Arminder S. Jassar

Gemcitabine Selectively Eliminates Splenic Gr-1+/CD11b+ Myeloid Suppressor Cells in Tumor-Bearing Animals and Enhances Antitumor Immune Activity

Purpose: Myeloid suppressor (Gr-1+/CD11b+) cells accumulate in the spleens of tumor-bearing mice where they contribute to immunosuppression by inhibiting the function of CD8+ T cells and by promoting tumor angiogenesis. Elimination of these myeloid suppressor cells may thus significantly improve antitumor responses and enhance effects of cancer immunotherapy, although to date few practical options exist. Experimental Design: The effect of the chemotherapy drug gemcitabine on the number of (Gr-1+/CD11b+) cells in the spleens of animals bearing large tumors derived from five cancer lines grown in both C57Bl/6 and BALB/c mice was analyzed. Suppressive activity of splenocytes from gemcitabine-treated and control animals was measured in natural killer (NK) cell lysis and Winn assays. The impact of myeloid suppressor cell activity was determined in an immunogene therapy model using an adenovirus expressing IFN-β. Results: This study shows that the chemotherapeutic drug gemcitabine, given at a dose similar to the equivalent dose used in patients, was able to dramatically and specifically reduce the number of myeloid suppressor cells found in the spleens of animals bearing large tumors with no significant reductions in CD4+ T cells, CD8+ T cells, NK cells, macrophages, or B cells. The loss of myeloid suppressor cells was accompanied by an increase in the antitumor activity of CD8+ T cells and activated NK cells. Combining gemcitabine with cytokine immunogene therapy using IFN-β markedly enhanced antitumor efficacy. Conclusions: These results suggest that gemcitabine may be a practical strategy for the reduction of myeloid suppressor cells and should be evaluated in conjunction with a variety of immunotherapy approaches.

Quantitative mitral valve modeling using real-time three-dimensional echocardiography: technique and repeatability.

BACKGROUND Real-time three-dimensional (3D) echocardiography has the ability to construct quantitative models of the mitral valve (MV). Imaging and modeling algorithms rely on operator interpretation of raw images and may be subject to observer-dependent variability. We describe a comprehensive analysis technique to generate high-resolution 3D MV models and examine interoperator and intraoperator repeatability in humans. METHODS Patients with normal MVs were imaged using intraoperative transesophageal real-time 3D echocardiography. The annulus and leaflets were manually segmented using a TomTec Echo-View workstation. The resultant annular and leaflet point cloud was used to generate fully quantitative 3D MV models using custom Matlab algorithms. Eight images were subjected to analysis by two independent observers. Two sequential images were acquired for 6 patients and analyzed by the same observer. Each pair of annular tracings was compared with respect to conventional variables and by calculating the mean absolute distance between paired renderings. To compare leaflets, MV models were aligned so as to minimize their sum of squares difference, and their mean absolute difference was measured. RESULTS Mean absolute annular and leaflet distance was 2.4±0.8 and 0.6±0.2 mm for the interobserver and 1.5±0.6 and 0.5±0.2 mm for the intraobserver comparisons, respectively. There was less than 10% variation in annular variables between comparisons. CONCLUSIONS These techniques generate high-resolution, quantitative 3D models of the MV and can be used consistently to image the human MV with very small interoperator and intraoperator variability. These data lay the framework for reliable and comprehensive noninvasive modeling of the normal and diseased MV.

Ischemic Mitral Regurgitation: A Quantitative Three-Dimensional Echocardiographic Analysis

BACKGROUND A comprehensive three-dimensional echocardiography based approach is applied to preoperative mitral valve (MV) analysis in patients with ischemic mitral regurgitation (IMR). This method is used to characterize the heterogeneous nature of the pathologic anatomy associated with IMR. METHODS Intraoperative real-time three-dimensional transesophageal echocardiograms of 18 patients with IMR (10 with anterior, 8 with inferior infarcts) and 17 patients with normal MV were analyzed. A customized image analysis protocol was used to assess global and regional determinants of annular size and shape, leaflet tethering and curvature, relative papillary muscle anatomy, and anatomic regurgitant orifice area. RESULTS Both mitral annular area and MV tenting volume were increased in the IMR group as compared with patients with normal MV (mitral annular area=1,065±59 mm2 versus 779±44 mm2, p=0.001; and MV tenting volume=3,413±403 mm3 versus 1,696±200 mm3, p=0.001, respectively). Within the IMR group, patients with anterior infarct had larger annuli (1,168±99 mm2) and greater tenting volumes (4,260±779 mm3 versus 2,735±245 mm3, p=0.06) than the inferior infarct subgroup. Papillary-annular distance was increased in the IMR group relative to normal; these distances were largest in patients with anterior infarcts. Whereas patients with normal MV had very consistent anatomic determinants, annular shape and leaflet tenting distribution in the IMR group were exceedingly variable. Mean anatomic regurgitant orifice area was 25.8±3.0 mm2, and the number of discrete regurgitant orifices varied from 1 to 4. CONCLUSIONS Application of custom analysis techniques to three-dimensional echocardiography images allows a quantitative and systematic analysis of the MV, and demonstrates the extreme variability in pathologic anatomy that occurs in patients with severe IMR.

Gemcitabine has significant immunomodulatory activity in murine tumor models independent of its cytotoxic effects

Given reports that the chemotherapeutic agent gemcitabine (GEM) does not block T-lymphocyte recall responses and is not detrimental to specific anti-tumor immunity, studies to evaluate the use of GEM in combination with immunotherapy were initiated. When we tested the therapeutic effects of GEM as a single agent in various murine tumor models, we found that a single dose of GEM had impressive anti-tumor activity in a specific subset of tumors. Surprisingly, efficacy was not related to in vitro drug sensitivity, but instead, correlated with the immunogenicity of the tumor. A key role of the immune system in GEM’s action was demonstrated in experiments showing that the anti-tumor effects of GEM were lost in nude mice. In addition, we saw equivalent anti-tumor effects of GEM in animals bearing tumors that were extremely resistant to the in vitro cytotoxic effects of GEM versus parental GEM-sensitive cells. This therapeutic efficacy was thus not due to direct cytotoxic effect on tumor cells, but rather to an enhancement of T-cell mediated anti-tumor immune effects. These data raise the exciting possibility that GEM may be a useful agent in combination with various types of tumor immunotherapy.

Three-Dimensional Echocardiographic Analysis of Mitral Annular Dynamics Implication for Annuloplasty Selection

Background— Proponents of flexible annuloplasty rings have hypothesized that such devices maintain annular dynamics. This hypothesis is based on the supposition that annular motion is relatively normal in patients undergoing mitral valve repair. We hypothesized that mitral annular dynamics are impaired in ischemic mitral regurgitation and myxomatous mitral regurgitation. Methods and Results— A Philips iE33 echocardiographic module and X7–2t probe were used to acquire full-volume real-time 3-dimensional transesophageal echocardiography loops in 11 normal subjects, 11 patients with ischemic mitral regurgitation and 11 patients with myxomatous mitral regurgitation. Image analysis was performed using Tomtec Image Arena, 4D-MV Assessment, 2.1 (Munich, Germany). A midsystolic frame was selected for the initiation of annular tracking using the semiautomated program. Continuous parameters were normalized in time to provide for uniform systolic and diastolic periods. Both ischemic mitral regurgitation (9.98±155 cm2) and myxomatous mitral regurgitation annuli (13.29±3.05 cm2) were larger in area than normal annuli (7.95±1.40 cm2) at midsystole. In general, ischemic mitral regurgitation annuli were less dynamic than controls. In myxomatous mitral regurgitation, annular dynamics were also markedly abnormal with the mitral annulus dilating rapidly in early systole in response to rising ventricular pressure. Conclusions— In both ischemic mitral regurgitation and myxomatous mitral regurgitation, annular dynamics and anatomy are abnormal. Flexible annuloplasty devices used in mitral valve repair are, therefore, unlikely to result in either normal annular dynamics or normal anatomy.

Intrapulmonary IFN-β Gene Therapy Using an Adenoviral Vector Is Highly Effective in a Murine Orthotopic Model of Bronchogenic Adenocarcinoma of the Lung

Given previous work showing that an adenoviral vector expressing IFN-beta (Ad.IFNbeta) was highly effective in eradicating i.p. mesothelioma tumors, the antitumor efficacy of this agent was evaluated in an orthotopic model of bronchogenic adenocarcinoma of the lung. These transgenic mice have a conditionally expressed, oncogenic K-rasG12D allele that can be activated by intratracheal administration of an adenovirus expressing Cre recombinase (Ad.Cre). K-rasG12D mutant mice were given Ad.Cre intranasally to activate the oncogene. Mice were then given 10(9) plaque-forming units of a control vector (Ad.LacZ) or Ad.IFNbeta intranasally 3 and 4 weeks later, a time when lung tumors had been established. Cells derived from K-ras-mutated lung tumors were also grown in the flanks of mice to study mechanisms of therapeutic responses. In two separate experiments, untreated tumor-bearing mice all died by day 57 (median survival, 49 days). Ad.LacZ-treated mice all died by day 71 (median survival, 65 days). In contrast, 90% to 100% of mice treated with Ad.IFNbeta were long-term survivors (>120 days; P < 0.001). In addition, immunity to re-challenge with tumor cells was induced. In vitro and flank tumor studies showed that Ad.IFNbeta induced direct tumor cell killing and that depleting natural killer or CD8+ T cells, but not CD4+ T cells, with antibodies attenuated the effect of Ad.IFNbeta. These studies, showing remarkable antitumor activity in this orthotopic lung cancer model, provide strong preclinical support for a trial of Ad.IFNbeta to treat human non-small cell lung cancer.

Graft Selection for Aortic Root Replacement in Complex Active Endocarditis: Does It Matter?

BACKGROUND Endocarditis affecting the aortic valve, with abscess formation and root destruction, remains a challenge to treat. Aortic root homografts have been advocated because of a perceived lower risk of infective complications than with other root replacement grafts. However, the theoretical advantage of homografts has not been re-evaluated in the modern era. This report is based on an examination of our results for all aortic root replacements in complex, active endocarditis affecting the aortic valve. METHODS From 2000 to 2010, 134 patients (70.9% male; mean age 58.3±14.8 years) at our institution underwent aortic root replacement for active endocarditis. Ninety of the patients (67.2%) had a previously implanted prosthetic aortic valve. Our findings for these patients included one or more of the following: abscess (n=110, 82.1%), valve vegetation (n=98, 73.1%), and pseudoaneurysm or rupture or both (n=62, 46.3%). We retrospectively reviewed data for the patients from hospital records and the social security data base. RESULTS A mechanical composite graft (MC) was used in 43 of the patients (32.1%), a non-homograft biologic valve conduit (BC) in 55 patients (41.0%), and a homograft (HG) valve in 36 patients (26.9%). There was no significant difference among the groups in the incidence of major complications or in-hospital mortality. During a mean follow-up of 32.1±29.4 months, the rates of readmission, reinfection, and reoperation were similar for the three groups. The mean 5-year survival in the study was 58±9% for the MC group, 62±7% for the BC group, and 58 ± 9% for the HG group, respectively (p=0.48). CONCLUSIONS Aortic root replacement in the presence of complex active infection is associated with significant morbidity and mortality. We report that the rates of major complications and late mortality were similar among MC, BC, and HG groups in our study.

Semi-automated mitral valve morphometry and computational stress analysis using 3D ultrasound

In vivo human mitral valves (MV) were imaged using real-time 3D transesophageal echocardiography (rt-3DTEE), and volumetric images of the MV at mid-systole were analyzed by user-initialized segmentation and 3D deformable modeling with continuous medial representation, a compact representation of shape. The resulting MV models were loaded with physiologic pressures using finite element analysis (FEA). We present the regional leaflet stress distributions predicted in normal and diseased (regurgitant) MVs. Rt-3DTEE, semi-automated leaflet segmentation, 3D deformable modeling, and FEA modeling of the in vivo human MV is tenable and useful for evaluation of MV pathology.

Using macrophage activation to augment immunotherapy of established tumours

Background:Successful immunotherapy will require alteration of the tumour microenvironment and/or decreased immune suppression. Tumour-associated macrophages (TAMs) are one major factor affecting tumour microenvironment. We hypothesised that altering TAM phenotype would augment the efficacy of immunotherapy.Methods:We and others have reported that 5,6-Dimethylxanthenone-4-acetic-acid (DMXAA, Vadimezan) has the ability to change TAM phenotypes, inducing a tumour microenvironment conducive to antitumour immune responses. We therefore combined DMXAA with active immunotherapies, and evaluated anti-tumour efficacy, immune cell phenotypes (flow cytometry), and tumour microenvironment (RT–PCR).Results:In several different murine models of immunotherapy for lung cancer, DMXAA-induced macrophage activation significantly augmented the therapeutic effects of immunotherapy. By increasing influx of neutrophils and anti-tumour (M1) macrophages to the tumour, DMXAA altered myeloid cell phenotypes, thus changing the intratumoural M2/non-M2 TAM immunoinhibitory ratio. It also altered the tumour microenvironment to be more pro-inflammatory. Modulating macrophages during immunotherapy resulted in increased numbers, activity, and antigen-specificity of intratumoural CD8+ T cells. Macrophage depletion reduced the effect of combining immunotherapy with macrophage activation, supporting the importance of TAMs in the combined effect.Conclusion:Modulating intratumoural macrophages dramatically augmented the effect of immunotherapy. Our observations suggest that addition of agents that activate TAMs to immunotherapy should be considered in future trials.

Cardiac Surgery in Jehovah's Witness Patients: Ten-Year Experience

BACKGROUND Cardiac surgery in Jehovahs Witnesses poses unique challenges. We have developed a comprehensive multimodality program for these patients and have obtained excellent results. METHODS Ninety-one Jehovahs Witness patients underwent cardiac surgery between 2000 and 2010. Preoperative, intraoperative, and postoperative considerations in the conduct of bloodless surgery in the Jehovahs Witness population are discussed. Mortality for isolated coronary artery bypass graft surgery and isolated aortic valve replacement was compared with predicted mortality from The Society of Thoracic Surgeons (STS) risk models. Perioperative outcomes were stratified by urgent and elective status of operations. RESULTS Mean age was 65±12.4 years. Comorbid conditions included hypertension (84.6%), diabetes mellitus (48.4%), previous myocardial infarction (23.1%), chronic lung disease (38.5%), peripheral vascular disease (20.9%), and renal failure (11%). In-hospital mortality was 5.5% (n=5). Mortality for isolated coronary artery bypass graft surgery and isolated aortic valve replacement was 2.2% (observed to expected ratio=1.05, 95% confidence interval: 0 to 3.02) and 5.6% (observed to expected=1.46, 95% confidence interval: 0 to 3.76), respectively. Other complications included reoperation (all=8.8%, cardiac=2.2%), sepsis (2.2%), sternal wound infection (1.1%), transient ischemic attack (1.1%), renal failure requiring dialysis (1.1%), and prolonged ventilation (18.7%). Major complication rates were not significantly different between the elective group and the urgent group. CONCLUSIONS Bloodless cardiac surgery in Jehovahs Witness patients can be performed with excellent outcomes in both elective and urgent situations. Mortality rates for isolated coronary artery bypass graft surgery and isolated aortic valve replacement are within the expected 95% confidence intervals of STS predicted mortality.