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Non-Genetic Mechanisms of Therapeutic Resistance in Cancer

Therapeutic resistance has repeatedly proved itself to be an unconquerable barrier for curative cancer treatment. However, recent investigations have challenged the common belief that genetic evolution is the main cause of tumour resistance. A recent study, published by Nature, explores a range of non-genetic adaptive mechanisms which contribute to this process.

Introduction

Dreams of achieving personalised cancer care draw closer each day as we learn more about the molecular pathogenesis of cancer. These therapies seek to eradicate cancer cells by interfering with specific target molecules essential for tumour development. In recent decades, a plethora of targeted therapies have been developed and have revolutionised patient outcomes. However, many patients with cancer, especially those at later stages of the disease experience therapeutic resistance and disease recurrence.

This resistance often arises from genetic evolution. A group of malignant cells may carry or mutate and acquire specific genetic alterations which provide these cells with a clonal advantage, allowing them to escape the therapeutic pressure. However, in many cancers, there is an absence of a clear genetic cause for this resistance.

Although non-genetic mechanisms of tumour resistance remain largely elusive, a recent study highlights potential non-genetic sources and how these converge with known molecular mechanisms to allow drug adaptation and resistance.

Therapeutic Resistance in Cancer

Acquired resistance is defined as a recurrence of malignancy following an initial clinical response to therapy. Acquired resistance is often portrayed as the genetic evolution of cancer in response to a therapeutic challenge. However, there is growing evidence that non-genetic adaptation of cancer cells contributes significantly to resistance to cancer therapies.

For example, single cell sequencing of tumour cells has suggested that transcriptional or metabolic adaptation may be more common than genetic evolution in many of these tumours. For example, the expression of a particular transcriptional programme may provide intrinsic resistance to a particular drug. Similarly, a cancer cell may adapt by rewiring its gene expression to programmes which offer a selective advantage without the acquisition of novel genetic mutations.

It is likely that most, if not all cancers leverage multiple different processes for therapeutic evasion. It is also important to note that these processes are not mutually exclusive. Thus, therapeutic strategies which only target genetic events are likely to be ineffective. Instead, we should focus our attention on identifying the common and cell type-specific mechanisms of resistance and use this to inform new therapeutic strategies.

Non-Genetic Mechanisms of Evolution:

Single cancer cells may be able to produce multiple different phenotypes and switch between them without genomic alterations. For example, chemotherapy has been demonstrated to induce phenotype-switching in various epithelial tumours, such as colon, lung and breast cancers. This is known as the epithelial-to-mesenchymal transition (EMT).

Re-treating patients with the same therapy after relapse is often ineffective. This remains true even if the patient has not been exposed to the therapy for months or years. For example, mice models and human data for acute myeloid leukaemia have shown evidence of non-genetic therapeutic resistance. Cancer cells which caused patient relapse were present before therapy began and did not acquire novel mutations over the course of treatment. Instead, these cells adopted a transcriptional state which was drug tolerant and allowed them to actively proliferate under therapeutic pressure. Compared to drug-tolerant persister cells, these cells can maintain their resistance phenotype even when withdrawn from drug pressure for long periods of time. 

This transcriptional plasticity is called ‘enhancer switching’ and is used by stem and progenitor cells to maintain the expression of important genes. Cancer cells can hijack these developmental programmes and the associated enhancer remodelling machinery, allowing non-genetic adaptive resistance to a range of anticancer therapeutics. These cancer stem-like cells likely have important contributions to therapy resistance.

The Interaction of Genetic and Non-Genetic Factors

The interplay between genetic mutations and the evolving non-genetic landscape is widespread. Thus, it is highly important that these processes are understood collectively, rather than in isolation.

For example, there is evidence that drug tolerant cells in may jumpstart classic mutation-based somatic evolution. Specifically, colorectal cancer cells showed repression of genes required for repair pathways and instead increased the expression of error-prone DNA polymerases. This demonstrates paradoxical enhancement of adaptation at a competing intrinsic fitness cost, allowing a rapid expansion of genetic diversity. 

Lineage plasticity also plays a key role. Genetic mutations can provide the cue for cellular identity and direct differentiation of cancer cells. Thus, cellular plasticity can enable immune evasion in cancer. Multiple studies have identified the role of transcription factors that govern lineage plasticity. For example, the inhibition of enhancer of zeste homologue 2 (EZH2), the catalytic component of Polycomb repressive complex 2 (PRC2), was able to reverse the transformation of certain cancer cells. This suggests that processes mediated by the chromatin complex are essential for the maintenance of the trans-differentiated state. 

Monitoring Mechanisms of Resistance

An understanding of the mechanisms used by resistant cells which persist even when clinical remission is reached is essential. Single-cell RNA sequencing analyses of tumour samples before and after therapy have shown to be informative for highlighting resistance in several tumour types. These techniques have also been applied to circulating tumour DNA and cells within the tumour microenvironment.

Unfortunately, the sensitivity, resolution and range of proteins and metabolites detected by these methods remains limited. They also require access to serial tumour samples, which is often not practical as this involves an invasive procedure. 

Furthermore, at clinical remission, the sampling of MRD cells is not usually technically and ethically feasible. Additionally, in the context of multifocal disease, a single tumour-biopsy sample is often not representative of the global tumour burden. Significant technical and conceptual advances will be needed before these approaches can inform clinical care in real time. 

Targeting Non-Genetic Resistance

It is clear that there will not be a single universal or uniform effective clinical approach for targeting non-genetic resistance. The combination of current treatments may be required and adapted on a case-by-case basis. 

Drug-tolerant persister cells could be targeted by directed phenotype switching. This describes the leveraging of cell plasticity for therapeutic benefit, to transform a drug-resistant population into one that is drug-sensitive. Alternatively, these resistant cells could be diverted into a permanently dormant state. This will experimentally require the barcoding and lineage tracing approaches to understand the mechanisms that underpin the re-emergence of  these cells. 

Furthermore, developing epigenetic therapies which restrain cellular plasticity is a major challenge. It is difficult to predict which tumours undergo therapeutic evasion and there are few detailed molecular insights into the process of this trans-differentiation. However, PRC2 which is required for cell fate decisions appears to have a central role in enabling and maintaining the trans differentiated state. PRC2 inhibitors could be used in patients after remission has been reached in an attempt to prevent relapse.

The development of curative therapies will require further study, especially at single-cell resolution to gauge the challenge of mechanisms which drive resistance and relapse. Together, these technological and conceptual advances will facilitate both the development of innovative therapies and more informed therapeutic approaches to improve outcomes for patients with cancer.


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Cancer / Chemoresistance / Epigenetics