Tumour response to hypoxia: understanding the hypoxic tumour microenvironment to improve treatment outcome in solid tumours

Abstract

Hypoxia is a common feature of solid tumours affecting their biology and response to therapy. One of the main transcription factors activated by hypoxia is hypoxia-inducible factor (HIF), which regulates the expression of genes involved in various aspects of tumourigenesis including proliferative capacity, angiogenesis, immune evasion, metabolic reprogramming, extracellular matrix (ECM) remodelling, and cell migration. This can negatively impact patient outcomes by inducing therapeutic resistance. The importance of hypoxia is clearly demonstrated by continued research into finding clinically relevant hypoxia biomarkers, and hypoxia-targeting therapies. One of the problems is the lack of clinically applicable methods of hypoxia detection, and lack of standardisation. Additionally, a lot of the methods of detecting hypoxia do not take into consideration the complexity of the hypoxic tumour microenvironment (TME). Therefore, this needs further elucidation as approximately 50% of solid tumours are hypoxic. The ECM is important component of the hypoxic TME, and is developed by both cancer associated fibroblasts (CAFs) and tumour cells. However, it is important to distinguish the different roles to develop both biomarkers and novel compounds. Fibronectin (FN), collagen (COL) and hyaluronic acid (HA) are important components of the ECM that create ECM fibres. These fibres are crosslinked by specific enzymes including lysyl oxidase (LOX) which regulates the stiffness of tumours and induces fibrosis. This is partially regulated by HIFs. The review highlights the importance of understanding the role of matrix stiffness in different solid tumours as current data shows contradictory results on the impact on therapeutic resistance. The review also indicates that further research is needed into identifying different CAF subtypes and their exact roles; with some showing pro-tumorigenic capacity and others having anti-tumorigenic roles. This has made it difficult to fully elucidate the role of CAFs within the TME. However, it is clear that this is an important area of research that requires unravelling as current strategies to target CAFs have resulted in worsened prognosis. The role of immune cells within the tumour microenvironment is also discussed as hypoxia has been associated with modulating immune cells to create an anti-tumorigenic environment. Which has led to the development of immunotherapies including PD-L1. These hypoxia-induced changes can confer resistance to conventional therapies, such as chemotherapy, radiotherapy, and immunotherapy. This review summarizes the current knowledge on the impact of hypoxia on the TME and its implications for therapy resistance. It also discusses the potential of hypoxia biomarkers as prognostic and predictive indictors of treatment response, as well as the challenges and opportunities of targeting hypoxia in clinical trials.

Keywords: cancer associated fibroblasts; extracellular matrix; hypoxia; immune cells; tumour microenvironment.

Figure 1

The hypoxic tumour microenvironment (TME). Hypoxia plays an important role in the development of the TME. The TME is composed of hypoxic tumour cells, cancer stem cells, tumour cells, immune cells, cytokines/chemokines, collagen, fibronectin, cancer associated fibroblasts (CAFs), endothelial cells and, blood vessels. As the tumour cells grow, the tumour cells further away from the blood supply have limited access to oxygen and become hypoxic tumour cells. Additionally, the TME undergoes a metabolic switch to meet the demands of the TME which involves increased glucose uptake and production of lactate resulting in an acidic TME characterised by a decreasing pH. These changes result in a change in cytokine/chemokine release, a change in immune cell phenotype, modification of the extracellular matrix, and activation of CAFs. Together, these form a more pro-tumorigenic environment prone to increased invasive and metastatic potential, as well as increased resistance to chemoradiotherapy. Created with BioRender.com.

Figure 2

Hypoxia influences the development of a cancerous ECM. Hypoxia in the ECM increases collagen (COL) and fibronectin (FN) deposition, as well as secretion of metalloproteinases (MMPs), lysyl oxidases (LOX) and prolyl 4-hydroxylase subunit alpha (P4HA) 1 and 2. Increased MMPs, LOX and P4HA1/2 promote the generation of organised aligned COL and FN fibre tracks in the ECM, enhancing cell migration. Hypoxia also induces the secretion of growth factors and cytokines, which are also released due to ECM remodelling, establishing a synergistic effect. Release of growth factors (e.g. transforming growth factor β [TGF-β], endothelial growth factor [EGF], fibroblast growth factor [FGF]) enhance not only cancer cell growth and survival, but also recruitment of cancer-associated fibroblasts (CAFs) and tumour associated macrophages (TAMs). TAMs and CAFs participate in the secretion of growth factors, ECM remodelling and COL/FN deposition, increasing the synergistic effect. Under hypoxic stress, angiogenesis is activated through secretion of angiogenic growth factors (e.g. vascular endothelial growth factor [VEGF], angiopoietin [ANG]). The angiogenic process allows for the development of new blood vessels, enhancing ECM remodelling during the process. Additionally, an organised hypoxic ECM provides migratory tracks directing cells towards blood vessels and enhancing intravasation. Pro-tumorigenic growth factors and enzymes (e.g. TGF-β, LOX) can intravasate and travel to distant healthy tissues, generating pre-metastatic niches through ECM remodelling. The same migratory tracks enhance cancerous cell migration and intravasation, allowing them to circulate and eventually colonise the pre-metastatic niches and seed new tumour cells. Created with BioRender.com.

Figure 3

CAF activation pathways and downstream effects. CAFs are derived from multiple cell types and activated by multiple molecules including hedgehog (Hh), transforming growth factor-β (TGF-β), reactive oxygen species (ROS), interleukin-1β (IL-1β), fibroblast growth factor (FGF), platelet-derived growth factor (PDGR), stromal cell-derived factor 1 (SDF-1), heparin binding growth factor (HBGF) which can be driven by hypoxia. Precursor cells including mesenchymal stem cells (MSCs) are a source of CAFs activated by CXCL-12 and TGF-β derived from tumour cells. Pericytes, fibrocytes, stellate cells and adipocytes are also recruited by tumours by CXCL-12 and TGF-β, and are activated by TGF-β and PDGF. CAFs can derive from mature epithelial cells that differentiate into functional CAFs by TGF-β mediated epithelial-mesenchymal transition (EMT). Endothelial cells undergo EndoMT to differentiate into CAFs through TGF-β and SMAD signalling. HIF and TGF-β have a role in the function of CAFS. Genes associated with changes in ECM remodelling, metabolic reprogramming, angiogenesis, immune response and metastasis are direct transcriptional targets of HIF in CAFs or cancer cells. This creates bi-directional communication between CAFs and cancer cells through release of cytokines and chemokines (blue dots) which promotes proliferation of both cells, and further enhancement of pro-tumorigenic pathways (Adapted from (181, 182)). Created with BioRender.com.

Figure 4

Tumour reoxygenation. Normoxic tumour cells are killed by irradiation which induces a reduction in tumour bulk. Oxygen consumption is reduced by normoxic cells which enables oxygen to diffuse to hypoxic regions. This induces vessel regrowth and reoxygenation of hypoxic cells. The reoxygenated cells are more sensitive for reirradiation. Created with BioRender.com.