSTAR), a powerful computational method, to elucidate EC transcriptomic states under low shear stress (LSS), physiological s...

Introduction Mutations activating RAS signaling, typically in NRAS or KRAS, are common in acute myeloid leukemia (AML). These alterations hyperactivate downstream RAF/MEK/ERK kinases and are typically associated with poor outcome, especially in children. We recently developed a structure-based dynamic RAS pathway model that successfully predicted R...

Introduction GLP-1 analogues can stimulate apoptosis and autophagy in endometrial cancer cell lines. We sought to assess the effect of liraglutide on endometrial cancer development and local microenvironment in the BDII/HAN rat model. Methods 38 BDII/Han rats were randomised into two groups at 12 months of age - a standard group (n=17) and a lirag...

Background Pancreatic ductal adenocarcinoma (PDAC) continues to be a lethal disease. It has been established that 90% of PDAC patients harbor somatic oncogenic point mutations in KRAS, which leads to the activation of the RAS-to-ERK signaling pathway. This pathway is resistant to almost all known small molecule inhibitors and often gets reactivated...

Pancreatic ductal adenocarcinoma (PDAC) presents significant challenges for targeted clinical interventions due to prevalent KRAS mutations, rendering PDAC resistant to RAF and MEK inhibitors (RAFi and MEKi). In addition, responses to targeted therapies vary between patients. Here, we explored the differential sensitivities of PDAC cell lines to RA...

Understanding signaling patterns of transformation and controlling cell phenotypes is a challenge of current biology. Here we applied a cell State Transition Assessment and Regulation (cSTAR) approach to a perturbation dataset of single cell phosphoproteomic patterns of multiple breast cancer (BC) and normal breast tissue-derived cell lines. Follow...

Necrosis of TB granulomas and TB susceptibility in a mouse model are mediated by the sst1 locus encoding interferon-inducible nuclear proteins Sp110 and Sp140 • The sst1 mutant (sst1S) macrophages display an aberrant response to TNF featuring unresolving oxidative and proteotoxic stress, and escalating type I interferon pathway hyperactivity • We s...

A quarter of human population is infected with Mycobacterium tuberculosis, but less than 10% of those infected develop clinical, mostly pulmonary, TB. To dissect mechanisms of susceptibility in immunocompetent individuals, we developed a genetically defined sst1-susceptible mouse model that uniquely reproduces a defining feature of human TB: develo...

Increased endothelial cell (EC) permeability significantly contributes to peripheral arterial disease and its severe manifestation, critical limb ischemia (CLI). CLI remains refractory to current pharmacological interventions, often requiring limb amputation and showing how limited our knowledge is about signaling pathways that control EC states. H...

Morphogenesis of new arteries as a result of flow-driven capillary remodeling, also termed arteriogenesis, is determined by a process of fate specification to acquire arterial identity by capillary endothelial cells (ECs). Although a rich pool of studies in the past two decades suggesting the role of VEGF in promoting Notch activation and arterial...

Fluid shear stress (FSS) from blood flow is sensed by vascular endothelial cells (ECs) to determine vessel stability, remodeling and susceptibility to atherosclerosis and other inflammatory diseases but the regulatory networks that govern these behaviors are only partially understood. We used cSTAR, a powerful new computational method, to define EC...

Understanding cell state transitions and purposefully controlling them to improve therapies is a longstanding challenge in biological research and medicine. Here, we identify a transcriptional signature that distinguishes activated macrophages from the tuberculosis (TB) susceptible and resistant mice. We then apply the cSTAR (cell state transition...

Citation: Imoto, H.; Rauch, N.; Neve, A.J.; Khorsand, F.; Kreileder, M.; Alexopoulos, L.G.; Rauch, J.; Okada, M.; Kholodenko, B.N.; Rukhlenko, O.S. A Combination of Conformation-Specific RAF Inhibitors Overcome Drug Resistance Brought about by RAF Overexpression. Biomolecules 2023, 13, 1212. https://doi. Abstract: Cancer cells often adapt to target...

Acquisition of omics data advances at a formidable pace. Yet, our ability to utilize these data to control cell phenotypes and design interventions that reverse pathological states lags behind. Here, we posit that cell states are determined by core networks that control cell-wide networks. To steer cell fate decisions, core networks connecting geno...

Understanding cell state transitions and purposefully controlling them to improve therapies is a longstanding challenge in biological research and medicine. Here, we identify a transcriptional signature that distinguishes activated macrophages from TB-susceptible and TB-resistant mice. We then apply the cSTAR (cell State Transition Assessment and R...

This protocol illustrates a pipeline for modeling the nonlinear behavior of intracellular signaling pathways. At fixed spatial points, nonlinear signaling dynamics are described by ordinary differential equations (ODEs). At constant parameters, these ODEs may have multiple attractors, such as multiple steady states or limit cycles. Standard optimiz...

Macrophages contribute to host immunity and tissue homeostasis via alternative activation programs. M1-like macrophages control intracellular bacterial pathogens and tumor progression. In contrast, M2-like macrophages shape reparative microenvironments that can be conducive for pathogen survival or tumor growth. An imbalance of these macrophages ph...

Macromolecular protein assemblies govern many cellular processes and are disturbed in many diseases including cancer. Often seen as static molecular machines, protein complexes involved in signal transduction networks exhibit intricate dynamics that are critical for their function. Using the RAS-RAF-MEK-ERK pathway as example we discuss recent prog...

Tuberculosis remains a critical infectious disease world-wide. The development of novel therapeutic strategies requires greater understanding of host factors that contribute to disease susceptibility. A major unknown in TB pathogenesis is the mechanism of necrosis in TB granulomas that leads to the massive lung tissue damage and cavity formation ne...

Migrating cells need to coordinate distinct leading and trailing edge dynamics but the underlying mechanisms are unclear. Here, we combine experiments and mathematical modeling to elaborate the minimal autonomous biochemical machinery necessary and sufficient for this dynamic coordination and cell movement. RhoA activates Rac1 via DIA and inhibits...

Migrating cells need to coordinate distinct leading and trailing edge dynamics but the underlying mechanisms are unclear. Here, we combine experiments and mathematical modeling to elaborate the minimal autonomous biochemical machinery necessary and sufficient for this dynamic coordination and cell movement. RhoA activates Rac1 via DIA and inhibits...

Migrating cells need to coordinate distinct leading and trailing edge dynamics but the underlying mechanisms are unclear. Here, we combine experiments and mathematical modeling to elaborate the minimal autonomous biochemical machinery necessary and sufficient for this dynamic coordination and cell movement. RhoA activates Rac1 via DIA and inhibits...

Protein-protein-interaction networks (PPINs) organize fundamental biological processes, but how oncogenic mutations impact these interactions and their functions at a network-level scale is poorly understood. Here, we analyze how a common oncogenic KRAS mutation (KRASG13D) affects PPIN structure and function of the Epidermal Growth Factor Receptor...

One of the great challenges in biology is to reconstruct the structure of signaling networks from experimental data. Modular Response Analysis (MRA) is a precise method to reconstruct and quantify activating or inhibitory strengths of the connections between network modules called nodes. Standard MRA assumes that each node is unique and no componen...

A rapid increase of new nanomaterial products poses new challenges for their risk assessment. Current traditional methods for estimating potential adverse health effect of nanomaterials (NMs) are complex, time consuming and expensive. In order to develop new prediction tests for nanotoxicity evaluation, a systems biology approach and data from high...

Receptor tyrosine kinases (RTKs) typically contain multiple autophosphorylation sites in their cytoplasmic domains. Once activated, these autophosphorylation sites can recruit downstream signaling proteins containing Src homology 2 (SH2) and phosphotyrosine-binding (PTB) domains, which recognize phosphotyrosine-containing short linear motifs (SLiMs...

Rank ordering of IGF1R binding partners in cell lines according to each of five metrics. Contains ranking tables for all cell lines evaluated in this study. The five metrics for ranking recruitment were 1) numerical simulations with the restructured model, 2) exact results from the analytical equilibrium binding model, 3) copy number, 4) KD, and 5)...

Derivation of the analytical approximation of equilibrium binding of SH2/PTB-containing proteins to an RTK. We derive analytical equations to predict binding of signaling proteins to an RTK and compare results to those of the numerical simulation described in the main text. Furthermore, we present a simplified set of equations, which provides a rul...

Summary of rank ordering of IGF1R binding partners across 45 cell line-specific models. (XLSX)

Comparison of quantitative predictions from numerical simulations and the analytical approximation for HeLa S3 and HeLa Kyoto cell lines. Plots show the number of molecules of each protein bound at steady state predicted by either numerical simulations (x-axis) or the analytical approximation (y-axis). A dashed gray line on the diagonal illustrates...

Cell-specific IGF1R signaling models. A compressed archive containing the files for 45 cell line-specific models. Both natural and restructured models are included for HeLa S3. Only the restructured models are included for the other cell lines. BNGL models are plain-text files with the filename extension .bngl, intended for processing by BioNetGen....

Parameter estimation files with BioNetFit. This compressed directory contains the plain-text files that were processed by BioNetFit to estimate the parameters given in Table 1. The configuration file (filename extension .conf) provides instructions for parameter estimation. The .bngl file contains the IGF1-IGF1R binding model. The .exp files contai...

Population models. This compressed directory contains the files and scripts needed to generate models for a clonal population of cells with variability in protein copy number. The Python script “population_model.py” generates BioNetGen input files. These files can then be processed by BioNetGen to generate steady-state recruitment data for all cell...

Pairwise correlations for IGF1R signaling protein recruitment in lung, colon, renal, liver, melanoma, leukemia, and mouse cell lines. Red indicates a negative Pearson’s r, blue indicates a positive Pearson’s r, and white indicates no correlation between each pair of proteins. (TIFF)

Pairwise correlations for IGF1R signaling protein recruitment in breast, cervical, prostate, central nervous system, and bone cell lines. Red indicates a negative Pearson’s r, blue indicates a positive Pearson’s r, and white indicates no correlation between each pair of proteins. (TIFF)

A tutorial overview of model restructuration. Includes historical background and detailed discussion of decoupling, bunching, and scaling with examples. (PDF)

Protein copy numbers in all cell lines. This table summarizes protein copy number values reported in the literature [63,68–70] and used in this study. (XLSX)

KD values for IGF1R binding partners. This table summarizes equilibrium dissociation constant values reported in the literature [45,46,89] and used in this study. (XLSX)

Exactness is maintained in the restructured formulation of the IGF1R signaling model. Plots show time courses of bound phosphotyrosine sites and bound signaling proteins from simulations of the HeLa S3 model in the natural formulation and the restructured formulation. (TIFF)

Illustration of model restructuration. Cartoons of (A) bunching (B) decoupling, and (C-D) scaling are shown. (A) We can couple an S1 site from one IGF1R monomer and the S2 site from the other IGF1R monomer into one binding pocket, P. In the natural formulation, four different binding sites can be either free or bound to IGF1. In the restructured fo...

Modular Response Analysis (MRA) is a method to reconstruct signalling networks from steady-state perturbation data which has frequently been used in different settings. Since these data are usually noisy due to multi-step measurement procedures and biological variability, it is important to investigate the effect of this noise onto network reconstr...

Background Carbon nanotubes (CNTs) usage has rapidly increased in the last few decades due to their unique properties, exploited in various industrial and commercial products. Certain types of CNTs cause adverse health effects, including chronic inflammation and fibrosis. Despite the large number of in vitro and in vivo studies evaluating these eff...

RAS is the most frequently mutated gene across human cancers, but developing inhibitors of mutant RAS has proven to be challenging. Given the difficulties of targeting RAS directly, drugs that impact the other components of pathways where mutant RAS operates may potentially be effective. However, the system-level features, including different local...

In this review we discuss the origination and evolution of Modular Response Analysis (MRA), which is a physics-based method for reconstructing quantitative topological models of biochemical pathways. We first focus on the core theory of MRA, demonstrating how both the direction and the strength of local, causal connections between network modules c...

The efficiency of anti-tumour drug strongly depends on its dose. Higher drug doses and exposure times usually result in better treatment. It is why the implementation of high-dose treatment is always attractive. However, most of the drug delivery techniques meet essential limitations. In isolated regional perfusion a tumour can be exposed to high-d...

The RAS/RAF/MEK/MAPK kinase pathway has been extensively studied for more than 25 years, yet we continue to be puzzled by its intricate dynamic control and plasticity. Different spatiotemporal MAPK dynamics bring about distinct cell fate decisions in normal vs cancer cells and developing organisms. Recent modelling and experimental studies provided...

The intricate dynamic control and plasticity of RAS to ERK mitogenic, survival and apoptotic signalling has mystified researches for more than 30 years. Therapeutics targeting the oncogenic aberrations within this pathway often yield unsatisfactory, even undesired results, as in the case of paradoxical ERK activation in response to RAF inhibition....

Increased shear stress such as observed at local stenosis may cause drastic changes in the permeability of the vessel wall to procoagulants and thus initiate intravascular blood coagulation. In this paper we suggest a mathematical model to investigate how shear stress-induced permeability influences the thrombogenic potential of atherosclerotic pla...

Effects of a chronic combined unpredictable stress on activities of two cell death-related proteases, calpain and cathepsin B, were studied along with indices of nitrergic system in rat brain structures. Male Wistar rats were subjected to a 2-week-long combined stress (combination of unpaired flash light and moderate footshock associated with a whi...

The mechanisms of hydrodynamical activation of blood coagulation system are investigated in stenosed vessels for a wide range of Reynolds number values (from 10 up to 500). It is assumed that the vessel wall permeability for procoagulant factors rapidly increases when wall shear stress exceeds specific threshold value. A number of patterns of blood...