Angiogenesis In Lung Cancer Research Articles

EGPI-1, a novel eIF4E/eIF4G interaction inhibitor, inhibits lung cancer cell growth and angiogenesis through Ras/MNK/ERK/eIF4E signaling pathway

eIF4E plays an important role in regulating tumor growth and angiogenesis, and eIF4E is highly expressed in a variety of lung cancer cell lines. siRNA eIF4E can significantly inhibit the proliferation of lung cancer cells, indicating that inhibition of eIF4E may become a novel anti-tumor target. In the previous study, we synthesized a series of small molecule compounds with the potential to inhibit eIF4E. Among them, the compound EGPI-1 significantly inhibited the proliferation of a variety of lung cancer cells such as A549, NCI-H460, NCI-H1650 and 95D without inhibiting the proliferation of HUVEC cells. Further studies found that EGPI-1 interfered with the eIF4E/eIF4G interaction and inhibited the phosphorylation of eIF4E in NCI-H460 cells. The results of flow cytometry showed that EGPI-1 induced apoptosis and G0/G1 cycle arrest in NCI-H460 cell. Interestingly, we also found that EGPI-1 induced autophagy and DNA damage in NCI-H460 cells. The mechanism results showed that EGPI-1 inhibited the Ras/MNK/ERK/eIF4E signaling pathway. Moreover, EGPI-1 inhibited tube formation of HUVECs, as well as inhibited the neovascularization of CAM, proving the anti-angiogenesis activity of EGPI-1. The NCI-H460 xenograft studies showed that EGPI-1 inhibited tumor growth and angiogenesis in vivo by regulating Ras/MNK/ERK/eIF4E pathway. Our studies proved that eIF4E was a novel target for regulating tumor growth, and the eIF4E/eIF4G interaction inhibitor EGPI-1 was promising to develop into a novel anti-lung cancer drug.

Protein S-mediated signal transduction pathway regulates lung cancer cell proliferation, migration and angiogenesis

Objective/BackgroundProtein S (PS; encoded by the PROS1 gene), a key vitamin K-dependent anticoagulant protein, is emerging as a key structural and functional protein that is overexpressd in various malignancies, but how PS signals to promote lung cancer progression is unclear. MethodsWe used immortalized, nontumorigenic human lung epithelial cell line NL-20, A549 cells as experimental cellular models for lung cancer, and human microvascular endothelial cells (HMEC-1) as a model system for angiogenesis. A loss- and gain-of-function approach was then used to analyze the role of tumor-derived PS and their natural TAM receptors Tyro3 and MerTK in regulating cell proliferation, migration, anchorage-independent growth, and capillary-like tube formation, all prominent attributes of the metastatic phenotype of tumor cells. ResultsEvidence is now provided that regulation of PROS1 gene expression using either stable cell lines expressing lentiviral-short hairpin RNA (shRNAs) or a replication-incompetent adenovirus alters the phosphorylation of several major signaling pathways, including Erk, PKB/Akt, p38, and focal adhesion kinase (FAK), and modulates PS-dependent Tyro3- and MerTK-mediated cell migration, proliferation, and anchorage-independent growth of lung cancer cells, and endothelial cell capillary-like tube formation. ConclusionThese finding suggest that the PS-Tyro3 and -MerTK axis mediates important signaling pathways to promote lung cancer progression. Genetic inhibition of endogenous PS may serve as a promising target for anticancer drug development.

Circulating mRNA Expression of Astrocyte-Elevated Gene-1 Associated with Treatment Response and Survival in Non-Small Cell Lung Cancer Patients Treated with Pemetrexed

Background: Astrocyte-elevated gene-1 (AEG-1) functions as an oncogene and regulates angiogenesis in non-small cell lung cancer (NSCLC). In this prospective study, we assessed the values of plasma AEG-1 mRNA expression by liquid biopsy associated with tumour response and survival in NSCLC patients treated with pemetrexed. Methods: Patients diagnosed with advanced NSCLC were enrolled to be treated with pemetrexed combined with platinum as first-line chemotherapy. All patients underwent blood sampling before any cancer treatment (C0) and at first response evaluation after two cycles (C2) of treatments. Response to chemotherapy and survival were assessed. Plasma mRNA was extracted from peripheral blood mononuclear cell (PBMC) and quantification of RNA was performed by real-time PCR. Results: A total of 50 patients with advanced NSCLC were included and 13 of 50 patients combined with bevacizumab. In patient groups of stable disease (SD) (n = 13) and progressive disease (PD) (n = 10), the plasma mRNA of AEG-1, thymidylate synthase (TS), and CK19 were elevated significantly at C2 compared to patients in treatment response group (PR, n = 27) (PR vs. SD or PD, AEG-1: 1.22 ± 0.80 vs. 4.51 ± 15.45, p = 0.043). NSCLC patients who had elevated AEG-1 (AEG-1 ≥ 2) after two cycles of chemotherapy had shorter PFS and OS (high AEG-1 vs. low AEG-1, median, PFS: 5.5 vs. 11.9 months, p = 0.021; OS: 25.9 vs. 40.8 months, p = 0.019, respectively). In a Cox regression analysis, increased plasma mRNA expression of AEG-1indicated poor prognosis in survival. Conclusions: Circulating mRNA concentration of AEG-1 could be a predictive and prognostic biomarker in NSCLC patients treated with pemetrexed. Increased expression of AEG-1 contributed to the chemoresistance and caused lung cancer progression.

PIGF and Flt-1 on the surface of macrophages induces the production of TGF-β1 by polarized tumor-associated macrophages to promote lung cancer angiogenesis

BackgroundThe interaction between tumor cells and tumor microenvironment is a necessary condition for promoting the metastasis of malignant tumors. MethodsTwo different transwell culture systems were interfered with by recombinant factor placental growth factor (re-PIGF) and the re-PIGF + transforming growth factor-β1 (TGF-β1)-neutralizing antibody (anti-TGF-β1). We performed immunofluorescence, flow cytometry and enzyme linked immunosorbent assay (ELISA) to analyze the expression of PIGF, fms-like tyrosine kinase-1 (Flt-1), macrophage marker F4/80 +, macrophage M2 marker CD163+ and TGF-β1 in vitro. Meanwhile, cell viability assay and optical microscope assay were conducted to explore the cell viability and vascularization ability of human umbilical vein endothelial cells (HUVECs). ResultsRe-PIGF increased the expression of PIGF in A549 cells and the expression of Flt-1 in BM-Mac cells, and significantly enhanced the ability of bone marrow-derived macrophages (BM-Mac) to transform into macrophages. At the same time, re-PIGF increased the expression of cytokine TGF-β1 in A549 cells/BM-Mac transwell culture system. On the contrary, re-PIGF + anti-TGF-β1 inhibited the expression of Flt-1 in BM-Mac cells and inhibited the ability of BM-Mac cells to transform into macrophages. Finally, re-PIGF + anti-TGF-β1 reduced the cell viability and angiogenesis of HUVECs. ConclusionThe surface molecule PIGF of lung cancer cells could bind to the receptor Flt-1 on the surface of macrophages, thereby increasing the production of TGF-β1, and ultimately promoting the formation of angiogenesis in lung cancer.

P70.11 Prevalence and Clinicopathological Characteristics of ARID1A Mutations in East Asian Patients With Non-Small Cell Lung Cancer

The AT-rich interacting domain-containing protein 1A gene (ARID1A) encodes ARID1A, a member of the SWI/SNF chromatin remodeling complex. Notably, ARID1A is mutated in over 50% of ovarian clear cell carcinomas and 30% of ovarian endometrioid carcinomas. ARID1A mutation is a known genetic driver of ovarian cancer. However, the role of ARID1A in the angiogenesis of human non-small cell lung cancer (NSCLC) remains unclear. The aim of this study is to investigate mutations and prognosis of NSCLC harboring ARID1A mutations A total of 234 patients with non-small-cell lung cancer were recruited between July 2012 and December 2018. The status of ARID1A mutations and other genes were detected by next generation sequencing. ARID1A gene mutation rate was 2.56% (6/234) in non-small cell lung cancer, including Q909*(1 patient), S634*(1 patient), E1104*(1 patient), H1384Tfs*97(1 patient), S1791L(1 patient) and G1711A plus Q611*(1 patient), and median overall survival (OS) for these patients was 20.0 months. Among them, all patients were ARID1A gene with co-occurring mutations. Briefly, patients with (n=2) or without (n=4) co-occurring ALK mutations had a median OS of 23.0 months and 11.0 months respectively (P=0.35); patients with (n=2) or without (n=4) co-occurring BRCA2 mutations had a median OS of 23.0 months and 11.0 months respectively (P=0.35); patients with (n=3) or without (n=13) co-occurring SMARCA4 mutations had a median OS of 23.0 months and 11.0 months respectively (P=0.35); patients with (n=2) or without (n=4) co-occurring KEAP1 mutations had a median OS of 18.0 months and 16.0 months respectively (P=0.73). Our data reveal ARID1A mutations represent a distinct subset of NSCLC, and improved understanding of the implications of ARID1A aberrations is critical for the identification of therapeutic target candidates.

Pulmonary MicroRNA Changes Alter Angiogenesis in Chronic Obstructive Pulmonary Disease and Lung Cancer

The pulmonary endothelium is dysfunctional in chronic obstructive pulmonary disease (COPD), a known risk factor for lung cancer. The pulmonary endothelium is altered in emphysema, which is disproportionately affected by cancers. Gene and microRNA expression differs between COPD and non-COPD lung. We hypothesised that the alteration in microRNA expression in the pulmonary endothelium contributes to its dysfunction. A total of 28 patients undergoing pulmonary resection were recruited and endothelial cells were isolated from healthy lung and tumour. MicroRNA expression was compared between COPD and non-COPD patients. Positive findings were confirmed by quantitative polymerase chain reaction (qPCR). Assays assessing angiogenesis and cellular migration were conducted in Human Umbilical Vein Endothelial Cells (n = 3–4) transfected with microRNA mimics and compared to cells transfected with negative control RNA. Expression of miR-181b-3p, miR-429 and miR-23c (all p < 0.05) was increased in COPD. Over-expression of miR-181b-3p was associated with reduced endothelial sprouting (p < 0.05). miR-429 was overexpressed in lung cancer as well and exhibited a reduction in tubular formation. MicroRNA-driven changes in the pulmonary endothelium thus represent a novel mechanism driving emphysema. These processes warrant further study to determine if they may be therapeutic targets in COPD and lung cancer.

The Relationship Between UBE2C and AGGF1 Overexpression and Tumor Angiogenesis in Non-Small Cell Lung Cancer.

BackgroundTumor infiltration and metastasis are the leading causes of death for patients with tumors. Angiogenesis is a prerequisite for tumor growth and metastasis. Angiogenic factor with G patch and FHA domains 1 (AGGF1) is an angiogenic factor, whereas ubiquitin-conjugating enzyme E2C (UBE2C) functions in protein ubiquitination. Microvessel density (MVD) is the most common indicator of tumor microvessels, and vasculogenic mimicry (VM) facilitates blood supply to tumors. This study explored UBE2C and AGGF1 expression in non-small cell lung cancer (NSCLC) and their relationship with angiogenesis and prognosis to identify biological factors that might predict NSCLC infiltration, metastasis, and prognosis.MethodsThe specimens and clinical pathological data of patients with NSCLC confirmed by pathology after surgical resection between January 2013 and December 2015 were collected. UBE2C and AGGF1 expression, as well as microvessel formation and VM in NSCLC, was observed using immunohistochemistry. The relationships between UBE2C, AGGF1, MVD, VM, and clinical pathological parameters and their relationships with overall survival (OS) and disease-free survival (DFS) were analyzed.ResultsUBE2C and AGGF1 levels in NSCLC tissues were significantly higher than those in corresponding normal tissues (57.1% vs 15.6 and 59.7% vs 25.3%, respectively; P < 0.05). UBE2C, AGGF1, MVD, and VM were positively correlated with each other (P < 0.05) and were all related to tumor size, lymph node metastasis, and tumor-node-metastasis stage (P < 0.05). Kaplan–Meier analysis showed that patient OS and DFS in the UBE2C, AGGF1, VM-positive, and high-MVD groups were reduced (all P < 0.001). Univariate and multivariate analyses showed that UBE2C, AGGF1, VM, and MVD were independent risk factors for NSCLC prognosis.ConclusionUBE2C and AGGF1 overexpression is associated with angiogenesis and poor prognosis and may be important for predicting NSCLC invasion, metastasis, and prognosis.

MicroRNA-320b suppresses HNF4G and IGF2BP2 expression to inhibit angiogenesis and tumor growth of lung cancer

We examined the effect of microRNA-320b (miR-320b) on tumor growth and angiogenesis in lung cancer and also determined its downstream molecular mechanisms. Lung cancer tissues and adjacent non-cancerous tissues were collected from 66 patients with lung cancer. miR-320b expression was experimentally determined to be expressed at low level in cancer tissues. The results of gain-of-function experiments suggested that miR-320b overexpression suppressed cancer cell invasion, tube formation, tumor volume and angiogenesis in xenografted nude mice. Hepatocyte nuclear factor 4 gamma (HNF4G) was identified as a target of miR-320b based on in silico analysis. Dual-luciferase reporter gene assays further identified the binding relationship between HNF4G and miR-320b. Lung cancer tissues exhibited increased expression of HNF4G and insulin-like growth factor 2 mRNA-binding protein 2 (IGF2BP2). Meanwhile, HNF4G knockdown suppressed IGF2BP2 expression, thereby repressing cancer cell invasion and tube formation. Furthermore, IGF2BP2 modified m6A to increase the expression of thymidine kinase 1 (TK1), thus promoting angiogenesis. In nude mice, restoration of TK1 reversed the suppressive effect of miR-320b overexpression on tumor growth rate and CD31 expression. In conclusion, miR-320b suppresses lung cancer growth and angiogenesis by inhibiting HNF4G, IGF2BP2 and TK1.

Role of hypoxia‑inducible factor‑2α in lung cancer (Review).

Hypoxia is a common phenomenon during tumorigenesis and tumour development. In recent years, studies have found that hypoxia‑inducible factor (HIF)‑2α, also referred to as endothelial PAS domain protein‑1, plays an important role in tumours. HIF‑2α is an important oncogene and a critical prognostic indicator in non‑small cell lung cancer. However, no unified conclusion can be drawn concerning HIF‑2α and small cell lung cancer, since few studies to date have focused on their association. An increasing number of studies have confirmed that HIF‑2α is involved in tumorigenesis, cell proliferation, angiogenesis, metastasis, drug resistance and radiotherapy failure in lung cancer. Of note, HIF‑2α plays a crucial role in lung cancer to maintain cancer cell stemness. Based on the importance of HIF‑2α in lung cancer, HIF‑2α‑targeted therapy has been attracting increasing attention. Although this strategy currently appears to be promising invitro, it has never been assessed as a therapy for lung cancer. The aim of the present review was to summarize the contribution of HIF‑2α to various aspects of lung cancer, as well as its potential as targeted therapy.

Analysis of Molecular Mechanism of YiqiChutan Formula Regulating DLL4-Notch Signaling to Inhibit Angiogenesis in Lung Cancer.

In order to explore the specific mechanism of YiqiChutan formula (YQCTF) in inhibiting the angiogenesis of lung cancer and its relationship with delta-like ligand 4- (DLL4-) Notch signaling, 30 healthy BALB/c-nu/nu rats were selected and divided into three groups: A549 group (implanted with lung adenocarcinoma cell line A549), NCI-H460 group (implanted with human lung large-cell carcinoma cell line NCI-H460), and NCI-H446 group (implanted with human lung small cell carcinoma cell line NCI-H446) for constructing lung cancer transplanted tumor models. After modeling, the group treated with normal saline was taken as control group, 200 mg/kg of YQCTF was adopted for intervention, and the tumor volume and growth inhibition rate were compared with the vascular targeted inhibitor Sorafenib. HE staining, CD31 fluorescent antibody staining, and microelectron microscopy were adopted to observe the neovascular endothelial cells of the transplanted tumor. The expression of VEGF, HIF-1α, DLL4, and Notch-1 in the transplanted tumors in each group was detected by Western blot and RT-PCR at the protein level or mRNA level. Compared with the control group, the YQCTF-treated group had obvious inhibitory effect on lung cancer transplanted tumor and lung cancer angiogenesis. In the YQCTF-treated group, the density of angiogenesis decreased significantly and the vascular lumen structure also decreased, and the expression levels of VEGF, HIF-1α, DLL4, and Notch-1 in the YQCTF-treated group were all lower than those in the control group. YQCTF could inhibit the growth of lung cancer transplanted tumor through antiangiogenesis, and it could also reduce the amount of angiogenesis in lung cancer transplanted tumor. In addition, the generation of lumen structure was also hindered, which was realized through the VEGF signaling pathway and DLL4-Notch signaling pathway.

Toll-Like Receptor 7 Mediates Inflammation Resolution and Inhibition of Angiogenesis in Non-Small Cell Lung Cancer

Simple SummaryThe progression of cancer is strictly linked to the formation of new blood vessels responsible for nutrition supply of the tumor. We identified TLR7 as an inhibitor of lung cancer vascularization. TLR7 is part of a large family of immune receptors that function as “sensors” of pathogen- and damage-derived signals. We found that TLR7 exerts antitumor functions in non-small cell lung cancer by inducing the production of specific molecules with inhibitory properties against new blood vessel formation. These molecules are known as specialized pro-resolving mediators (SPMs) and are derived from ω-3 and ω-6 fatty acids. We believe that the results obtained suggest novel potential targets and strategies to treat lung cancer.Pattern recognition receptors (PRR) promote inflammation but also its resolution. We demonstrated that a specific PRR—formyl peptide receptor 1 (FPR1)—sustains an inflammation resolution response with anti-angiogenic and antitumor potential in gastric cancer. Since toll-like receptor 7 (TLR7) is crucial in the physiologic resolution of airway inflammation, we asked whether it could be responsible for pro-resolving and anti-angiogenic responses in non-small cell lung cancer (NSCLC). TLR7 correlated directly with pro-resolving and inversely with angiogenic mediators in NSCLC patients, as revealed by a publicly available RNAseq analysis. In NSCLC cells, depletion of TLR7 caused an upregulation of angiogenic mediators and a stronger vasculogenic response of endothelial cells compared to controls, assessed by qPCR, ELISA, protein array, and endothelial cell responses. TLR7 activation induced the opposite effects. TLR7 silencing reduced, while its activation increased, the pro-resolving potential of NSCLC cells, evaluated by qPCR, flow cytometry, and EIA. The increased angiogenic potential of TLR7-silenced NSCLC cells is due to the lack of pro-resolving mediators. MAPK and STAT3 signaling are responsible for these activities, as demonstrated through Western blotting and inhibitors. Our data indicate that TLR7 sustains a pro-resolving signaling in lung cancer that inhibits angiogenesis. This opens new possibilities to be exploited for cancer treatment.

Tumor-derived exosomal miRNA-141 promote angiogenesis and malignant progression of lung cancer by targeting growth arrest-specific homeobox gene (GAX)

ABSTRACT Previous researches have suggested that exosomal miRNA-141 has association with metastatic lung cancer, however, its role and regulatory mechanism require further study. In this study, exosomes were isolated from lung cancer patients and normal human serum and identified. We found that the expression of miRNA-141 was up-regulated in the lung cancer serum exosomes compared with the normal serum exosomes. When the exosomes were extracted for co-culture with HUVECs, they were absorbed and distributed around the nucleus by confocal microscopy. Moreover, exosomal miRNA-141 from A549 significantly not only promoted the migration and invasion of A549 but also increased the cell proliferation, tube formation of HUVECs. In order to reveal the mechanism of exosomal miRNA-141, bioinformatics analysis revealed that miRNA-141 targeted the binding of Growth arrest-specific homeobox gene (GAX) in the 3ʹUTR region, and confirmed by MS2-RIP assay and dual-luciferase assay. Exosome miRNA-141 could down-regulate the expression of GAX. Taken together, our results demonstrate that tumor-derived exosomal miRNA-141 promote angiogenesis and malignant progression of lung cancer by targeting GAX. It provides a new possibility for the treatment of lung cancer.

A bivalent cyclic RGD–siRNA conjugate enhances the antitumor effect of apatinib via co-inhibiting VEGFR2 in non-small cell lung cancer xenografts

The vascular endothelial growth factor receptor 2 (VEGFR2) is considered to be a pivotal target for anti-tumor therapy against angiogenesis of non-small cell lung cancer (NSCLC). However, effective and low-toxicity targeted therapies to inhibit VEGFR2 are still lacking. Here, biRGD–siVEGFR2 conjugate comprising murine VEGFR2 siRNA and [cyclo(Arg-Gly-Asp-D-Phe-Lys)-Ahx]2-Glu-PEG-MAL (biRGD) peptide which selectively binds to integrin αvβ3 receptors expressing on neovascularization endothelial cell was synthesized. The anti-tumor activity and renal toxicity of biRGD–siVEGFR2 or its combination therapy with low-dose apatinib were investigated on NSCLC xenografts. The immunogenicity of biRGD–siVEGFR2 was also evaluated in C57BL/6J mice. In vivo, intravenously injected biRGD–siVEGFR2 substantially inhibited NSCLC growth with a marked reduction of vessels and a down-regulation of VEGFR2 in tumor tissue. Furthermore, biRGD–siVEGFR2 in combination with low-dose apatinib achieved powerful anti-tumor effect with less nephrotoxicity compared with the regular dose of apatinib. Besides, no obvious immunogenicity of biRGD–siVEGFR2 was found. These findings demonstrate that biRGD–siVEGFR2 conjugate can be used as a new candidate for the treatment of NSCLC and its combination therapy with apatinib may also provide a novel strategy for cancer treatment in clinic.

The role of angiotensin-(1-7) on acquired platinum resistance-induced angiogenesis in non-small cell lung cancer in vitro and in vivo.

Renin-angiotensin system (RAS) signaling has been implicated in the development of cancer. The new RAS ACE2/Ang-(1-7)/Mas axis antagonizes the classical ACE/Ang II/AT1R axis. Ang-(1-7) has pleiotropic roles in lung cancer including suppressing proliferation, angiogenesis, and metastasis. This research was designed to investigate the effect of Ang-(1-7) on tumor-associated angiogenesis in DDP-resistant lung cancer cell lines. We first established acquired DDP-resistant cell lines A549 (A549-DDP) and LLC (LLC-DDP). We next performed RT-qPCR, western blot, ELISA, tube formation, microvessel density detection, immunohistochemistry, and tumor formation assays. The results showed that the mRNA and protein levels of RAS components and vascular endothelial growth factor A (VEGFa) were lessened in the A549/LLC-DDP+Ang-(1-7) group compared with the A549/LLC-DDP group. This effect could be blocked by the MAS receptor antagonist A779. The data revealed that Ang-(1-7) could perform its antiangiogenic function by PI3K/AKT and MAPK pathways. Furthermore, the impact of Ang-(1-7) on tumor-associated angiogenesis has been confirmed in lung cancer xenograft model with acquired DDP resistance. These results provide a theoretical basis for designing therapeutic strategies for targeting Ang-(1-7) in the treatment of NSCLC.

Cytochalasin H isolated from mangrove-derived endophytic fungus inhibits epithelial-mesenchymal transition and cancer stemness via YAP/TAZ signaling pathway in non-small cell lung cancer cells.

Our previous studies have isolated cytochalasin H (CyH) from endophytic fungus derived from mangrove and found that CyH induced apoptosis and inhibited migration and angiogenesis in non-small cell lung cancer (NSCLC) cells. In this study, we further investigated the effect of CyH on epithelial-mesenchymal transition (EMT) and cancer stemness of A549 and NCI-H460 NSCLC cells and the underlying mechanisms, especially the role of YAP/ TAZ signaling pathway in the process. Our results showed that CyH significantly inhibited invasive ability and the sphere formation of NSCLC cells. The expression of E-cadherin, an EMT epithelial marker, was obviously up-regulated, while the expression of Vimentin and N-cadherin, the EMT mesenchymal markers, was dramatically down-regulated by CyH treatment in NSCLC cells. Moreover, the expression of EMT-associated transcription factors including Slug, Twist1, and Snail1 and stemness markers including Nanog, Sox-2, and Oct-4 was significantly down-regulated by CyH treatment in NSCLC cells. Additionally, CyH significantly down-regulated YAP and TAZ expression and up-regulated LAST1/2 and MST1/2 expression, and CyH inhibited the interaction between YAP and TEAD. Furthermore, YAP knockdown abolished the effect of CyH on the expression of EMT- and stemness-related markers in NSCLC cells. Taken together, these results suggest that CyH inhibits EMT and cancer stemness of NSCLC cells via the regulation of YAP/TAZ signaling pathway.

Profilin 2 promotes growth, metastasis, and angiogenesis of small cell lung cancer through cancer-derived exosomes

Small cell lung cancer (SCLC) is highly aggressive and prone to hypervascular metastases. Recently, we found profilin 2 (PFN2) expression in SCLC but not in normal tissues. Furthermore, PFN2 expression had been shown to promote angiogenesis through exosomes. However, it remains unclear whether PFN2 contributes to the progression and metastasis of SCLC through angiogenesis. We report here that overexpression (OE) of PFN2 increased, whereas its knockdown (KD) decreased the proliferation, migration, and invasion of SCLC cell H446. The exosomes from OE-H446 (SCLC-OE-exo) exhibited similar effects on H446 properties. Culturing of endothelial cells (ECs) in SCLC-OE conditioned medium (CM) or SCLC-OE-exo increased the migration and tube formation ability of ECs, whereas SCLC-KD-CM and SCLC-KD-exo had inhibitory effects. Interestingly, both SCLC- and EC-derived exosomes were internalized in H446 more rapidly than in ECs. More importantly, OE-PFN2 dramatically elevated SCLC growth and vasculature formation as well as lung metastasis in tumor xenograft models. Finally, we found that PFN2 activated Smad2/3 in H446 and pERK in ECs, respectively. Taken together, our study revealed the role of PFN2 in SCLC development and metastasis, as well as tumor angiogenesis through exosomes, providing a new molecular target for SCLC treatment.

ILT3 promotes tumor cell motility and angiogenesis in non-small cell lung cancer.

Immunoglobulin-like transcript (ILT) 3 is an immunosuppressive molecule that negatively regulates myeloid cell activation. ILT3 overexpression in tumor cells induces immune escape of solid tumors and facilitates invasion of monocytic acute myeloid leukemia cells. However, the expression and function of ILT3 in non-small cell lung cancer (NSCLC) cells remain elusive. Herein, we found that ILT3 was enriched in human NSCLC cells, and predicted advanced disease and poor overall survival. ILT3 overexpression enhanced the migration and invasion of NSCLC cells and tubule formation of human umbilical vein endothelial cells by upregulating and interacting with its ligand apolipoprotein E (ApoE) in vitro. Mechanistically, ILT3 recruited SHP2 and SHIP1, and subsequently activated ERK1/2 signaling mediating epithelial-mesenchymal transition (EMT) and increasing vascular endothelial growth factor (VEGF)-A expression in NSCLC cells, which are responsible for tumor cell motility and angiogenesis, respectively. Using murine metastasis models, we further confirmed ILT3 promoted NSCLC metastasis and explored the exact correlation of ILT3 with ApoE, EMT, and VEGF-A in vivo. These results unraveled novel mechanisms for ILT3-induced tumor progression and proposed ILT3 as a potential therapeutic target and prognostic biomarker for NSCLC patients.

Beyond Conventional: The New Horizon of Anti-Angiogenic microRNAs in Non-Small Cell Lung Cancer Therapy.

GLOBOCAN 2018 identified lung cancer as the leading oncological pathology in terms of incidence and mortality rates. Angiogenesis is a key adaptive mechanism of numerous malignancies that promotes metastatic spread in view of the dependency of cancer cells on nutrients and oxygen, favoring invasion. Limitation of the angiogenic process could significantly hamper the disease advancement through starvation of the primary tumor and impairment of metastatic spread. This review explores the basic molecular mechanisms of non-small cell lung cancer (NSCLC) angiogenesis, and discusses the influences of the key proangiogenic factors—the vascular endothelial growth factor-A (VEGF-A), basic fibroblast growth factor (FGF2), several matrix metalloproteinases (MMPs—MMP-2, MMP-7, MMP-9) and hypoxia—and the therapeutic implications of microRNAs (miRNAs, miRs) throughout the entire process, while also providing critical reviews of a number of microRNAs, with a focus on miR-126, miR-182, miR-155, miR-21 and let-7b. Finally, current conventional NSCLC anti-angiogenics—bevacizumab, ramucirumab and nintedanib—are briefly summarized through the lens of evidence-based medicine.

Epigenetic regulation of angiogenesis in lung cancer.

Lung cancer is the leading cause of cancer-related deaths worldwide, in which angiogenesis is highly required for lung cancer cell growth and metastasis. Genetic regulation of this multistep process is being studied extensively, however, relatively less is known about the epigenetic regulation of angiogenesis in lung cancer. Several epigenetic alterations contribute to regulating angiogenesis, such as epimodifications of DNA, posttranslational modification of histones, and expression of noncoding RNAs. Here, we review the current knowledge of the epigenetic regulation of angiogenesis and discuss the potential clinical applications of epigenetic-based anticancer therapy in lung cancer. Overall, epigenetic-based therapy will likely emerge as a prominent approach to treat lung cancer in the future.

Midkine Is a Potential Therapeutic Target of Tumorigenesis, Angiogenesis, and Metastasis in Non-Small Cell Lung Cancer.

Hypoxia-inducible factors (HIFs) induced by reduced O2 availability activate the transcription of target genes encoding proteins that play important roles in communication between cancer and stromal cells. Cancer cells were incubated under hypoxic conditions: H1299, A549 (NSCLC); Hep3B, HepG2 (HCC); HCT116, CT26 (Colon cancer); MCF-7, MDAMB231 (Breast cancer); MKN1, MKN5 (Gastric cancer); U87MG, SHSY5Y (Brain cancer); and SKOV3, SNU840 (Ovary cancer). All cells expressed HIF-1α and HIF-2α mRNA and proteins. However, cell proliferation of NSCLC, breast, gastric, and brain cancer cells under hypoxia was more dependent on HIF-1α except for HCC cells where it was more dependent on HIF-2α. Among HIF-1α dependent cells H1299 was the most affected in terms of cell proliferation by HIF-1α knockdown. To examine which cytokines are secreted in NSCLC cells by HIF-1α to communicate with stromal cells, we performed a cytokine-profiling array with H1299. We screened the top 14 cytokines which were dependent on the HIF-1α expression pattern. Among them, midkine (MDK) expression was affected the most in response to HIF-1α. MDK is a heparin-binding growth factor that promotes angiogenesis and carcinogenesis. Indeed, MDK significantly increased HUVEV endothelial cell migration and neo- vascularization in chick chorioallantoic membrane assay (CAM) assay via paracrine signaling. In addition, MDK secreted from NSCLC cells interacted with Notch2 which activated the Notch signaling pathway and induced EMT, upregulated NF-κB, and increased cancer promotion. However, in response to MDK knock down, siRNA or the MDK inhibitor, iMDK treatment not only decreased MDK-induced migration and angiogenesis of endothelial cells but also abrogated the progression and metastasis of NSCLC cells in in vitro and in vivo orthotopic and spontaneous lung metastasis models. Consequently, iMDK treatment significantly increased mice survival rates compared with the control or MDK expression group. MDK plays a very important role in the progression and metastasis of NSCLC cells. Moreover, the MDK targeting strategy provides a potential therapeutic target for the treatment of MDK-expressing lung cancers.