Ling Cai

Diagnosis of lung cancer in individuals with solitary pulmonary nodules by plasma microRNA biomarkers

BackgroundMaking a definitive preoperative diagnosis of solitary pulmonary nodules (SPNs) found by CT has been a clinical challenge. We previously demonstrated that microRNAs (miRNAs) could be used as biomarkers for lung cancer diagnosis. Here we investigate whether plasma microRNAs are useful in identifying lung cancer among individuals with CT-detected SPNs.MethodsBy using quantitative reverse transcriptase PCR analysis, we first determine plasma expressions of five miRNAs in a training set of 32 patients with malignant SPNs, 33 subjects with benign SPNs, and 29 healthy smokers to define a panel of miRNAs that has high diagnostic efficiency for lung cancer. We then validate the miRNA panel in a testing set of 76 patients with malignant SPNs and 80 patients with benign SPNs.ResultsIn the training set, miR-21 and miR-210 display higher plasma expression levels, whereas miR-486-5p has lower expression level in patients with malignant SPNs, as compared to subjects with benign SPNs and healthy controls (all P ≤ 0.001). A logistic regression model with the best prediction was built on the basis of miR-21, miR-210, and miR-486-5p. The three miRNAs used in combination produced the area under receiver operating characteristic curve at 0.86 in distinguishing lung tumors from benign SPNs with 75.00% sensitivity and 84.95% specificity. Validation of the miRNA panel in the testing set confirms their diagnostic value that yields significant improvement over any single one.ConclusionsThe plasma miRNAs provide potential circulating biomarkers for noninvasively diagnosing lung cancer among individuals with SPNs, and could be further evaluated in clinical trials.

Combination immunotherapy using adoptive T-cell transfer and tumor antigen vaccination on the basis of hTERT and survivin after ASCT for myeloma

In a phase 1/2 two-arm trial, 54 patients with myeloma received autografts followed by ex vivo anti-CD3/anti-CD28 costimulated autologous T cells at day 2 after transplantation. Study patients positive for human leukocyte antigen A2 (arm A, n = 28) also received pneumococcal conjugate vaccine immunizations before and after transplantation and a multipeptide tumor antigen vaccine derived from the human telomerase reverse transcriptase and the antiapoptotic protein survivin. Patients negative for human leukocyte antigen A2 (arm B, n = 26) received the pneumococcal conjugate vaccine only. Patients exhibited robust T-cell recoveries by day 14 with supraphysiologic T-cell counts accompanied by a sustained reduction in regulatory T cells. The median event-free survival (EFS) for all patients is 20 months (95% confidence interval, 14.6-24.7 months); the projected 3-year overall survival is 83%. A subset of patients in arm A (36%) developed immune responses to the tumor antigen vaccine by tetramer assays, but this cohort did not exhibit better EFS. Higher posttransplantation CD4(+) T-cell counts and a lower percentage of FOXP3(+) T cells were associated with improved EFS. Patients exhibited accelerated polyclonal immunoglobulin recovery compared with patients without T-cell transfers. Adoptive transfer of tumor antigen vaccine-primed and costimulated T cells leads to augmented and accelerated cellular and humoral immune reconstitution, including antitumor immunity, after autologous stem cell transplantation for myeloma. This study was registered at www.clinicaltrials.gov as NCT00499577.

Combination Immunotherapy after ASCT for Multiple Myeloma Using MAGE-A3/Poly-ICLC Immunizations Followed by Adoptive Transfer of Vaccine-Primed and Costimulated Autologous T Cells

Purpose: Myeloma-directed cellular immune responses after autologous stem cell transplantation (ASCT) may reduce relapse rates. We studied whether coinjecting the TLR-3 agonist and vaccine adjuvant Poly-ICLC with a MAGE-A3 peptide vaccine was safe and would elicit a high frequency of vaccine-directed immune responses when combined with vaccine-primed and costimulated autologous T cells. Experimental Design: In a phase II clinical trial (NCT01245673), we evaluated the safety and activity of ex vivo expanded autologous T cells primed in vivo using a MAGE-A3 multipeptide vaccine (compound GL-0817) combined with Poly-ICLC (Hiltonol), granulocyte macrophage colony-stimulating factor (GM-CSF) ± montanide. Twenty-seven patients with active and/or high-risk myeloma received autografts followed by anti-CD3/anti-CD28–costimulated autologous T cells, accompanied by MAGE-A3 peptide immunizations before T-cell collection and five times after ASCT. Immune responses to the vaccine were evaluated by cytokine production (all patients), dextramer binding to CD8+ T cells, and ELISA performed serially after transplant. Results: T-cell infusions were well tolerated, whereas vaccine injection site reactions occurred in >90% of patients. Two of nine patients who received montanide developed sterile abscesses; however, this did not occur in the 18 patients who did not receive montanide. Dextramer staining demonstrated MAGE-A3–specific CD8 T cells in 7 of 8 evaluable HLA-A2+ patients (88%), whereas vaccine-specific cytokine-producing T cells were generated in 19 of 25 patients (76%). Antibody responses developed in 7 of 9 patients (78%) who received montanide and only weakly in 2 of 18 patients (11%) who did not. The 2-year overall survival was 74% [95% confidence interval (CI), 54%–100%] and 2-year event-free survival was 56% (95% CI, 37%–85%). Conclusions: A high frequency of vaccine-specific T-cell responses were generated after transplant by combining costimulated autologous T cells with a Poly-ICLC/GM-CSF–primed MAGE-A3 vaccine. Clin Cancer Res; 20(5); 1355–65. ©2013 AACR.

Analysis of MicroRNAs in Sputum to Improve Computed Tomography for Lung Cancer Diagnosis

Introduction: Computed tomography (CT) plays a central role in lung cancer diagnosis. However, CT has relatively low specificity, presenting a challenge in clinical settings. We previously identified 12 microRNAs (miRNAs) whose expressions in tumor tissues were associated with lung cancer. Methods: Using quantitative reverse transcriptase polymerase chain reaction, we aimed to identify miRNA biomarkers in sputum that could complement CT for diagnosis of lung cancer. Results: In a training set consisting of 66 lung cancer patients and 68 cancer-free smokers, 10 of the 12 miRNAs were differentially expressed between the cases and controls (p ⩽ 0.01). From the miRNAs, a logistic regression model was built on the basis of miR-31 and miR-210, both of which had the best prediction for lung cancer, producing an area under receiver operating characteristic curve of 0.83. Combined use of the two miRNAs yielded 65.2% sensitivity and 89.7% specificity, CT had 93.9% sensitivity and 83.8% specificity for lung cancer diagnosis. Notably, combined analysis of the miRNA biomarkers and CT produced a higher specificity than does CT used alone (91.2% versus 83.8%; p < 0.05). The diagnostic performance of the biomarkers was confirmed in a testing set comprising 64 lung cancer patients and 73 cancer-free smokers. Conclusion: The sputum miRNA biomarkers might be useful in improving CT for diagnosis of lung cancer, but further independent validation on an external and prospective cohort of patients is required.

The Novel BCR-ABL and FLT3 Inhibitor Ponatinib Is a Potent Inhibitor of the MDR-Associated ATP-Binding Cassette Transporter ABCG2

Ponatinib is a novel tyrosine kinase inhibitor with potent activity against BCR-ABL with mutations, including T315I, and also against fms-like tyrosine kinase 3. We tested interactions between ponatinib at pharmacologically relevant concentrations of 50 to 200 nmol/L and the MDR-associated ATP-binding cassette (ABC) proteins ABCB1, ABCC1, and ABCG2. Ponatinib enhanced uptake of substrates of ABCG2 and ABCB1, but not ABCC1, in cells overexpressing these proteins, with a greater effect on ABCG2 than on ABCB1. Ponatinib potently inhibited [125I]-IAAP binding to ABCG2 and ABCB1, indicating binding to their drug substrate sites, with IC50 values of 0.04 and 0.63 μmol/L, respectively. Ponatinib stimulated ABCG2 ATPase activity in a concentration-dependent manner and stimulated ABCB1 ATPase activity at low concentrations, consistent with it being a substrate of both proteins at pharmacologically relevant concentrations. The ponatinib IC50 values of BCR-ABL–expressing K562 cells transfected with ABCB1 and ABCG2 were approximately the same as and 2-fold higher than that of K562, respectively, consistent with ponatinib being a substrate of both proteins, but inhibiting its own transport, and resistance was also attenuated to a small degree by ponatinib-induced downregulation of ABCB1 and ABCG2 cell-surface expression on resistant K562 cells. Ponatinib at pharmacologically relevant concentrations produced synergistic cytotoxicity with ABCB1 and ABCG2 substrate chemotherapy drugs and enhanced apoptosis induced by these drugs, including daunorubicin, mitoxantrone, topotecan, and flavopiridol, in cells overexpressing these transport proteins. Combinations of ponatinib and chemotherapy drugs warrant further testing. Mol Cancer Ther; 11(9); 2033–44. ©2012 AACR.

Sputum microRNA biomarkers for identifying lung cancer in indeterminate solitary pulmonary nodules

Purpose: The early detection of lung cancer in heavy smokers by low-dose CT (LDCT) can reduce the mortality. However, LDCT screening increases the number of indeterminate solitary pulmonary nodules (SPN) in asymptomatic individuals, leading to overdiagnosis. Making a definitive preoperative diagnosis of malignant SPNs has been a clinical challenge. We have demonstrated that sputum miRNAs could provide potential biomarkers for lung cancer. Here, we aimed to develop sputum miRNA biomarkers for diagnosis of malignant SPNs. Experimental Design: Using quantitative RT-PCR, we evaluated expressions of 13 sputum miRNAs, previously identified sputum miRNA signatures of lung cancer, in a training set of 122 patients with either malignant (n = 60) or benign SPNs (n = 62) to define a panel of biomarkers. We then validated the biomarker panel in an internal testing set of 136 patients with either malignant (n = 67) or benign SPNs (n = 69), and an external testing cohort of 155 patients with either malignant (n = 76) or benign SPNs (n = 79). Results: In the training set, a panel of three miRNA biomarkers (miRs21, 31, and 210) was developed, producing 82.93% sensitivity and 87.84% specificity for identifying malignant SPNs. The sensitivity and specificity of the biomarkers in the two independent testing cohorts were 82.09% and 88.41%, 80.52% and 86.08%, respectively, confirming the diagnostic value. Conclusions: Sputum miRNA biomarkers may improve LDCT screening for lung cancer in heavy smokers by preoperatively diagnosing malignant SPNs. Nevertheless, a prospective study in a large population to validate the biomarkers is needed. Clin Cancer Res; 21(2); 484–9. ©2015 AACR.

Patient-reported outcomes in women with breast cancer enrolled in a dual-center, double-blind, randomized controlled trial assessing the effect of acupuncture in reducing aromatase inhibitor-induced musculoskeletal symptoms.

Aromatase inhibitors (AIs) have been associated with decrements in patient‐reported outcomes (PROs). The objective of this study was to assess whether real acupuncture (RA), compared with sham acupuncture (SA), improves PROs in patients with breast cancer who are receiving an adjuvant AI.

Platelet glycoprotein Ibα ectodomain shedding and non-surgical bleeding in heart failure patients supported by continuous-flow left ventricular assist devices

BACKGROUND Non-surgical bleeding (NSB) is a major complication among heart failure (HF) patients supported by continuous-flow left ventricular assist devices (CF-LVADs). Understanding the hemostatic defects contributing to NSB after CF-LVAD implantation is crucial for prevention of this adverse event. The aim of this study was to examine the link between platelet glycoprotein Ibα (GPIbα) ectodomain shedding and NSB in CF-LVAD recipients and to identify a potential biomarker of NSB. METHODS Serial blood samples were collected from 35 HF patients supported with CF-LVADs. Platelet function was evaluated by a platelet function analysis device and thromboelastography (TEG). Platelet GPIbα shedding, von Willebrand factor (vWF) antigen and vWF collagen binding capacity were determined using enzyme-linked immunosorbent assays (ELISAs). The structural analysis of vWF was performed by gel electrophoresis. These platelet function measures with vWF parameters of the patients who had NSB between 4 and 32 days after CF-LVAD implantation (bleeder) were analyzed against those without NSB (non-bleeder). Blood samples from 7 healthy individuals were collected to obtain healthy reference values for the laboratory assays. RESULTS Elevated GPIbα shedding was found to be a pre-existing condition in all HF patients prior to CF-LVAD implantation. Post-operative level of GPIbα shedding increased and remained elevated in the bleeder group, whereas a consistent decrease was found in the non-bleeder group. A receiver operating characteristic (ROC) analysis indicated that the level of GPIbα shedding had a predictive power of NSB in patients on CF-LVAD support. CONCLUSIONS Platelet GPIbα ectodomain shedding which attenuates platelet reactivity is associated with NSB. Plasma GPIbα level may potentially be used to refine bleeding risk stratification in CF-LVAD patients.

Density functional calculations of chemical shielding of backbone 15N in helical residues of protein G

We performed density functional calculations of backbone 15N chemical shielding tensors in selected helical residues of protein G. Here we describe a computationally efficient methodology to include most of the important effects in the calculation of chemical shieldings of backbone 15N. We analyzed the role of long-range intra-protein electrostatic interactions by comparing models with different complexity in vacuum and in charge field. Our results show that the dipole moment of the α-helix can cause significant deshielding of 15N; therefore, it needs to be considered when calculating 15N chemical shielding. We found that it is important to include interactions with the side chains that are close in space when the charged form for ionizable side chains is adopted in the calculation. We also illustrate how the ionization state of these side chains can affect the chemical shielding tensor elements. Chemical shielding calculations using a 8-residue fragment model in vacuum and adopting the charged form of ionizable side chains yield a generally good agreement with experimental data.

The effect of computer-aided detection on radiologist performance in the detection of lung cancers previously missed on a chest radiograph.

Purpose: The purpose of the study was to determine whether computer-aided detection (CAD) can improve a radiologist’s ability to detect lung cancers previously missed on a chest radiograph (CXR). Materials and Methods: Eighty-one cases of lung cancer previously missed on CXR were collected, along with the CXRs of 215 age-matched and emphysema-matched controls without lung cancer. Tumor subtlety was scored from 1 (very subtle) to 5 (very obvious) by expert thoracic radiologists. All 297 CXRs were processed using a CAD system (OnGuard, Version 5.1 Riverain Medical, Miamisburg, OH) to create a set of 2 images for each patient, 1 with and 1 without CAD. Eleven general radiologists took part in a reader study. Each radiologist viewed the CXR without CAD and then the one with CAD for each patient sequentially. Areas of concern, if present, were marked. The degree of confidence in diagnosis was scored on a scale of 0 (no cancer) to 100 (definite) in succession for CXRs without CAD and then for those with CAD. Localization receiver operating characteristic analysis was used for evaluation of the observers’ performance. Results: Of the 81 cancer cases, OnGuard correctly detected 40 tumors with a sensitivity of 49.4%. In the reader study, there was a significant increase in the area under the localization receiver operating characteristic curve with the aid of OnGuard, which increased from 0.38 to 0.43. Aggregate reader sensitivity improved significantly from 0.44 to 0.5 with the use of OnGuard. Conclusion: The use of OnGuard improves reader accuracy and sensitivity for the detection of lung cancers previously missed on CXR.