A rapid and robust tri-color flow cytometry assay for monitoring malaria parasite development

Microscopic examination of Giemsa-stained thin blood smears remains the gold standard method used to quantify and stage malaria parasites. However, this technique is tedious, and requires trained microscopists. We have developed a fast and simple flow cytometry method to quantify and stage, various malaria parasites in red blood cells in whole blood or in vitro cultured Plasmodium falciparum. The parasites were stained with dihydroethidium and Hoechst 33342 or SYBR Green I and leukocytes were identified with an antibody against CD45. Depending on the DNA stains used, samples were analyzed using different models of flow cytometers. This protocol, which does not require any washing steps, allows infected red blood cells to be distinguished from leukocytes, as well as allowing non-infected reticulocytes and normocytes to be identified. It also allows assessing the proportion of parasites at different developmental stages. Lastly, we demonstrate how this technique can be applied to antimalarial drug testing

CPE Hepatitis C therapy: Looking toward interferon-sparing regimens

Abstract Objective: To describe chronic hepatitis C virus (HCV) infection, including its epidemiology and pathophysiology; review current treatment options for HCV infection; recognize investigational agents being studied as part of interferon-free therapy; and summarize clinical trials for the new agents. Data sources: PubMed for 2004 through August 2014 using search terms hepatitis C, American Association for the Study of Liver Diseases, sofosbuvir, simeprevir, and as needed specific names of other agents in development during this time; news articles and news releases about company actions with regard to clinical trials and filings for marketing approval in the United States. Study selection: At the discretion of the author based on clinical relevance of study and relevance to national guidelines for HCV therapy. Results: HCV infection is an important medical and public health problem in the United States and worldwide that can cause cirrhosis, hepatocellular carcinoma, and liver failure. The advent of newly developed targeted therapies is changing the treatment paradigm for this disease. Although traditional therapy with pegylated interferon and ribavirin remain therapeutic options, direct-acting agents such as sofosbuvir (Sovaldi-Gilead) and simeprevir (Olysio-Janssen) are producing faster, earlier, and improved treatment response with fewer adverse effects. The combination of anti-HCV agents and the duration of treatment are based on genotype, patient treatment status, and patient risk factors. The dramatic and sustained clearance of the virus with these drugs makes sustained virologic response a reality for patients who are unable to tolerate pegylated interferon. The downside is their high cost, which may make them economically unsustainable. However, for patients infected with HCV, the potential for a cure and improved quality of life may now be a reality. Conclusion: HCV, a well-known blood-borne disease associated with significant morbidity and mortality worldwide, can be effectively and safely treated with new anti-HCV agents such as SOF. While these new medications are in their early days of real-world practice, they offer hope that cure is truly possible. Learning objectives At the conclusion of this knowledge-based activity, the pharmacist will be able to: ■ Describe chronic hepatitis C virus (HCV) infection, including its epidemiology and pathophysiology. ■ Review current treatment options for HCV infection. ■ Recognize investigational agents being studied as part of interferon-free therapy. ■ Summarize clinical trials for the new agents. There is no fee associated with this activity for members of the American Pharmacists Association. There is a $25 fee for nonmembers. Accreditation informatio

Program in Personalized Health Care

Abstract: Background: The increasing number of available cancer therapies render medical decision-making (MDM) less straightforward. Patients want to know about the outcomes of similarly treated patients. Objective: The goal of this study was to design a breast cancer dashboard (BCD) tool that presents survival information to support MDM activities. Methods: Clinical variables during the clinic visit were determined via provider meetings and evaluated for accessibility from medical databases. Women with breast cancer (BC) were interviewed about their health care experiences after cancer diagnosis. We created a cohort of BC adult women treated at our institution from 1995 to 2012, from which clinical scenarios were defined and used to test survival outcomes. For the BCD, a simple, graphical user interface was built to present point-of-care clinical and survival data. Results: It is feasible to build the BCD using our institution's databases and generate survival plots to facilitate MDM activities. Patients with early-stage BC had the highest survival rate (82.3%) and the longest mean life years of 7.0 (SD 4.5) years. In late-stage BC, poor prognosis outweighs the influence of number of comorbidities on mortality. The BCD tool promotes more predictive, personalized, and collaborative health care. ABOUT THE AUTHORS The authors work across the University of Utah Health Sciences in oncology clinical practice, bioinformatics, and pharmacotherapy outcomes research. Our work is aimed at improving patient care via outcomes research and assessment. The Center's personnel have expertise in health economics, modeling, various clinical subspecialties (including oncology), drug information, statistical analysis and programming, and database management. PUBLIC INTEREST STATEMENT As more breast cancer treatment options become available, the shared medical decision-making relationship between physician and patient is becoming less straightforward. To support this important interaction, we have created an institution-specific medical communication tool. Decision aids have been shown to improve the health care experience of patients and ultimately lead to the achievement of higher-quality decisions. Our communication tool, named the breast cancer dashboard (BCD), was designed to be used during the clinic visit. It specifically address patients' needs to know about the survival outcomes of similar patients treated at our cancer specialty hospital. The BCD brings together comprehensive and breast cancerspecific clinical information. It allows both the provider and patient to navigate the information using a patient-friendly graphic interface. By enhancing shared decision-making, there would be a shift toward patient care that is more predictive, preventive, personalized, and collaborative

Predictive genomic markers of response to VEGF targeted therapy in metastatic renal cell carcinoma.

BACKGROUND:First-line treatment for metastatic renal cell carcinoma (mRCC) is rapidly changing. It currently includes VEGF targeted therapies (TT), multi-target tyrosine kinase inhibitors (TKIs), mTOR inhibitors, and immunotherapy. To optimize outcomes for individual patients, genomic markers of response to therapy are needed. Here, we aim to identify tumor-based genomic markers of response to VEGF TT to optimize treatment selection. METHODS:From an institutional database, primary tumor tissue was obtained from 79 patients with clear cell mRCC, and targeted sequencing was performed. Clinical outcomes were obtained retrospectively. Progression-free survival (PFS) on first-line VEGF TT was correlated to genomic alterations (GAs) using Kaplan-Meier methodology and Cox proportional hazard models. A composite model of significant GAs predicting PFS in the first-line setting was developed. RESULTS:Absence of VHL mutation was associated with inferior PFS on first-line VEGF TT. A trend for inferior PFS was observed with GAs in TP53 and FLT1 C/C variant. A composite model of these 3 GAs was associated with inferior PFS in a dose-dependent manner. CONCLUSION:In mRCC, a composite model of TP53 mutation, wild type VHL, and FLT1 C/C variant strongly predicted PFS on first-line VEGF TT in a dose-dependent manner. These findings require external validation

Phospholipid Membrane-Mediated Hemozoin Formation: The Effects of Physical Properties and Evidence of Membrane Surrounding Hemozoin

Phospholipid membranes are thought to be one of the main inducers of hemozoin formation in Plasmodia and other blood-feeding parasites. The "membrane surrounding hemozoin" has been observed in infected cells but has not been observed in in vitro experiments. This study focused on observing the association of phospholipid membranes and synthetic β-hematin, which is chemically identical to hemozoin, and on a further exploration into the mechanism of phospholipid membrane-induced β-hematin formation. Our results showed that β-hematin formation was induced by phospholipids in the fluid phase but not in the gel phase. The ability of phospholipids to induce β-hematin formation was inversely correlated with gel-to-liquid phase transition temperatures, suggesting an essential insertion of heme into the hydrocarbon chains of the phospholipid membrane to form β-hematin. For this study, a cryogenic transmission electron microscope was used to achieve the first direct observation of the formation of a monolayer of phospholipid membrane surrounding β-hematin