Tumour plasticity

Topic description

Specific Challenge:

The last decade has seen tremendous advances in the development of effective targeted therapies as well as in immuno-oncology to more effectively treat cancer. Despite this, cures are still rare in the metastatic setting. In most cases, an initial response to treatment is followed by the eventual emergence of drug resistance . Drug resistance in cancer is one of the greatest causes of mortality and despite increasing success with targeted therapies in the clinic (including immunotherapy), the mechanisms by which cancer cells evade cell death are still not well understood. Drug combinations are likely to be critical to overcoming drug resistance but are dependent on identifying the cellular programmes that cancer cells use to resist therapeutic agents.

In tumours that initially respond to treatment, rare cancer cells can survive and withstand therapy (‘Drug Tolerant Persister’ cells, DTPs) and can act as a reservoir for the eventual emergence of drug resistance. Furthermore, these studies have shown that these cells are able to survive drug treatment by altering the transcriptional state of specific signalling pathways, and that in the early stages such changes are plastic and reversible but that over time these changes become stable and fixed.

Recent technological advances in single-cell sequencing have revolutionised the study of individual cells within cancer populations and, importantly, would allow the characterisation of DTPs, something previously impossible with bulk sequencing technologies . Single-cell sequencing provides information that is not confounded by genotypic or phenotypic heterogeneity of bulk samples. Importantly, it has confirmed the existence of DTPs in patients following treatment response and, more importantly, the characterisation of the transcriptionally altered pathways in DTPs. Characterising the transcriptionally altered pathways in persister cells, the biological processes they regulate and their druggability will be critical to future drug combination strategies, with the goal of preventing or significantly delaying the development of drug resistance.

There are numerous challenges in applying single cell sequencing to arguably one of the most important barriers to curing in cancer today – drug resistance, and specifically:

• Defining best sequencing protocols – single-cell RNA-sequencing (scRNA-seq) is a fast-moving field with a recent benchmarking paper comparing 13 different methods.
• Computational approaches to big data – as with sequencing methods, the analysis framework is constantly evolving and there are challenges in integrating data across studies and platforms
• Standardisation of data formats
• Best practice single cell collection from in vitro and in vivo model systems
• Application of single-cell sequencing to clinical samples
• Spatial imaging technologies
• Biological interpretation of data, including novel target identification

This topic proposes to apply state-of-the-art single-cell sequencing technologies to characterise cancer cell populations pre-treatment, at minimal residual disease (for DTPs) and upon the acquisition of drug resistance and from a variety of pre-clinical human and mouse models as well as clinical samples.

The overall objective of the call topic is to use state-of-the-art single-cell sequencing to understand and overcome drug resistance in cancer by characterising the biology of drug tolerant persister cells, building the capability for such studies across Europe.

The call topic will address primarily adult tumours, with the provision to include childhood tumours where appropriate models are available at a later stage of the program. To optimise our ability to determine the role of tissue lineage on the biological processes observed in single-cells, we propose that the majority (>80%) of the single-cells should be provided from drug treatments in 3 adult cancers:

• non-small cell lung cancer (NSCLC)
• breast cancer
• colorectal cancer

The goals of the call topic are:

• To characterise the biology of drug tolerant persister cells - defining the signalling pathways and cellular processes that enable DTPs to survive drug treatment and thereby identify novel drug targets to overcome this – using state-of-the-art single-cell sequencing and spatial transcriptomics in a range of cancer models.
• To better understand the tumour microenvironment – to avoid solely focusing on cell intrinsic drug resistance programmes, a key element of the work packages should be to use spatial imaging techniques to explore the interaction between cancer cells and the microenvironment.
• Generation of single cell RNAseq data from adult and childhood cancers – although the pre-clinical models used to explore the biology of drug treatment in cancer are predominantly based on adult cancers, drug resistance is equally a major problem in childhood tumours. The applicants should anticipate that from year 3 of the funded project, specific childhood cancers could be considered for inclusion where the appropriate models are accessible and where there is a hypothesis relationship with drugs or tumours being investigated by the consortium.
• To develop best practice in clinical validation and single-cell sequencing – clinical validation will be key to translation of any findings and a change in clinical practice. To include informed patient consent forms that cover all intended uses, including clinical outcome data and sharing of data inside the consortium and with 3rd parties. GDPR-compliant tracking of patient data, samples and PDXs.
• To create gold standard protocols for single cell collection – across a range of models and to include differing methods for isolating single cells from human (organoids, clinical biopsies) and mouse (PDX, genetically engineered mouse models (GEMM) and syngeneic mice) model systems.
• To develop core analytical methods – use pre-treatment, on-treatment and post-treatment single-cell sequencing data to develop novel computational approaches to identify the different subtypes of cancer cells present and the biological processes that are complicit in maintaining their survival following drug treatment.

To build EU capability in single-cell sequencing – in the process of developing the protocols for single cell collection, sequencing and analysis, the funded project will put in place infrastructure to enable other groups in the EU to carry out similar single-cell sequencing studies in both cancer and non-cancer models.

A comprehensive database, profiling DTPs across a range of cancers and therapies would enable a deeper understanding of the biology of DTPs and allow cross-tumour studies.

Impact for Patients

• Identification of novel drug targets in DTPs and resulting drug combinations that delay or prevent the emergence of drug resistance in cancer
• Better understanding of the contribution of tumour heterogeneity and plasticity to disease outcome, progression and relapse

Impact for Academia and SMEs

• Harmonisation of protocols for single cell experiments
• Enhanced infrastructure in the EU for single cell sequencing
• Development of gold standards for the analysis of single-cell sequencing data
• Access to comparative data on different pre-clinical and clinical models and better understanding of the biology of DTPs in cancer with a high likelihood of spin-off projects
• Improvements in single cell sequencing and spatial imaging with potential for commercial development
• Better understanding of drug development post-novel target identification

Impact for Industry

• Access to a data source for further functional studies (e.g. KO, knock-out, knock-in, target perturbation) that will lead to opportunities for identification of novel targets in DTP space - pointing to new targets or rational drug combinations that alter the drug resistance paradigm
• Access to single cell measurement data combined with outcome data (models) and clinical outcome data
• Development of expertise in the analysis of single-cell sequencing data

Gold standard methods for the delivery of single cell projects