Jianghao Chen

The role of human papillomavirus infection in breast cancer

Breast cancer is the leading female cancer and the third most common cause of cancer deaths worldwide. Many studies have suggested a possible link between breast cancer pathogenesis and viral infection, particularly mouse mammary tumour virus, simian virus 40, Epstein–Barr virus, and human papillomavirus (HPV). A significant number of recent studies have reported that approximately 29% of human breast cancer tissues were positive for high-risk HPV subtypes, especially HPV subtypes 16, 18, or 33. In contrast, several other investigations did not detect any HPV subtypes in either breast cancer tissue or normal breast tissue from patients diagnosed with breast cancer. Given these conflicting data and the established complexity of the association between HPV with other cancers, a definitive relationship between human breast cancer and HPV infection has not been determined. Recent advances in laboratory methodologies aim to overcome the inherent challenges in detecting HPV in breast cancer tissue. There is an urgent need to obtain additional evidence in order to assess the possibility of breast cancer prevention using HPV vaccines.

Drug concentrations in axillary lymph nodes after lymphatic chemotherapy on patients with breast cancer

BackgroundLymph node status is one of the decisive prognostic factors in breast cancer. Chemotherapy targeting regional lymphatic tissues has emerged as a promising therapy for the treatment of malignancies with a high tendency to disseminate lymphatically. The present study determined the drug concentrations in axillary lymph nodes after lymphatic chemotherapy (LC) in patients with breast cancer and compared the results with those receiving intravenous chemotherapy (VC) to investigate whether LC could improve the accumulation of anticancer drug in regional lymph nodes.MethodsSixty patients with breast carcinoma confirmed by preoperative puncture-biopsy were divided into two groups at random. The LC group (n = 30) received a subcutaneous injection of 4 ml of carboplatin-activated carbon suspension, containing 20 mg of carboplatin, adjacent to the primary tumour. The VC group (n = 30) received an intravenous administration of an equal dose of aqueous carboplatin. At 1, 12, 24, 36 and 48 hours after administration, modified radical mammectomies were performed on 12 patients at each time point, with 6 from each group. Axillary lymph nodes were removed for pathological examination. The platinum concentrations in nodes were determined by Zeeman atomic absorption spectrometry.ResultsA total of 275 axillary lymph nodes were resected, with 154 in the LC group and 121 in the VC group. Of the 275 lymph nodes, 136 (49.5%) from 23 patients (38.3%) had histopathologically detected metastases. At 1, 12, 24, 36 and 48 hours after injection, the carboplatin concentrations in the LC group were 11.82 ± 3.50, 23.58 ± 7.34, 18.22 ± 4.93, 16.70 ± 5.15 and 14.62 ± 4.29 μg/g (means ± SD), respectively, whereas those in the VC group were 0.06 ± 0.02, 0.11 ± 0.05, 0.10 ± 0.02, 0.05 ± 0.02 and 0 μg/g, respectively. Significant differences were found in each corresponding comparison (P < 0.001). Lymph node metastasis was uncorrelated with drug concentration (P > 0.05).ConclusionLC can effectively and continuously improve the drug concentrations in axillary lymph nodes in patients with breast cancer, in comparison with VC.

Prognostic significance of USP22 as an oncogene in papillary thyroid carcinoma

Ubiquitin-specific protease 22 (USP22), a novel deubiquitinating enzyme, has been associated with metastasis, therapy resistance, and cell cycle progression. The purpose of this study was to investigate the expression level of USP22 in papillary thyroid carcinoma (PTC) samples and to evaluate its clinical significance in PTC patients. USP22 expression was examined in 30 fresh PTC tissues and paired adjacent noncancerous tissues by real-time quantitative RT-PCR. Immunohistochemistry for USP22 was performed on additional 156 PTC tissues. The clinical significance of USP22 expression was analyzed. We found that the expression levels of USP22 mRNA and protein in PTC tissues were both significantly higher than those in noncancerous tissues. Clinicopathological analysis showed that USP22 expression was significantly correlated with tumor size (p = 0.036), extracapsular invasion (p = 0.012), multifocality (p = 0.014), lymph node metastasis (p = 0.022), distant metastasis (p = 0.005), and TNM stage (p = 0.002). The Kaplan–Meier survival curves revealed that USP22 expression was associated with poor prognosis in PTC patients. USP22 expression was an independent prognostic marker of overall patient survival in a multivariate analysis. Our findings suggest that USP22 is an independent predictor of poor prognosis of PTC patients.

The significance of expression of autophagy-related gene Beclin, Bcl-2, and Bax in breast cancer tissues

The purpose of this study was to detect the expression of autophagy-related gene Beclin1 and apoptosis-related genes Bcl-2 and Bax in breast cancer tissues, to investigate their relationship and significance to the occurrence and development of breast cancer, and to provide an experimental basis for the biological treatment of breast cancer in the future. Human breast cancer tissues and relatively healthy breast tissue adjacent to the tumor were collected during surgical resection. By using RT–PCR and western blot, the mRNA and protein expressions of Beclin1, Bcl-2, and Bax were detected in the breast cancer tissues and the relatively healthy, adjacent tissues. The correlations of these expressions with the occurrence, development, and clinicopathology of breast cancer were analyzed. The mRNA and protein expressions of Beclin1 and Bcl-2 in breast cancer tissues were significantly lower than those in the relatively healthy, adjacent breast tissues (p < 0.05); the lower the degree of tumor differentiation, the lower the mRNA and protein expressions of Beclin1 and Bcl-2 (p < 0.05); the mRNA and protein expressions of Beclin1 and Bcl-2 in breast cancer tissues from patients positive for lymph node metastasis were significantly lower than those negative for lymph node metastasis (p < 0.05); the mRNA and protein expressions of Beclin1 and Bcl-2 in breast cancer tissues from patients positive for distant metastasis were significantly lower than those negative for distant metastasis (p < 0.05); the mRNA and protein expressions of Beclin1 and Bcl-2 in breast cancer tissues from patients positive for ki67 were significantly lower than those negative for ki67 (p < 0.05). The mRNA and protein expressions of Bax were different from those of Beclin1 and Bcl-2. In breast cancer tissues, the mRNA and protein expressions of Bax were up-regulated (p < 0.05); the lower the degree of tumor differentiation, the higher the mRNA and protein expressions of Bax (p < 0.05); the mRNA and protein expressions of Bax in breast cancer tissues from patients positive for lymph node metastasis were significantly higher than those negative for lymph node metastasis (p < 0.05); and the mRNA and protein expressions of Bax in breast cancer tissues from patients positive for distant metastasis were significantly higher than those in patients negative for distant metastasis (p < 0.05). However, the mRNA and protein expressions of these three genes were not correlated with patient age, tumor size, progesterone receptor positivity, or human epidermal growth factor positivity (p > 0.05). The correlation of Bcl-2 and Bax mRNA with Beclin1 mRNA expressed in breast cancer tissues were both statistically significant (p < 0.05). The activity change of autophagy and apoptosis is associated with the tumorigenesis and tumor progression of breast cancer. The joint detection of these three genes (Beclin1, Bcl-2, and Bax) contributes to the early diagnosis of and predicts prognosis for breast cancer, and this also provides an experimental basis for the biological therapy of breast cancer.

Correlation between hybrid 18F-FDG PET/CT and apoptosis induced by neoadjuvant chemotherapy in breast cancer.

Quantitative or semi-quantitative analysis of fluorine-18 fluorodeoxyglucose positron emission tomography (18F-FDG PET) has been reported to correlate with the clinical and pathological response of tumors to preoperative treatment. This study was conceived to evaluate the correlation between hybrid 18F-FDG PET/CT and apoptosis in breast cancer after neoadjuvant chemotherapy. Three cycles of neoadjuvant chemotherapy were given to forty-five patients with primary breast cancer proven by core needle biopsy. Hybrid PET/CT was performed before and after treatment and tumor to non-tumor radioactivity ratio (T/N) was calculated. The apoptotic index (AI) in core-cut and surgically resected samples was determined using TUNEL techniques. The mean T/N pre- and post-chemotherapy was 3.23 ± 0.63 and 2.31 ± 0.49, respectively (P = 0.006), with the mean reduction rate below baseline of 31.18 ± 13.18% (range, 6.4-50.8%). The mean AI pre- and post-chemotherapy was 2.81 ± 0.76% and 17.31 ± 6.85%, respectively (P < 0.0001). The mean AI change was 14.34 ± 5.36% (range, 1.9-41.3%). A positive correlation was detected between the T/N reduction rate and AI change (rs = 0.850, P < 0.0001). At a threshold of 20% decrease from baseline in T/N, the mean AI change in the tumors with a reduction of 20% or more in T/N was 20.86 ± 4.29%, while that in the tumors with a reduction of less than 20% in T/N was 8.59 ± 2.87% (P < 0.0001). The sensitivity, specificity, positive and negative predictive values, and the accuracy of PET/CT for the prediction of clinical response were 90.9%, 83.3%, 93.8%, 76.9%, and 92%, respectively. These data suggested that neoadjuvant chemotherapy may effectively induce apoptosis in breast tumors and decrease their glucose uptake. Hybrid PET/CT imaging appeared to be positively related to apoptosis level and therefore to be of value in predicting the response of breast cancer to neoadjuvant chemotherapy.

Full-length single-cell RNA-seq applied to a viral human cancer: applications to HPV expression and splicing analysis in HeLa S3 cells

BackgroundViral infection causes multiple forms of human cancer, and HPV infection is the primary factor in cervical carcinomas. Recent single-cell RNA-seq studies highlight the tumor heterogeneity present in most cancers, but virally induced tumors have not been studied. HeLa is a well characterized HPV+ cervical cancer cell line.ResultWe developed a new high throughput platform to prepare single-cell RNA on a nanoliter scale based on a customized microwell chip. Using this method, we successfully amplified full-length transcripts of 669 single HeLa S3 cells and 40 of them were randomly selected to perform single-cell RNA sequencing. Based on these data, we obtained a comprehensive understanding of the heterogeneity of HeLa S3 cells in gene expression, alternative splicing and fusions. Furthermore, we identified a high diversity of HPV-18 expression and splicing at the single-cell level. By co-expression analysis we identified 283 E6, E7 co-regulated genes, including CDC25, PCNA, PLK4, BUB1B and IRF1 known to interact with HPV viral proteins.ConclusionOur results reveal the heterogeneity of a virus-infected cell line. It not only provides a transcriptome characterization of HeLa S3 cells at the single cell level, but is a demonstration of the power of single cell RNA-seq analysis of virally infected cells and cancers.

Steroidal saponin of Trillium tschonoskii. Reverses multidrug resistance of hepatocellular carcinoma

Combating with multidrug resistance (MDR) is a major part of hepatocellular carcinoma (HCC) chemotherapy. Steroidal saponin from Trillium tschonoskii (TTS) could be a potential weapon. We found TTS could reverse the MDR in HCC cells and significantly enhance chemosensitization. TTS inhibited HepG2 and R-HepG2 cells survival in a dose-dependent manner by 75% and 76%, respectively (p<0.01), as well as colony formation 77% and 81% (p<0.01). Moreover, TTS induced sensitization of R-HepG2 to anti-cancer drugs, indicated by significantly reduced IC50. On the other hand, TTS suppressed expression of P-glucoprotein in MDR HCC cells, and thereby increased accumulation of doxorubicin from 126 ng/10(5)cells to 752 ng/10(5)cells (p<0.01). TTS also repressed expression of many other MDR genes, such as MRP1, MRP2, MRP3, MRP5, MVP and GST-π. In vivo, TTS dose-dependently reduced R-HepG2 cells xenografts tumour formation by inhibiting tumour cells proliferation in mice. Consistence with in vitro finding, TTS induced R-HepG2 sensitization to doxorubicin and therefore reduced tumour formation in vivo.

Subanesthetic Isoflurane Reduces Zymosan-Induced Inflammation in Murine Kupffer Cells by Inhibiting ROS-Activated p38 MAPK/NF-κB Signaling

Volatile anesthetic isoflurane (ISO) has immunomodulatory effects. The fungal component zymosan (ZY) induces inflammation through toll-like receptor 2 or dectin-1 signaling. We investigated the molecular actions of subanesthetic (0.7%) ISO against ZY-induced inflammatory activation in murine Kupffer cells (KCs), which are known as the resident macrophages within the liver. We observed that ISO reduced ZY-induced cyclooxygenase 2 upregulation and prostaglandin E2 release, as determined by western blot and radioimmunoassay, respectively. ISO also reduced the production of tumor necrosis factor-α, interleukin-1β, IL-6, high-mobility group box-1, macrophage inflammatory protein-1α, macrophage inflammatory protein-2, and monocyte chemoattractant protein-1 as assessed by enzyme-linked immunosorbent assays. ISO blocked the ZY-induced nuclear translocation and DNA-binding activity of nuclear factor- (NF)-κB p65. Moreover, ISO attenuated ZY-induced p38 mitogen-activated protein kinase (MAPK) activation partly by scavenging reactive oxygen species (ROS); the interregulation that ROS activated p38 MAPK followed by NF-κB activation was crucial for the ZY-induced inflammatory responses in KCs. An in vivo study by peritoneal injection of ZY into BALB/C mice confirmed the anti-inflammatory properties of 0.7% ISO against ZY in KCs. These results suggest that ISO ameliorates ZY-induced inflammatory responses in murine KCs by inhibiting the interconnected ROS/p38 MAPK/NF-κB signaling pathways.

Lymphatic chemotherapy induces apoptosis in lymph node metastases in a rabbit breast carcinoma model

The purpose of the study was to evaluate the potential of lymphatic chemotherapy in inducing apoptosis in axillary lymph node metastases in a rabbit breast cancer model. A total of 30 female New Zealand rabbits with mammary implantation of VX2 carcinomas were divided into three groups randomly, with ten in each. Treatment was carried out once axillary lymph node reached 5 mm in the maximum diameter. Group A received a subcutaneous injection of liposomal adriamycin (LADR) adjacent to the breast tumor. Group B received free adriamycin (FADR) administered into the auricular vein. Group C received a sham treatment. The dose of adriamycin in each administration was 1 mg/kg in groups A and B. Treatment was repeated every 48 h. Axillary lymph nodes were dissected out 48 h after the third treatment. The nodal sizes before and after the treatment were measured. The therapeutic effect was evaluated in terms of the node volume ratio and apoptotic index (AI) of metastatic cells in nodes identified with TUNEL technique. The significance of difference was determined with one-way ANOVA followed by the Fischer LSD test. Compared to group C, the enlargement of lymph nodes was sufficiently slowed down in both groups A and B, and group A showed a further strong inhibitory effect than group B (P=0.002). Apparent VX2 cell apoptosis was detected in the lymph nodes of groups A and B. The average AI in group B (15.31%) was significantly higher than in group C (5.16%). The highest AI was found in animals of group A (21.73%), with a further significant difference from group B (P=0.000). These data suggest that lymphatic chemotherapy appears to be a promising method to induce apoptosis in lymph node metastases and holds potential in the treatment of advanced breast cancer.

Multiplex PCR/mass spectrometry screening of biological carcinogenic agents in human mammary tumors.

BACKGROUND While many studies have suggested a possible link between breast cancer pathogenesis and infection by viruses, the role of viruses in breast carcinogenesis remains controversial. OBJECTIVES We analyzed the prevalence of 30 oncogenic human papillomaviruses (HPVs), Epstein-Barr virus (EBV), Kaposis sarcoma herpes virus (KSHV) and six polyomaviruses in breast tumor specimens. STUDY DESIGN We analyzed breast specimens from 100 breast cancer patients (group 1) and 50 benign breast disease patients (group 2) from Shaanxi Province in China. We also screened for the viruses in blood samples from the patients and 96 female blood donor volunteers (group 3). RESULTS EBV, Merkel cell polyomavirus (MCPyV) and HPV-18 were detected in 60, 14 and 2 breast cancer patients, respectively, and EBV and MCPyV were detected in 16 and 1 benign breast disease patients, respectively. EBV and MCPyV were more prevalent in group 1 than in group 2 (EBV: 60.0% vs. 32.0%, p = 0.0012; MCPyV: 14.0% vs. 2.0%; p = 0.02). In contrast, there was no difference in the prevalence of EBV and MCPyV in blood samples between group 1 and group 2, group 1 and group 3. EBV was detected in malignant breast tissue and its presence was confined to the malignant cells using in situ hybridization. CONCLUSIONS We found that EBV and MCPyV were more prevalent in the tumors of women with breast cancer than in samples from women with benign breast disease. Our results support an etiologic role for EBV in breast cancer pathogenesis in Chinese patients.