Weifeng Zhu

Immunoassay for LMP1 in nasopharyngeal tissue based on surface-enhanced Raman scattering

Background: Previous studies have shown that Epstein–Barr virus (EBV)-encoded latent membrane protein 1 (LMP1) is closely associated with the occurrence and development of nasopharyngeal carcinoma, and can be used as a tumor marker in screening for the disease. Here we report a new methodology based on highly specific and sensitive surface-enhanced Raman scattering (SERS) technology to detect LMP1 in nasopharyngeal tissue sections directly with no need of tedious procedures as with conventional immunohistochemistry methods. Methods: LMP1-functionalized 4-mercaptobenzoic acid (4-MBA)-labeled Au/Ag core-shell bimetallic nanoparticles were prepared first and then applied for analyzing LMP1 in formalin-fixed paraffin-embedded nasopharyngeal tissue sections obtained from 34 cancer patients and 20 healthy controls. SERS spectra were acquired from a 25 × 25 spot square area on each tissue section and used to generate SERS images. Results: Data from SERS spectra and images show that this new SERS-based immunoassay detected LMP1 in formalin-fixed paraffin-embedded nasopharyngeal tissue sections with high sensitivity and specificity. The results from the new LMP1-SERS probe method are superior to those of conventional immunohistochemistry staining for LMP1, and in excellent agreement with those of in situ hybridization for EBV-encoded small RNA (EBER). Conclusion: This new SERS technique has the potential to be developed into a new clinical tool for detection and differential diagnosis of nasopharyngeal carcinoma as well as for predicting metastasis and immune-targeted treatment of nasopharyngeal carcinoma.

Label-Free Detection of Breast Masses Using Multiphoton Microscopy

Histopathology forms the gold standard for the diagnosis of breast cancer. Multiphoton microscopy (MPM) has been proposed to be a potentially powerful adjunct to current histopathological techniques. A label-free imaging based on two- photon excited fluorescence and second-harmonic generation is developed for differentiating normal breast tissues, benign, as well as breast cancer tissues. Human breast biopsies (including human normal breast tissues, benign as well as breast cancer tissues ) that are first imaged (fresh, unfixed, and unstained) with MPM and are then processed for routine H-E histopathology. Our results suggest that the MPM images, obtained from these unprocessed biopsies, can readily distinguish between benign lesions and breast cancers. In the tissues of breast cancers, MPM showed that the tumor cells displayed marked cellular and nuclear pleomorphism. The tumor cells, characterized by irregular size and shape, enlarged nuclei, and increased nuclear-cytoplasmic ratio, infiltrated into disrupted connective tissue, leading to the loss of second-harmonic generation signals. For breast cancer, MPM diagnosis was 100% correct because the tissues of breast cancers did not have second-harmonic generation signals in MPM imaging. On the contrary, in benign breast masses, second-harmonic generation signals could be seen easily in MPM imaging. These observations indicate that MPM could be an important potential tool to provide label-free noninvasive diagnostic impressions that can guide surgeon in biopsy and patient management.

Loss of expression rather than cytoplasmic mislocalization of RUNX3 predicts worse outcome in non-small cell lung cancer

Functional inactivation of human runt-related transcription factor 3 (RUNX3) through mutation or epigenetic silencing has been well-documented in many cancerous entities. In addition to gene mutation and promoter hypermethylation, cytoplasmic mislocalization has emerged as another major manifestation of RUNX3 dysfunction in malignancies including breast, colorectal and gastric cancers. The aim of the present study was to investigate whether patients with non-small cell lung cancer (NSCLC) and different RUNX3 expression patterns would have different overall survival (OS), and the associations between different patterns of clinicopathological parameters and clinical outcome. Expressions of RUNX3 and Ki-67 were immunohistochemically detected in normal lung tissue (n=5) and surgically resected tissues from NSCLC patients (n=188). The optimal cutoff of RUNX3 was determined by X-tile software associated with their survival. Apoptotic index in cancerous tissue was evaluated using the terminal deoxynucleotidyl transferase mediated dUTP-biotin nick end labelling method. The prognostic significance of different expression patterns of RUNX3 was determined by means of Kaplan-Meier survival estimates and log-rank tests. It was revealed that loss of RUNX3 expression in NSCLC was correlated with a low cancerous apoptotic index (P<0.001), shorter OS and worse prognosis (P=0.0142), while no statistical difference of apoptotic index (P=0.73) or survival (P=0.3781) was determined between patient subgroups with different localization of RUNX3 expression, which was quite different from the situation demonstrated in other malignancies. In conclusion, loss of expression rather than cytoplasmic mislocalization of RUNX3 predicted worse outcome in NSCLC, which was quite different from what manifested in other cancer types, and thus, the underlying mechanism may deserve further investigation.

TP53 mutations predict for poor survival in ALK rearrangement lung adenocarcinoma patients treated with crizotinib

Background Advanced non-small cell lung cancer (NSCLC) patients who harbor anaplastic lymphoma kinase (ALK) rearrangement are sensitive to an ALK inhibitor (crizotinib), but not all ALK-positive patients benefit equally from crizotinib treatment. We analyze the impact of TP53 mutations on response to crizotinib in patients with ALK rearrangement NSCLC. Methods Sixty-six ALK rearrangement NSCLC patients receiving crizotinib were analyzed. 21 cases were detected successfully by the next generation sequencing validation FFPE before crizotinib. TP53 mutations were evaluated in 8 patients in relation to disease control rate (DCR), objective response rate (ORR), progression-free survival (PFS) and overall survival (OS). Results TP53 mutations were observed in 2 (25.00%), 1 (12.50%), 1 (12.50%) and 4 (50.00%) patients in exons 5, 6, 7 and 8, respectively. The majority of patients were male (75.00%, 6/8), less than 65 years old (62.50%, 5/8) and never smokers (75.00%, 6/8). ORR and DCR for crizotinib in the entire case series were 61.90% and 71.43%, respectively. Statistically significant difference was observed in terms of PFS and OS between TP53 gene wild group and mutation group patients (P=0.038, P=0.021, respectively). Conclusions TP53 mutations reduce responsiveness to crizotinib and worsen prognosis in ALK rearrangement NSCLC patients.

Clinicopathologic features of 112 cases with mantle cell lymphoma.

Objective This study aims to explore the clinicopathologic features of 112 patients with mantle cell lymphoma (MCL). Methods Data from 112 MCL cases were collected, and immunohistochemical assay was conducted. Fluorescence in situ hybridization (FISH) detected a break in the CCND1 gene. The t-test was used in the statistical analysis. Results All tumor cells in the 112 cases expressed B cell-related antigen, including 1 blastoid subtype and 1 polymorphic subtype. Among all cases, 106 expressed CD5 and 104 expressed cyclin D1. A break in the CCND1 gene was not found in 3 cases with CD5-MCL. IgH/CCND1 polyploid was observed in 2 classic cases. Conclusion MCL is a type of special immunophenotypic B-cell lymphoma. The prognoses of blastoid and polymorphic subtypes are poor. Special subtypes should be classified during diagnosis.