Correction to: Circular RNA circFGFR1 promotes progression and anti-PD-1 resistance by sponging miR-381-3p in non-small cell lung cancer cells.

After the publication of this work [1], an error was found in Fig. 1b. As described in the Results section that circ0084003 has 13 exons, it should be formed by 5-17 (13exons) exons, authors have described "formed by 5-19 exons". The authors extend their apology for the mistake caused by their action. With that mistake, the correct version of Fig. 1b is provided below.

Circular RNA circFGFR1 promotes progression and anti-PD-1 resistance by sponging miR-381-3p in non-small cell lung cancer cells.

BACKGROUND: Immune system evasion, distance tumor metastases, and increased cell proliferation are the main reasons for the progression of non-small cell lung cancer (NSCLC) and the death of NSCLC patients. Dysregulation of circular RNAs plays a critical role in the progression of NSCLC; therefore, further understanding the biological mechanisms of abnormally expressed circRNAs is critical to discovering novel, promising therapeutic targets for NSCLC treatment. METHODS: The expression of circular RNA fibroblast growth factor receptor 1 (circFGFR1) in NSCLC tissues, paired nontumor tissues, and cell lines was detected by RT-qPCR. The role of circFGFR1 in NSCLC progression was assessed both in vitro by CCK-8, clonal formation, wound healing, and Matrigel Transwell assays and in vivo by a subcutaneous tumor mouse assay. In vivo circRNA precipitation, RNA immunoprecipitation, and luciferase reporter assays were performed to explore the interaction between circFGFR1 and miR-381-3p. RESULTS: Here, we report that circFGFR1 is upregulated in NSCLC tissues, and circFGFR1 expression is associated with deleterious clinicopathological characteristics and poor prognoses for NSCLC patients. Forced circFGFR1 expression promoted the migration, invasion, proliferation, and immune evasion of NSCLC cells. Mechanistically, circFGFR1 could directly interact with miR-381-3p and subsequently act as a miRNA sponge to upregulate the expression of the miR-381-3p target gene C-X-C motif chemokine receptor 4 (CXCR4), which promoted NSCLC progression and resistance to anti-programmed cell death 1 (PD-1)- based therapy. CONCLUSION: Taken together, our results suggest the critical role of circFGFR1 in the proliferation, migration, invasion, and immune evasion abilities of NSCLC cells and provide a new perspective on circRNAs during NSCLC progression.

The Effect of and Mechanism Underlying Autophagy in Hepatocellular Carcinoma Induced by CH12, a Monoclonal Antibody Directed Against Epidermal Growth Factor Receptor Variant III.

BACKGROUND/AIMS: Epidermal growth factor receptor variant III (EGFRvIII), the most frequent EGFR variant, is constitutively activated without binding to EGF and is correlated with a poor prognosis. CH12, a human-mouse chimeric monoclonal antibody, has been developed in our laboratory and selectively binds to overexpressed EGFR and EGFRvIII. A previous study had reported that EGFR could influence autophagic activity, and autophagy is closely related to tumor development and the response to drug therapy. In this study, we aimed to elucidate the effect of CH12 on autophagy and efficacy of combining CH12 with an autophagy inhibitor against EGFRvIII-positive tumors. METHODS: EGFRvIII was overexpressed in liver cancer, glioblastoma and breast cancer, and the change in the autophagy-relevant protein levels was analyzed by western blot assays, LC3 punctate aggregation was analyzed by immunofluorescence. The interaction of Beclin-1 and Rubicon was assessed by co-immunoprecipitation (Co-IP) after CH12 treatment. The efficacy of ATG7 or Beclin-1 siRNA in combination with CH12 in Huh-7-EGFRvIII cells was assessed by CCK-8 assays. The autophagy and apoptosis signaling events in Huh-7-EGFRvIII cells upon treatment with control, CH12, siRNA or combination for 48 h were assessed by western blot assays. RESULTS: Our results showed that, in cancer cell lines overexpressing EGFRvIII, only the liver cancer cell lines Huh-7 and PLC/PRF/5 suggested autophagy activation. We then investigated the mechanism of autophagy activation after EGFRvIII overexpression. The results showed that EGFRvIII interacted with Rubicon, an autophagy inhibition protein, and released Beclin-1 to form the inducer complex, thus contributing to autophagy. In addition, CH12, via inhibiting the phosphorylation of EGFRvIII, promoted the interaction of EGFRvIII with Rubicon, further inducing autophagy. In vitro assays suggested that knocking down the expression of the key proteins ATG7 or Beclin-1 in the autophagy pathway with siRNA inhibits tumor cell proliferation. Combining autophagy-related proteins 7 (ATG7) or Beclin-1 siRNA with CH12 in Huh-7-EGFRvIII cells showed better inhibition of cell proliferation. CONCLUSION: EGFRvIII could induce autophagy, and CH12 treatment could improve autophagy activity in EGFRvIII-positive liver cancer cells. The combination of CH12 with an autophagy inhibitor or siRNA against key proteins in the autophagy pathway displayed more significant efficacy on EGFRvIII-positive tumor cells than monotherapy, and induced cell apoptosis.

Development of T cells carrying two complementary chimeric antigen receptors against glypican-3 and asialoglycoprotein receptor 1 for the treatment of hepatocellular carcinoma.

Adoptive immunotherapy leveraging chimeric antigen receptor-modified T (CAR-T) cells holds great promise for the treatment of cancer. However, tumor-associated antigens often have low expression levels in normal tissues, which can cause on-target, off-tumor toxicity. Recently, we reported that GPC3-targeted CAR-T cells could eradicate hepatocellular carcinoma (HCC) xenografts in mice. However, it remains unknown whether on-target, off-tumor toxicity can occur. Therefore, we proposed that dual-targeted CAR-T cells co-expressing glypican-3 (GPC3) and asialoglycoprotein receptor 1 (ASGR1) (a liver tissue-specific protein)-targeted CARs featuring CD3ζ and 28BB (containing both CD28 and 4-1BB signaling domains), respectively, may have reduced on-target, off-tumor toxicity. Our results demonstrated that dual-targeted CAR-T cells caused no cytotoxicity to ASGR1+GPC3- tumor cells, but they exhibited a similar cytotoxicity against GPC3+ASGR1- and GPC3+ASGR1+ HCC cells in vitro. We found that dual-targeted CAR-T cells showed significantly higher cytokine secretion, proliferation and antiapoptosis ability against tumor cells bearing both antigens than single-targeted CAR-T cells in vitro. Furthermore, the dual-targeted CAR-T cells displayed potent growth suppression activity on GPC3+ASGR1+ HCC tumor xenografts, while no obvious growth suppression was seen with single or double antigen-negative tumor xenografts. Additionally, the dual-targeted T cells exerted superior anticancer activity and persistence against single-targeted T cells in two GPC3+ASGR1+ HCC xenograft models. Together, T cells carrying two complementary CARs against GPC3 and ASGR1 may reduce the risk of on-target, off-tumor toxicity while maintaining relatively potent antitumor activities on GPC3+ASGR1+ HCC.

An EpCAM/CD3 bispecific antibody efficiently eliminates hepatocellular carcinoma cells with limited galectin-1 expression.

There have been several studies suggesting that cancer stem cells (CSCs) contribute to the high rates of recurrence and resistance to therapies observed in hepatocellular carcinoma (HCC). Epithelial cell adhesion molecule (EpCAM) has been demonstrated to be a biomarker of CSCs and a potential therapeutic target in HCC. Here, we prepared two anti-EpCAM monoclonal antibodies (1H8 and 2F2) and an anti-EpCAM bispecific T cell engager (BiTE) 1H8/CD3, which was derived from 1H8, and used them to treat HCC in vitro and in vivo. The results demonstrated that all of the developed anti-EpCAM antibodies specifically bound to EpCAM. Neither anti-EpCAM monoclonal antibody had obvious anti-HCC activities in vitro or in vivo. However, anti-EpCAM BiTE 1H8/CD3 induced strong peripheral blood mononuclear cell-dependent cellular cytotoxicity in Huh-7 and Hep3B cells but not EpCAM-negative SK-Hep-1 cells. Notably, 1H8/CD3 completely inhibited the growth of Huh-7 and Hep3B xenografts in vivo. Treatment of the Huh-7 HCC xenografts with 1H8/CD3 significantly suppressed tumor proliferation and reduced the expression of most CSC biomarkers. Intriguingly, galectin-1 (Gal-1) overexpression inhibited 1H8/CD3-induced lymphocytotoxicity in HCCs while knockdown of Gal-1 increased the lymphocytotoxicity. Collectively, these results indicate that anti-EpCAM BiTE 1H8/CD3 is a promising therapeutic agent for HCC treatment. Gal-1 may contribute to the resistance of HCC cells to 1H8/CD3-induced lysis.

Longitudinal qualitative study on the psychological experiences of COVID-19 patients based on timing it right framework.

Timing it right framework was used as a framework to explore the illness experiences of patients infected with COVID-19 and to analyze the patients' perceptions of the disease and their true inner feelings to provide a reference for the control of infectious diseases. This research adopted a phenomenological research approach to develop a longitudinal qualitative study. A purposive sampling method was used to select participants and 37 patients were recruited. Depending on the principle that participants should have maximum variation and sampling should cease when interviews content saturation is achieved, 16 COVID-19 patients in an isolation ward in Ningbo City, Zhejiang Province were finally included. Data were collected using semi-structured interviews, and the content of the interviews was analyzed by Colaizzi's 7-step method. The themes of COVID-19 patients' experiences at various phase were presented as follows: multiple emotions intertwined at the time of diagnosis (anxiety, stressful panic, facing the diagnosis calmly), multiple pressures during the hospitalization period (concerns about the disease, unable to adapt to the ward environment, worrying about future hardship), growth of positive illness experience during the isolation and observation period (sublimated outlook on life, affirmation of the government's anti-epidemic policy, more concerned about their own health), adjustment after returning to society (stigma, loss of previous living environment, problems caused by nucleic acid testing), and adaptation to social life (return to normal life, avoidance of illness experience, post-covid-19 syndrome). The illness experience of COVID-19 patients changed dynamically with time, but a sense of shame and uncertainty about recovery was present throughout the process. Interventions should be developed according to the needs of the patients at different times to inform subsequent optimization of care and management of infectious diseases.

Circular RNA drives resistance to anti-PD-1 immunotherapy by regulating the miR-30a-5p/SOX4 axis in non-small cell lung cancer.

Aim: Circular RNAs are widely and abnormally expressed in human cancer cells, and they participate in cancer progression. However, they have rarely been investigated in the immune evasion of non-small cell lung cancer (NSCLC). Here, we elucidated the function and molecular mechanism of hsa_circ_0020714 in promoting the resistance to anti-PD-1 immunotherapy of NSCLC. Methods: The expression of hsa_circ_0020714 were examined by qRT-PCR. In vivo experiments were executed to investigate the biological function of hsa_circ_0020714 in the sensitivity of NSCLC to anti-PD-1 immunotherapy. The qRT-PCR, fluorescence in situ hybridization, RNA pulldown, RNA immunoprecipitation, and western blot were carried out to investigate the potential regulatory mechanisms of hsa_circ_0020714 in NSCLC immune evasion. Results: The expression of hsa_circ_0020714 was upregulated in NSCLC tissues compared to the paired adjacent non-tumor tissues, and an increased expression of hsa_circ_0020714 was significantly associated with a bad prognosis and resistance to anti-PD-1 immunotherapy in patients with NSCLC. Mechanistically, hsa_circ_0020714 functions as an endogenous miR-30a-5p sponge to enhance SOX4 expression, thereby promoting immune evasion and anti-PD-1 resistance in NSCLC patients. Conclusion: Hsa_circ_0020714 induces the immune evasion and resistance to anti-PD-1 immunotherapy of NSCLC via the miR-30a-5p/SOX4 axis, and may be an promising immunotherapeutic target in NSCLC.

Alternative splicing events in tumor immune infiltration in renal clear cell carcinomas.

Alternative splicing (AS) is a gene regulatory mechanism that drives protein diversity and dysregulation of AS plays a significant role in tumorigenesis. This study aimed to develop a prognostic signature based on AS and elucidate the role in tumor immune microenvironment (TIME) in clear cell renal cell carcinoma (ccRCC). The prognosis-related AS events were analyzed by univariate Cox regression analysis. Gene set enrichment analyses (GSEA) were performed for functional annotation. Prognostic signatures were identified and validated using univariate and multivariate Cox regression, LASSO regression, Kaplan-Meier survival analysis, and proportional hazards model. The context of TIME in ccRCC was also analyzed. Gene and protein expression data of C4orf19 were obtained from ONCOMINE website and Human Protein Altas. Splicing factors (SFs) regulatory networks were visualized. 4431 survival-related AS events in ccRCC were screened. Based on splicing subtypes, eight AS prognostic signatures were constructed. A nomogram with good prognostic prediction was generated. Furthermore, the prognostic signatures were significantly correlated with TIME diversity and immune checkpoint inhibitor (ICI)-related genes. C4orf19 was the only gene whose expression levels were downregulated among the prognostic AS-related genes, which is considered as a promising prognostic factor in ccRCC. Potential functions of SFs were determined by splicing regulatory networks. In our study, AS patterns of novel indicators for prognostic prediction of ccRCC were explored. The AS-SF networks provide information of regulatory mechanisms. Players of AS events related to TIME were investigated, which contribute to prognosis monitoring of ccRCC.

Rapamycin inhibits lung squamous cell carcinoma growth by downregulating glypican-3/Wnt/ß-catenin signaling and autophagy.

PURPOSE: There is not much progress in the treatment for lung squamous cell carcinoma LSCC in the past few years. Rapamycin Rapa, an inhibitor of mammalian target of rapamycin mTOR, has exhibited antitumor efficacy in a variety of malignant tumors. It has recently been reported that Rapamycin can induce autophagy signaling pathway in lung cancer and Glypican-3GPC3 can promote the growth of hepatocellular carcinoma by stimulating canonical Wnt signaling pathway. The aim of this study is to investigate the mechanisms of rapamycin's antitumor efficacy in relation to GPC3/Wnt/ß-catenin pathway and autophagy in LSCC. METHODS: SK-MES-1 cells, a LSCC cell line, were treated with various concentrations of rapamycin with or without Glypican-3 GPC3-targeting siRNA. SK-MES-1 cell proliferation was determined by MTT assay. Protein expression levels of GPC3, ß-catenin, Beclin-1 were checked via western blotting. We established the xenograft mice model to investigate the suppression effect of rapamycin on LSCC. In addition, we further testified the metabolism protein of autophagy process using the xenograft tumor tissue. RESULTS: Rapamycin could inhibit the SK-MES-1 cell proliferation in a concentration-dependent manner both in vitro and in vivo by decreasing the GPC3 expression and downregulating the glypican-3/Wnt/ß-catenin signaling pathway. In addition, we found that GPC3 silencing can activate the glypican-3/Wnt/ß-catenin pathway and autophagy, which contribute to the suppression of tumor growth both in vitro and in vivo. CONCLUSION: Rapamycin suppresses the growth of lung cancer through down-regulating glypican-3/Wnt/ß-catenin signaling, which mediates with activation of autophagy. This study suggests GPC3 is a new promising target for rapamycin in the treatment of lung cancer.

Development of GPC3 and EGFR-dual-targeting chimeric antigen receptor-T cells for adoptive T cell therapy.

Adoptive transfer of T cells expressing specific anti-glypican-3 (GPC3) chimeric antigen receptors (CARs) has demonstrated therapeutic potential against hepatocellular carcinoma (HCC). However, normal tissues with low expression of neoplasm-associated antigens often show on-target, off-tumor toxicity. Previous studies have revealed that the development of HCC xenografts in mice could be inhibited effectively by GPC3-targeting CAR-T cells. However, these studies did not provide information regarding on-target, off-tumor toxicity. We hypothesized that on-target, off-tumor toxicity may decrease in dual-targeting CAR-T cells that co-express GPC3 with epidermal growth factor receptor (EGFR)-targeted CARs characterized by CD3ζ and 28BB expression. Our research confirmed that dual-targeting CAR-T (CARgpc3-egfr) cells exhibited similar proliferative ability and cytotoxicity to CARgpc3 T cells against GPC3+EGFR+ HCC in vitro. However, EGFR-targeting CAR-T (CARegfr) cells showed poor proliferation activity and cytotoxicity against GPC3+EGFR+ HCC cells, similar to mock CAR-T cells. CARgpc3 and CARgpc3-egfr T cells showed enhanced cytokine secretion compared to CARegfr and mock CAR-T cells in vitro. In vivo, tumor growth suppression was better for CARgpc3-egfr T cells than for CARgpc3 T cells in GPC3+EGFR+ HCC, while it was not observed for CARegfr or mock CAR-T cells. Taken together, our data indicated that dual-targeting CAR-T cells with two CARs against GPC3 and EGFR may maintain relatively effective anti-neoplasm functions in GPC3+EGFR+ HCC in vitro and in vivo, a strategy that may reduce off-tumor toxicity.

The lipid platform increases the activity of STING agonists to synergize checkpoint blockade therapy against melanoma.

The response rate to PD-1/PD-L1 immune checkpoint inhibition (ICI) therapy in melanoma remains low due to the immunosuppressive tumor microenvironment. Novel strategies synergizing ICI treatment are urgently sought after. Activation of the stimulator of interferon genes (STING) has recently emerged as a critical pathway to overcome immunosuppression. Herein, 2'3' cyclic guanosine monophosphate-adenosine monophosphate (cGAMP), a universal STING agonist, was encapsulated into lipid nanoparticles conjugated with mannose (LP-cGAMP) for dendritic cell (DC)-specific cytosolic delivery. LP-cGAMP induced STING-related pro-inflammatory and intratumoral injections of LP-cGAMP increased DC maturation and CD8+ T cell infiltration more efficiently compared to free cGAMP. Given the upregulation of PD-L1 on tumor cells in response to STING activation, we further tested the combination therapy of LP-cGAMP and anti-PD-L1 and observed a superior antitumor effect in B16F10 and BRAF-mutated murine melanoma models. Our findings prove that targeted delivery of cGAMP can synergize PD-L1 blockade therapy in melanoma and the combinational immune therapy has a great potential to produce a long-lasting anti-tumor effect.

CK1α-targeting inhibits primary and metastatic colorectal cancer in vitro, ex vivo, in cell-line-derived and patient-derived tumor xenograft mice models.

BACKGROUND: Colorectal cancer (CRC) remains a leading cause of cancer-related deaths globally. Despite improved understanding of its initiation and progression, and advances in diagnostic or therapeutic strategies, the treatment of metastatic CRC remains a clinical challenge, necessitating identification of novel efficacious therapeutics with little/no toxicity to non-tumor colorectal cells. The present study investigated the effect of Epiblastin A, an adenosine triphosphate (ATP)-mediated competitive inhibitor of casein kinase 1α (CK1α) on the viability, proliferation, and oncogenicity of CRC cells. METHODS: Comparative evaluation of the effect of Epiblastin A on CK1α in fetal human normal colonic mucosa (FHC) and CRC (HCT116, HT29, DLD1) cell lines, using western blot, immunohistochemical staining, real-time polymerase chain reaction (RT-PCR), and sulforhodamine B (SRB) cytotoxicity assays. Primary culture cells, patient-derived xenograft (PDX), and tumor xenograft mice CRC models were also employed. Kaplan-Meier plots were used for survival analysis of our CRC cohort. RESULTS: CRC cells aberrantly express CK1α at mRNA and protein levels. This overexpression of CK1α is strongly associated with worse 5-year overall survival (OS) in patients with CRC. Epiblastin A inhibits CK1α and compared to its apparent non-effect on FHC cells regardless of concentration, it elicits significant dose-dependent inhibition of the viability of HT29, HCT116, and DLD1 cells with a 48 h IC50 of 6.8, 5.0, and 3.2 µM, respectively. The expression of CK1α in CRC primary cultures and PDX samples, significantly correlated with Ki-67 expression, and both were attenuated by Epiblastin A. We also observed that the effect of 5 mg/kg Epiblastin A on tumor volume, and body weight in the CRC PDX mice models, was similar to that of 5 mg/kg Cetuximab over the time-course of our in vivo study. In DLD1-derived tumor xenograft mice, Epiblastin A with very mild effect on mice body weight, suppressed tumor volume and tumor weight in a CK1α-dependent manner (P=0.024). CONCLUSIONS: Our results demonstrate the efficacy of Epiblastin A in CRC and its potential as a putative small-molecule inhibitor of CK1α and Ki-67 signaling, which are relevant in the CRC initiation, progression and prognosis.

Corrigendum: Knockdown of lncRNA LINC01234 Suppresses the Tumorigenesis of Liver Cancer via Sponging miR-513a-5p.

[This corrects the article DOI: 10.3389/fonc.2020.571565.].

Knockdown of lncRNA LINC01234 Suppresses the Tumorigenesis of Liver Cancer via Sponging miR-513a-5p.

BACKGROUND: Liver cancer is a frequent malignancy with poor prognosis and high mortality all over the world. It has been reported many lncRNAs could modulate the tumorigenesis of liver cancer. To identify novel potential targets for liver cancer, the differential expressed lncRNAs between liver cancer and adjacent normal tissues was analyzed with bioinformatics tool. METHODS: The differential expressed lncRNAs between liver cancer and adjacent normal tissues were analyzed with bioinformatics tool. Cell viability and proliferation was tested by CCK8 and Ki67, respectively. Apoptosis of liver cancer cells was tested by flow cytometry. Gene and protein expressions in liver cancer cells were measured by qRT-PCR and western blot, respectively. In vivo model of liver cancer was established to detect the effect of LINC01234 on liver cancer in vivo. RESULTS: LINC01234 was found to be negatively correlated with the survival rate of patients with liver cancer. Moreover, knockdown of LINC01234 significantly suppressed the proliferation and invasion of liver cancer cells via inducing the apoptosis. Meanwhile, miR-513a-5p was sponged by LINC01234, and USP4 was found to be a direct target of miR-513a-5p. In addition, LINC01234 knockdown inhibited the tumorigenesis of liver cancer via inactivating TGF-ß signaling. Furthermore, silencing of LINC01234 notably inhibited the tumor growth of liver cancer in vivo. CONCLUSION: Downregulation of LINC01234 could inhibit the tumorigenesis of liver cancer via mediation of miR-513a-5p/USP4/TGF-ß axis. Thus, LINC01234 might serve as a new target for the treatment of liver cancer.

SNHG22 overexpression indicates poor prognosis and induces chemotherapy resistance via the miR-2467/Gal-1 signaling pathway in epithelial ovarian carcinoma.

Recently, an increasing number of studies have reported that dysregulation of long noncoding RNAs (lncRNAs) plays an important role in cancer initiation and progression, including in epithelial ovarian carcinoma (EOC). However, little is known about the detailed biological functions of the lncRNA small nucleolar RNA host gene 22 (SNHG22) during the progression of EOC. Here, we found that SNHG22 was significantly increased in EOC tissues and was significantly associated with a low level of differentiation. Forced SNHG22 expression promoted chemotherapy resistance in EOC cells. Knockdown of SNHG22 expression increased the sensitivity of EOC cells to cisplatin and paclitaxel. Importantly, we found that SNHG22 could directly interact with miR-2467 and lead to the release of miR-2467-targeted Gal-1 mRNA. Moreover, SNHG22 overexpression induced EOC cell resistance to chemotherapy agents via PI3K/AKT and ERK cascade activation. In summary, our findings demonstrate that SNHG22 plays a critical role in the chemotherapy resistance of EOC by mediating the miR-2467/Gal-1 regulatory axis.

Combination of an anti-EGFRvIII antibody CH12 with Rapamycin synergistically inhibits the growth of EGFRvIII+PTEN-glioblastoma in vivo.

There are still unmet medical needs for the treatment of glioblastoma (GBM), the most frequent and aggressive brain tumor worldwide. EGFRvIII, overexpressed in approximately 30% of GBM, has been regarded as a potential therapeutic target. In this study, we demonstrated that CH12, an anti-EGFRvIII monoclonal antibody, could significantly suppress the growth of EGFRvIII+ GBM in vivo; however, PTEN deficiency in GBM reduced the efficacy of CH12 by attenuating its effect on PI3K/AKT/mTOR pathway. To overcome this problem, CH12 was combined with the mTOR inhibitor rapamycin, leading to a synergistic inhibitory effect on EGFRvIII+PTEN- GBM in vivo. Mechanistically, the synergistic antitumor effect was achieved via attenuating EGFR and PI3K/AKT/mTOR pathway more effectively and reversing the STAT5 activation caused by rapamycin treatment. Moreover, the combination therapy suppressed angiogenesis and induced cancer cell apoptosis more efficiently. Together, these results indicated that CH12 and rapamycin could synergistically suppress the growth of EGFRvIII+PTEN- GBM, which might have a potential clinical application in the future.

Adoptive immunotherapy using T lymphocytes redirected to glypican-3 for the treatment of lung squamous cell carcinoma.

There are unmet medical needs for patients with lung squamous cell carcinoma (LSCC). Therefore, in this study, we explored the antitumor potential of third-generation glypican 3 (GPC3)-redirected chimeric antigen receptor (CAR)-engineered T lymphocytes (CARgpc3 T cells) in tumor models of LSCC. First, we demonstrated by immunohistochemistry (IHC) that GPC3 was expressed in 66.3% of LSCC samples and in 3.3% of lung adenocarcinoma (LAD) samples but not in normal lung tissues. In the presence of GPC3-positive LSCC cells, CARgpc3 T cells were highly activated and increased in number. CARgpc3 T cells could specifically lyse GPC3-positive LSCC cells in vitro. In two established LSCC xenograft models, CARgpc3 T cells could almost completely eliminate the growth of GPC3-positive cells. Additionally, the CARgpc3 T cells were able to persist in vivo and efficiently infiltrate the cancerous tissues. Taken together, these findings indicate that CARgpc3 T cells might be a novel potential therapeutic agent for the treatment of patients with LSCC.

Synergistic antitumor efficacy against the EGFRvIII+HER2+ breast cancers by combining trastuzumab with anti-EGFRvIII antibody CH12.

Although Trastuzumab, an anti-HER2 antibody, benefits certain patients with HER2-overexpressing breast cancer, de novo or acquired trastuzumab resistance remains a haunting issue. EGFRvIII, co-expressing with HER2 in some breast tumors, indicates a poor clinical prognosis. However, the role of EGFRvIII in the function of trastuzumab is not clear. Here, we demonstrated that EGFRvIII overexpression contributed to de novo trastuzumab resistance and the feedback activation of STAT3 caused by trastuzumab also resulted in acquired resistance in EGFRvIII(+)HER2(+) breast cancers. CH12, a highly effective anti-EGFRvIII monoclonal antibody that preferentially binds to EGFRvIII, significantly suppressed the growth of EGFRvIII+HER2(+) breast cancer cells in vitro and in vivo. Importantly, CH12 in combination with trastuzumab had a synergistic inhibitory effect on EGFRvIII(+)HER2(+) breast cancers in vitro and in vivo via attenuating the phosphorylation of EGFR and HER2 and their downstream signal pathways more effectively and reversing STAT3 feedback activation. Moreover, the combination therapy suppressed angiogenesis and induced cell apoptosis significantly. Together, these results suggested a synergistic efficacy of the combination of trastuzumab with CH12 against EGFRvIII(+)HER2(+) breast cancers, which might be a potential clinical application in the future.

Development of T cells redirected to glypican-3 for the treatment of hepatocellular carcinoma.

PURPOSE: The aim of our study is to elucidate whether T cells expressing GPC3-targeted chimeric antigen receptor (CAR) can efficiently eliminate GPC3-positive HCC cells and their potential in the treatment of HCC. EXPERIMENTAL DESIGN: T cells expressing a first-generation and third-generation GPC3-targeted CAR were prepared using lentiviral vector transduction. The in vitro and in vivo cytotoxic activities of the genetically engineered CAR T cells were evaluated against various HCC cell lines. RESULTS: GPC3-targeted CAR T cells could efficiently kill GPC3-positive HCC cells but not GPC3-negative cells in vitro. These cytotoxic activities seemed to be positively correlated with GPC3 expression levels in the target cells. In addition, T cells expressing the third-generation GPC3-targeted CAR could eradicate HCC xenografts with high level of GPC3 expression and efficiently suppress the growth of HCC xenografts with low GPC3 expression level in vivo. The survival of the mice bearing established orthotopic Huh-7 xenografts was significantly prolonged by the treatment with the third-generation GPC3-targeted CAR T cells. CONCLUSIONS: GPC3-targeted CAR T cells could potently eliminate GPC3-positive HCC cells, thereby providing a promising therapeutic intervention for GPC3-positive HCC.

A novel EGFR isoform confers increased invasiveness to cancer cells.

As a validated therapeutic target in several human cancers, the EGF receptor (EGFR) provides a focus to gain deeper insights into cancer pathophysiology. In this study, we report the identification of a naturally occurring and widely expressed EGFR isoform termed EGFRvA, which substitutes a Ser/Thr-rich peptide for part of the carboxyl-terminal regulatory domain of the receptor. Intriguingly, EGFRvA expression relates more closely to histopathologic grade and poor prognosis in patients with glioma. Ectopic expression of EGFRvA in cancer cells conferred a higher invasive capacity than EGFR in vitro and in vivo. Mechanistically, EGFRvA stimulated expression of STAT3, which upregulated heparin-binding EGF (HB-EGF). Reciprocally, HB-EGF stimulated phosphorylation of EGFRvA at Y845 along with STAT3, generating a positive feedback loop that may reinforce invasive function. The significance of EGFRvA expression was reinforced by findings that it is attenuated by miR-542-5p, a microRNA that is a known tumor suppressor. Taken together, our findings define this newfound EGFR isoform as a key theranostic molecule.