Mechanisms of Cisplatin Resistance in HPV Negative Head and Neck Squamous Cell Carcinomas

Abstract

Head and neck squamous cell carcinomas (HNSCCs) are the eighth most common cancers worldwide. While promising new therapies are emerging, cisplatin-based chemotherapy remains the gold standard for advanced HNSCCs, although most of the patients relapse due to the development of resistance. This review aims to condense the different mechanisms involved in the development of cisplatin resistance in HNSCCs and highlight future perspectives intended to overcome its related complications. Classical resistance mechanisms include drug import and export, DNA repair and oxidative stress control. Emerging research identified the prevalence of these mechanisms in populations of cancer stem cells (CSC), which are the cells mainly contributing to cisplatin resistance. The use of old and new CSC markers has enabled the identification of the characteristics within HNSCC CSCs predisposing them to treatment resistance, such as cell quiescence, increased self-renewal capacity, low reactive oxygen species levels or the acquisition of epithelial to mesenchymal transcriptional programs. In the present review, we will discuss how cell intrinsic and extrinsic cues alter the phenotype of CSCs and how they influence resistance to cisplatin treatment. In addition, we will assess how the stromal composition and the tumor microenvironment affect drug resistance and the acquisition of CSCs' characteristics through a complex interplay between extracellular matrix content as well as immune and non-immune cell characteristics. Finally, we will describe how alterations in epigenetic modifiers or other signaling pathways can alter tumor behavior and cell plasticity to induce chemotherapy resistance. The data generated in recent years open up a wide range of promising strategies to optimize cisplatin therapy, with the potential to personalize HNSCC patient treatment strategies.

Keywords: HNSCC; cancer stem cells; cell plasticity; cisplatin; epigenetics; tumor microenvironment.

Figure 1

Classical mechanisms of cisplatin resistance in HNSCC. (A) Alterations in cellular import and export of cisplatin. Downregulation of VRAC and OCT3 transporters reduces cisplatin cytotoxicity by decreasing its intracellular concentration. While CTR1/2, ATP7A/B and ABC transporters are promising candidates in cisplatin resistance, their functional role in HNSCC is still unknown. (B) Activation of DNA repair pathways. Upregulation of NER pathway components ERCC1 and XPF, enhances the removal of DNA intra-strand adducts caused by cisplatin. If NER fails, it will produce DNA double-strand breaks. In this case, a deficit in the FA/HR pathway, which carries out the repair of this type of DNA damage, is related to cisplatin resistance but only at low cisplatin levels. On the other hand, MGMT expression increases the removal of cisplatin-adducts by directly binding to them. (C) Enhanced oxidative stress management. Cisplatin induces ROS accumulation, which is counteracted by the overexpression of NRF2. NRF2 upregulates the expression of ROS scavenging proteins, promoting cell survival and, thus, cisplatin resistance. Among these proteins, GLRX5 and SLC7A11 are specifically mentioned, as they are involved in ferroptosis inhibition. Furthermore, NRF2 expression is controlled by EpCAM-IL6/p62 and c-MYC expression. Altogether, these three classical mechanisms converge in cisplatin resistance. Created with BioRender.com (accessed on 1 February 2022).

Figure 2

HNSCC CSCs’ characteristics and their role in cisplatin resistance and tumor relapse. (A) HNSCCs resemble the architecture of an epithelial tissue, with a basal layer enriched in proliferative cells, potentially containing CSCs. Basal cells give rise to differentiated cells that ultimately form terminally differentiated keratin pearls or necrotic regions. (B) HNSCCs present intra-tumoral heterogeneity, containing CSCs pools with different properties, which increases the chances of persisting after cisplatin treatment. CSCs surviving after chemotherapy could repopulate the affected zone causing tumor relapses that are usually poorly differentiated. (C) CSCs’ properties include increased self-renewal promoted by IL-6-induced BMI1 expression, enhanced ROS detoxification driven by NRF2 overexpression and the induction of quiescence or mesenchymal programs through TGFβ. Created with BioRender.com.

Figure 3

Epigenetic reprogramming in cisplatin-resistant HNSCC. (A) In physiological conditions, tissue homeostasis is supported by the expression of the TFs PITX1, TP63 and SOX2. (B) In tumor initiation events, two different landscapes can occur. On one hand, overexpression of oncogenic TFs promotes chromatin accessibility to cancer-specific enhancers, which results in self-renewal, EMT and drug resistance genes increase. On the other hand, the loss of active marks (H3K4me1) and the gain of repressive marks (H3K27me3) near tissue-specific enhancers lead to the silencing of tissue differentiation and pro-inflammatory genes. (C) Tumors adapt to cisplatin by replacing SOX2 with SOX9 in EMT enhancers as well as the deposition of the H3K27Ac mark, allowing the expression of mesenchymal and drug resistance genes. Created with BioRender.com.

Figure 4

TME involvement in chemoresistance development. Tumor cell-extrinsic influence on therapy response can be divided into physical (1) and biological (2) factors. HNSCC cells can recruit surrounding fibroblasts (middle section) through TGFβ signaling, which induces an activation of the CAF phenotype (2.1). CAFs then further produce TGFβ establishing a positive feedback loop between cancer and stromal cells. TGFβ pathway activation promotes the deposition of matrix proteins (COL8A1, COL11A1, FN, HA) and remodeling enzymes (LOX/LOXL, MMPs), which help to reshape and shift the ECM composition. As a result, the matrix becomes stiffer (A), promoting cytoskeletal reorganization within HNSCC cells and thus cell–ECM interactions through increased FAP and integrin expression. Increased cell–ECM adhesion favors the activation of integrin-dependent signaling and mechanosensitive pathways (such as YAP/TAZ), which downregulate cell–cell interactions, enhancing motility and the acquisition of an EMT-resistant phenotype (B). CAFs can also secrete inflammatory cytokines such as IL-6, which can both induce CSC-like properties in HNSCCs (B) and recruit other TME populations, such as immune cells. TAMs (2.2) become active in response to proinflammatory cytokines to further secrete TGFβ, supporting the activation of this pathway. Together with infiltrated T regulatory lymphocytes (2.3), they help to maintain an immunosuppressed tumoral microenvironment which is likely to evade immunotherapy (C). Created with BioRender.com.

Figure 5

Cell-intrinsic (1) and -extrinsic (2) cisplatin resistance mechanisms converge to promote aggressive, invasive and chemoresistant phenotype selection in HPV-negative HNSCC (4). Classic drug resistance mechanisms ((1), also in Figure 1) include efficient detoxification through deregulated transporter expression (1.1), enhanced NER DNA repair (1.2) and increased antioxidant mechanisms via the upregulation of NRF2 (1.3). CSCs displaying this innate chemoresistant behavior ((3.1), also in Figure 2C) may additionally intensify or boost these properties in response to cisplatin treatment (3.2), contributing to resistant clone selection which proliferates and gives way to relapses ((4), also in Figure 2B). Cell-extrinsic factors (2) can be divided into physical/mechanical factors (2.1) and biological components of the TME (2.2). As tumors grow, shear pressure makes the distribution of chemotherapy inefficient and nutrient supply becomes insufficient, triggering hypoxia response programs, which include the upregulation of efflux transporters (1.1) and ECM remodeling components (LOX/LOXL enzymes, collagens and MMPs), which cleave and restructure the ECM, releasing embedded factors such as VEGF, involved in neoangiogenesis. Other factors contributing to ECM remodeling are non-tumoral cells within the TME (2.2), which establish an intricate crosstalk with the neighboring tumor. Important interactions include bidirectional TGFβ and IL-6 feedback loops between tumoral and non-tumoral populations (CAFs, TAMs and Tregs), which induce ECM protein deposition and stiffening (Figure 4). This modifies the adhesive distribution within tumoral cells, increasing cell–ECM interactions and triggering mechanosensitive pathways. These ultimately promote EMT expression programs, favoring invasive and resistant behavior (4). Created with BioRender.com.