publications

2024

1. Epigenome and early selection determine the tumour-immune evolutionary trajectory of colorectal cancer

   Eszter Lakatos, Vinaya Gunasri, Luis Zapata, and 19 more authors

   bioRxiv, 2024

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   Immune system control is a major hurdle that cancer evolution must circumvent. The relative timing and evolutionary dynamics of subclones that have escaped immune control remain incompletely characterized, and how immune-mediated selection shapes the epigenome has received little attention. Here, we infer the genome- and epigenome-driven evolutionary dynamics of tumour-immune coevolution within primary colorectal cancers (CRCs). We utilise our existing CRC multi-region multi-omic dataset that we supplement with high-resolution spatially-resolved neoantigen sequencing data and highly multiplexed imaging of the tumour microenvironment (TME). Analysis of somatic chromatin accessibility alterations (SCAAs) reveals frequent somatic loss of accessibility at antigen presenting genes, and that SCAAs contribute to silencing of neoantigens. We observe that strong immune escape and exclusion occur at the outset of CRC formation, and that within tumours, including at the microscopic level of individual tumour glands, additional immune escape alterations have negligible consequences for the immunophenotype of cancer cells. Further minor immuno-editing occurs during local invasion and is associated with TME reorganisation, but that evolutionary bottleneck is relatively weak. Collectively, we show that immune evasion in CRC follows a “Big Bang” evolutionary pattern, whereby genetic, epigenetic and TME-driven immune evasion acquired by the time of transformation defines subsequent cancer-immune evolution.Competing Interest StatementT.A.G. and A.M.B. are listed as coinventors on patent application GB2305655.9 that concerns T-cell receptor sequencing of cancers, and T.A.G. is a coinventor on patent application GB2317139.0 that concerns measurement of cancer evolutionary dynamics. T.A.G. has received an honorarium from Genentech Inc.

2. Homopolymer switches mediate adaptive mutability in mismatch repair-deficient colorectal cancer

   Hamzeh Kayhanian, William Cross, Suzanne E. M. Horst, and 34 more authors

   Nature Genetics, 2024

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   Mismatch repair (MMR)-deficient cancer evolves through the stepwise erosion of coding homopolymers in target genes. Curiously, the MMR genes MutS homolog 6 (MSH6) and MutS homolog 3 (MSH3) also contain coding homopolymers, and these are frequent mutational targets in MMR-deficient cancers. The impact of incremental MMR mutations on MMR-deficient cancer evolution is unknown. Here we show that microsatellite instability modulates DNA repair by toggling hypermutable mononucleotide homopolymer runs in MSH6 and MSH3 through stochastic frameshift switching. Spontaneous mutation and reversion modulate subclonal mutation rate, mutation bias and HLA and neoantigen diversity. Patient-derived organoids corroborate these observations and show that MMR homopolymer sequences drift back into reading frame in the absence of immune selection, suggesting a fitness cost of elevated mutation rates. Combined experimental and simulation studies demonstrate that subclonal immune selection favors incremental MMR mutations. Overall, our data demonstrate that MMR-deficient colorectal cancers fuel intratumor heterogeneity by adapting subclonal mutation rate and diversity to immune selection.

3. The genomic landscape of 2,023 colorectal cancers

   Alex J. Cornish, Andreas J. Gruber, Ben Kinnersley, and 33 more authors

   Nature, 2024

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   Colorectal carcinoma (CRC) is a common cause of mortality1, but a comprehensive description of its genomic landscape is lacking2–9. Here we perform whole-genome sequencing of 2,023 CRC samples from participants in the UK 100,000 Genomes Project, thereby providing a highly detailed somatic mutational landscape of this cancer. Integrated analyses identify more than 250 putative CRC driver genes, many not previously implicated in CRC or other cancers, including several recurrent changes outside the coding genome. We extend the molecular pathways involved in CRC development, define four new common subgroups of microsatellite-stable CRC based on genomic features and show that these groups have independent prognostic associations. We also characterize several rare molecular CRC subgroups, some with potential clinical relevance, including cancers with both microsatellite and chromosomal instability. We demonstrate a spectrum of mutational profiles across the colorectum, which reflect aetiological differences. These include the role of Escherichia colipks+ colibactin in rectal cancers10 and the importance of the SBS93 signature11–13, which suggests that diet or smoking is a risk factor. Immune-escape driver mutations14 are near-ubiquitous in hypermutant tumours and occur in about half of microsatellite-stable CRCs, often in the form of HLA copy number changes. Many driver mutations are actionable, including those associated with rare subgroups (for example, BRCA1 and IDH1), highlighting the role of whole-genome sequencing in optimizing patient care.

4. Lower incidence of small intestinal adenocarcinomas may be due to constrained copy number karyotype and increased immune surveillance

   Salpie Nowinski, Vickna Balarajah, Ann-Marie Baker, and 7 more authors

   In CANCER RESEARCH, 2024

2023

1. O44 Co-evolution of mismatch repair loss and the immune response in Lynch Syndrome

   Ottilie Swinyard, Ann-Marie Baker, Kevin Monahan, and 6 more authors

   Gut, 2023

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   Background Lynch syndrome (LS) is a hereditary condition characterised by heterozygous pathogenic germline mutation or hypermethylation of one of the mismatch repair (MMR) genes, predisposing carriers to a variety of cancers, predominantly colorectal cancer (CRC). The somatic loss of the remaining functional allele results in complete MMR deficiency (MMR-d), however the sequence and timing of events in the trajectory towards LS lesions are not fully understood. Additionally, how immune surveillance responds to single MMR-d crypt foci (MMR-dcf) has not been examined. We explored the earliest interactions between MMR-dcf and the immune microenvironment in the LS bowel to characterise the co-evolution of MMR-d clones and the immune system.Methods Prospective and retrospective colon tissue samples were collected from 32 confirmed LS patients. The samples were classified as normal from a LS colon free of cancer (n=13), normal from a LS colon with a distant cancer (n=6) or adenoma adjacent normal tissue (n=13). The frequency and location of MMR-dcf were identified through immunohistochemical staining for MMR proteins. Multiplexed, whole-slide cyclic immunofluorescence (CyCIF) was performed to characterise the immune microenvironment surrounding MMR-dcf using a panel of 21 markers, including those identifying T cells, B cells, macrophages, neutrophils, and immune checkpoints. Single cell segmentation and marker quantification compared the immune landscape of MMR proficient (MMR-p) and MMR-d patches in non-dysplastic and diseased tissue.Results Morphologically normal MMR-d crypts were primarily found in adenoma adjacent areas (n = 194/7256 crypts (2.7%), found in 6/13 patients) compared to in the healthy LS mucosa (n = 1/4087 crypts (0.02%), found in 1/13 patients, p = 0.0276 t test), with the patch size increasing with patient age. This suggests that MMR-dcf occur very rarely in the non-dysplastic LS colon, or that they are effectively removed by negative selection or immune predation. Interestingly, non-neoplastic MMR-d patches are only seen next to MMR-d polyps. Our CyCIF analysis of the immune microenvironment revealed that MMR-d and MMR-p crypts have distinct immune neighbourhoods, with MMR-dcf characterised by increased infiltration of cytotoxic T cells and B cells, but no change in regulatory T cells or macrophages. This suggests an active inflammatory response to MMR-d, but no evidence of negative selection of these populations was seen. Staining and genomic sequencing showed a loss of HLA expression in a subset of MMR-d patches, a possible immune evasion mechanism that could explain the lack of immune eradication of these early mutated populations.Conclusion We present evidence that patches of morphologically normal MMR-dcf are more prevalent adjacent to adenomas, but rare in the healthy LS bowel. These foci appear to evoke a cytotoxic immune response, whilst evading immune elimination. Together our data indicate a critical role for immunosurveillance in the genesis of LS-associated CRC, highlighting potential opportunities for chemoprevention.

2. Evolutionary and immune microenvironment dynamics during neoadjuvant treatment of oesophagael adenocarcinoma.

   Melissa Barroux, Jacob Househam, Eszter Lakatos, and 22 more authors

   Res Sq, Apr 2023

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   Locally advanced oesophageal adenocarcinoma (EAC) remains difficult to treat because of common resistance to neoadjuvant therapy and high recurrence rates. The ecological and evolutionary dynamics responsible for treatment failure are incompletely understood. Here, we performed a comprehensive multi-omic analysis of samples collected from EAC patients in the MEMORI clinical trial, revealing major changes in gene expression profiles and immune microenvironment composition that did not appear to be driven by changes in clonal composition. Multi-region multi-timepoint whole exome (300x depth) and paired transcriptome sequencing was performed on 27 patients pre-, during and after neoadjuvant treatment. EAC showed major transcriptomic changes during treatment with upregulation of immune and stromal pathways and oncogenic pathways such as KRAS, Hedgehog and WNT. However, genetic data revealed that clonal sweeps were rare, suggesting that gene expression changes were not clonally driven. Additional longitudinal image mass cytometry was performed in a subset of 15 patients and T-cell receptor sequencing in 10 patients, revealing remodelling of the T-cell compartment during treatment and other shifts in microenvironment composition. The presence of immune escape mechanisms and a lack of clonal T-cell expansions were linked to poor clinical treatment response. This study identifies profound transcriptional changes during treatment with limited evidence that clonal replacement is the cause, suggesting phenotypic plasticity and immune dynamics as mechanisms for therapy resistance with pharmacological relevance.

3. Adaptive therapy achieves long-term control of chemotherapy resistance in high grade ovarian cancer

   Helen Hockings, Eszter Lakatos, Weini Huang, and 8 more authors

   bioRxiv, Apr 2023

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   Drug resistance results in poor outcomes for most patients with metastatic cancer. Adaptive Therapy (AT) proposes to address this by exploiting presumed fitness costs incurred by drug-resistant cells when drug is absent, and prescribing dose reductions to allow fitter, sensitive cells to re-grow and re- sensitise the tumour. However, empirical evidence for treatment-induced fitness change is lacking. We show that fitness costs in chemotherapy-resistant ovarian cancer cause selective decline and apoptosis of resistant populations in low-resource conditions. Moreover, carboplatin AT caused fluctuations in sensitive/resistant tumour population size in vitro and significantly extended survival of tumour-bearing mice. In sequential blood-derived cell-free DNA and tumour samples obtained longitudinally from ovarian cancer patients during treatment, we inferred resistant cancer cell population size through therapy and observed it correlated strongly with disease burden. These data have enabled us to launch a multicentre, phase 2 randomised controlled trial (ACTOv) to evaluate AT in ovarian cancer.Competing Interest StatementThe authors have declared no competing interest.

4. Immune selection determines tumor antigenicity and influences response to checkpoint inhibitors

   Luis Zapata, Giulio Caravagna, Marc J. Williams, and 12 more authors

   Nature Genetics, Apr 2023

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   In cancer, evolutionary forces select for clones that evade the immune system. Here we analyzed >10,000 primary tumors and 356 immune-checkpoint-treated metastases using immune dN/dS, the ratio of nonsynonymous to synonymous mutations in the immunopeptidome, to measure immune selection in cohorts and individuals. We classified tumors as immune edited when antigenic mutations were removed by negative selection and immune escaped when antigenicity was covered up by aberrant immune modulation. Only in immune-edited tumors was immune predation linked to CD8 T cell infiltration. Immune-escaped metastases experienced the best response to immunotherapy, whereas immune-edited patients did not benefit, suggesting a preexisting resistance mechanism. Similarly, in a longitudinal cohort, nivolumab treatment removes neoantigens exclusively in the immunopeptidome of nonimmune-edited patients, the group with the best overall survival response. Our work uses dN/dS to differentiate between immune-edited and immune-escaped tumors, measuring potential antigenicity and ultimately helping predict response to treatment.

2022

1. Phenotypic plasticity and genetic control in colorectal cancer evolution

   Jacob Househam, Timon Heide, George D. Cresswell, and 30 more authors

   Nature, Apr 2022

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   Genetic and epigenetic variation, together with transcriptional plasticity, contribute to intratumour heterogeneity1. The interplay of these biological processes and their respective contributions to tumour evolution remain unknown. Here we show that intratumour genetic ancestry only infrequently affects gene expression traits and subclonal evolution in colorectal cancer (CRC). Using spatially resolved paired whole-genome and transcriptome sequencing, we find that the majority of intratumour variation in gene expression is not strongly heritable but rather ‘plastic’. Somatic expression quantitative trait loci analysis identified a number of putative genetic controls of expression by cis-acting coding and non-coding mutations, the majority of which were clonal within a tumour, alongside frequent structural alterations. Consistently, computational inference on the spatial patterning of tumour phylogenies finds that a considerable proportion of CRCs did not show evidence of subclonal selection, with only a subset of putative genetic drivers associated with subclone expansions. Spatial intermixing of clones is common, with some tumours growing exponentially and others only at the periphery. Together, our data suggest that most genetic intratumour variation in CRC has no major phenotypic consequence and that transcriptional plasticity is, instead, widespread within a tumour.

2. The co-evolution of the genome and epigenome in colorectal cancer

   Timon Heide, Jacob Househam, George D. Cresswell, and 28 more authors

   Nature, Apr 2022

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   Colorectal malignancies are a leading cause of cancer-related death1 and have undergone extensive genomic study2,3. However, DNA mutations alone do not fully explain malignant transformation4–7. Here we investigate the co-evolution of the genome and epigenome of colorectal tumours at single-clone resolution using spatial multi-omic profiling of individual glands. We collected 1,370 samples from 30 primary cancers and 8 concomitant adenomas and generated 1,207 chromatin accessibility profiles, 527 whole genomes and 297 whole transcriptomes. We found positive selection for DNA mutations in chromatin modifier genes and recurrent somatic chromatin accessibility alterations, including in regulatory regions of cancer driver genes that were otherwise devoid of genetic mutations. Genome-wide alterations in accessibility for transcription factor binding involved CTCF, downregulation of interferon and increased accessibility for SOX and HOX transcription factor families, suggesting the involvement of developmental genes during tumourigenesis. Somatic chromatin accessibility alterations were heritable and distinguished adenomas from cancers. Mutational signature analysis showed that the epigenome in turn influences the accumulation of DNA mutations. This study provides a map of genetic and epigenetic tumour heterogeneity, with fundamental implications for understanding colorectal cancer biology.

3. The mutational signatures of formalin fixation on the human genome

   Qingli Guo, Eszter Lakatos, Ibrahim Al Bakir, and 3 more authors

   Nature Communications, Apr 2022

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   Clinical archives of patient material near-exclusively consist of formalin-fixed and paraffin-embedded (FFPE) blocks. The ability to precisely characterise mutational signatures from FFPE-derived DNA has tremendous translational potential. However, sequencing of DNA derived from FFPE material is known to be riddled with artefacts. Here we derive genome-wide mutational signatures caused by formalin fixation. We show that the FFPE-signature is highly similar to signature 30 (the signature of Base Excision Repair deficiency due to NTHL1 mutations), and chemical repair of DNA lesions leads to a signature highly similar to signature 1 (clock-like signature due to spontaneous deamination of methylcytosine). We demonstrate that using uncorrected mutational catalogues of FFPE samples leads to major mis-assignment of signature activities. To correct for this, we introduce FFPEsig, a computational algorithm to rectify the formalin-induced artefacts in the mutational catalogue. We demonstrate that FFPEsig enables accurate mutational signature analysis both in simulated and whole-genome sequenced FFPE cancer samples. FFPEsig thus provides an opportunity to unlock additional clinical potential of archival patient tissues.

4. Fluctuating methylation clocks for cell lineage tracing at high temporal resolution in human tissues

   Calum Gabbutt, Ryan O. Schenck, Daniel J. Weisenberger, and 14 more authors

   Nature Biotechnology, Apr 2022

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   Molecular clocks that record cell ancestry mutate too slowly to measure the short-timescale dynamics of cell renewal in adult tissues. Here, we show that fluctuating DNA methylation marks can be used as clocks in cells where ongoing methylation and demethylation cause repeated ‘flip–flops’between methylated and unmethylated states. We identify endogenous fluctuating CpG (fCpG) sites using standard methylation arrays and develop a mathematical model to quantitatively measure human adult stem cell dynamics from these data. Small intestinal crypts were inferred to contain slightly more stem cells than the colon, with slower stem cell replacement in the small intestine. Germline APC mutation increased the number of replacements per crypt. In blood, we measured rapid expansion of acute leukemia and slower growth of chronic disease. Thus, the patterns of human somatic cell birth and death are measurable with fluctuating methylation clocks (FMCs).

2021

1. LiquidCNA: Tracking subclonal evolution from longitudinal liquid biopsies using somatic copy number alterations.

   Eszter Lakatos, Helen Hockings, Maximilian Mossner, and 3 more authors

   iScience, Aug 2021

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   Cell-free DNA (cfDNA) measured via liquid biopsies provides a way for minimally invasive monitoring of tumor evolutionary dynamics during therapy. Here we present liquidCNA, a method to track subclonal evolution from longitudinally collected cfDNA samples sequenced through cost-effective low-pass whole-genome sequencing. LiquidCNA utilizes somatic copy number alteration (SCNA) to simultaneously genotype and quantify the size of the dominant subclone without requiring B-allele frequency information, matched-normal samples, or prior knowledge on the genetic identity of the emerging clone. We demonstrate the accuracy of liquidCNA in synthetically generated sample sets and in vitro mixtures of cancer cell lines. In vivo application in patients with metastatic lung cancer reveals the progressive emergence of a novel tumor subpopulation. LiquidCNA is straightforward to use, is computationally inexpensive, and enables continuous monitoring of subclonal evolution to understand and control-therapy-induced resistance.

2. O15 Evolutionary characteristics of neoantigens in inflammatory bowel disease and colorectal cancer

   Jatinder Stanley, Eszter Lakatos, Ann-Marie Baker, and 3 more authors

   Gut, Aug 2021

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   Introduction The immune system plays an active role in fighting growing tumours via recognising tumour-specific neoantigens and initiating an immune response. Consequently, the abundance and diversity of tumour neoantigens is shaped by the interaction with immune cells. Colonic mucosa in patients with inflammatory bowel disease (IBD) has a high immune cell presence, and we hypothesised this would cause increased immune predation on neoantigen-bearing epithelial cells. To test this, we compared neoantigen burdens in ulcerative colitis-associated colorectal cancers (CA-CRCs) and sporadically arising colorectal cancers (SPCRCs).Methods Existing multi-region whole-exome and whole-genome sequencing data1 2 from CA-CRCs (n=15) and SPCRCs (n=10) was used to computationally predict the abundance and diversity of immunogenic neoantigens using NeoPredPipe.3 Variant call data was filtered to retain high confidence variants. Neoantigen burden was compared between groups using a normalised measure, representing the proportion of non-synonymous mutations predicted to produce >=1 immunogenic neoantigen. Multi-region data from normal, histologically normal adjacent-to-tumour (NAT) and tumour samples was used to calculate the clonality and subclonality of neoantigens.Results The neoantigen burden of CA-CRCs was lower than SPCRCs (figure 1). Excluding cancers with microsatellite instability, CA-CRCs had relatively higher numbers of subclonal neoantigens per clonal neoantigens (p=0.029, Wilcoxon test), suggesting a greater degree of intra-tumour heterogeneity in CA-CRCs. In a subset of patients with CA-CRCs, 50–100% of clonal neoantigens found in tumour samples were shared with NAT samples in the same patient, revealing evidence of field cancerisation at the neoantigen level.Abstract O15 Figure 1 Comparison of neoantigen burden between CA-CRCs and SPCRCsConclusions These novel results support the hypothesis of increased immune surveillance in CA-CRCs compared to SPCRCs. Subclonal neoantigens accrue following immune escape, and so the higher burden of subclonal neoantigens in CA-CRCs points to the early evolution of effective immune escape mechanisms in these tumours. These data have implications for prognostication and immunotherapy treatment decisions for patients with CA-CRCs.ReferencesBaker AM, et al. Evolutionary history of human colitis-associated colorectal cancer. Gut (2018).Cross W, et al. The evolutionary landscape of colorectal tumorigenesis. Nat. Ecol. Evol. (2018).Schenck RO, Lakatos E, Gatenbee C, Graham TA & Anderson ARA. NeoPredPipe: high-throughput neoantigen prediction and recognition potential pipeline. BMC Bioinformatics (2019).

2020

1. Evolutionary dynamics of neoantigens in growing tumors

   Eszter Lakatos, Marc J. Williams, Ryan O. Schenck, and 10 more authors

   Nature Genetics, Aug 2020

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   Cancers accumulate mutations that lead to neoantigens, novel peptides that elicit an immune response, and consequently undergo evolutionary selection. Here we establish how negative selection shapes the clonality of neoantigens in a growing cancer by constructing a mathematical model of neoantigen evolution. The model predicts that, without immune escape, tumor neoantigens are either clonal or at low frequency; hypermutated tumors can only establish after the evolution of immune escape. Moreover, the site frequency spectrum of somatic variants under negative selection appears more neutral as the strength of negative selection increases, which is consistent with classical neutral theory. These predictions are corroborated by the analysis of neoantigen frequencies and immune escape in exome and RNA sequencing data from 879 colon, stomach and endometrial cancers.

2019

1. 25P - Low pass whole genome sequencing can be used to calculate the relative proportion of chemotherapy resistant disease in high grade serous ovarian cancer

   H. A. Hockings, M. Mossner, E. Lakatos, and 2 more authors

   Annals of Oncology, Aug 2019

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   Background Survival in high grade serous ovarian cancer (HGSOC) is impacted by the emergence of resistance to platinum chemotherapy. The traditional ethos of killing the greatest number of cancer cells with the maximum tolerated dose of chemotherapy may be flawed in evolutionary terms, facilitating the growth of resistant subclones and accelerating decreased treatment efficacy. Adaptive therapy is a new model of cancer treatment that exploits the competitive interactions between drug-sensitive and resistant subclones. The aim is to maintain a stable tumour burden by keeping a sufficient population of sensitive cells which suppress the proliferation of the ‘less fit’ resistant cells. A challenge in bringing adaptive therapy to the clinical setting is quantifying the relative proportions of sensitive and resistant disease to guide the personalised scheduling of treatment. In HGSOC, chemotherapy resistance is not associated with any common, measurable point mutations, but is correlated with a higher burden of copy number aberrations. We have utilised this to develop a method of quantifying the relative proportion of chemotherapy resistant disease. Methods An in silico dilution series was created using known copy number profiles of paired sensitive and resistant HGSOC OVCAR4 cells. Low pass whole genome sequencing (1×) was performed on cell mixtures of known ratios and bioinformatician-blinded ratios, combined with varied amounts of ‘normal’ DNA extracted from donor leucocytes. Results Our proof of principle experiment verified the method of quantifying copy number aberrations as a biomarker of the resistant population. The technique is now being applied to circulating tumour DNA isolated from serial plasma samples collected from 10 HGSOC patients, using the copy number profile from a diagnostic biopsy as a baseline for sensitive disease. Conclusions We have developed a method to monitor the relative drug-sensitive/resistant composition of tumours that can be applied to liquid biopsies. This will enable real-time serial tracking of tumour composition allowing for tumour heterogeneity, and guide personalised treatment scheduling in adaptive therapy. Legal entity responsible for the study The authors. Funding Cancer Research UK. Disclosure All authors have declared no conflicts of interest.

2. NeoPredPipe: high-throughput neoantigen prediction and recognition potential pipeline

   Ryan O. Schenck, Eszter Lakatos, Chandler Gatenbee, and 2 more authors

   BMC Bioinformatics, Aug 2019

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   Next generation sequencing has yielded an unparalleled means of quickly determining the molecular make-up of patient tumors. In conjunction with emerging, effective immunotherapeutics for a number of cancers, this rapid data generation necessitates a paired high-throughput means of predicting and assessing neoantigens from tumor variants that may stimulate immune response.

2018

1. Connecting the Microenvironmental Niche to Treatment Response in Ovarian Cancer

   Maximilian Strobl, Matthew Wicker, Vikram Adhikarla, and 11 more authors

   bioRxiv, Aug 2018

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   Ovarian cancer has the highest mortality rate of all gynecologic cancers, which may be attributed to an often late stage diagnosis, when the cancer is already metastatic, and rapid development of treatment resistance. We propose that the metastatic disease could be better characterized by observing interactions within the microenvironmental niche of the primary site that shapes the tumor’s early phenotypic progression. We present a mechanistic mathematical model of ovarian cancer that considers spatial interactions between tumor cells and several key stromal components. We demonstrate how spatial biomarker imaging data from the primary tumor can be analyzed to define a patient-specific microenvironment in the mathematical model. We then show preliminary results, using this model, that demonstrate how differences in the niche composition of a tumor affects phenotypic evolution and treatment response.

2. Transition state characteristics during cell differentiation

   Rowan D Brackston, Eszter Lakatos, and Michael P H Stumpf

   PLoS computational biology, Sep 2018

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   Models describing the process of stem-cell differentiation are plentiful, and may offer insights into the underlying mechanisms and experimentally observed behaviour. Waddington’s epigenetic landscape has been providing a conceptual framework for differentiation processes since its inception. It also allows, however, for detailed mathematical and quantitative analyses, as the landscape can, at least in principle, be related to mathematical models of dynamical systems. Here we focus on a set of dynamical systems features that are intimately linked to cell differentiation, by considering exemplar dynamical models that capture important aspects of stem cell differentiation dynamics. These models allow us to map the paths that cells take through gene expression space as they move from one fate to another, e.g. from a stem-cell to a more specialized cell type. Our analysis highlights the role of the transition state (TS) that separates distinct cell fates, and how the nature of the TS changes as the underlying landscape changes-change that can be induced by e.g. cellular signaling. We demonstrate that models for stem cell differentiation may be interpreted in terms of either a static or transitory landscape. For the static case the TS represents a particular transcriptional profile that all cells approach during differentiation. Alternatively, the TS may refer to the commonly observed period of heterogeneity as cells undergo stochastic transitions.

3. Topology of interaction between titin and myosin thick filaments

   Miklós Kellermayer, Dominik Sziklai, Zsombor Papp, and 3 more authors

   Journal of Structural Biology, Sep 2018

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   Titin is a giant protein spanning between the Z- and M-lines of the sarcomere. In the A-band titin is associated with the myosin thick filament. It has been speculated that titin may serve as a blueprint for thick-filament formation due to the super-repeat structure of its A-band domains. Accordingly, titin might provide a template that determines the length and structural periodicity of the thick filament. Here we tested the titin ruler hypothesis by mixing titin and myosin at in situ stoichiometric ratios (300 myosins per 12 titins) in buffers of different ionic strength (KCl concentration range 100–300 mM). The topology of the filamentous complexes was investigated with atomic force microscopy. We found that the samples contained distinct, segregated populations of titin molecules and myosin thick filaments. We were unable to identify complexes in which myosin molecules were regularly associated to either mono- or oligomeric titin in either relaxed or stretched states of the titin filaments. Thus, the electrostatically driven self-association is stronger in both myosin and titin than their binding to each other, and it is unlikely that titin functions as a geometrical template for thick-filament formation. However, when allowed to equilibrate configurationally, long myosin thick filaments appeared with titin oligomers attached to their surface. The titin meshwork formed on the thick-filament surface may play a role in controlling thick-filament length by regulating the structural dynamics of myosin molecules and placing a mechanical limit on the filament length.

2017

1. Protein degradation rate is the dominant mechanism accounting for the differences in protein abundance of basal p53 in a human breast and colorectal cancer cell line

   Eszter Lakatos, Ali Salehi-Reyhani, Michael Barclay, and 2 more authors

   PLOS ONE, May 2017

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   We determine p53 protein abundances and cell to cell variation in two human cancer cell lines with single cell resolution, and show that the fractional width of the distributions is the same in both cases despite a large difference in average protein copy number. We developed a computational framework to identify dominant mechanisms controlling the variation of protein abundance in a simple model of gene expression from the summary statistics of single cell steady state protein expression distributions. Our results, based on single cell data analysed in a Bayesian framework, lends strong support to a model in which variation in the basal p53 protein abundance may be best explained by variations in the rate of p53 protein degradation. This is supported by measurements of the relative average levels of mRNA which are very similar despite large variation in the level of protein.

2. Control mechanisms for stochastic biochemical systems via computation of reachable sets

   Eszter Lakatos, and Michael P. H. Stumpf

   Royal Society Open Science, May 2017

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   Controlling the behaviour of cells by rationally guiding molecular processes is an overarching aim of much of synthetic biology. Molecular processes, however, are notoriously noisy and frequently nonlinear. We present an approach to studying the impact of control measures on motifs of molecular interactions that addresses the problems faced in many biological systems: stochasticity, parameter uncertainty and nonlinearity. We show that our reachability analysis formalism can describe the potential behaviour of biological (naturally evolved as well as engineered) systems, and provides a set of bounds on their dynamics at the level of population statistics: for example, we can obtain the possible ranges of means and variances of mRNA and protein expression levels, even in the presence of uncertainty about model parameters.

2016

1. MEANS: python package for Moment Expansion Approximation, iNference and Simulation

   Sisi Fan, Quentin Geissmann, Eszter Lakatos, and 5 more authors

   Bioinformatics, May 2016

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   Motivation: Many biochemical systems require stochastic descriptions. Unfortunately these can only be solved for the simplest cases and their direct simulation can become prohibitively expensive, precluding thorough analysis. As an alternative, moment closure approximation methods generate equations for the time-evolution of the system’s moments and apply a closure ansatz to obtain a closed set of differential equations; that can become the basis for the deterministic analysis of the moments of the outputs of stochastic systems.Results: We present a free, user-friendly tool implementing an efficient moment expansion approximation with parametric closures that integrates well with the IPython interactive environment. Our package enables the analysis of complex stochastic systems without any constraints on the number of species and moments studied and the type of rate laws in the system. In addition to the approximation method our package provides numerous tools to help non-expert users in stochastic analysis.Availability and implementation:https://github.com/theosysbio/meansContacts:m.stumpf@imperial.ac.uk or e.lakatos13@imperial.ac.ukSupplementary information:Supplementary data are available at Bioinformatics online.

2015

1. Multivariate moment closure techniques for stochastic kinetic models

   Eszter Lakatos, Angelique Ale, Paul D. W. Kirk, and 1 more author

   The Journal of Chemical Physics, May 2015

2013

1. Structure of Synthetic Vertebrate Myosin Thick Filaments Explored with High-Resolution AFM

   Brennan Decker, Eszter Lakatos, and Miklós Kellermayer

   Biophysical Journal, Jan 2013