Spatio-temporal Stochastic Models for Phenotype and Gene Regulation by the Human Papillomavirus

Project Information

STEPV

Grant agreement ID: 101033459

DOI

10.3030/101033459

Project closed

EC signature date 24 March 2021

Start date 7 February 2022

End date 6 August 2023

Funded under

• EXCELLENT SCIENCE - Marie Skłodowska-Curie Actions

Total cost

€ 159 700,32

EU contribution

€ 159 700,32

Coordinated by

THE CHANCELLOR MASTERS AND SCHOLARS OF THE UNIVERSITY OF CAMBRIDGE
United Kingdom

Periodic Reporting for period 1 - STEPV (Spatio-temporal Stochastic Models for Phenotype and Gene Regulation by the Human Papillomavirus)

Reporting period: 2022-02-07 to 2023-08-06

Summary of the context and overall objectives of the project

In this project systems biology was used to develop temporal and spatiotemporal models to understand how high risk Human Papillomaviruses (HPV-16,) affect cell fate choices through 16E6 viral protein in order to affect tissue homeostasis, hence the balance between cell proliferation and differentiation.
This is an important problem for society because even though vaccines exist against HPVs, they are prophilactic vaccines and therefore the development of antiviral therapies is of pivotal importance. Understanding how HPVs affect cell fate choices in a phenomenological way under tissue biology approaches as well as a mechanistic perspective under a pathway understanding is of pivotal importance in order to properly understand viral perstistence and reservior. This will help to better understand under a rigorous approach the key regulators that has to be targeted and how to optimally target them in order to develop novel antiviral effective therapies.
The objectives were to understand the important question about how HPV affects tissue homeostasis through E6 both phenomenologically through the development of a cell state model describing cell states (proliferative, differentiation) in the infected epithelium. The objectives were also related to study how E6 could affect Notch pathway in order to mechanistically understand, through a pathway understanding, how cell fate choices in terms of cell proliferation and differentiation are controlled as well as explore antiviral therapies hypothesis through model simulations.
In conclusion, STEPV project reached its major goals, and it was possible to validate with biological experiments a stochastic mathematical model explaining the competitve advantage of HPV over wild type (healthy) cells in terms of cell fate choices (i.e. symmetric/asymmetric division, commitment to differentiation). The model was expanded into a 2D stochastic lattice grid model capable to reproduce in space and time the HPV competitve advantage in terms of the balance between cell proliferation and differentiation at a tissue level. The model was specialized with Notch/DLL1 pathway in order to understand how HPV E6 is altering such a pathway to make stronger hypothesis about HPV competitive advantage. The model was also used as a simulator to test different biological hypothesis also regarding antiviral therapies.

Work performed from the beginning of the project to the end of the period covered by the report and main results achieved so far

During the fellowship the fellow worked on two in vitro tissue culture systems: i) a mosaic assay where red (mCherry) cells (HPV cells) and green (eGFP) cells (wild type keratinocytes) compete to each other for an observation time of two weeks: ii) a single clone assay where we are targeting with green (eGFP) cells for 2-3 weeks in order to explain the stochastic cell fate choices of a progeny of cells derived by a single progenitor cell. It was possible to develop, with the aid of the mosaic assay, an idea of how HPV cells and wild type cells compete to each other under an average perspective and we could confirm the HPV competitive advantage over wild type cells. A deterministic minimal model was built to describe cell kinetics. Single clone assay was used to study stochastic cell fate choice in terms of symmetric/asymmetric division and cell differentiation. A stochastic model was built and validated with the theory of master equation on the single clone data collected as time varying probability distributions. The validated model allowed to rigorously and quantitatively describe the biology and allowed to estimate, for HPV positive cells, a 5% increase in symmetric cell division and 10% decrease in asymmetric cell division (Figure). The model was then extended to a spatial model based on hexagonal lattice grid by extending the single clone model to a spatial context in order to simulate clone dynamics in space and time. A model of Notch/DLL1 pathway affected by HPV 16E6 was developed as to deeply understand, through systems biology, how E6 can alter the cell-cell lateral inhibition mechanism through DLL1 by increasing DLL1 transcription and knocking down Notch. This helped understanding how to possibly design novel antiviral drugs targeting DLL1/E6 interaction. The model was used to better understand the phenotype induced by E6 in space and time and understand in a quantitiavive way how E6 can decrease cell differentiation and consequently increase symmetric and asymmetric cell dvision. Model simulations of Notch pathway perturbed by E6 under different conditions were run by simulating an antiviral therapy targeting DLL1.
In conclusion, biological experiments and systems biology were merged to understand under both a phenomenological and a mechanistic approach, through pathways, how HPV 16E6 is affecting cell fate choices related to cell proliferation and differentiation in order to affect tissue homeostasis.

Progress beyond the state of the art and expected potential impact (including the socio-economic impact and the wider societal implications of the project so far)

The projects results went much beyond the state of the art. Indeed, currently there is no systems virology understanding of human papillomaviruses validated with data and there is no deep and rigorous understanding through systems biology of how HPVs are affectting cell fate choices through E6 during tissue homeostasis. Moreover, it can be a representative case scenario where for the first time we have extended systems virology frameworks, validated with experiments, to DNA tumor viruses as well as the first time to merge sstems virology with tissue biology and biophysics/systems biology of tissues. With this project the fellow could lay the foundations of a new paradigm shifting approach to systems virology and biophysics of tissues. Morover, with STEPV the fellow laid the foundations for the first time to understand tissues under subtle optimized perturbations induced by viruses. This will help the scientific community to filling the gaps in tissue biology to deeper understand how to pass from a healthy condition to a tumorigenic condition in the future. Moreover, unveiling both phenomenologically and mechnistically how HPV is affecting cell fate choices during homeostasis will help start a new paradigm shift in developing novel antiviral therapies targeting the concept of tissue homeostasis hence thinking to develop the antiviral therapy by addressing tissue level features and not only the single cell mechanisms.

Single clones of HPV16 and validated mathematical model of cell fate choices