Yuping Sun

Construction and evaluation of a novel humanized HER2-specific chimeric receptor

IntroductionThe human epidermal growth factor receptor 2 (HER2) represents one of the most studied tumor-associated antigens (TAAs) for cancer immunotherapy. The monoclonal antibody (mAb) trastuzumab has improved the outcomes of patients with HER2+ breast cancer. However, a large number of HER2+ tumors are not responsive to, or become resistant to, trastuzumab-based therapy, and thus more effective therapies targeting HER2 are needed.MethodsHER2-specific T cells were generated by the transfer of genes that encode chimeric antigen receptor (CAR). Using a multistep overlap extension PCR method, we constructed a novel, humanized HER2 CAR-containing, chA21 single-chain variable fragment (scFv) region of antigen-specific mAb and T-cell intracellular signaling chains made up of CD28 and CD3ζ. An interferon γ and interleukin 2 enzyme-linked immunosorbent assay and a chromium-51 release assay were used to evaluate the antitumor immune response of CAR T cells in coculture with tumor cells. Furthermore, SKBR3 tumor–bearing nonobese diabetic/severe combined immunodeficiency (NOD/SCID) mice were treated with HER2 CAR T cells to evaluate antitumor activity. Human CD3+ T cell accumulation in tumor xenograft was detected by immunohistochemistry.ResultschA21-28z CAR was successfully constructed, and both CD4+ and CD8+ T cells were transduced. The expanded HER2 CAR T cells expressed a central memory phenotype and specifically reacted against HER2+ tumor cell lines. Furthermore, the SKBR3 tumor xenograft model revealed that HER2 CAR T cells significantly inhibited tumor growth in vivo. Immunohistochemical analysis showed robust accumulation of human CD3+ T cells in regressing SKBR3 lesions.ConclusionsThe results of this study show that novel chA21 scFv-based, HER2-specific CAR T cells not only recognized and killed HER2+ breast and ovarian cancer cells ex vivo but also induced regression of experimental breast cancer in vivo. Our data support further exploration of the HER2 CAR T-cell therapy for HER2-expressing cancers.

Podoplanin: a novel regulator of tumor invasion and metastasis

Podoplanin, a small mucin-type sialoglycoprotein, was recently shown to be involved in tumor progression. Podoplanin is overexpressed in cancer cells of various human malignancies, and recently, it is also detected in intratumoral stromal cells. We now appreciate that podoplanin plays a dual role in cancer: it can not only suppress tumor growth but also promote tumor progression. Researchers have identified several potential pathways invoked by podoplanin, which participate in the epithelial-to-mesenchymal transition, collective-cell migration, platelet activation and aggregation, and lymphangiogenesis, and thus regulate the tumor invasion and metastasis. Here, we discuss the current experimental and human clinical data on podoplanin to validate the multiple context-dependent functions in different microenvironments and to delineate the diverse regulatory mechanisms.

B7-H3 expression in ductal and lobular breast cancer and its association with IL-10

Aberrant tumor cell expression of B7-H3, a member of the B7-family that stimulates interleukin-10 (IL-10) secretion, contributes to tumor immune evasion and tumor progression. The aim of this study was to investigate the expression of B7-H3 and IL-10 in ductal and lobular breast cancer tissues. Using immunohistochemistry, B7-H3 and IL-10 protein expression in tumor specimens of primary human breast cancer was investigated. The association between B7-H3 or IL-10 expression and clinicopathological variables was analyzed. The correlation between the expression of B7-H3 and IL-10 was also evaluated. In tumor tissues, the expression of B7-H3 and IL-10 was identified on the cell membrane and in the cytoplasm. Expression of B7-H3 was observed in 90.60% (106/117) of the specimens and 80.34% (94/117) expressed IL-10. Patients with a positive B7-H3 or high IL-10 expression were more likely to have positive lymph node metastasis (N1-3; P=0.018 or 0.035, respectively) and advanced disease (stagexa0II-IV; P=0.011 or 0.039, respectively) compared to those with a negative or low expression. Furthermore, B7-H3 expression was correlated with IL-10 in tumor cells (R=0.545, P=0.000). High B7-H3 expression in human breast cancer tissues may be important in tumor progression and invasiveness. This expression appeared to be correlated with the ability of B7-H3 to promote IL-10 secretion.

Expression of Ig-Like Transcript 4 Inhibitory Receptor in Human Non-small Cell Lung Cancer

BACKGROUNDnHuman Ig-like transcript 4 (ILT-4) is a member of the inhibitory receptor family for immune function. Little is known about the expression levels of ILT-4 in tumor cells.nnnMETHODSnWe have studied the expression levels of ILT-4 both in vitro in cancer cell lines and in vivo in tumor tissues from 70 patients with non-small cell lung cancer (NSCLC) by reverse transcriptase-polymerase chain reaction, fluorescence-activated cell sorting, and immunohistochemical analysis.nnnRESULTSnThree cancer cell lines (H1299, A549, and U1810) express ILT-4 messenger RNA, and only two cell lines (H1299 and A549) express ILT-4 protein on the cell surface. Approximately 37.1% of 70 tumor tissue samples express ILT-4, which is localized in the cell membrane and cytoplasm. In addition, tumor cells and stromal and plasma cells also express ILT-4. The number of infiltrating lymphoid cells in the tumor tissues that express B7-H3 was much lower than those that did not, but there is no significant correlation between ILT-4 expression and disease progression including nodal metastasis.nnnCONCLUSIONSnThese findings suggest that ILT-4 is frequently expressed in both tumor and stromal cells of NSCLC, and it might play an important role in regulation of the host immune system.

ILT4 drives B7-H3 expression via PI3K/AKT/mTOR signalling and ILT4/B7-H3 co-expression correlates with poor prognosis in non-small cell lung cancer.

Immunoglobulin‐like transcript (ILT) 4 is critical for the inhibitory function of certain immune cells. We previously demonstrated that ILT4 is over‐expressed in human non‐small cell lung cancer (NSCLC) cells and is involved in tumour evasion via an unknown mechanism. In this report, we demonstrate that ILT4 increases the expression of the co‐inhibitory molecule B7‐H3 through PI3K/AKT/mTOR signalling. In primary human NSCLC tissues, a significant positive relationship is observed between ILT4 and B7‐H3 expression. ILT4/B7‐H3 co‐expression is significantly associated with a reduction in T infiltrating lymphoid cells and lower overall survival. In summary, ILT4 increases B7‐H3 expression and ILT4/B7‐H3 co‐expression may be involved in NSCLC progression.

Inhibitory receptor immunoglobulin-like transcript 4 was highly expressed in primary ductal and lobular breast cancer and significantly correlated with IL-10

BackgroundImmunoglobulin-like transcript 4 (ILT4) is an inhibitory molecule involved in immune response and has recently been identified to be strongly inducible by IL-10. The aim of the present study was to examine the associations of ILT4 expression with clinicopathological characteristics and IL-10 expression in primary ductal and lobular breast cancer.MethodsWe studied the expression of ILT4 in 4 cancer cell lines, 117 primary tumor tissues and 97 metastatic lymph nodes from patients with primary ductal and lobular breast cancer by reverse transcription-polymerase chain reaction, western blot or immunohistochemistry analysis. Additionally, IL-10 expression was also investigated using immunohistochemistry in primary tumor tissues. Then the relationship between ILT4 expression and clinicopathological characteristics/IL-10 expression was evaluated.ResultsILT4 was highly expressed in all 4 human breast cancer cell lines on both mRNA and protein levels. In primary tumor tissues, ILT4 or IL-10 was expressed in the cell membrane, cytoplasm, or both; the positive rate of ILT4 and IL-10 expression was 60.7% (71/117) and 80.34% (94/117), respectively. ILT4 level was significantly correlated with IL-10 (r =0.577; pu2009<u20090.01). Furthermore, the expression of ILT4 or IL-10 was associated with less number of Tumor Infiltrating Lymphocytes (TILs) (pu2009=u20090.004 and 0.018, respectively) and more lymph node metastasis (pu2009=u20090.046 and 0.035, respectively).ConclusionOur data demonstrated the association of ILT4 and IL-10 expression in human breast cancer, suggesting their important roles in immune dysfunction and lymph node metastases.Virtual SlidesThe virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/1692652692107916

Co-expression of immunoglobulin-like transcript 4 and angiopoietin-like proteins in human non-small cell lung cancer

The development of strategies for the inhibition of non‑small cell lung cancer (NSCLC) progression and metastasis have been mainly unsuccessful, in part due to insufficient mechanistic understanding of the disease. In the current study, the critical role of the co‑expression of immunoglobulin‑like transcript 4 (ILT4) and its ligands, angiopoietin‑like proteins (ANGPTLs), in the development of NSCLC was demonstrated. ILT4 and ANGPTL2 or ANGPTL5 were found to be co‑expressed in the five NSCLC cell lines that were investigated at the mRNA and protein level. Upon up‑ or downregulation of ILT4, the expression of ANGPTL2 was increased or reduced, respectively, while the expression of ANGPTL5 was unaffected. The co‑expression of ILT4 and ANGPTL2/ANGPTL5 was detected in human primary NSCLC tissues using immunohistochemical analysis. In total, 114 lung cancer specimens were included in the study; high expression of ILT4, ANGPTL2 and ANGPTL5 was observed in 58.8, 45.6 and 55.3%, respectively. The expression of ILT4 was found to be significantly correlated with a high expression level of ANGPTL2 (R=0.466, P=0.004); however, it was not correlated with the expression of ANGPTL5 (R=0.142, P=0.131). In ILT4‑positive samples, cases with ANGPTL2‑positive expression levels presented greater levels of lymph node metastasis (P=0.011) and shorter overall survival times (P=0.045). In addition, cases with ANGPTL5‑positive expression presented poor overall survival rates (P=0.040). By contrast, in the ILT4‑negative cases, no statistically significant differences were identified in the overall survival rates between samples with high and low expression of ANGPTL2 or ANGPTL5. In conclusion, the present study demonstrated the presence of interaction among ILT4 and ANGPTLs, which may be important in NSCLC progression. Therefore, the blockade of ANGPTLs or ILT4 may be an effective therapeutic approach for NSCLC treatment.

Clinicopathological significance of fibroblast growth factor 1 in non-small cell lung cancer.

Fibroblast growth factor (FGF) 1 is identified as a candidate cancer biomarker in several kinds of cancer. However, little is known about its expression and function in non-small cell lung cancer (NSCLC). The aim of this study is to identify the expression of FGF1 in primary human NSCLC tissues and evaluate its clinical significance for NSCLC patients. Archived tissues from NSCLC (n = 113) and adjacent normal lung tissues (n = 71) were examined for the immunohistochemical expression of FGF1; then we analyzed the correlations of FGF1 expression with clinicopathological factors and overall survival of the patients. FGF1 expression was identified in the cytoplasm or both cytoplasm and nucleus of NSCLC cells. Immunoreactive scores of FGF1 were significantly higher in NSCLC specimens than in peritumoral normal tissues. High expression of FGF1 (immunoreactive score >3) was detected in 61.9% (70/113) of NSCLC specimens, and high FGF1 expression in cancer cells was significantly correlated with larger primary tumor size, squamous cell carcinoma (SQCC), and vascular invasion. In addition, FGF1 expression was correlated with intratumoral microvessel density in both SQCC and adenocarcinoma subgroups. Moreover, NSCLC patients with high FGF1 expression had a significantly lower overall survival rate, compared with those with low FGF1 expression. Furthermore, subgroup analyses showed that FGF1 expression was associated with poor prognosis in lung SQCC, but not in adenocarcinoma. These findings suggest that the presence of FGF1 in NSCLC cells may serve as a prognostic indicator and a potential therapeutic target for NSCLC patients, especially for lung SQCC.

Overexpression of both platelet-derived growth factor-BB and vascular endothelial growth factor-C and its association with lymphangiogenesis in primary human non-small cell lung cancer

BackgroundMetastatic spread of tumor through lymphatic vasculature is an important adverse prognostic factor in a variety of human cancer and tumor lymphangiogenesis requires the interplay of several growth factors. Platelet-derived growth factor (PDGF)-BB and vascular endothelial growth factor (VEGF)-C are two important molecules involving in tumor metastasis and lymphangiogenesis. Therefore, the aim of this study was to investigate the coexpression of PDGF-BB and VEGF-C in primary human non-small cell lung cancer (NSCLC) and its association with lymphangiogenesis.MethodsUsing immunohistochemical staining, PDGF-BB and VEGF-C expression were detected in 109 primary NSCLC tissues, while the lymphatic micro-vessel density (LMVD) was counted.ResultsOf 109 cases, PDGF-BB and VEGF-C overexpression was 66.97% (73/109) and 65.14% (71/109), respectively. 52 (47.7%) had overexpression of both PDGF-BB and VEGF-C (Pu2009+u2009V+), 21 (19.3%) overexpression of PDGF-BB but low expression of VEGF-C (Pu2009+u2009V-), 19(17.4%) overexpression of VEGF-C but low expression of PDGF-BB (P-V+) and 17(15.6%) low expression of both PDGF-BB and VEGF-C (P-V-). PDGF-BB expression was positively related to that of VEGF-C (ru2009=u20090.451, pu2009=u20090.034). LMVD in cases with Pu2009+u2009Vu2009+u2009was much higher than those with P-V- (pu2009=u20090.004). In addition, the patients with Pu2009+u2009Vu2009+u2009were younger and also had larger tumor size, more likely lymph node metastasis and worse histological differentiation than those with P-V-. Moreover, the overall survival (OS) of patients with Pu2009+u2009Vu2009+u2009was shorter than those with P-V- (pu2009=u20090.015).ConclusionCoexpression of both PDGF-BB and VEGF-C was associated with lymphangiogenesis and poor prognosis in NSCLC, and might play a critical role in NSCLC progression.Virtual SlidesThe virtual slide(s) for this article can be found here:http://www.diagnosticpathology.diagnomx.eu/vs/2261801312571320

A Significant Statistical Advancement on the Predictive Values of ERCC1 Polymorphisms for Clinical Outcomes of Platinum-Based Chemotherapy in Non-Small Cell Lung Cancer: An Updated Meta-Analysis

Background. There is no definitive conclusion so far on the predictive values of ERCC1 polymorphisms for clinical outcomes of platinum-based chemotherapy in non-small cell lung cancer (NSCLC). We updated this meta-analysis with an expectation to obtain some statistical advancement on this issue. Methods. Relevant studies were identified by searching MEDLINE, EMBASE databases from inception to April 2015. Primary outcomes included objective response rate (ORR), progression-free survival (PFS), and overall survival (OS). All analyses were performed using the Review Manager version 5.3 and the Stata version 12.0. Results. A total of 33 studies including 5373 patients were identified. ERCC1 C118T and C8092A could predict both ORR and OS for platinum-based chemotherapy in Asian NSCLC patients (CT + TT versus CC, ORR: OR = 0.80, 95% CI = 0.67–0.94; OS: HR = 1.24, 95% CI = 1.01–1.53) (CA + AA versus CC, ORR: OR = 0.76, 95% CI = 0.60–0.96; OS: HR = 1.37, 95% CI = 1.06–1.75). Conclusions. Current evidence strongly indicated the prospect of ERCC1 C118T and C8092A as predictive biomarkers for platinum-based chemotherapy in Asian NSCLC patients. However, the results should be interpreted with caution and large prospective studies are still required to further investigate these findings.