Non-coding RNAs in lung cancer: emerging regulators of angiogenesis

Lung cancer ranks second in incidence among all tumor types worldwide and causes almost one-fourth of cancer deaths. Non-small cell lung cancer (NSCLC) accounts for more than 80% of lung cancers and is the most common histological type of lung cancer [1, 2]. Owing to the highly aggressive nature of NSCLC and the difficulty in diagnosing it at an early stage, the majority of NSCLC patients already have advanced-stage disease at diagnosis. The standard treatment for advanced NSCLC relies on systemic chemotherapy, targeted therapy, immunotherapy, and radiotherapy [3]. The overall survival of lung cancer patients has improved with advancements in treatment methods and earlier diagnosis in recent decades. However, due to distant metastasis and tumor recurrence after treatment, the outcomes of lung cancer patients are still poor [4].

Angiogenesis refers to the formation of new blood vessels from preexisting capillaries or postcapillary venules and is one of the hallmarks of cancer [5]. Unlike physiological conditions, such as wound healing, tumor angiogenesis is persistently aberrant in growing tumors, as they require oxygen and nutrients delivered through the blood circulation system to survive and proliferate [6]. In addition, cancer cells can enter the blood circulation through angiogenesis, resulting in hematogenous tumor metastasis [7, 8]. Over the past few decades, accumulating evidence has revealed that angiogenesis is a vital cancer hallmark related to poor prognosis in various solid tumors [9, 10]. Anti-angiogenic agents can affect the tumor microenvironment to regress existing tumor vessels while inhibiting tumor angiogenesis [11‐13]. However, anti-angiogenic drugs targeting vascular endothelial growth factor (VEGF) or the vascular endothelial growth factor receptor (VEGFR) rarely result in durable responses and have had a limited effect on improving the overall survival of patients with lung cancer [14]. Consequently, to exploit new therapeutics to overcome this poor efficacy, further research is needed to discover the mechanisms of angiogenesis in lung cancer.

Angiogenesis is a complex process regulated by many pro-angiogenic and anti-angiogenic genes, and numerous studies have confirmed that angiogenesis is crucial for lung cancer cell proliferation and metastasis [15]. Accumulating evidence suggests that non-coding RNAs (ncRNAs), especially microRNAs (miRNAs) and long non-coding RNAs (lncRNAs), can be involved in tumor angiogenesis by controlling different genes and pathways, resulting in distant metastasis and tumor recurrence [16]. Vascular homeostasis is managed by many different pro- and anti-angiogenic genes, and VEGF has been identified as a critical gene in inducing lung cancer angiogenesis [17‐19]. The binding of VEGF and VEGFR initiates various intracellular signaling pathways and mediates the survival, proliferation, and migration of vascular endothelial cells (ECs), in turn promoting angiogenesis and enhancing vascular permeability in lung cancer [20]. Additionally, VEGF can also be produced by tumor immune cells and regulate the functions of innate and adaptive immune cells towards immunosuppression [21]. Hypoxia is the most crucial inducer of angiogenesis, and hypoxia-inducible factors (HIFs) have a crucial function in physiological adaptation to hypoxic states [22]. HIF proteins, especially HIF-1 alpha (HIF-1α), are closely associated with lung cancer occurrence, metastasis, and angiogenesis [23‐25]. Multiple studies have indicated that the activation and expression levels of HIF-1α are closely correlated with the outcome of lung cancer [26, 27]. HIF-1α can target VEGFA and regulate its expression at the transcriptional level. Targeting the HIF-1α/VEGFA axis could be a promising strategy against tumor angiogenesis [28].

To date, the US Food and Drug Administration (FDA) has approved various drugs targeting VEGF/VEGFR for tumor anti-vascular therapy (Fig. 1) [29]. Although these drugs have demonstrated efficacy in NSCLC patients, the application of drugs is still affected by clinically significant bleeding events, such as major hemoptysis. On the other hand, long-term VEGF/VEGFR inhibitors probably lead cancer cells to exploit different angiogenic mechanisms and develop drug resistance [30]. Hence, full understanding of the mechanism of tumor angiogenesis will facilitate the discovery of novel therapeutic approaches to improve the prognosis of patients. NcRNAs have become promising biomarkers and potential therapeutic tools in many cancers because of their high stability and good biocompatibility [31‐33]. Since ncRNAs act as pro- or anti-angiogenesis factors, their function in angiogenesis in lung cancer can be divided into enhancing or inhibiting, which is recapitulated in Table 1.

NcRNAs are crucial regulatory factors in tumorigenesis that activate or inhibit the oncogenic process. Therefore, advancing practical therapeutic approaches to suppress oncogenic ncRNAs or overexpress cancer-associated ncRNAs is becoming a hot research field [93]. Eleven ncRNA-based therapies, all of which are small interfering RNAs (siRNAs) or antisense oligonucleotides (ASOs) targeting specific genes, have been approved for clinical treatment. Moreover, many ncRNA-based therapeutics are in clinical development with potential applications in various tumors, including lung cancer [94]. The ncRNA-based anti-angiogenesis therapies in lung cancer presented in Table 2.

Nucleic acid aptamers are small, single-stranded DNA or RNA molecules chemically synthesized for binding to a specific target. As a therapeutic tool, aptamers have several advantages, such as their small physical size and lack of immunogenicity, and are thus an invaluable targeted delivery carrier for siRNAs, miRNAs, and chemotherapeutic agents [95]. Lai et al. constructed two nucleolin aptamer-siRNA chimeras targeting snail family zinc finger 2 (SLUG) and neuropilin 1 (NRP1). Combined treatment with these two aptamer siRNAs significantly silenced SLUG and NRP1 expression in lung cancer cells, leading to specific inhibition of tumor invasion and angiogenesis [96]. Liao et al. constructed a bivalent cyclic RGD–siVEGFR2 conjugate delivery system that can silence VEGFR2 expression by targeting neovascularization in endothelial cells. Furthermore, biRGD–siVEGFR2 exhibited synergistic antitumor activity with apatinib in NSCLC, which may represent a new strategy for clinical NSCLC treatment [97]. Activation of CXCR4 and STAT3 results in the initiation of multiple signaling pathways, leading to metastasis and angiogenesis in lung cancer; thus, CXCR4 and STAT3 are potential targets for anti-angiogenic therapy [98]. Li et al. developed a fluorinated polymeric CXCR4 antagonist-stabilized perfluorocarbon (PFC) to deliver therapeutic siSTAT3. These nanoemulsions suppressed tumor proliferation and angiogenesis by simultaneously down-regulating CXCR4 and STAT3 in lung metastases [99].

Dicer, a type III cytoplasmic endoribonuclease, is required for the maturation of the vast majority of miRNAs [100]. Chen et al. generated miRNA-deficient tumors by knocking out Dicer1 only and found that the depletion of all miRNAs notably suppressed NSCLC angiogenesis. Mechanistic studies suggested that miRNAs promote tumor responses to hypoxia and angiogenesis by repressing FIH1, an inhibitor of HIF-1α [101]. Previous research has shown that malaria infection could suppress Lewis lung cancer cell proliferation by inducing innate and adaptive antitumor immune responses [102]. Yang et al. found that intratumoral injection of exosomes derived from the plasma of Plasmodium-infected mice significantly suppressed Lewis lung cancer growth. High levels of miR-16/322/497/17 were detected in exosomes isolated from the plasma of mice infected with Plasmodium, and up-regulated expression of these miRNAs in ECs corresponded with down-regulated expression of VEGFR2 [103]. This study helped to advance potential exosome-based anti-angiogenic tumor therapeutics.

Muramatsu et al. found that miR-125b inhibits EC tube formation by inhibiting the translation of VE-cadherin. Then, they used non-viral vectors composed of the cationic polymer polyethylenimine to package miR-125b and directly injected it into subcutaneous tumors. By targeting VE-cadherin, miR-125b induced the formation of non-functional blood vessels and inhibited tumor growth in vivo, which suggested a therapeutic potential for tumor angiogenesis [104]. Li et al. designed a therapeutic strategy against orthotopic NSCLC tumors that can intelligently co-deliver siVEGF and the chemotherapeutic etoposide (ETO) through multi-functional nanoparticles (NPs). Compared with monotherapy, combination therapy in orthotopic NSCLC causes more significant tumor growth and metastasis by simultaneously inhibiting tumor proliferation and angiogenesis [105]. Therefore, ncRNA-based anti-angiogenic therapy may be a promising strategy.

Circulating miRNAs may potentially act as biomarkers for predicting the response to anti-angiogenic therapy. M. Joerger’s study revealed that circulating miRNAs could predict the efficacy of angiogenic drugs combined with targeted therapies. Pretreatment circulating miRNA profiling indicated that the expression of 12 miRNAs was significantly associated with tumor shrinkage after bevacizumab/erlotinib treatment, with miR-665 being the strongest predictive marker. Moreover, miR-223 was related to time to progression (TTP) after bevacizumab/erlotinib treatment andafter second-line chemotherapy [106].

Tumor metastasis and chemotherapeutic resistance lead to the poor prognosis of lung cancer. Current research suggests that ncRNAs are crucial players in tumorigenesis and tumor angiogenesis in lung cancer. Even though the function of ncRNAs has been determined in the tumor angiogenesis, less attention has been given to research on the clinical application of ncRNAs, suggesting that the requirement for ncRNA studies is unmet. Nucleic acid therapeutics targeting angiogenesis, such as modified siRNAs and miRNA mimics, are promising therapeutics for lung cancer. In conclusion, despite the emerging application of ncRNAs in tumor diagnosis and therapy and the challenges in this field, targeting ncRNAs is still an innovative and promising strategy to improve therapeutic outcomes for lung cancer patients.