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Background: Accurately identifying patients with axillary pathologic complete response (pCR) after neoadjuvant chemotherapy (NAC) in breast cancer patients remains challenging., Purpose: To compare the feasibility of shear wave elastography (SWE) performed on breast tumors and axillary lymph nodes (LNs) in predicting the axillary status after NAC., Materials and Methods: This prospective study included a total of 319 breast cancer patients with biopsy-proven positive node who received NAC followed by axillary lymph node dissection from 2019 to 2022. The correlations between shear wave velocity (SWV) and pathologic characteristics were analyzed separately for both breast tumors and LNs after NAC. We compared the performance of SWV between breast tumors and LNs in predicting the axillary status after NAC. Additionally, we evaluated the performance of the most significantly correlated pathologic characteristic in breast tumors and LNs to investigate the pathologic evidence supporting the use of breast or axilla SWE., Results: Axillary pCR was achieved in 51.41% of patients with node-positive breast cancer. In breast tumors, there is a stronger correlation between SWV and collagen volume fraction (CVF) (r = 0.52, p < 0.001) compared to tumor cell density (TCD) (r = 0.37, p < 0.001). In axillary LNs, SWV was weakly correlated with CVF (r = 0.31, p = 0.177) and TCD (r = 0.29, p = 0.213). No significant correlation was found between SWV and necrosis proportion in breast tumors or axillary LNs. The predictive performances of both SWV and CVF for axillary pCR were found to be superior in breast tumors (AUC = 0.87 and 0.85, respectively) compared to axillary LNs (AUC = 0.70 and 0.74, respectively)., Conclusion: SWE has the ability to characterize the extracellular matrix, and serves as a promising modality for evaluating axillary LNs after NAC. Notably, breast SWE outperform axilla SWE in determining the axillary status in breast cancer patients after NAC., (© 2024. The Author(s).)

Background: To develop a convenient modality to predict axillary response to neoadjuvant chemotherapy (NAC) in breast cancer patients., Materials and Methods: In this multi-center study, a total of 1019 breast cancer patients with biopsy-proven positive lymph node (LN) receiving NAC were randomly assigned to the training and validation groups at a ratio of 7:3. Clinicopathologic and ultrasound (US) characteristics of both primary tumors and LNs were used to develop corresponding prediction models, and a nomogram integrating clinicopathologic and US predictors was generated to predict the axillary response to NAC., Results: Axillary pathological complete response (pCR) was achieved in 47.79% of the patients. The expression of estrogen receptor, human epidermal growth factor receptor -2, Ki-67 score, and clinical nodal stage were independent predictors for nodal response to NAC. Location and radiological response of primary tumors, cortical thickness and shape of LNs on US were also significantly associated with nodal pCR. In the validation cohort, the discrimination of US model (area under the curve [AUC], 0.76) was superior to clinicopathologic model (AUC, 0.68); the combined model (AUC, 0.85) demonstrates strong discriminatory power in predicting nodal pCR. Calibration curves of the nomogram based on the combined model demonstrated that substantial agreement can be observed between the predictions and observations. This nomogram showed a false-negative rates of 16.67% in all patients and 10.53% in patients with triple negative breast cancer., Conclusion: Nomogram incorporating routine clinicopathologic and US characteristics can predict nodal pCR and represents a tool to aid in treatment decisions for the axilla after NAC in breast cancer patients., Competing Interests: Disclosure The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Background: Gastric cancer (GC) is the most common malignant tumor and ranks third for cancer-related deaths among the worldwide. The disease poses a serious public health problem in China, ranking fifth for incidence and third for mortality. Knowledge of the invasive depth of the tumor is vital to treatment decisions., Aim: To evaluate the diagnostic performance of double contrast-enhanced ultrasonography (DCEUS) for preoperative T staging in patients with GC by comparing with multi-detector computed tomography (MDCT)., Methods: This single prospective study enrolled patients with GC confirmed by preoperative gastroscopy from July 2021 to March 2023. Patients underwent DCEUS, including ultrasonography (US) and intravenous contrast-enhanced ultrasonography (CEUS), and MDCT examinations for the assessment of preoperative T staging. Features of GC were identified on DCEUS and criteria developed to evaluate T staging according to the 8 th edition of AJCC cancer staging manual. The diagnostic performance of DCEUS was evaluated by comparing it with that of MDCT and surgical-pathological findings were considered as the gold standard., Results: A total of 229 patients with GC (80 T1, 33 T2, 59 T3 and 57 T4) were included. Overall accuracies were 86.9% for DCEUS and 61.1% for MDCT ( P < 0.001). DCEUS was superior to MDCT for T1 (92.5% vs 70.0%, P < 0.001), T2 (72.7% vs 51.5%, P = 0.041), T3 (86.4% vs 45.8%, P < 0.001) and T4 (87.7% vs 70.2%, P = 0.022) staging of GC., Conclusion: DCEUS improved the diagnostic accuracy of preoperative T staging in patients with GC compared with MDCT, and constitutes a promising imaging modality for preoperative evaluation of GC to aid individualized treatment decision-making., Competing Interests: Conflict-of-interest statement: All authors declare that they have no conflict of interest., (©The Author(s) 2024. Published by Baishideng Publishing Group Inc. All rights reserved.)

Clinical trials frequently include multiple end points that mature at different times. The initial report, typically based on the primary end point, may be published when key planned co-primary or secondary analyses are not yet available. Clinical Trial Updates provide an opportunity to disseminate additional results from studies, published in JCO or elsewhere, for which the primary end point has already been reported. We previously reported comparable 3-year regional relapse-free survival (RRFS) using elective upper-neck irradiation (UNI) in N0-1 nasopharyngeal carcinoma (NPC) compared with standard whole-neck irradiation (WNI). Here, we present the prespecified 5-year overall survival (OS), RRFS, late toxicity, and additional analyses. In this randomized trial, patients received UNI (n = 224) or WNI (n = 222) for an uninvolved neck. After a median follow-up of 74 months, the UNI and WNI groups had similar 5-year OS (95.9% v 93.1%, hazard ratio [HR], 0.63 [95% CI, 0.30 to 1.35]; P = .24) and RRFS (95.0% v 94.9%, HR, 0.96 [95% CI, 0.43 to 2.13]; P = .91) rates. The 5-year disease-free survivors in the UNI group had a lower frequency of hypothyroidism (34% v 48%; P = .004), neck tissue damage (29% v 46%; P < .001), dysphagia (14% v 27%; P = .002), and lower-neck common carotid artery stenosis (15% v 26%; P = .043). The UNI group had higher postradiotherapy circulating lymphocyte counts than the WNI group (median: 400 cells/μL v 335 cells/μL, P = .007). In conclusion, these updated data confirmed that UNI of the uninvolved neck is a standard of care in N0-1 NPC, providing outstanding efficacy and reduced long-term toxicity, and might retain more immune function.

Purpose To compare the tissue adequacy and diagnostic accuracy of US-guided biopsies of peripheral pulmonary lesions (PPLs) with and without contrast agents. Materials and Methods A retrospective study was conducted at four medical centers in patients with PPLs who underwent US-guided percutaneous transthoracic needle biopsy (PTNB) between January 2017 and October 2022. The patients were divided into contrast-enhanced US (CEUS) and US groups based on whether prebiopsy CEUS evaluation was performed. Tissue adequacy and the diagnostic accuracy of PTNB, stratified by lesion size, were analyzed and compared between groups. A propensity score matching (PSM) analysis was conducted using the nearest-neighbor matching method. Results A total of 1027 lesions were analyzed, with 634 patients (mean age, 59.4 years ± 13.0 [SD]; 413 male) in the US group and 393 patients (mean age, 61.2 years ± 12.5; 270 male) in the CEUS group. The CEUS group produced more acceptable samples than the US group (98.2% vs 95.7%; P = .03) and achieved higher diagnostic accuracy (96.9% vs 94.2%; P = .04), with no evidence of a difference in sensitivity (96.7% vs 94.0%; P = .06). PSM and stratified analyses ( n = 358 per group) indicated higher tissue adequacy (99.0% vs 95.7%; P = .04) and diagnostic accuracy (98.5% vs 92.9%; P = .006) in the CEUS group compared with the US group for 2-7-cm PPLs but not for lesions larger than 7 cm. Conclusion PTNB with prebiopsy CEUS evaluation demonstrated significantly better tissue adequacy and diagnostic accuracy compared with US guidance alone for PPLs ranging from 2 to 7 cm, with similar biopsy performance achieved between groups for lesions larger than 7 cm. Keywords: Contrast Material, Thoracic Diseases, Ultrasonography, Image-Guided Biopsy © RSNA, 2024.

Rationale and Objectives: To develop a monitoring model using radiomics analysis based on longitudinal B-mode ultrasound (BUS) and shear wave elastography (SWE) to early predict pathological response to neoadjuvant chemotherapy (NAC) in breast cancer patients., Materials and Methods: In this prospective study, 112 breast cancer patients who received NAC between September 2016 and March 2022 were included. The BUS and SWE data of breast cancer were obtained prior to treatment as well as after two and four cycles of NAC. Radiomics features were extracted followed by measuring the changes in radiomics features compared to baseline after the second and fourth cycles of NAC (△R [C2], △R [C4]), respectively. The delta radiomics signatures were established using a support vector machine classifier., Results: The area under receiver operating characteristic curve (AUC) values of △R BUS (C2) and △R BUS (C4) for predicting the response to NAC were 0.83 and 0.84, while those of △R SWE (C2) and △R SWE (C4) were 0.88 and 0.90, respectively. △R SWE exhibited significantly superior performance to △R BUS for predicting NAC response (Delong test, p < 0.01). No significant differences were observed in the performances between △R (C2) and △R (C4) based on BUS or SWE data. The longitudinal dual-modal ultrasound radiomics (LDUR) model had an excellent discrimination, good calibration and clinical usefulness, with the AUC, sensitivity and specificity of 0.97, 95.52% and 91.11%, respectively., Conclusion: The LDUR model achieved excellent performance in predicting the pathological response to chemotherapy during the early stages of NAC for breast cancer., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Association of University Radiologists. Published by Elsevier Inc. All rights reserved.)

Objective: Abdominal ultrasonography after transrectal filling with contrast agent (AU-TFCA) was retrospectively evaluated with respect to determination of T stage and lesion length in patients with colorectal cancer (CRC) who had previously failed colonoscopy because of severe intestinal stenosis., Methods: The population comprised 83 patients with CRC with intestinal stenosis and previously failed colonoscopy who underwent AU-TFCA, and in addition contrast-enhanced computed tomography (CECT) and/or magnetic resonance imaging (MRI), 2 wk before surgery. The diagnostic performance of AU-TFCA and CECT/MRI was evaluated relative to the post-operative pathological results (PPRs) by paired sample t-test, receiver operator characteristic (ROC) curve, Pearson's χ 2 -test and κ and intraclass correlation coefficients., Results: The T staging identified via AU-TFCA, but not CECT/MRI, was relatively consistent with that of the PPRs (linearly weighted κ coefficient: 0.558, p < 0.001, and linearly weighted κ coefficient: 0.237, p < 0.001, respectively). The overall diagnostic accuracy of T staging based on AU-TFCA (83.1%) was significantly higher than that based on CECT/MRI (50.6%). Regarding lesion length, the results of AU-TFCA and PPRs were comparable (t = 1.852, p = 0.068), but those of CECT/MRI and PPRs were significantly different (t = 8.450, p < 0.001)., Conclusion: AU-TFCA is effective in evaluation of lesion length and T stage in patients with severely stenotic CRC lesions who previously failed colonoscopy. The diagnostic accuracy of AU-TFCA is significantly better compared with that of CECT/MRI., Competing Interests: Conflict of interest The authors declare no competing interests., (Copyright © 2023 World Federation for Ultrasound in Medicine & Biology. Published by Elsevier Inc. All rights reserved.)

Rationale and Objectives: To carry out radiomics analysis/deep convolutional neural network (CNN) based on B-mode ultrasound (BUS) and shear wave elastography (SWE) to predict response to neoadjuvant chemotherapy (NAC) in breast cancer patients., Materials and Methods: In this prospective study, 255 breast cancer patients who received NAC between September 2016 and December 2021 were included. Radiomics models were designed using a support vector machine classifier based on US images obtained before treatment, including BUS and SWE. And CNN models also were developed using ResNet architecture. The final predictive model was developed by combining the dual-modal US and independently associated clinicopathologic characteristics. The predictive performances of the models were assessed with five-fold cross-validation., Results: Pretreatment SWE performed better than BUS in predicting the response to NAC for breast cancer for both the CNN and radiomics models (P < 0.001). The predictive results of the CNN models were significantly better than the radiomics models, with AUCs of 0.72 versus 0.69 for BUS and 0.80 versus 0.77 for SWE, respectively (P = 0.003). The CNN model based on the dual-modal US and molecular data exhibited outstanding performance in predicting NAC response, with an accuracy of 83.60% ± 2.63%, a sensitivity of 87.76% ± 6.44%, and a specificity of 77.45% ± 4.38%., Conclusion: The pretreatment CNN model based on the dual-modal US and molecular data achieved excellent performance for predicting the response to chemotherapy in breast cancer. Therefore, this model has the potential to serve as a non-invasive objective biomarker to predict NAC response and aid clinicians with individual treatments., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Association of University Radiologists. Published by Elsevier Inc. All rights reserved.)

Phenylalanine ammonia lyase (PAL) catalyzes the reversible conversion of l-phenylalanine into the corresponding trans -cinnamic acid, providing a route to optically pure α-amino acids. We explored the catalytic function of all five PALs encoded in the genome of lettuce ( Lactuca sativa L.) that are previously known to be involved in wound browning. All Ls PALs were active toward l-phenylalanine in the ammonia elimination reaction and displayed maximum activity at 55-60 °C and pH 9.0-9.5. However, four of them, Ls PAL1- Ls PAL4, showed significantly higher activity and thermal stability than Ls PAL5, as well as a broader substrate spectrum including some challenging substrates with steric demanding or electron-donating substituents. The best one Ls PAL3 was subjected to the kinetic resolution of a panel of 21 rac -phenylalanine derivatives, as well as the ammonia addition of 21 cinnamic acid derivatives. It showed excellent enantioselectivity in most cases and significantly better activity than previously described PALs for a number of challenging non-natural substrates, demonstrating its great potential in biocatalysis.

Objective: This study compared the performance between ultrasound (US)- and contrast-enhanced US (CEUS)-guided liver biopsies and evaluated the benefit of CEUS in percutaneous biopsy for focal liver lesions (FLLs)., Methods: We performed a retrospective study of 820 patients with FLLs, who underwent percutaneous liver biopsy in our center between 2017 and 2019. The patients were divided into two groups based on whether US (n = 362) or CEUS (n = 458) used before a biopsy. The two groups were compared based on specimen adequacy for pathological diagnosis and diagnostic accuracy of liver biopsy. Stratification analysis was performed based on lesion and protocol characteristics to provide detailed information for selecting the imaging guidance for biopsy., Results: Compared with the US group, the CEUS group yielded more acceptable samples (97.6% vs. 99.4%, p < 0.05) and improved diagnostic accuracy (92.6% vs. 96.4%, p < 0.05), and achieved better sensitivity (92.5% vs. 96.2%, p < 0.05) for liver biopsies, especially in FLLs ≥ 5 cm, heterogeneous hypoechoic FLLs, or FLLs with an obscure boundary. The CEUS group showed significantly higher accuracy compared with the US group pertaining to single-puncture biopsies (100% vs. 92.7%, p < 0.05) or biopsies with punctures ≤ 2 (97.6% vs. 94.3%, p < 0.05)., Conclusion: CEUS achieved an enhanced success rate for sampling and diagnostic accuracy of liver biopsies, especially in FLLs ≥ 5 cm, heterogeneous hypoechoic FLLs, or FLLs with an obscure boundary. CEUS can be used to decrease the number of punctures needed, which might increase the safety of liver biopsy., Key Points: • CEUS can help confirm an adequate biopsy site, increasing the sampling success rate and diagnostic accuracy of the liver biopsy. • CEUS can be used to decrease the number of punctures needed to improve the safety of liver biopsy. • It is recommended to use CEUS guidance for liver biopsies, especially with FLLs ≥ 5 cm, heterogeneous hypoechoic FLLs, or FLLs with an obscure boundary., (© 2022. The Author(s), under exclusive licence to European Society of Radiology.)

Objective: To determine the ability of conventional ultrasound (US) combined with shear wave elastography (SWE) to reveal axillary status after neoadjuvant chemotherapy (NAC) in breast cancer patients., Methods: From September 2016 to December 2021, 201 patients with node-positive breast cancer who underwent NAC were enrolled in this prospective study. Conventional US features of axillary lymph nodes and SWE characteristics of breast lesions after NAC were analyzed. The diagnostic performances of US, SWE, and their combination were assessed using multivariate logistic regression and receiver operator characteristic curve (ROC) analyses., Results: The area under the ROC curve (AUC) for the ability of conventional US features to determine axillary status after NAC was 0.82, with a sensitivity of 85.23%, a specificity of 67.39%, and an accuracy of 76.11%. Shear wave velocity (SWV) displayed moderate performance for predicting axilla status after NAC with SWV mean demonstrating an AUC of 0.85. Cortical thickness and shape of axillary nodes and SWV mean of breast tumors were independently associated with axillary nodal metastasis after NAC. Compared to conventional US, the combination of conventional US of axillary lymph nodes with SWE of breast lesions achieved a significantly higher AUC (0.90 vs 0.82, p < 0.01, Delong's test) with a sensitivity of 87.50%, improved specificity of 82.61% and accuracy of 85.00%., Conclusions: Breast SWE was independently associated with residual metastasis of axillary node after NAC in patients with initially diagnosed positive axilla. Combining SWE with conventional US showed good diagnostic performance for axillary node disease after NAC., Key Points: • Breast SWE can serve as a supplement to axilla US for the evaluation of the axilla after NAC. • The combination of axilla US with breast SWE may be a promising method to facilitate less-invasive treatment in patients receiving NAC., (© 2022. The Author(s), under exclusive licence to European Society of Radiology.)

Protein stability is crucial in enzymatic catalysis. To improve the efficiency in the searching for thermostablizing mutations, we applied a sequence consensus approach focusing on dimeric interface residues of ketoreductase ChKRED20. The strategy returned a success rate of 43%, revealing 9 beneficial mutations from 21 candidates with improved kinetic or thermodynamic stability. Several combinatorial mutants were then constructed, and mutant M8K displayed the highest thermostability, with a melting temperature (T m ) of 89 °C and a half-inactivation temperature (T 50 ) of 93.4 °C, both of over 35 °C increase compared to the wild-type. M8K could remain stable for at least 7 days at its optimal reaction temperature of 55 °C. Its inactivation half-life (t 1/2 ) was 110 min at 90 °C, while the wild-type was 18.6 min at 60 °C. The results were interpreted in the context of structural and molecular dynamic simulation analysis, which revealed the addition of intramolecular interactions, decreased conformational flexibility and increased compactness, all in agreement with the observed effect., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Asymmetric epoxidation catalyzed with styrene monooxygenase (SMO) is a powerful enzymatic process producing enantiopure styrene epoxide derivatives. To establish a more diversified reservoir of SMOs, a new SMO from Bradyrhizobium sp. ORS 375, named BrSMO, was mined from the database and characterized. BrSMO was constituted of an epoxygenase component of 415 amino acid residues and an NADH-dependent flavin reductase component of 175 residues. BrSMO catalyzed the epoxidation of styrene and 7 more styrene derivatives, yielding the corresponding (S)-epoxides with excellent enantiomeric excesses (95- > 99% ee), with the highest activity achieved for styrene. BrSMO also catalyzed the asymmetric sulfoxidation of 7 sulfides, producing the corresponding (R)-sulfoxides (20-90% ee) with good yields.

Ethyl ( S )-3-hydroxy-3-(2-thienyl)propanoate(( S )-HEES)acts as a key chiral intermediate for the blockbuster antidepressant drug duloxetine, which canbe achieved viathe stereoselective bioreduction ofethyl 3-oxo-3-(2-thienyl) propanoate (KEES) that containsa 3-oxoacyl structure.The sequences of the short-chain dehydrogenase/reductases from Chryseobacterium sp. CA49 were analyzed, and the putative3-oxoacyl-acyl-carrier-protein reductase, Ch KRED12, was able to stereoselectivelycatalyze theNADPH-dependent reduction to produce ( S )-HEES.The reductase activity of Ch KRED12 towardsothersubstrates with 3-oxoacyl structure were confirmed with excellent stereoselectivity (>99% enantiomeric excess) in most cases. When coupled with a cofactor recycling system using glucose dehydrogenase, the Ch KRED12 was able to catalyze the complete conversion of 100 g/l KEES within 12h, yielding the enantiopure product with >99% ee, showing a remarkable potential to produce ( S )-HEES.

Background: Accurate preoperative pathologic diagnosis is very important for making appropriate therapeutic decisions for patients with rectal lesions. This study aimed (I) to determine diagnostic value and safety of endoscopic forceps biopsy (EFB) and transrectal ultrasound (TRUS)-guided core needle biopsy (CNB), and (II) to analyze the risk factors for their histopathologic discrepancies, with a particular focus in identifying the indicators for re-biopsy using TRUS-guided CNB after EFB., Methods: We retrospectively reviewed the records of 102 patients who received EFB and TRUS-guided CNB before surgery. The histopathologic concordance and risk factors for underdiagnosis by EFB and TRUS-guided CNB were analyzed., Results: Compared with postoperative pathology, the histopathologic discrepancy rate of EFB and TRUS-guided CNB was 51.0% (52/102 lesions) and 8.8% (9/102 lesions), respectively. The kappa value for consistency with postoperative pathology findings was 0.420 for EFB and 0.876 for TRUS-guided CNB. The multivariate analyses and receiver operating characteristic (ROC) curve indicated that lesions thickness ≥13.5 mm [OR 1.080 (95% CI: 1.021-1.142), P=0.007] and flat/depressed shape [OR 0.206 (95% CI: 0.076-0.564), P=0.002] were significantly associated with histopathologic discrepancies in EFB., Conclusions: EFB was of limited clinical value in identifying the preoperative diagnosis of rectal lesions. Lesions thickness and flat/depressed shape at EFB were independent risk factors for pathologic discrepancies. TRUS-guided CNB may serve as a safe and effective supplement to routine EFB., Competing Interests: Conflicts of Interest: The authors have no conflicts of interest to declare., (2019 Annals of Translational Medicine. All rights reserved.)

ChKRED20 is a robust NADH-dependent ketoreductase identified from the genome of Chryseobacterium sp. CA49 that can use 2-propanol as the ultimate reducing agent. The wild-type can reduce over 100 g/l ketones for some pharmaceutical relevant substrates, exhibiting a remarkable potential for industrial application. In this work, to overcome the limitation of ChKRED20 to aryl ketoesters, we first refined the X-ray crystal structure of ChKRED20/NAD + complex at a resolution of 1.6 Å, and then performed three rounds of iterative saturation mutagenesis at critical amino acid sites to reshape the active cavity of the enzyme. For methyl 2-oxo-2-phenylacetate and ethyl 3-oxo-3-phenylpropanoate, several gain-of-activity mutants were achieved, and for ethyl 2-oxo-4-phenylbutanoate, improved mutants were achieved with k cat /K m increasing to 196-fold of the wild-type. All three substrates were completely reduced at 100 g/l loading catalyzed with selected ChKRED20 mutants, and deliver the corresponding chiral alcohols with >90% isolated yield and 97 - >99%ee., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Objectives: To determine the methodology of non-invasive test for evaluation of liver stiffness (LS) with tumours using two-dimensional (2D) shear wave elastography (SWE)., Methods: One hundred and twenty-seven patients with liver tumours underwent 2D-SWE before surgery to measure liver and spleen stiffness (SS). Two-dimensional SWE values were obtained in the liver at 0-1 cm, 1-2 cm and >2 cm from the tumour edge (PLS-1, PLS-2 and RLS, respectively). The influence of tumour-associated factors was evaluated. The area under the receiver operating characteristic curve (AUC) for each value was analysed to diagnose cirrhosis., Results: PLS-1 was higher than PLS-2, which was even higher than RLS (p < 0.001). The AUCs of PLS-1, PLS-2, RLS and SS for diagnosing cirrhosis were 0.760, 0.833, 0.940 and 0.676, with the specificity of 75.7%, 67.6%, 90.3% and 77.4%, respectively. Tumour sizes, locations or types showed no apparent influence on 2D-SWE values except for RLS, which was higher in patients with primary hepatic carcinomas (p < 0.05)., Conclusions: LS with tumours is best measured at >2 cm away from the tumour edge. SS measurement could be used as an alternative to LS measurement in the event of no available liver for detection., Key Points: • Tumour-associated factors impact background liver stiffness assessment. • Background liver stiffness is best measured at >2 cm from tumour edge. • Spleen stiffness can be an alternative to assess background liver stiffness.

Objective:: The purpose of this study is to compare contrast-enhanced ultrasound (CEUS) to MRI for evaluating local invasion of cervical cancer., Methods:: A total of 108 patients with cervical cancer were included in this study. All the enrolled patients were Stage IIA2-IVB according to the International Federation of Obstetrics and Gynecology and treated with volumetric modulated arc therapy. Tumour size in different dimensions was compared between MRI and CEUS. The correlation coefficients (r) between MRI and CEUS for diagnosing local invasion, parametrial extension, and invasion to vagina, uterine corpus and adjacent organs were assessed., Results:: Measurements by MRI and CEUS were strongly correlated in the three dimensions: left-right r = 0.84, craniocaudal r = 0.86 and anteroposterior r = 0.88. Vaginal and parametrial invasion were detected by both MRI and CEUS with moderate concordance, and invasion of uterine corpus, bladder and rectum with good concordance., Conclusion:: CEUS is comparable to MRI for measuring tumour size, with good concordance for evaluating invasion of cervical cancer., Advances in Knowledge:: CEUS is a less expensive non-invasive modality for assessment of tumour size and invasion of cervical cancer.

Objective: This study investigated the feasibility of using strain elastography (SE) and real time shear wave elastography (RT-SWE) to evaluate early tumor response to cytotoxic chemotherapy in a murine xenograft breast cancer tumor model., Methods: MCF-7 breast cancer-bearing nude mice were treated with either cisplatin 2 mg kg -1 plus paclitaxel 10 mg kg -1 (treatment group) or sterile saline (control group) once daily for 5 days. The tumor elasticity was measured by SE or RT-SWE before and after therapy. Tumor cell density was assessed by hematoxylin and eosin staining, and the ratio of collagen fibers in the tumor was evaluated by Van Gieson staining. The correlation between tumor elasticity, as determined by SE and SWE, as well as the pathological tumor responses were analyzed., Results: Chemotherapy significantly attenuated tumor growth compared to the control treatment (p < 0.05). Chemotherapy also significantly increased tumor stiffness (p < 0.05) and significantly decreased (p < 0.05) tumor cell density compared with the control. Moreover, chemotherapy significantly increased the ratio of collagen fibers (p < 0.05). Tumor stiffness was positively correlated with the ratio of collagen fibers but negatively correlated with tumor cell density., Conclusion: The study suggests that ultrasound elastography by SE and SWE is a feasible tool for assessing early responses of breast cancer to chemotherapy in our murine xenograft model. Advances in knowledge: This study showed that the tumor elasticity determined by ultrasound elastography could be a feasible imaging biomarker for assessing very early therapeutic responses to chemotherapy.

Docetaxel-based chemotherapy remains the first-line treatment for patients with metastatic castration-resistant prostate cancer (mCRPC) in China; however, the prognostic factors associated with effects in these patients are still controversial. In this study, we retrospectively reviewed the data from 71 eligible Chinese patients who received docetaxel chemotherapy from 2009 to 2016 in our hospital and experienced a reduction of prostate-specific antigen (PSA) level ≥50% during the treatment and investigated the potential role of time to nadir (TTN) of PSA. TTN was defined as the time from start of chemotherapy to the nadir of PSA level during the treatment. Multivariable Cox regression models and Kaplan-Meier analysis were used to predict overall survival (OS). In these patients, the median of TTN was 17 weeks. Patients with TTN ≥17 weeks had a longer response time to chemotherapy compared to TTN <17 weeks (42.83 vs 21.50 weeks, P < 0.001). The time to PSA progression in patients with TTN ≥17 weeks was 11.44 weeks compared to 5.63 weeks when TTN was <17 weeks. We found several factors to be associated with OS, including TTN (hazard ratio [HR]: 3.937, 95% confidence interval [CI]: 1.502-10.309, P = 0.005), PSA level at the diagnosis of cancer (HR: 4.337, 95% CI: 1.616-11.645, P = 0.004), duration of initial androgen deprivation therapy (HR: 2.982, 95% CI: 1.104-8.045, P = 0.031), neutrophil-to-lymphocyte ratio (HR: 3.963, 95% CI: 1.380-11.384, P = 0.011), and total PSA response (Class 1 [<0 response] compared to Class 2 [0-50% response], HR: 3.978, 95% CI: 1.278-12.387, P = 0.017). In conclusion, TTN of PSA remains an important prognostic marker in predicting therapeutic outcome in Chinese population who receive chemotherapy for mCRPC and have >50% PSA remission.

Objectives: This study assessed the efficacy and safety of transvaginal ultrasound (US)-guided core needle biopsy (CNB) for obtaining adequate pelvic mass samples for histologic analysis and evaluated factors that may affect biopsy success., Methods: Two hundred cases underwent transvaginal US-guided CNBs for primary inoperable tumors, suspicion of metastases to the ovaries or peritoneum, recurrence, or other solid lesions in the pelvis. Biopsy samples were obtained from the pelvic cavity (67.0%), vaginal cuff or vaginal wall (17.5%), or peritoneal cake (15.5%). The potential influences of the biopsy site (pelvic cavity, vaginal cuff or vaginal wall, or peritoneal cake), vascularization, ascites, tumor size, and tumor type (inoperable, metastases, recurrence, or solid pelvic tumor) on the success of transvaginal US-guided CNB were evaluated by a univariate analysis., Results: Adequate samples were obtained in 192 of 200 biopsies (96.0%), of which 190 yielded successful diagnoses (95.0%). The biopsy site had a significant effect on biopsy adequacy, as there was a significantly lower probability of obtaining satisfactory specimens for histologic verification from the peritoneal cake compared to pelvic tumors and the vaginal cuff or vaginal wall (P < .01). Adequacy was also affected by tumor size (P < .05) but not by vascularization, ascites, or tumor type. No complications occurred during the biopsy procedures., Conclusions: Transvaginal US-guided CNB is a safe and effective alternative to more invasive methods for evaluating pelvic lesions, such as laparoscopy and laparotomy., (© 2017 by the American Institute of Ultrasound in Medicine.)

Background: The aim of this study was to investigate the survival of patients with signet ring cell carcinoma (SRCC) based on primary tumor location., Methods: Patient data were obtained from the Surveillance, Epidemiology, and End Results database (1988-2012) with ≥200 cases per tumor location. Cox regression analysis was used to investigate prognostic factors of cause-specific survival (CSS)., Results: We identified 24,171 patients. Of the patients, 63.4% had gastric SRCC, followed by colon (18.2%), esophageal (5.0%), rectal (3.5%), lung (3.1%), pancreatic (1.8%), breast (1.5%), bladder (1.3%), small intestine (1.1%), and gallbladder SRCC (1.0%). The 5-year CSS was 22.1%, 69.0%, 33.2%, 28.1%, 24.8%, 16.1%, 13.6%, 12.6%, 11.0%, 6.4% in patients with gastric, breast, colon, rectum, bladder, small intestine, esophageal, gallbladder, lung, and pancreatic SRCC, respectively (P < 0.001). Multivariate analyses showed that the primary tumor location was an independent prognostic factor of survival. Patients with lung, small intestine, and bladder SRCC had a comparable CSS to gastric SRCC, while breast and colorectal SRCC had better survival than gastric SRCC. Esophageal, gallbladder, and pancreatic SRCC were significantly associated with poor CSS compared with gastric SRCC., Conclusion: Our study suggests a major difference in survival of SRCC based on the primary tumor locations.

Aim: The aim of this article is to test the hypothesis that remote ischaemic preconditioning (RIPC) increases circulating endogenous local and systemic plasma (nitrite) during RIPC and ischaemia-reperfusion (IR) as a potential protective mechanism against ischaemia-reperfusion injury (IRI)., Methods: Six healthy male volunteers (mean age 29.5 ± 7.6 years) were randomized in a crossover study to initially receive either RIPC (4 × 5 min cycles) to the left arm, or no RIPC (control), both followed by an ischaemia-reperfusion (IR) sequence (20 min cuff inflation to 200 mmHg, 20 min reperfusion) to the right arm. The volunteers returned at least 7 days later for the alternate intervention. The primary outcome was the effect of RIPC vs. control on local and systemic plasma (nitrite)., Results: RIPC did not significantly change plasma (nitrite) in either the left or the right arm during the RIPC sequence. However, compared to control, RIPC decreased plasma (nitrite) during the subsequent IR sequence by ~26% (from 118 ± 9 to 87 ± 5 nmol l -1 ) locally in the left arm (P = 0.008) overall, with an independent effect of -58.70 nmol l -1 (95% confidence intervals -116.1 to -1.33) at 15 min reperfusion, and by ~24% (from 109 ± 9 to 83 ± 7 nmol l -1 ) systemically in the right arm (P = 0.03)., Conclusions: RIPC had no effect on plasma (nitrite) during the RIPC sequence, but instead decreased plasma (nitrite) by ~25% during IR. This would likely counteract the protective mechanisms of RIPC, and contribute to RIPC's lack of efficacy, as observed in recent clinical trials. A combined approach of RIPC with nitrite administration may be required., (© 2017 The British Pharmacological Society.)

Objective: To evaluate the potential role of neutrophil-to-lymphocyte ratio (NLR) with therapeutic response in patients who were treated with docetaxel for mCRPC., Materials and Methods: We retrospectively analyzed the clinical data from 111 consecutive patients who were treated with docetaxel for mCRPC from 2009 to 2016 in a single center from Northwestern China. Pretreatment baseline and follow-up data including age, PSA response, Gleason score, and cycle number were reviewed, and multivariable Cox regression models and Kaplan-Meier analysis were used to predict overall survival (OS) and progression-free survival (PFS)., Results: In Kaplan-Meier analyses, the NLR (optimal threshold 3.3), total PSA response, number of chemotherapy cycles, stage T, baseline of PSA, albumin, presence of visceral metastases, and PSA level at the diagnosis of cancer were significantly associated with OS, respectively. In multivariable analyses, higher NLR (>3.3), PSA level at the diagnosis of cancer (≥162 ng/ml), number of chemotherapy cycles, and albumin (<40.5 g/l) were associated with increased risk of death, respectively. Meanwhile, young age, higher NLR, number of chemotherapy cycles, presence of visceral metastases, and poor PSA response were associated with shorter PFS., Conclusion: NLR combined with PSA level at the diagnosis of cancer remains an important prognostic marker in predicting therapeutic outcome in Chinese men who receive chemotherapy for mCRPC.

(1S)-2-chloro-1-(3, 4-difluorophenyl) ethanol ((S)-CFPL) is an intermediate for the drug ticagrelor, and is manufactured via chemical approaches. To develop a biocatalytic solution to (S)-CFPL, an inventory of ketoreductases from Chryseobacterium sp. CA49 were rescreened, and ChKRED20 was found to catalyze the reduction of the ketone precursor with excellent stereoselectivity (>99 % ee). After screening an error-prone PCR library of the wild-type ChKRED20, two mutants, each bearing a single amino acid substitution of H145L or L205M, were identified with significantly increased activity. Then, the two critical positions were each randomized by constructing saturation mutagenesis libraries, which delivered several mutants with further enhanced activity. Among them, the mutant L205A was the best performer with a specific activity of 178 μmol/min/mg, ten times of that of the wild-type. Its k cat /K m increased by 15 times and half-life at 50 °C increased by 70 %. The mutant catalyzed the complete conversion of 150 and 200 g/l substrate within 6 and 20 h, respectively, to yield enantiopure (S)-CFPL with an isolated yield of 95 %.

Objective: To evaluate the feasibility of quantitative contrast-enhanced ultrasonography (CEUS) for predicting and assessing cervical tumor response to neoadjuvant chemotherapy (NACT)., Methods: Thirty-eight cases with stage IB2 or IIA cervical cancer were studied using CEUS before and after one cycle of NACT. The quantitative CEUS parameters maximum intensity (IMAX), rise time (RT), time to peak (TTP), and mean transit time (MTT) were compared between cervical tumors and myometrium (reference zone) using Sonoliver software. Absolute and relative changes in quantitative CEUS parameters were also compared among complete response, partial response, and non-responsive groups. Correlations between pre-treatment IMAX and changes in quantitative parameters were assessed after one cycle of NACT., Results: There were significant changes in cervical tumor IMAX (P<0.001), RT (P<0.05), and TTP (P<0.05) after one cycle of NACT. According to the Response Evaluation Criteria In Solid Tumors guidelines, the enrollments were divided into complete response, partial response, stable disease and progressive disease groups. There were no significant differences in quantitative CEUS parameters among complete response, partial response, and non-responsive groups (P>0.05). In the stable disease group (n=17), cervical tumor IMAX, RT, and TTP decreased significantly after NACT (P<0.001). The absolute and percentage changes in IMAX were positively correlated with pre-treatment IMAX in all 38 patients (r=0.576, P<0.001 and r=0.429, P<0.001)., Conclusion: Quantitative CEUS analysis can reveal changes in tumor perfusion following NACT. Tumor perfusion values changes likely precede size changes during the NACT course, and pre-treatment IMAX may be a valuable predictor of cervical tumor perfusion response to NACT with a great decrease in IMAX correlated with better perfusion response., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

Objective: This study aimed to investigate the use of contrast-enhanced ultrasonography (CEUS) and time-intensity curves to assess angiogenesis in cervical cancer., Methods: 60 patients who were scheduled to undergo radical surgery for biopsy-proven cervical cancers underwent CEUS. Surgical tissue sections from 32 patients who did not receive neoadjuvant chemotherapy were analyzed with CD34 staining to estimate intratumoral microvessel density (MVD). CEUS images were analyzed for maximum intensity (IMAX), rise time (RT), time to peak (TTP) and mean transit time., Results: Cervical lesions had a higher IMAX and shorter RT and TTP (p < 0.001) than reference regions. There was a linear association between the IMAX of the cervical lesion and the mean intratumoral MVD (r = 0.624, p < 0.001). There were no significant differences in CEUS variables according to histological type, grade and stage., Conclusion: Quantitative CEUS variables have potential use for monitoring perfusion changes in tumours after non-surgical therapy for advanced cervical cancer., Advances in Knowledge: The article demonstrates the capability and value of quantitative CEUS as a non-invasive strategy for detecting the perfusion and angiogenic status of cervical cancer. Quantitative CEUS variables have potential use for monitoring tumour response to non-surgical therapy.

Trimethylation of lysine 27 on histone H3 (H3K27ME3) is a transcription-suppressive histone mark mediated by enhancer of zeste homolog 2 (EZH2). We have previously suggested that EZH2-mediated H3K27ME3 plays a critical oncogenic role in human hepatocellular carcinoma (HCC) aggressiveness. However, the direct downstream targets of EZH2-H3K27ME3 and the molecular mechanisms by which regulates HCC pathogenesis remain unclear. In this study, we used chromatin immunoprecipitation together with high-throughput sequencing (ChIP-seq) and gene expression profiling by microarray analysis to assess genome-wide chromatin occupancy of H3K27ME3 in HCC cells. We identified that claudin14 (CLDN14) is a potentially direct target for EZH2-mediated H3K27ME3 in HCC. In a large cohort of clinical HCC tissues, we found that low expression of CLDN14 was significantly associated with advanced tumor stage and determined to be an independent predictor of shortened survival of HCC patients. Next, functional experiment demonstrated that depletion of CLDN14 substantially restored EZH2-silenced HCC cells motility and invasive capacities and supported cell epithelial-mesenchymal transition (EMT). Furthermore, downregulation of CLDN14 dramatically re-enhanced the wnt/β-catenin signaling activity in EZH2-silenced HCC cells by increasing the levels of active β-catenin and promoting the nuclear localization of β-catenin. These results, collectively, uncover that CLDN14 is a novel direct target of EZH2-mediated H3K27ME3, and provide an explanation for the aggressive nature of HCC with downregulation of CLDN14 and the underling mechanism that links the tumor suppressor CLDN14 to the wnt/β-catenin signaling pathway., (© The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Ethyl (S)-4-chloro-3-hydroxybutanoate ((S)-CHBE) is an important chiral intermediate for the synthesis of "blockbuster" drug statins. The carbonyl reductase ChKRED20 from Chryseobacterium sp. CA49 was found to catalyze the bio-reductive production of (S)-CHBE with excellent stereoselectivity (>99.5 % ee). Perceiving a capacity for improvement, we sought to increase the thermostability of ChKRED20 to allow a higher reaction temperature. After one round of error-prone PCR (epPCR) library screening followed by the combination of beneficial mutations, a triple-mutant MC135 was successfully achieved with substantially enhanced thermostablity. The activity of MC135 at 50 °C was similar to the wild type. However, at its temperature optima of 65 °C, the mutant displayed 63 % increase of activity compared to the wild type and remained >95 % activity after being incubated for 15 days, while the wild type had a half-life of 11.9 min at 65 °C. At a substrate/catalyst ratio of 100 (w/w), the mutant catalyzed the complete conversion of 300 g/l substrate within 1 h to yield enantiopure (S)-CHBE with an isolated yield of 95 %, corresponding to a space-time yield of 1824 mM/h.

The aim of this study was to develop a prognostic classifier and subdivided the M1 stage for nasopharyngeal carcinoma patients with synchronous metastases (mNPC). A retrospective cohort of 347 mNPC patients was recruited between January 2000 and December 2010. Thirty hematological markers and 11 clinical characteristics were collected, and the association of these factors with overall survival (OS) was evaluated. Advanced machine learning schemes of a support vector machine (SVM) were used to select a subset of highly informative factors and to construct a prognostic model (mNPC-SVM). The mNPC-SVM classifier identified ten informative variables, including three clinical indexes and seven hematological markers. The median survival time for low-risk patients (M1a) as identified by the mNPC-SVM classifier was 38.0 months, and survival time was dramatically reduced to 13.8 months for high-risk patients (M1b) (P < 0.001). Multivariate adjustment using prognostic factors revealed that the mNPC-SVM classifier remained a powerful predictor of OS (M1a vs. M1b, hazard ratio, 3.45; 95% CI, 2.59 to 4.60, P < 0.001). Moreover, combination treatment of systemic chemotherapy and loco-regional radiotherapy was associated with significantly better survival outcomes than chemotherapy alone (the 5-year OS, 47.0% vs. 10.0%, P < 0.001) in the M1a subgroup but not in the M1b subgroup (12.0% vs. 3.0%, P = 0.101). These findings were validated by a separate cohort. In conclusion, the newly developed mNPC-SVM classifier led to more precise risk definitions that offer a promising subdivision of the M1 stage and individualized selection for future therapeutic regimens in mNPC patients.

The microRNA, miR-200c, is involved in the tumorigenesis and progression of a variety of cancers. The purpose of this study was to investigate the expression, mechanism and prognostic roles of miR-200c in breast cancer. We found that miR-200c was downregulated in both breast cancer tissue and cell lines using quantitative real-time PCR (qRT-PCR). In situ hybridization (ISH) and microarrays showed that low miR-200c expression was associated with poor patient overall survival (OS) and disease free survival (DFS). We used luciferase reporter plasmids to find that miR-200c inhibited the AKT and ERK pathways by directly targeting KRAS. Repression of KRAS by miR-200c suppressed the proliferation and survival of breast cancer cells in vitro and in vivo. miR-200c also had an anti-tumor effect by negatively regulating KRAS in a xenograft mouse model. Our findings provide clues regarding the role of miR-200c as a tumor suppressor in breast cancer through the inhibition of KRAS translation both in vitro and in vivo. miR-200c could be a potential therapeutic target in breast cancer.

Nasopharyngeal carcinoma (NPC) has the highest metastasis rate among head and neck cancers with unclear mechanism. WNT5A belongs to the WNT family of cysteine-rich secreted glycoproteins. Our previous high-throughput gene expression profiling revealed that WNT5A was up-regulated in highly metastatic cells. In the present study, we first confirmed the elevated expression of WNT5A in metastatic NPC tissues at both the mRNA and protein levels. We then found that WNT5A promoted epithelial-mesenchymal transition (EMT) in NPC cells, induced the accumulation of CD24-CD44+ cells and side population, which are believed to be cancer stem cell characteristics. Moreover, WNT5A promoted the migration and invasion of NPC cells in vitro, while in vivo treatment with recombinant WNT5A promoted lung metastasis. Knocking down WNT5A diminished NPC tumorigenesis in vivo. When elevated expression of WNT5A coincided with the elevated expression of vimentin in the primary NPC, the patients had a poorer prognosis. Among major signaling pathways, protein kinase C (PKC) signaling was activated by WNT5A in NPC cells. A positive feedback loop between WNT5A and phospho-PKC to promote EMT was also revealed. Taken together, these data suggest that WNT5A is an important molecule in promoting stem cell characteristics in NPC, leading to tumorigenesis and metastasis.

A putative enoate reductase, Achr-OYE4, was mined from the genome of Achromobacter sp. JA81, expressed in Escherichia coli, and was characterized. Sequence analysis and spectral properties indicated that Achr-OYE4 is a typical flavin mononucleotide-dependent protein; it preferred NADH over NADPH as a cofactor. The heterologously expressed protein displayed good activity and excellent stereoselectivity toward some activated alkenes in the presence of NADH, NADPH, or their recycling systems. The glucose dehydrogenase-based recycling system yielded the best results in most cases, with a product yield of up to 99 % and enantiopurity of >99 % ee. Achr-OYE4 is an important addition to the asymmetric reduction reservoir as an "old yellow enzyme" from Achromobacter.

Objective: To explore the value of transrectal ultrasonography (TRUS) for tumor node metastasis (TNM) restaging for patients with locally advanced rectal cancer after neoadjuvant chemoradiotherapy (neo-CRT)., Methods: One hundred and forty-nine patients with locally advanced rectal cancer (cT3-4 or cN+) who underwent TRUS after neo-CRT were retrospectively reviewed. TRUS restaging was compared with the results of post-operative pathological TNM findings., Results: After neo-CRT, the accuracy of TRUS for diagnosing T-staging was 30.9%, with 60.4% (90/149) of cases overestimated. The sensitivity of TRUS for T-staging (T0 vs T1 vs T2 vs T3 vs T4) were 16.3%, 0%, 12.5%, 42.6% and 75.0%, respectively. The accuracy of TRUS for diagnosing N-staging after neo-CRT was 81.2%, with the sensitivities of N0 and N+ were 93.3% and 31.0%, respectively. After neo-CRT, 27.5% (41/149) of patients achieved pathologically complete response (pCR). The sensitivity, specificity, positive predictive value and negative predictive values of TRUS for pCR were 17.1%, 99.1%, 87.5% and 75.9%, respectively., Conclusions: TRUS can be applied for restaging T4 and N0, and has potential for screening out patients with pCR in those with locally advanced rectal cancer after neo-CRT, although some stages are overestimated for T-staging and its sensitivity for predicting pCR is low.

Objective: To explore the potential of quantitative analysis of contrast-enhanced ultrasonography (CEUS) in differentiating focal nodular hyperplasia (FNH) from hepatocellular carcinoma (HCC)., Methods: 34 cases of FNH and 66 cases of HCC (all lesions <5 cm) were studied using CEUS to evaluate enhancement patterns and using analytic software Sonoliver® (Image-Arena™ v.4.0, TomTec Imaging Systems, Munich, Germany) to obtain quantitative features of CEUS in the region of interest. The quantitative features of maximum of intensity (IMAX), rise slope (RS), rise time (RT) and time to peak (TTP) were compared between the two groups and applied to further characterise both FNH and HCC with hypoenhancing patterns in the late phase on CEUS., Results: The sensitivity and specificity of CEUS for diagnosis of FNH were 67.6% and 93.9%, respectively. For quantitative analysis, IMAX and RS in FNHs were significantly higher than those in HCCs (p<0.05), while RT and TTP in FNHs were significantly shorter (p<0.05). Both the 11 FNHs and 62 HCCs with hypo-enhancing patterns in the late phase were further characterised with their quantitative features, and the sensitivity and specificity of IMAX for diagnosis of FNH were 90.9% and 43.5%, RS 81.8% and 80.6%, RT 90.9% and 71.0%, and TTP 90.9% and 71.0%, respectively., Conclusion: The quantitative features of CEUS in FNH and HCC were significantly different, and they could further differentiate FNH from HCC following conventional CEUS., Advances in Knowledge: Our findings suggest that quantitative analysis of CEUS can improve the accuracy of differentiating FNH from HCC.

Three design strategies, based on rational and semi-rational approaches, were employed to investigate the functional impact of thermostability-related amino acid substitutions in the β-glycosidase BglY from Thermus thermophilus. Five beneficial mutations were identified, of which 1 mutation was located in the active cavity of the enzyme and contributed to the released substrate inhibition. Combining all 5 beneficial substitutions resulted in the mutant HF5 with a 4.7-fold increase in half-life, with thermal inactivation at 93 °C, and complete lack of substrate inhibition toward the substrate p-nitrophenyl-β-D-glucopyranoside at lower reaction temperatures. The results of this study provide valuable information on amino acid substitutions related to thermostability and substrate inhibition of BglY., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Objective: The quantitative parameters in the contrast-enhanced ultrasonography time-intensity curve of hepatocellular carcinoma (HCC) were studied to explore their possible implication for histological grading of HCC., Methods: A total of 130 HCC patients (115 males and 15 females; age: 48.13±11.00 years) were studied using contrast-enhanced ultrasonography time-intensity curve and histological pathology. The quantification software Sonoliver® (TomTec Imaging Systems, Unterschleissheim, Germany) was applied to derive time-intensity curves of regions of interest in the interior of HCCs and in reference. Quantitative parameters of 115 patients were successfully obtained, including maximum of intensity (IMAX), rise time (RT), time to peak (TTP), rise slope (RS) and washout time (WT). Histological grading of HCC was performed using haematoxylin-eosin staining, and monoclonal antibodies specific for smooth muscle actin were used to observe unpaired arteries (UAs)., Results: There were significant differences among WTs in the three differentiated HCC groups (p<0.05). However, there were no significant differences among RT, TTP, RS and IMAX in the differentiated HCC groups. Moreover, the number of UAs in the differentiated HCC groups showed no statistical significance., Conclusion: WT plays an important role in predicting well, moderately and poorly differentiated HCC.

Feruloyl esterase A from Aspergillus niger (AnFaeA) is one of the most important feruloyl esterases of industrial relevance. Previous work aided by the PoPMuSiC algorithm has identified two beneficial mutants (D93G and S187F) with thermostabilization effect. In this work, twelve additional amino acid substitutions were identified to be beneficial to the thermostability of AnFaeA after screening a random mutagenesis library constructed in Pichia pastoris. Combination of these mutations resulted in a mutant with 80% residual activity after heat treatment at 90 °C for 15 min and a half-life increasing from 15 min to >4000 min at 55 °C. The thermostabilized mutant displayed significantly enhanced performance compared to the parental AnFaeA when applied to the treatment of steam-exploded corn stalk at 60 °C together with an xylanase, demonstrating its great potential for industrial application., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

The strain Achromobacter sp. JA81, which produced enoate reductase, was applied in the asymmetric reduction of activated alkenes. The strain could catalyze the bioreduction of alkenes to form enantiopure (R)-β-aryl-β-cyano-propanoic acids, a precursor of (R)-γ-amino butyric acids, including the pharmaceutically active enantiomer of the chiral drug (R)-baclofen with excellent enantioselectivity. It could catalyze as well the stereoselective bioreduction of other activated alkenes such as cyclic imides, β-nitro acrylates, and nitro-alkenes with up to >99 % ee and >99 % conversion. The draft genome sequencing of JA81 revealed six putative old yellow enzyme homologies, and the transcription of one of them, Achr-OYE3, was detected using reverse transcription polymerase chain reaction. The recombinant Escherichia coli expressing Achr-OYE3 displayed enoate reductase activity toward (Z)-3-cyano-3-phenyl-propenoic acid (2a).

Objective: The quantitative parameters in contrast-enhanced ultrasonography-time-intensity curve of hepatocellular carcinoma (HCC) were studied to explore their potential importance in monitoring the effects of anti-angiogenic therapy for HCC., Methods: 115 HCC patients were studied with contrast-enhanced ultrasonography-time-intensity curve (CEUS-TIC) and with immunohistochemical analysis of tissue sections. The CEUS images were analyzed off-line to obtained quantitative parameters including maximum of intensity (IMAX), rise time (RT), time to peak (TTP), mean transit time (mTT), rise slope (RS), and washout time (WT). Monoclonal antibodies specific for smooth muscle actin and anti-CD34 were used to observe unpaired arteries (UAs) and microvessel area (MVA) of sinusoidal capillarization, respectively. The UAs and MVA of 82 HCC cases were successfully stained., Results: The number of UAs had moderate correlation with RT (r=-0.446), TTP (r=-0.432), and RS (r=0.431) (P<0.05), and it had mild correlation with IMAX (r=0.303) and WT (r=0.285) (P<0.05). MVA of sinusoidal capillarization had no correlation with perfusion parameters., Conclusion: Quantitative CEUS-TIC parameters reflecting hemodynamics of tumors are correlated with UAs, but not with MVA, and they might be used to monitor the effects of anti-angiogenic therapy on HCC., (Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.)

Background: ZEB2 has been suggested to mediate EMT and disease aggressiveness in several types of human cancers. However, the expression patterns of ZEB2 in hepatocellular carcinoma (HCC) and its effect on prognosis of HCC patients treated with hepatectomy are unclear., Methodology/principal Findings: In this study, the methods of tissue microarray and immunohistochemistry (IHC) were utilized to investigate ZEB2 expression in HCC and peritumoral liver tissue (PLT). Receiver operating characteristic (ROC), spearman's rank correlation, Kaplan-Meier plots and Cox proportional hazards regression model were used to analyze the data. Up-regulated expression of cytoplasmic/nuclear ZEB2 protein was observed in the majority of PLTs, when compared to HCCs. Further analysis showed that overexpression of cytoplasmic ZEB2 in HCCs was inversely correlated with AFP level, tumor size and differentiation (P<0.05). Also, overexpression of cytoplasmic ZEB2 in PLTs correlated with lower AFP level (P<0.05). In univariate survival analysis, a significant association between overexpression of cytoplasmic ZEB2 by HCCs/PLTs and longer patients' survival was found (P<0.05). Importantly, cytoplasmic ZEB2 expression in PLTs was evaluated as an independent prognostic factor in multivariate analysis (P<0.05). Consequently, a new clinicopathologic prognostic model with cytoplasmic ZEB2 expression (including HCCs and PLTs) was constructed. The model could significantly stratify risk (low, intermediate and high) for overall survival (P = 0.002)., Conclusions/significance: Our findings provide a basis for the concept that cytoplasmic ZEB2 expressed by PLTs can predict the postoperative survival of patients with HCC. The combined cytoplasmic ZEB2 prognostic model may become a useful tool for identifying patients with different clinical outcomes.

Background/aims: A high prevalence of serum IL-6 has been associated with the pathogenesis of hepatocellular carcinoma (HCC) in both animals and humans. However, it is not clear how the levels of serum IL-6 influence the prognosis of HCC patients. This study was carried out in order to attempt to answer this question., Methodology: A total of 156 adults were selected and categorized into four groups: healthy subjects (n=18), those with tumor recurrence (n=26), those initially diagnosed with HCC (n=32), and those with HCC (n=80) who received curative resection between 2002 and 2004 with five years of follow-up. Serum IL-6 levels were determined in all subjects by the same ELISA method., Results: IL-6 was found in high levels in the serum of patients initially diagnosed with HCC (8.47±5.92, p<0.0001) and in patients with HCC and tumor recurrence (12±31.90, p=0.001) compared with healthy subjects (0.89±1.51). This includes all patients who received therapy between 2007 and 2008. The levels of serum IL-6 were positively correlated with tumor size (p=0.002) in the HCC patients who received curative resection between 2002 and 2004 with five years of follow-up., Conclusions: High levels of serum IL-6 correlated positively with tumor size and with poor prognosis in HCC patients.

Thermostability of β-glucosidase was enhanced by family shuffling, site saturation mutagenesis, and site-directed mutagenesis. Family shuffling was carried out based on β-glucosidase BglC from Thermobifida fusca and β-glucosidase BglB from Paebibacillus polymxyxa with the help of synthetic primers. High-throughput screening revealed mutants with higher thermostability than both parental enzymes. The most thermostable mutant VM2 containing three key amino acid changes in L444Y/G447S/A433V had a 144-fold increase in half-life of inactivation as compared to the parental enzyme BglC. The mutant VM2 showed 28% and 94% increase in k(cat) towards p-nitrophenyl-β-D-glucopyranoside (pNPG) and cellobiose, respectively. The mutant with enhanced stability would facilitate the recycle of β-glucosidase in the bioconversion of cellulosic biomass., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

A saturation mutagenesis library was constructed at the position 329 of the endoglucanase CelA from Clostridium thermocellum based on previous results (Yi and Wu, 2010), and one mutation, S329G, was identified to contribute to the enhanced thermostability. The result inspired a rational design approach focusing on the introduction of Gly or Pro residue onto the protein surface, which led to the identification of two additional beneficial mutations, H194G and S269P. Combination of these three mutations resulted in a mutant with a 10-fold increase in half-life of inactivation (60 min) at 86°C without compromising activity compared with the wild-type. Its reaction temperature for maximum activity increased from 75 to 85°C. The results provide valuable thermostability-related structural information on this thermophilic enzyme., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

It has been suggested that trimethylation of lysine 27 on histone H3 (H3K27me3) is a crucial epigenetic process in tumorigenesis. However, the expression dynamics of H3K27me3 and its clinicopathological/prognostic significance in hepatocellular carcinoma (HCC) are unclear. In this study, immunohistochemical analysis (IHC) was used to examine protein expression of H3K27me3 in HCC tissues from two independent cohorts and corresponding nontumorous hepatocellular tissues by tissue microarray. The optimal cutpoint of H3K27me3 expression was assessed by the X-tile program. Our results showed that the cutpoint for high expression of H3K27me3 in HCCs was determined when more than 70% of the tumor cells showed positive staining. High expression of H3K27me3 was observed in 134 of 212 (63.2%) and 76 of 126 (60.4%) of HCCs in the testing and validation cohorts, respectively. Correlation analysis demonstrated that high expression of H3K27me3 in HCCs was significantly correlated with large tumor size, multiplicity, poor differentiation, advanced clinical stage and vascular invasion (P < 0.05). In addition, high expression of H3K27me3 in HCC patients was associated closely with shortened survival time, independent of serum α-fetoprotein levels, tumor size and multiplicity, clinical stage, vascular invasion and relapse as evidenced by univariate and multivariate analysis in both cohorts (P < 0.05). In different subsets of HCC patients, H3K27me3 expression was also a prognostic indicator in patients with stage II tumors (P < 0.05). Thus, these findings provide evidence that a high expression of H3K27me3, as detected by IHC, correlates closely with vascular invasion of HCCs and is an independent molecular marker for poor prognosis in patients with HCC.

Protein engineering of the thermostable xylanase XT6 from Geobacillus stearothermophilus was performed to obtain enzymes with improved thermal tolerance. Mutants producing such enzymes were obtained after several rounds of directed evolution using error-prone PCR and sequence family shuffling, in combination with a consensus-based semi-rational approach. The most thermostable mutant enzyme contained 13 amino acid substitutions and its half-life of inactivation was 52-fold of that of the wild-type. Its reaction temperature for maximum activity increased from 77 degrees C to 87 degrees C, and catalytic efficiency (k(cat)/K(m)) increased by 90%. The mutant is of potential interest for industrial applications., (2010 Elsevier Ltd. All rights reserved.)

To elucidate the effects of codon optimization and chaperone coexpression on the heterologous expression of mammalian cytochrome P450s (P450) in Escherichia coli, the expression of P450s 2B1, 2S1, 2U1, 2W1, and 27C1 were investigated. With codon optimization for N-terminus or the entire gene, the expression levels of P450 27C1, 2U1 and 2W1 increased 22-fold, 3.6-fold and 2.1-fold, respectively, while those for P450s 2B1 and 2S1 remained unchanged. With coexpression of E. coli molecular chaperones GroEL/ES, the expression level increased up to 14-fold for P450 27C1, and 3- to 5-fold for P450s 2B1, 2S1, and 2W1. Simultaneous application of these two techniques resulted in synergetic effects.

This study was to assess the changes in the haemodynamic parameters of the hepatic artery and vein in the diagnosis of liver metastases by contrast-enhanced ultrasound (CEUS). 52 patients with proven liver metastases (patient group) and 23 normal volunteers (control group) were recruited in this study. Each subject was administered an intravenous bolus injection of SonoVue (0.6 ml). The arrival time in the hepatic artery (AT HA), time to peak in the hepatic artery (TTP HA), peak intensity of the hepatic artery (PI HA), arrival time in the hepatic vein (AT HV), time to peak in the hepatic vein (TTP HV) and peak intensity of the hepatic vein (PI HV) were measured with the use of time-intensity curve software. The hepatic artery to vein transit time (HAVTT) was calculated as the difference between the arrival times in the hepatic artery and the hepatic vein. AT HA, TTP HA, AT HV, TTP HV and HAVTT in the patient group were significantly shorter than those of the control group (P<0.01). PI HA and PI HV in the patient group were significantly higher than those of the control group (P<0.01). These results suggest that CEUS assessment of changes in the haemodynamic parameters of the hepatic artery and vein help to diagnose liver metastases. This functional imaging technique may contribute to the early detection of micrometastases in the liver.