Adoptive Transfer of NKG2D CAR mRNA-Engineered Natural Killer Cells in Colorectal Cancer Patients.

By fusing the extracellular domain of the natural killer (NK) cell receptor NKG2D to DAP12, we constructed a chimeric antigen receptor (CAR) to improve NK cell tumor responses. An RNA electroporation approach that provides transient expression of the CAR was adopted as a risk mitigation strategy. Expression of the NKG2D RNA CAR significantly augmented the cytolytic activity of NK cells against several solid tumor cell lines in vitro and provided a clear therapeutic benefit to mice with established solid tumors. Three patients with metastatic colorectal cancer were then treated with local infusion of the CAR-NK cells. Reduction of ascites generation and a marked decrease in number of tumor cells in ascites samples were observed in the first two patients treated with intraperitoneal infusion of low doses of the CAR-NK cells. The third patient with metastatic tumor sites in the liver was treated with ultrasound-guided percutaneous injection, followed by intraperitoneal infusion of the CAR-NK cells. Rapid tumor regression in the liver region was observed with Doppler ultrasound imaging and complete metabolic response in the treated liver lesions was confirmed by positron emission tomography (PET)- computed tomographic (CT) scanning. Our results highlight a promising therapeutic potential of using RNA CAR-modified NK cells to treat metastatic colorectal cancer.

Renal cell carcinoma: predicting RUNX3 methylation level and its consequences on survival with CT features.

PURPOSE: To investigate associations between CT imaging features, RUNX3 methylation level, and survival in clear cell renal cell carcinoma (ccRCC). MATERIALS AND METHODS: Patients were divided into high RUNX3 methylation and low RUNX3 methylation groups according to RUNX3 methylation levels (the threshold was identified by using X-tile). The CT scanning data from 106 ccRCC patients were retrospectively analyzed. The relationship between RUNX3 methylation level and overall survivals was evaluated using the Kaplan-Meyer analysis and Cox regression analysis (univariate and multivariate). The relationship between RUNX3 methylation level and CT features was evaluated using chi-square test and logistic regression analysis (univariate and multivariate). RESULTS: ß value cutoff of 0.53 to distinguish high methylation (N = 44) from low methylation tumors (N = 62). Patients with lower levels of methylation had longer median overall survival (49.3 vs. 28.4) months (low vs. high, adjusted hazard ratio [HR] 4.933, 95% CI 2.054-11.852, p < 0.001). On univariate logistic regression analysis, four risk factors (margin, side, long diameter, and intratumoral vascularity) were associated with RUNX3 methylation level (all p < 0.05). Multivariate logistic regression analysis found that three risk factors (side: left vs. right, odds ratio [OR] 2.696; p = 0.024; 95% CI 1.138-6.386; margin: ill-defined vs. well-defined, OR 2.685; p = 0.038; 95% CI 1.057-6.820; and intratumoral vascularity: yes vs. no, OR 3.286; p = 0.008; 95% CI 1.367-7.898) were significant independent predictors of high methylation tumors. This model had an area under the receiver operating characteristic curve (AUC) of 0.725 (95% CI 0.623-0.827). CONCLUSIONS: Higher levels of RUNX3 methylation are associated with shorter survival in ccRCC patients. And presence of intratumoral vascularity, ill-defined margin, and left side tumor were significant independent predictors of high methylation level of RUNX3 gene. KEY POINTS: • RUNX3 methylation level is negatively associated with overall survival in ccRCC patients. • Presence of intratumoral vascularity, ill-defined margin, and left side tumor were significant independent predictors of high methylation level of RUNX3 gene.

Presence of Intratumoral Calcifications and Vasculature Is Associated With Poor Overall Survival in Clear Cell Renal Cell Carcinoma.

PURPOSE: The objective of this study was to explore the prognostic significance of the preoperative computed tomography (CT) features in clear cell renal cell carcinoma. PATIENTS AND METHODS: The clinical data and CT data from 210 patients (1 grade 1, 84 grade 2, 92 grade 3, and 32 grade 4) generated with The Cancer Imaging Archive were reviewed. Overall survival was assessed using Kaplan-Meyer analysis. The relationship between CT features and survivals were evaluated using univariate and multivariable Cox regression analysis. RESULTS: The follow-up occurred between 13 and 3989 days (median, 1405 days; mean, 1434 days).On univariate Cox regressions, 4 preoperative CT features (intratumoral calcifications: yes vs no hazard ratio [HR], 2.054; 95% confidence interval [CI], 1.231-3.428; renal vein invasion: yes vs no HR, 2.013; 95% CI, 1.218-3.328; collecting system invasion: yes vs no HR, 2.139; 95% CI, 1.286-3.558; gross appearance of intratumoral vasculature: yes vs no HR, 2.385; 95% CI, 1.454-3.915) were significantly associated with overall survival (all P < 0.05). On multivariable Cox regression analysis, predictors of mortality in clear cell renal cell carcinoma were the presence of intratumoral calcifications (HR, 1.718; 95% CI, 1.014-2.911; P = 0.044) and gross appearance of intratumoral vasculature (HR, 2.137; 95% CI, 1.284-3.557; P = 0.003). CONCLUSIONS: Presence of intratumoral calcifications and vasculature can be potential prognostic features to screen patients for unfavorable prognosis.

Long noncoding RNA STXBP5-AS1 inhibits cell proliferation, migration, and invasion through inhibiting the PI3K/AKT signaling pathway in gastric cancer cells.

INTRODUCTION: Poor prognosis of gastric cancer (GC) has partly been a result of late diagnosis due to nonspecific symptoms in the early stages. The overall survival rate of patients with GC is quite low. Here, we presented the functional role and potential mechanism of long noncoding RNA STXBP5-AS1 in GC. MATERIALS AND METHODS: CCK-8, scratch wound healing and Transwell assays were conducted to analyze proliferation, migration, and invasion of SGC7901 and MKN45 cells. Real-time polymerase chain reaction (qPCR) and Western blot assays were performed to investigate the relationship between STXBP5-AS1 and STXBP5. Finally, the correlation between STXBP5-AS1 and phosphorylated AKT1 (p-AKT1) was explored to reveal the potential mechanism of STXBP5-AS1 in GC. Western blot assays were performed to analyze phosphorylated AKT1 (p-AKT1) and AKT levels. RESULTS: Our results suggested that STXBP5-AS1 suppressed proliferation, migration, and invasion, and the upregulation of STXBP5-AS1 significantly repressed STXBP5 expression, and knockdown of STXBP5-AS1 promoted STXBP5 expression. In addition, the p-AKT1 level decreased when STXBP5-AS1 was overexpressed and the p-AKT1 level increased with STXBP5-AS1 knockdown in SGC7901 and MKN45 cells. CONCLUSION: In summary, our results indicate that STXBP5-AS1 inhibits cell proliferation, migration, and invasion through PI3K/AKT in GC.

BI-RADS 3-5 microcalcifications: prediction of lymph node metastasis of breast cancer.

PURPOSE: To determine whether the clinicopathological parameters and Breast Imaging Reporting and Data System (BI-RADS) 3-5 microcalcifications differed between lymph node positive (LN (+)) and lymph node negative (LN (-)) invasive ductal carcinoma (IDC). RESULTS: For microcalcification-associated breast cancers, seven selected features (age, tumor size, Ki-67 status, lymphovascular invasion, calcification range, calcification diameter and calcification density) were significantly associated with LN status (all P < 0.05). Multivariate logistic regression analysis found that three risk factors (age: older vs. younger OR: 0.973 P = 0.006, tumor size: larger vs. smaller OR: 1.671, P < 0.001 and calcification density: calcifications > 20/cm2 vs. calcifications ≤ 20/cm2 OR: 1.698, P < 0.001) were significant independent predictors. This model had an area under the receiver operating characteristic curve (AUC) of 0.701. The nodal staging (N0 and N1 χ2 = 5.701, P = 0.017; N0 and N2 χ2 = 6.614, P = 0.013) was significantly positively associated with calcification density. The luminal B subtype had the highest risk of LN metastasis. Multivariate analysis demonstrated that calcification > 2 cm in range (OR: 2.209) and larger tumor size (OR: 1.882) were independently predictive of LN metastasis in the luminal B subtype (AUC = 0.667). MATERIALS AND METHODS: Mammographic images of 419 female breast cancer patients were included. Associations between the risk factors and LN status were evaluated using a Chi-square test, ANOVA and binary logistic regression analysis. CONCLUSIONS: This study found that age, tumor size and calcifications density can be conveniently used to facilitate the preoperative prediction of LN metastasis. The luminal B subtype has the highest risk of LN metastasis among the microcalcification-associated breast cancers.

BI-RADS 3-5 microcalcifications can preoperatively predict breast cancer HER2 and Luminal a molecular subtype.

PURPOSE: To investigate associations between breast cancer molecular subtype and the patterns of mammographically detected calcifications. RESULTS: Identified were 93 (19.1%) Luminal A, 242 (49.9%) Luminal B, 108 (22.2%) HER2 and 42 (8.7%) basal subtypes. In univariate analysis, the clinicopathological parameters and BI-RADS 3-5 microcalcifications, which consisted 9 selected features was significantly associated with breast cancer molecular subtype (all P < 0.05). Among subtypes, multivariate analysis showed that calcification >2 cm in range (OR: 1.878, 95% CI: 1.150 to 3.067) and calcification > 0.5 mm in diameter (OR:2.206, 95% CI: 1.235 to 3.323) was independently predictive of HER2 subtype. The model showed good discrimination for predicting HER2 subtype, with a C-index of 0.704. In addition, multivariate analysis showed that calcification morphology (amorphour or coarse heterogenous calcifications OR: 2.847, 95% CI: 1.526 to 5.312) was independently predictive of Luminal A subtype. The model showed good discrimination for predicting Luminal A subtype, with a C-index of 0.74. And we demonstrated that amorphour or coarse heterogenous calcifications were associated with a higher incidence of Luminal A subtype than pleomorphic or fine linear or branching calcifications. There was no significant difference between breast cancer subtypes (Luminal B vs. other; Basal vs. other) and the patterns of mammographically detected calcifications. MATERIALS AND METHODS: Mammographic images of 485 female patients were included. The correlation between mammographic imaging features and breast cancer subtype was analyzed using Chi-square test, univariate and binary logistic regression analysis. CONCLUSIONS: This study shows that BI-RADS 3-5 microcalcifications can be conveniently used to facilitate the preoperative prediction of HER2 and Luminal A molecular subtype in patients with infiltrating ductal carcinoma.

Reduced Expression of SARM in Mouse Spleen during Polymicrobial Sepsis.

Objective Immune dysfunction, including prominent apoptosis of immune cells and decreased functioning of the remaining immune cells, plays a central role in the pathogenesis of sepsis. Sterile α and HEAT/armadillo motif-containing protein (SARM) is implicated in the regulation of immune cell apoptosis. This study aimed to elucidate SARM contributes to sepsis-induced immune cell death and immunosuppression. Methods A mouse model of polymicrobial sepsis was generated by cecum ligation and puncture (CLP). SARM gene and protein expression, caspase 3 cleavage and intracellular ATP production were measured in the mouse spleens. Results CLP-induced polymicrobial sepsis specifically attenuated both the gene and protein expression of SARM in the spleens. Moreover, the attenuation of SARM expression synchronized with splenocyte apoptosis, as evidenced by increased caspase 3 cleavage and ATP depletion. Conclusions These findings suggest that SARM is a potential regulator of sepsis-induced splenocyte apoptosis.

Differential diagnosis between malignant and benign breast lesions using single-voxel proton MRS: a meta-analysis.

OBJECTIVES: We aim to investigate the diagnostic capability of single-voxel proton MR spectroscopy (MRS) for benign/malignant discrimination of focal breast lesions with a meta-analysis. MATERIALS AND METHODS: The meta-analysis included a total of 750 malignant breast lesions and 419 benign breast lesions from eighteen studies. RESULTS: The pooled sensitivity and specificity of MRS were 0.71 (95 % CI 0.68-0.74) and 0.85 (95 % CI 0.81-0.88), respectively. The positive likelihood ratio and negative LR were 4.11 (95 % CI 3.11-5.43) and 0.25 (95 % CI 0.17-0.36), respectively. The P value for χ(2) heterogeneity for all pooled estimates was <0.05. From the fitted summary receiver operating characteristics curve, AUC was 0.89 and Q* was 0.84. Asymmetrical in funnel plots indicated there may be publication bias (t = 2.85, P = 0.012). The meta-regression analysis indicated that neither threshold effect nor evaluated covariates that include strength of field, scanning technique (PRESS or STEAM), repetition time, NSA, and pre- or post-contrast agent were the sources of heterogeneity (all P value >0.05). CONCLUSIONS: Single-voxel proton MRS was useful for differentiation between malignant and benign breast lesions. However, pooled diagnostic measures might be overestimated. The standardization of the acquisition protocol for MRS across the multicenter trials is recommended.

Detection and analysis of 12 heavy metals in blood and hair sample from a general population of Pearl River Delta area.

To detect the content of 12 heavy metals in blood and hair sample from a general population of Pearl River Delta area, and to analyze the influence of duration of residence, gender, age, smoking and drinking on the heavy metal content. Use inductively coupled plasma mass spectrometry to detect the content of 12 heavy metals lead (Pb), mercury (Hg), cadmium (Cd), aluminum (Al), arsenic (As), copper (Cu), chrome (Cr), manganese (Mn), nickel (Ni), zinc (Zn), tin (Sn) and antimony (Sb) in blood and hair samples of a total of 50 subjects from a general population, collected by multistage stratified cluster random sampling method. The geometric mean of heavy metal content in blood samples of general population (µg/L): blood aluminum 214.00; blood chrome 92.82; blood manganese 21.43; blood nickel 20.59; blood copper 0.67; blood zinc 11.50; blood arsenic 0.55; blood cadmium 2.45; blood tin 0.00; blood antimony 1.92; blood lead 158.84; and blood mercury 1.19. The geometric mean of heavy metal content in hair samples of general population (µg/g): hair aluminum is 84.65; hair chrome 0.00; hair manganese 2.44; hair nickel 0.61; hair copper 28.49; hair zinc 136.65; hair arsenic 0.75; hair cadmium 0.46; hair tin 1.04; hair antimony 0.05; hair lead 8.97; and hair mercury 0.69. Some heavy metals were correlated with duration of residence, gender, age, smoking and drinking. This was the first time that simultaneously detecting heavy metal content in blood and hair was used to analyze the internal heavy metal burden in resident population of Pearl River Delta area. These data can serve as reference for further research.

Enhancement of specific cellular immune response induced by DNA vaccines encoding PML-RARalpha and hIL-2 genes.

A DNA vaccine encoding PML-RAR alpha fusion gene is thought to be a promising approach for acute promyelocytic leukemia patients to enhance immune responses after attaining complete remission. In this study, we sought to enhance cellular immunity by coexpressing human interleukin (hIL)-2 genes. Successfully constructed plasmids PML-RAR alpha-hIL-2-pIRES, PML-RAR alpha-pIRES and hIL-2-pIRES were delivered intramuscularly in BALB/C mice at 14-day intervals for three cycles. The cellular immune responses with respect to the specific cytotoxicity of spleen cells; interferon-gamma secretion in sera, and the T-cell receptor rearrangement excision circles of thymocyte were significantly increased from PML-RARalpha-hIL-2-pIRES immunized mice. Our results indicate that a DNA vaccine with PML fusion gene segment and hIL-2 together might elicit increased cellular immune responses in mice.

[Construction and identification of blood type B antigen mimetic polypeptide-Mip3beta double expression recombinant plasmid].

OBJECTIVE: To construct and identify blood type B antigen mimetic polypeptide-macrophage inflammatory protein 3beta (Mip3beta) double expression recombinant plasmid. METHODS: The positive phage clone P1 was obtained using phage random 12-mer peptide library. Specific primers were designed to amplify the phage DNA of P1 and transmembrane domain and inner segment of PBluscript-Fas gene. The products of the amplification were linked into Mip3betav21 to construct blood type B antigen mimetic polypeptide-Mip3beta double expression recombinant plasmid. The recombinant plasmid was transfected into human melanoma cell line B16 to identify its expression. RESULTS AND CONCLUSION: Blood type B antigen mimetic polypeptide-Mip3beta double expression recombinant plasmid is successfully obtained and expressed in human melanoma cell line B16.

Effects of contrast agent on water suppression and shimming of kidney single-volume proton MR spectroscopy at 3.0T.

RATIONALE AND OBJECTIVES: The aim of this study was to determine whether the administration of gadolinium diethylenetriamine penta-acetic acid (Gd-DTPA) significantly affects shimming and water suppression on kidney magnetic resonance spectroscopic prescanning and whether the impact of shimming and water suppression is changed with time after intravenous administration on a 3.0-T system. METHODS: Forty patients (two patients were excluded from analysis because of motion) were examined before and after the administration of Gd-DTPA (the interval between the right and left kidneys was approximately 40 seconds). Regions of interest were carefully positioned in the region of the corresponding location of both kidneys separately. Line widths (full width at half maximum) and water suppression were obtained. A paired t test for comparison of means was used. In addition, repeat measurements with a shorter time interval (obtained 120-130 seconds after the injection) and a longer time interval (obtained 150-160 seconds after the injection) were performed in five patients in the same regions of interest of the right kidney. Sequential ¹H magnetic resonance spectroscopic prescanning in the same region of interest was performed in one patient. RESULTS: The left kidney had slightly better shimming and water suppression effects than the right kidney after contrast agent administration (all P values < .01). The limiting resolution of both shimming and water suppression effects was decreased on enhanced images in both kidneys (all P values < .01). The longer time interval group had better shimming and water suppression effects than the shorter time interval group (all P values < .01). After the administration of Gd-DTPA in one patient, sequential values of shimming and water suppression in the right and left kidneys, respectively, were 13 Hz and 97% and 12 Hz and 97% prior to the examination, 34 Hz and 86% and 30 Hz and 88% at 5 minutes, 32 Hz and 89% and 27 Hz and 90% at 10 minutes, 28 Hz and 91% and 24 Hz and 91% at 15 minutes, and 24 Hz and 92% and 20 Hz and 92% at 25 minutes. CONCLUSIONS: Gd-DTPA exerts adverse effects on water suppression and shimming, both of which show a trend of becoming well gradually with time extension after the injection of Gd-DTPA. This phenomenon limits the diagnostic use of kidney magnetic resonance spectroscopy performed immediately after contrast-enhanced magnetic resonance imaging.