[Correlation Between Serological Indices and Liver Histological Pathology in Patients with HBV Infection].

OBJECTIVE: To analyze the correlation between serological indices and liver histological pathology in the patients with hepatitis B virus ( HBV) infection. METHODS: This study enrolled 301 patients with HBV infection. All of them received liver biopsy, serum biochemical examination, including alanineamino transferase (ALT), aspart ateaminotransferase (AST), albumin (ALB), globulin (GLB ), HBV DNA test and HBV genotype detection. RESULTS: ALT, AST and AST/ALT in G2/G(3+4) group were significantly higher than those in group G0/G1 (P < 0.05), and all showed positive correlation with liver inflammation (r = 0.487, 0.648, 0.509, P < 0.05). GLB in S2/S(3+4) group was significantly higher than that in group S0/S1 (P < 0.05), which had positive correlation with liver fibrosis (r = 0.674, P < 0.05). ALB/GLB (A/G) in S2/S(3+4) group was significantly lower than that in group S0/S1 (P < 0.05), it had negative correlation with liver fibrosis(r = -0.500, P < 0.05). The inflammation and fibrosis level in patients with C genotype was higher than that of B genotype (χ2 = 11.460, 12.729, P < 0.05). CONCLUSION: ALT, AST and AST/ALT show better diagnostic value in liver inflammation. GLB and A/G show better diagnostic value in liver fibrosis. The progress of this disease is relatively faster in the patients with C genotype HBV infection.

OTX1 promotes colorectal cancer progression through epithelial-mesenchymal transition.

Orthodenticle homeobox 1 (OTX1), a transcription factor containing a bicoid-like homeodomain, plays a role in brain and sensory organ development. In this study, we report that OTX1 is overexpressed in human colorectal cancer (CRC) and OTX1 overexpression is associated with higher stage. Functional analyses reveal that overexpression of OTX1 results in accumulation of CRC cell proliferation and invasion in vitro and tumor growth in vivo, whereas ablation of OTX1 expression significantly inhibits the proliferative and invasive capability of CRC cells in vitro. Together, our results indicate that OTX1 is involved in human colon carcinogenesis and may serve as a potential therapeutic target for human colorectal cancer.

[Value of fGGO in diagnosing stage I lung cancers].

OBJECTIVE: To determine the value of focal area of ground-glass opacity (fGGO) for early detection and diagnosis of lung cancers. METHODS: We reviewed clinical data of all patients whose chest CT images showed isolated lesions < or = 5 cm in diameter in the Department of Chest Surgery at West China Hospital, Sichuan University between 2007 and 2010. According to the volume of solid components, the lesions were classified as pure ground-glass opacity (pGGO), mixed ground-glass opacity (mGGO) or solid lesions. The malignant ratio and stage of lesions were calculated based on the postoperative pathological tests. The characteristics of CT signs were compared between the benign and malignant lesions. RESULTS: Of the 202 cases, 63 (included 15 pGGO and 48 mGGO) had fGGO with a malignant ratio of 71.4% (45/63). The percentage of malignant tumors in the mGGO, pGGO and solid lesions was 75.0%, 60.0% and 48.2% respectively. Stage I lung cancers had an occurrence of spiculation, lobulation and vascular convergence in fGGO of over 70%, higher than that of the benign tumors (P < 0.05). CONCLUSION: fGGO is an important indicator of lung cancer. mGGO is highly likely to be malignant, particularly when one or more signs of spiculation, lobulation and vascular convergence appear.

[Effect of doxorubicin on TRAIL resistance and TRAIL receptor expression in lymphoma cell line SNK-6 cells].

OBJECTIVE: To investigate the effect of doxorubicin on TRAIL resistance and TRAIL receptor expression in lymphoma cell line SNK-6 cells. METHODS: SNK-6 cells treated with doxorubicin at different concentrations alone or in combination with tumor necrosis factor related apoptosis inducing ligand (TRAIL). Cell proliferation was evaluated by MTT assay. Apoptosis and the expression of TRAIL receptors were determined by flow cytometry. RESULTS: MTT assay showed that treatment with 100 and 1000 ng/ml doxorubicin for 24 h, the survival rates of SNK-6 cells were (80.9 ± 7.2)% and (53.7 ± 2.8)%, significantly higher than that by treatment combined with 500 ng/ml TRAIL (64.9 ± 1.1)% and (34.0 ± 3.9)%, respectively (P < 0.05). Flow cytometry showed that after treatment with 100 and 1000 ng/ml doxorubicin for 48 h, the survival rates of SNK-6 cells were (69.9 ± 6.1)% and (31.1 ± 1.9)%, while treated in combination with 500 ng/ml TRAIL, the cell survival rates were (37.5 ± 6.4)% and (15.0 ± 1.8)%, respectively. The early apoptosis rate was (14.8 ± 0.6)% and (30.8 ± 1.5)%, significantly lower than that [(28.7 ± 0.6)% and (46.6 ± 2.8)%] after treatment in combination with TRAIL (P < 0.05). The expressions of TRAIL receptors and decoy receptors were increased when SNK-6 cells were treated with 100 ng/ml doxorubicin for 24 hours. CONCLUSIONS: Doxorubicin can overcome to a certain extent the TRAIL resistance of SNK-6 cells and induce upregulation of TRAIL death receptors and decoy receptors on the surface of SNK-6 cells. However, a higher dose is needed.

Multisystem ALK-positive histiocytosis: a multi-case study and literature review.

BACKGROUND: Anaplastic lymphoma kinase (ALK)-positive histiocytosis, a novel rare histiocytic proliferation, was first described in 2008; it occurs in early infancy with liver and hematopoietic involvement. The spectrum was subsequently broadened to include localized diseases in older children and young adults. However, its full clinicopathological features and molecular lineage have not been fully elucidated. RESULTS: Here, we report four cases of multisystem ALK-positive histiocytosis without hematopoietic involvement. Clinically, three patients were adults aged between 32 and 51 years. Two patients', whose main manifestations were intracranial mass and numerous micronodules in the thoracoabdominal cavity organs and skin papules respectively, had a partial response to ALK inhibitors after surgery. One patient presented with mediastinal neoplasm without surgical treatment, and progressive disease occurred after two years of ALK inhibitor therapy. The fourth patient was a 17-month-old male with a large intracranial mass and presented with a poor response to ALK inhibitor and chemoradiotherapy; he died eight months after surgery. Pathologically, the histiocytes were large, with abundant eosinophilic cytoplasm, and mixed with variable numbers of foamy cells and Touton giant cells. Interstitial fibrosis was also observed. Histiocytes were positive for macrophage markers (CD68 and CD163) and ALK. KIF5B-ALK fusions were detected in two cases, EML4-ALK in one, and both DCTN1-ALK and VRK2-ALK fusions were detected in one case. CONCLUSIONS: We observed that ALK inhibitors present robust and durable responses in adult patients but a poor response in young children with central nervous system involvement. There is no consensus on the optimal treatment regimen and long-term prognosis requires further observation. Moreover, every unusual histiocytic proliferative lesion, especially unresectable and multisystem involvement, should be routinely tested for ALK immunohistochemical staining to identify this rare disease.

[Langerhans cell histiocytosis: a clinicopathologic and immunohistochemical analysis of 258 cases].

OBJECTIVES: To observe the clinicopathologic features of Langerhans cell histiocytosis (LCH), and to evaluate the values of langerin, CD1a and S-100 protein expression in diagnosis of the tumor. METHODS: Total 258 cases of Langerhans cell histiocytosis in the past 18 years (from 1992 to 2008) were collected, morphologic review and immunohistochemical staining were performed. RESULTS: In all 258 cases, the ages of patients older than 16 years or younger than 2 years were 126 (48.8%) and 37 (14.3%), respectively, in the remaining 95 (36.8%) of the cases, the age of the patients ranged from 2 to 16 years. For all of 258 cases, there were 364 diseased sites. Bony lesions accounted for 77.2% (281 cases), especially the skull (112 cases, 39.9%), followed by lymph node (25 cases, 6.9%) and skin (14 cases, 3.8%). Clinically, unisystem or unifocal disease was predominant (201 cases, 77.9%), followed by unisystem and multifocal disease (21 cases, 8.1%), multi-system disease (26 cases, 10.1%), isolated pulmonary LCH (2 cases, 0.8%), and unclassified (8 cases, 3.1%). Histologically, variable number of Langerhans cells was present in 265 samples of 258 cases. Multinucleated giant cells were found in 166 (62.6%) of the samples. Eosinophils were the major infiltrating non-neoplastic cells, and eosinophilic abscess was seen in 57 cases (21.5%). Coagulative necrosis and dead bone were detected in 29 (10.9%) and 124 (46.8%) of the cases, respectively. Immunohistochemically, the expression of S-100 protein, CD1a and langerin was 99.1% (209/211), 100% (206/206) and 98.5% (193/196), respectively, and the sensitivity of them had no statistical difference. CONCLUSIONS: In this group of LCH cases, the ratio of adult patients is high, but the proportion of multi-organ lesion is low. No significant difference of the sensitivity is found among langerin, CD1a and S-100 expression in diagnosis of LCH.

Identification and characterization of oligonucleotides that inhibit Toll-like receptor 2-associated immune responses.

Toll-like receptors (TLRs) play important roles in the immune responses against invading microorganisms. Development of TLR antagonists is recognized as a promising direction in suppressing the associated inflammatory reactions of the TLRs. Aptamers are single-stranded RNA or DNA molecules isolated through an in vitro selection process. Using a novel molecular evolution strategy that combines immunoprecipitation (IP) with systematic evolution of ligands by exponential enrichment (SELEX), we developed an IP-SELEX selection method to facilitate the screening of high-affinity aptamers for the Toll-like receptor 2 (TLR2). Also, human TLR2 functional aptamers were identified and characterized using NF-kappaB reporter assays. Among the functional aptamers, the most effective, AP177, with a dissociation constant of 73 pM, was characterized with TLR2-expressing cells challenged with bacterial cells and purified ligands. The aptamer could effectively antagonize TLR2, significantly inhibit NF-kappaB activity, and suppress the secretion of the cytokines by >80%. In addition, the precise region within the functional aptamer that specifically bound TLR2 was resolved using aptamer microarray analysis. The results of functional assays showed that AP177 acted as a TLR2 antagonist and may hold therapeutic potential in the treatment of diseases related to dysregulated TLR2 immune responses.

[Lymphoplasmacytic lymphoma with Waldenström's macroglobulinemia: a clinicopathological and immunophenotypic study of 40 Chinese patients].

OBJECTIVE: To investigate the clinicopathologic features of lymphoplasmacytic lymphoma (LPL) with Waldenström's macroglobulinemia (WM) and to evaluate the usefulness of immunophenotype analysis in diagnosis and differential diagnosis of the tumor. METHODS: A total of 40 cases of LPL with WM diagnosed according to the 2008 World Health Organization classification of tumors of hematopoietic and lymphoid tissues were analyzed using immunophenotype and follow-up information. RESULTS: The mostly common initial clinical presentations were non-specific symptoms, such as fatigue, anemia and hemorrhage. Lymphadenopathy, splenomegaly and hepatomegaly were found in 42.5%, 20.0% and 12.5% of the patients respectively. The pattern of bone marrow involvement included mixed type (47.2%), diffuse type (41.7%) and interstitial type (11.1%). The nodal architecture was completely destroyed in one case and partially effaced with residual germinal centers and dilated sinuses in 8 cases. All of the neoplastic cells expressed CD20 and CD79a. Neoplastic plasma cells were positive for CD138 and CD79a. No cases expressed CD5. Four cases weakly expressed CD23. No significant prognosis related factors were identified in the survival analysis. CONCLUSIONS: LPL with WM is a rare indolent small B-cell lymphoma, which is commonly seen, in older male patients. The tumor frequently involves bone marrow and shows various clinical manifestations. Combination analyses of the bone marrow biopsy histology, immunophenotypic study and clinical data, especially the serum examination are important for the diagnosis of LPL with WM.

[Primary cutaneous anaplastic large cell lymphoma: a clinicopathologic analysis of 8 cases].

OBJECTIVE: To study the clinicopathologic features, immunophenotype and prognosis of primary cutaneous anaplastic large cell lymphoma (C-ALCL). METHODS: Eight cases of C-ALCL were enrolled into the study. The clinicopathologic features, immunohistochemical findings and results of in-situ hybridization for EBER 1/2 were analyzed. RESULTS: Three of the 8 patients were males and 5 were females. The median age was 49.5 years. C-ALCL often presented with solitary skin nodule, without systemic symptoms. Histologically, the lymphoma cells infiltrated the dermis and subcutis in a sheet-like pattern. They were of large size and showed conspicuous nuclear atypia. Immunohistochemical study showed that more than 75% of the lymphoma cells were positive for CD30. All cases expressed one to three T cell markers (CD3, CD5 or CD45RO) and cytotoxic granule-associated antigens (TIA-1, granzyme B or perforin). The staining for leukocyte common antigen was positive in all cases, while the expression of CD5, CD8, ALK-1 and epithelial membrane antigen was noted in 5, 1, 1 and 3 cases, respectively. The staining for CD15, CD20, CK and HMB45 was negative. In-situ hybridization for EBER 1/2 was also negative in all the cases studied. Follow-up information was available in 6 patients. Five of them were still alive and 1 died of unclear cause. CONCLUSIONS: C-ALCL has distinctive clinicopathologic and immunophenotypic features. It is not Epstein-Barr virus-related and often carries a favorable prognosis.

Significant benefits of osimertinib in treating acquired resistance to first-generation EGFR-TKIs in lung squamous cell cancer: A case report.

BACKGROUND: Lung squamous cell cancer (LSCC) rarely harbors epidermal growth factor receptor (EGFR) mutations, even much rarer for acquired T790M mutation. Although clinical trials of AURA series illustrated that non-small cell lung cancer (NSCLC) with EGFR T790M mutation can benefit from osimertinib, only five LSCC patients were enrolled in total; moreover, the efficacy for LSCC was not shown in the results. Therefore, the response of LSCC to osimertinib is still unclear to date. CASE SUMMARY: We report an LSCC case with T790M-related acquired resistance after treatments with first-generation EGFR-tyrosine kinase inhibitors (EGFR-TKIs) and benefited from osimertinib significantly. A 63-year-old Chinese man was diagnosed with stage IV (cT2N2M1b) LSCC harboring an EGFR exon 19-deletion mutation. Following disease progression after gefitinib and multi-line chemotherapy, re-biopsy was conducted. Molecular testing of EGFR by amplification refractory mutation system-polymerase chain reaction detected the exon 19-deletion without T790M mutation. Therefore, the patient was given erlotinib, but progression developed only 3 mo later. Then the frozen re-biopsy tissue was tested by next-generation sequencing (NGS), which detected an EGFR T790M mutation. However, he was very weak with symptoms of dysphagia and cachexia. Fortunately, osimertinib was started, leading to alleviation from the symptoms. Four months later, normal deglutition was restored and partial response was achieved. Finally, the patient achieved an overall survival time period of 29 mo. CONCLUSION: Our findings highlight that EGFR T790M mutation may also be an important acquired drug resistance mechanism for LSCC and offer direct evidence of the efficacy of osimertinib in LSCC with T790M mutation. NGS and better preservation conditions may contribute to higher sensitivity of EGFR T790M detection.

Distribution of PD-L1 expression and its relationship with clinicopathological variables: an audit from 1071 cases of surgically resected non-small cell lung cancer.

BACKGROUND: Programmed death ligand 1 (PD-L1) was reported to predict the response of immunotherapy; however, the association between PD-L1 expression and clinicopathologic characteristics has yet to be elucidated in non-small cell lung cancer (NSCLCs). MATERIALS AND METHODS: We reviewed PDL1 expression investigated by immunohistochemical analysis using FFPE tissue in a total of 1071 cases of primary or metastatic NSCLC tissues analyzed between 2015-2017, and evaluated the association between PD-L1 expression and the clinicopathologic characteristics. RESULTS: PD-L1 expression was observed in 361 (33.7%) cases with positive staining in at least 1% tumor cells and 116 (10.8%) cases had positive staining in ≥50% tumor cells. The PD-L1 positive prevalence was significantly higher in squamous cell carcinoma (SCC) than in adenocarcinoma (AD). In the AD subgroup, PD-L1 expression on tumors was higher in males and smokers, and with high histologic grade, relative high T, N, M status, advanced AJCC stage, and in ALK rearrangement patients. However, EGFR mutated patients showed relatively lower PD-L1 expression than wild type patients. CONCLUSION: This study revealed the unique distribution of PD-L1 expression with clinicopathologic features in East Asian NSCLCs in a single, large cohort of patients. Since immunohistochemistry of the PD-L1 protein (PD-L1 IHC) is the only clinically approved predictive biomarker for anti-PD-1/-PD-L1 therapy currently, our outcomes could help to stratify patients to ensure selection of those who would most benefit from PD-1/PD- L1 inhibitor therapy.

[Correlation between VEGF expression and clinical characteristic of patient with non-Hodgkin's lymphoma: an initial study].

OBJECTIVE: To study whether VEGF expression is associated with clinical characteristics such as gender, age, aggressive, stage, B-symptoms and complete remission (CR) rate of patients with non-Hodgkin's lymphoma. METHODS: Expression of VEGF had been tested in 98 biopsy samples by immunohistochemical method as LSAB. Chi-Square test was used to do the statistical analysis of ratio comparison between different groups. RESULTS: Comparing VEGF positive rates grouped by sexuality, age, aggressive grade or stage, the results suggested no significant correlation between them. VEGF positive rate of patients accompanied with B-symptoms was 86.1%, versus 51.6% of patients B-symptoms free, P = 0.001. Among 94 patients, their responses to chemotherapy could be estimated, delaminated by B-symptoms, and the CR rate of VEGF negative group was significantly higher, P values were 0.000 and 0.028. CONCLUSIONS: Expression of VEGF is associated with B-symptoms and poor response to chemotherapy in patients with non-Hodgkin's lymphoma, there was not any relationship to have been found between VEGF expression and sexuality, age, aggressive grade, stage.

Three-dimensional visualization of the functional fascicular groups of a long-segment peripheral nerve.

The three-dimensional (3D) visualization of the functional bundles in the peripheral nerve provides direct and detailed intraneural spatial information. It is useful for selecting suitable surgical methods to repair nerve defects and in optimizing the construction of tissue-engineered nerve grafts. However, there remain major technical hurdles in obtaining, registering and interpreting 2D images, as well as in establishing 3D models. Moreover, the 3D models are plagued by poor accuracy and lack of detail and cannot completely reflect the stereoscopic microstructure inside the nerve. To explore and help resolve these key technical problems of 3D reconstruction, in the present study, we designed a novel method based on re-imaging techniques and computer image layer processing technology. A 20-cm ulnar nerve segment from the upper arm of a fresh adult cadaver was used for acetylcholinesterase (AChE) staining. Then, 2D panoramic images were obtained before and after AChE staining under the stereomicroscope. Using layer processing techniques in Photoshop, a space transformation method was used to fulfill automatic registration. The contours were outlined, and the 3D rendering of functional fascicular groups in the long-segment ulnar nerve was performed with Amira 4.1 software. The re-imaging technique based on layer processing in Photoshop produced an image that was detailed and accurate. The merging of images was accurate, and the whole procedure was simple and fast. The least square support vector machine was accurate, with an error rate of only 8.25%. The 3D reconstruction directly revealed changes in the fusion of different nerve functional fascicular groups. IN CONCLUSION: The technique is fast with satisfactory visual reconstruction.

Mediastinal myelolipoma/extramedullary hematopoiesis presenting as a mass: rare differential diagnosis among mediastinal tumors.

OBJECTIVES: Mediastinal myelolipoma/extramedullary hematopoiesis presenting as a mass is infrequent and can lead to misdiagnosis. Here we describe a large series aiming to illustrate the clinicopathologic features. METHODS: We retrospectively searched mediastinal tumors and myelolipoma diagnosed at the Department of Pathology, West China Hospital from 2010 to 2015 and collected 14 mediastinal myelolipoma/extramedullary hematopoiesis cases presenting as an encapsulated mass among 1324 mediastinal mass diseases and 252 myelolipomas. RESULTS: There were 8 females and 6 males aged from 35 to 67 years old, most of whom were diagnosed incidentally. Cross-sectional imaging revealed encapsulated masses located in the posterior mediastinum with fat and soft tissue density showing heterogeneous enhancement. Radiologic diagnosis was neurogenic tumor for most cases. All but one patient underwent surgery and postoperative pathologic findings showed fat and hematologic elements. Considering the accompanying hematologic disorders, 5 patients were diagnosed as extramedullary hematopoiesis and the remaining 9 as myelolipoma. The average hematopoietic tissue percentage in extramedullary hematopoiesis was 70%, significantly higher than it was in myelolipoma. Patients showed no sign of recurrence or metastasis apart from the patient with hepatocellular carcinoma. CONCLUSIONS: Mediastinal myelolipoma/extramedullary hematopoiesis is a rare entity of solid tumors in the posterior mediastinum, affecting patients from their third decades, with no sex predilection and lacking unique clinical symptoms, and may be misdiagnosed as a malignant tumor on cross-sectional imaging. The final diagnosis relies on pathologic findings, and the precise classification of myelolipoma or extramedullary hematopoiesis relies on percentage of hematopoietic tissue and accompanying clinical symptoms. Surgery is the recommended treatment.

Synthesis and pharmacological evaluation of novel beta-nitrostyrene derivatives as tyrosine kinase inhibitors with potent antiplatelet activity.

Protein tyrosine kinases have been known to be involved in regulation of platelet aggregation, suggesting a potential target for antiplatelet therapy. Our previous study showed that 3,4-methylenedioxy-beta-nitrostyrene (MNS) prevented platelet aggregation caused by various stimulators, and this action was accompanied by inhibition of tyrosine kinases. In the present study, in order to examine the structural determinants required for the actions of MNS and to develop more potent tyrosine kinase inhibitors and antiplatelet agents, a new series of beta-nitrostyrene derivatives were synthesized and pharmacologically characterized. The beta-nitrostyrene derivatives inhibited thrombin- or collagen-induced human platelet aggregation, ATP secretion, GPIIb/IIIa activation and protein tyrosine phosphorylation. In recombinant enzyme assay, some beta-nitrostyrene derivatives also demonstrated potent inhibition of Src and/or Syk kinase activity. Furthermore, there was a good correlation between the inhibitory potency of these compounds on tyrosine kinases and on platelet activation/aggregation. Among them, a benzoyl ester derivative (compound 10) possess up to 8-fold greater potency than MNS and over two orders of magnitude greater potency than genistein or tyrphostin A47 in inhibiting platelet responses to thrombin. Our data suggest that beta-nitrostyrenes may represent a new class of tyrosine kinase inhibitors with potent antiplatelet activity.

Overexpression of Rad51 Predicts Poor Prognosis in Colorectal Cancer: Our Experience with 54 Patients.

BACKGROUND: Aberrant Rad51 expression is implicated in the progression of human malignancies. However, the role of Rad51 in colorectal cancer (CRC) remains undefined. This study aimed to establish a relationship between Rad51 and clinicopathologic features of CRC. METHODS: We retrospectively examined the paraffin-embedded tissue samples obtained from 54 patients with CRC who had received surgical therapies at our institution during 2006-2008. Rad51 expression in adenocarcinoma, paracancerous tissue, and normal colonic tissue was determined by immunohistochemistry. The correlation between Rad51 immunoreactivity and clinicopathologic features of these patients was evaluated. RESULTS: Rad51 immunoreactivity was detected in 67% of adenocarcinoma, 48% of paracancerous tissue, and 27% of normal colonic mucosa. Rad51 expression in adenocarcinoma was significantly higher than normal colonic tissue (p < 0.05). Rad51 was also overexpressed in poorly differentiated tumors and tumor samples from patients with lymph node metastasis (p < 0.05). Patients with Rad51 overexpression had a 69% two-year survival, 49% three-year survival, and 16% five-year survival, considerably worse than patients with negative Rad51 expression (p < 0.05). CONCLUSION: Our data suggest that Rad51 overexpression is correlated with malignant phenotypes of CRC and may predict poor prognosis for these patients.

[Diffuse large B-cell lymphoma with expression of anaplastic lymphoma kinase protein: clinicopathologic and immunohistochemical study of 5 cases].

OBJECTIVE: To study the clinicopathologic features of diffuse large B-cell lymphoma (DLBCL) with expression of anaplastic lymphoma kinase (ALK) protein. METHODS: Nine hundred and forty-five (945) cases of DLBCL (including 177 consultation cases) diagnosed according to the 2001 World Health Organization classification of tumors of hematopoietic and lymphoid tissues were enrolled into the study. Immunohistochemical study for anti-ALK-11 was performed using LSAB technique. The ALK-positive cases were further confirmed by immunohistochemical study using EnVision technique. Only ALK-positive cases by EnVision technique were further analyzed by immunostaining for antigens including CD20, CD3, CD30, EMA, granzyme-B, TIA-1 and PC. Immunoglobulin heavy chain gene rearrangement study was also performed and follow-up data collected. RESULTS: There were altogether 5 (4 males and 1 female) cases of DLBCL showing expression of ALK protein. The age of the patients ranged from 34 to 72 years. All were primary nodal DLBCL. One case belonged to clinical stage I, 2 in stage II and 2 in stage III. The duration of follow up ranged from 4 to 32 months. Three patients subsequently died and the longest survival was 32 months. Morphologic subtypes included centroblastic 2, anaplastic 1, immunoblastic with plasmacytoid differentiation 1 and plasmablastic 1. Immunohistochemically, 4 cases were CD20 positive (including 2 centroblastic, 1 anaplastic and 1 immunoblastic cases). The plasmablastic case expressed kappa light chain and was negative for CD20. Rearrangement of immunoglobulin heavy chain gene was demonstrated in all 5 cases studied. As for ALK protein staining, a mixed membranous and cytoplasmic (1 immunoblastic case), granular cytoplasmic (2 centroblastic and 1 anaplastic cases) and mixed nuclear and cytoplasmic (1 plasmablastic case) patterns were observed. CONCLUSIONS: Expression of ALK protein is a rare phenomenon in DLBCL and can be seen in centroblastic, anaplastic, immunoblastic and plasmablastic subtypes. It is often associated with aggressive clinical behavior and worse prognosis. A new pattern of ALK protein expression, mixed membranous and cytoplasmic, is reported.

EGFR mutations in non-small cell lung cancer: an audit from West China Hospital.

OBJECTIVES: To discover the incidence and characteristics of EGFR mutations in non-small cell lung cancer (NSCLC) in a single, large cohort as a part of routine diagnostic investigations. METHODS: We reviewed EGFR mutations investigated by Amplification Refractory Mutation System (ARMS) PCR (covering 29 known mutations) using DNA samples from FFPE tissue or cell clot specimens in a total of 3894 cases of NSCLC analysed between 2012-2014. RESULTS: EGFR mutations are preferentially associated with adenocarcinomaand adenosquamous histology, particularly those well to moderately differentiated, and were significantly more common in female than male patients irrespective of histological subtypes. Exon 19 deletion (45.7%) and exon 21 L858R (45.6%) accounted for the vast majority of the EGFR mutations detected, with the remaining mutations being infrequent (<2%). Compound mutations were seen in 51 (3%) of the mutant cases, the combination of these compound mutations could be classified into three subgroups according to the potential impact of individual mutations on EGFR TKI therapy. Accordingly, 7 cases had both sensitive mutations, 4 cases harboured one sensitive and one less responsive /uncertain mutation, 19 cases contained one sensitive and one resistant change, and a further 21 cases had two less responsive /uncertain mutations. CONCLUSION: Our data represents the largest EGFR mutation survey based on routine clinical diagnostic laboratory data from a single institution, it confirms the incidence and characteristics of EGFR mutations in NSCLC seen in Asian patients, and also unravels the combinatorial nature of rare compound EGFR mutations.

Noninvasive genotyping and monitoring of anaplastic lymphoma kinase (ALK) rearranged non-small cell lung cancer by capture-based next-generation sequencing.

Noninvasive genotyping of driver genes and monitoring of tumor dynamics help make better personalized therapeutic decisions. However, neither PCR-based assays nor amplicon-based targeted sequencing can detect fusion genes like anaplastic lymphoma kinase (ALK) rearrangements in blood samples. To investigate the feasibility and performance of capture-based sequencing on ALK fusion detection, we developed a capture-based targeted sequencing panel to detect and quantify rearrangement events, along with other driver mutation variants in plasma. In this perspective study, we screened 364 patients with advanced non-small cell lung cancer (NSCLC) for ALK rearrangements, and collected blood samples from 24 of them with confirmed ALK rearrangements based on their tissue biopsies. ALK rearrangements were successfully detected in 19 of 24 patients at baseline with 79.2% (95% CI 57.9%, 92.9%) sensitivity and 100% (36/36) specificity. Among the 24 patients, we obtained longitudinal blood samples from 7 of them after either chemotherapy and/or Crizotinib treatment for disease monitoring. The by-sample detection rate of ALK rearrangements after treatment drops to 69.2% (9 of 13). In addition to detecting ALK rearrangements, we also detected 3 Crizotinib resistant mutations, ALK L1152R, ALK I1171T and ALK L1196M from patient P4. ctDNA concentration correlates with responses and disease progression, reflecting its ability as a biomarker. Our findings suggest capture-based sequencing can detect and quantify ALK rearrangements as well as other somatic mutations, including mutations mediated drug resistance, in plasma with high sensitivity, paving the way for its application in identifying driver fusion genes and monitoring tumor dynamics in the clinic.