Characterization and comparison of novel adjuvants for a prefusion clamped MERS vaccine.

Various chemical adjuvants are available to augment immune responses to non-replicative, subunit vaccines. Optimized adjuvant selection can ensure that vaccine-induced immune responses protect against the diversity of pathogen-associated infection routes, mechanisms of infectious spread, and pathways of immune evasion. In this study, we compare the immune response of mice to a subunit vaccine of Middle Eastern respiratory syndrome coronavirus (MERS-CoV) spike protein, stabilized in its prefusion conformation by a proprietary molecular clamp (MERS SClamp) alone or formulated with one of six adjuvants: either (i) aluminium hydroxide, (ii) SWE, a squalene-in-water emulsion, (iii) SQ, a squalene-in-water emulsion containing QS21 saponin, (iv) SMQ, a squalene-in-water emulsion containing QS21 and a synthetic toll-like receptor 4 (TLR4) agonist 3D-6-acyl Phosphorylated HexaAcyl Disaccharide (3D6AP); (v) LQ, neutral liposomes containing cholesterol, 1.2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) and QS21, (vi) or LMQ, neutral liposomes containing cholesterol, DOPC, QS21, and 3D6AP. All adjuvanted formulations induced elevated antibody titers which where greatest for QS21-containing formulations. These had elevated neutralization capacity and induced higher frequencies of IFNƔ and IL-2-producing CD4+ and CD8+ T cells. Additionally, LMQ-containing formulations skewed the antibody response towards IgG2b/c isotypes, allowing for antibody-dependent cellular cytotoxicity. This study highlights the utility of side-by-side adjuvant comparisons in vaccine development.

Impact of Minimally Invasive Benign Prostatic Hyperplasia Therapies on 30- and 90-Day Postoperative Office Encounters.

OBJECTIVE: To compare the frequency of postoperative encounters in the 30-day and 90-day postoperative periods for various bladder outlet obstruction surgical therapies. MATERIALS AND METHODS: All patients who underwent transurethral resection of the prostate (TURP), GreenLight laser photovaporization of the prostate (GL-PVP) (American Medical Systems Inc.), and holmium laser enucleation of the prostate (HoLEP) from January 1, 2012 to December 31, 2014 were followed for 6 months postoperatively. All postoperative encounters such as patient calls or questions, catheter exchanges or removals, and hospital-based readmissions or emergency department visits were recorded in the electronic medical record. RESULTS: Two hundred and ninety-one consecutive patients underwent outlet procedures during the study period: TURP (N = 199; mean age, 71 years; mean body mass index [BMI], 28.5), HoLEP (N = 60; mean age, 68 years; mean BMI, 28.1), or GL-PVP (N = 32; mean age, 72 years; mean BMI, 29.3). No statistically significant difference was observed for age, BMI, preoperative American Urological Association symptom score, or preoperative maximum flow velocity between the 3 groups. Thirty-day postoperative encounters differed significantly between the 3 surgery types (P < .001). Specifically, there were fewer encounters within 30 days of surgery for TURP compared to both HoLEP (≥1 encounter: TURP = 48.7%, HoLEP = 66.7%; P = .006) and GL-PVP (≥1 encounter: TURP = 48.7%, GL-PVP = 93.7%; P < .001). The number of encounters within 90 days postoperatively was also significantly lower for TURP patients (P < .001). CONCLUSION: TURP results in fewer postoperative encounters in both the 30-day and 90-day postoperative periods compared to HoLEP and GL-PVP. Laser prostate therapies may place increased burden on clinic staff during the 30-day and 90-day postoperative periods.

A generic screening platform for inhibitors of virus induced cell fusion using cellular electrical impedance.

Fusion of the viral envelope with host cell membranes is an essential step in the life cycle of all enveloped viruses. Despite such a clear target for antiviral drug development, few anti-fusion drugs have progressed to market. One significant hurdle is the absence of a generic, high-throughput, reproducible fusion assay. Here we report that real time, label-free measurement of cellular electrical impedance can quantify cell-cell fusion mediated by either individually expressed recombinant viral fusion proteins, or native virus infection. We validated this approach for all three classes of viral fusion and demonstrated utility in quantifying fusion inhibition using antibodies and small molecule inhibitors specific for dengue virus and respiratory syncytial virus.

Recent advances in the development of subunit-based RSV vaccines.

Respiratory syncytial virus (RSV) is a major cause of lower respiratory tract infections causing pneumonia and bronchiolitis in infants. RSV also causes serious illness in elderly populations, immunocompromised patients and individuals with pulmonary or cardiac problems. The significant morbidity and mortality associated with RSV infection have prompted interest in RSV vaccine development. In the 1960s, a formalin-inactivated vaccine trial failed to protect children, and indeed enhanced pathology when naturally infected later with RSV. Hence, an alternative approach to traditional killed virus vaccines, which can induce protective immunity without serious adverse events, is desired. Several strategies have been explored in attempts to produce effective vaccine candidates including gene-based and subunit vaccines. Subunit-based vaccine approaches have shown promising efficacy in animal studies and several have reached clinical trials. The current stage of development of subunit-based vaccines against RSV is reviewed in this article.

Surface modified microprojection arrays for the selective extraction of the dengue virus NS1 protein as a marker for disease.

While advances in assay chemistry and detection continue to improve molecular diagnostics technology, blood samples are still collected using the 150-year-old needle/syringe method. Surface modified microprojection arrays have been developed as a novel platform for in vivo, needle-free biomarker capture. These devices are gold coated silicon arrays with >20,000 projections per cm(2), which can be applied to the skin for tunable penetration into the epidermis or dermis. The microprojection array conceptually offers several advantages over the current methods including: minimally invasive sample collection, no need for sample processing and concentration of specific markers at the device surface for sensitive detection. In this study, Microprojection arrays were coated with antibodies to capture an early marker of dengue virus infection, NS1, from the skin of live mice. We also developed a complementary "total IgG" assay which could be used as a positive control for adequate penetration of the projections. Surface modifications designed for selective extraction were tested against standard microtiter plate ELISA. We also investigated the use of Protein G-mediated antibody immobilization in order to orient capture antibodies. While we found that capture efficiency could be improved, the direct EDC-based antibody immobilization resulted in a significantly higher surface density leading to a higher degree of NS1 capture. Using mice intravenously injected with recombinant dengue virus type 2 NS1 as a pseudomodel for dengue infection, NS1 was successfully extracted using microprojection arrays sampling from skin fluid, with a detection limit of 8 µg/mL.

In silico screening of small molecule libraries using the dengue virus envelope E protein has identified compounds with antiviral activity against multiple flaviviruses.

The flaviviruses comprise a large group of related viruses, many of which pose a significant global human health threat, most notably the dengue viruses (DENV), West Nile virus (WNV) and yellow fever virus (YFV). Flaviviruses enter host cells via fusion of the viral and cellular membranes, a process mediated by the major viral envelope protein E as it undergoes a low pH induced conformational change in the endosomal compartment of the host cell. This essential entry stage in the flavivirus life cycle provides an attractive target for the development of antiviral agents. We performed an in silico docking screen of the Maybridge chemical database within a previously described ligand binding pocket in the dengue E protein structure that is thought to play a key role in the conformational transitions that lead to membrane fusion. The biological activity of selected compounds identified from this screen revealed low micromolar antiviral potency against dengue virus for two of the compounds. Our results also provide the first evidence that compounds selected to bind to this ligand binding site on the flavivirus E protein abrogate fusion activity. Interestingly, one of these compounds also has antiviral activity against both WNV (kunjin strain) and YFV.

Conservative management of incidental contrast-enhancing renal masses as safe alternative to invasive therapy.

OBJECTIVES: To describe small contrast-enhancing renal masses suggestive of cancer that were managed conservatively with watchful waiting and serial computed tomography scans. Advanced diagnostic imaging has led to the increased incidental detection of renal masses in patients whose multiple comorbid conditions preclude invasive management. METHODS: A retrospective review was conducted of 29 consecutive patients with 29 incidentally detected asymptomatic renal masses less than 3.5 cm in diameter that were managed conservatively with watchful waiting (because of patient wishes or multiple comorbid conditions) and serial computed tomography scans. RESULTS: The average patient age was 70 years (range 51 to 88), and the average duration of follow-up imaging was 32 months (range 10 to 89). The average number of follow-up computed tomography scans was 4.9 per patient (range 1 to 11). The average width of the renal masses at diagnosis was 1.83 cm (range 0.4 to 3.5), and the average change in size per year was 0.12 cm for all patients. Four patients underwent radical nephrectomy because of growth of the renal mass (n = 1) or patient wishes (n = 3). The histologic findings in 3 of these 4 patients were consistent with renal cell carcinoma. Two patients underwent radiofrequency ablation of the masses. At last follow-up, metastatic disease had not developed in any patient, and no patient had died of renal cell carcinoma. Two patients had died of other causes. CONCLUSIONS: The results of our study showed that when comorbid conditions or patient wishes preclude invasive treatment, contrast-enhancing renal masses less than 3.5 cm wide that are suggestive of cancer can be safely managed with watchful waiting and serial computed tomography scans.

Prediction of genitourinary tract morbidity after brachytherapy for prostate adenocarcinoma.

OBJECTIVE: To investigate whether preoperative genitourinary variables in patients undergoing brachytherapy for localized prostate adenocarcinoma could predict postoperative genitourinary tract morbidity. PATIENTS AND METHODS: We retrospectively reviewed medical records of 105 men who received either iodine 125 or palladium 103 radioactive seed implants with or without external beam radiotherapy or hormone blockade from January 1, 1998, through December 31, 2000, at the Mayo Clinic in Jacksonville, Fla. Patients with one or more of the following were classified as having a high risk of postoperative genitourinary tract morbidity: American Urological Association symptom scores greater than 15, maximum urinary flow rate less than 10 mL/s, postvoid residual urinary volume greater than 100 mL, or prostate volume greater than 40 cm3. Of the 105 men, 59 (56%) were classified as high risk and 46 (44%) as low risk. Mean follow-up after brachytherapy was 23.6 months. Modified Radiation Therapy Oncology Group scores were used to assess postoperative genitourinary tract morbidity. The term significant genitourinary tract morbidity was applied to patients with a Radiation Therapy Oncology Group grade of 3 or 4 after at least 6 months of follow-up. RESULTS: Significant morbidity occurred in 37% of high-risk vs 15% of low-risk patients (P = .006). In high-risk patients, transurethral resection or incision of the prostate was required in 5 patients, urethral dilation in 4, direct-vision internal urethrotomy in 1, and placement of a suprapubic catheter in 1. In low-risk patients, transurethral incision of the prostate was required in only 1 patient. Urinary flow rate was a significant individual predictor of postoperative morbidity (P = .03). CONCLUSIONS: A combination of urinary flow rate, prostate volume, postvoid residual urinary volume, and American Urological Association symptom score can help identify patients with underlying voiding dysfunction. Urinary flow rate was a statistically significant predictor of genitourinary tract morbidity after brachytherapy for localized prostate adenocarcinoma. Patients and physicians should consider these factors before a patient decides to undergo brachytherapy.