Combined targeting of MEK and PI3K/mTOR effector pathways is necessary to effectively inhibit NRAS mutant melanoma in vitro and in vivo.

Activating mutations in the neuroblastoma rat sarcoma viral oncogene homolog (NRAS) gene are common genetic events in malignant melanoma being found in 15-25% of cases. NRAS is thought to activate both mitogen activated protein kinase (MAPK) and PI3K signaling in melanoma cells. We studied the influence of different components on the MAP/extracellular signal-regulated (ERK) kinase (MEK) and PI3K/mammalian target of rapamycin (mTOR)-signaling cascade in NRAS mutant melanoma cells. In general, these cells were more sensitive to MEK inhibition compared with inhibition in the PI3K/mTOR cascade. Combined targeting of MEK and PI3K was superior to MEK and mTOR1,2 inhibition in all NRAS mutant melanoma cell lines tested, suggesting that PI3K signaling is more important for cell survival in NRAS mutant melanoma when MEK is inhibited. However, targeting of PI3K/mTOR1,2 in combination with MEK inhibitors is necessary to effectively abolish growth of NRAS mutant melanoma cells in vitro and regress xenografted NRAS mutant melanoma. Furthermore, we showed that MEK and PI3K/mTOR1,2 inhibition is synergistic. Expression analysis confirms that combined MEK and PI3K/mTOR1,2 inhibition predominantly influences genes in the rat sarcoma (RAS) pathway and growth factor receptor pathways, which signal through MEK/ERK and PI3K/mTOR, respectively. Our results suggest that combined targeting of the MEK/ERK and PI3K/mTOR pathways has antitumor activity and might serve as a therapeutic option in the treatment of NRAS mutant melanoma, for which there are currently no effective therapies.

Detection of GNAQ mutations and reduction of cell viability in uveal melanoma cells with functionalized gold nanoparticles.

BACKGROUND: Uveal melanoma (UM) is the most common primary intraocular malignancy in adults. Early treatment may improve any chances of preventing metastatic disease, but diagnosis of small UM is challenging. Up to 95 % of all UMs carry somatic mutations in the G-coupled proteins GNAQ and GNA11 promoting anchorage-independent growth and proliferation. About 50 % of UMs are fatal. Once metastatic, patients have limited options for successful therapy. METHODS: We have developed functionalized gold nanoparticles (AuNPs) to visualize transcripts of mutant GNAQ mRNA in living cells. In addition to their suitability as a specific tool for GNAQ mutation detection, we have developed a novel linker that enables conjugation of siRNAs to AuNPs allowing for greater and more rapid intracellular release of siRNAs compared to previously described approaches. RESULTS: Binding of modified AuNPs to matching target mRNA leads to conformational changes, resulting in a detectable fluorescent signal that can be used for mutation detection in living cells. Knockdown of GNAQ with siRNA-AuNPs effectively reduced downstream signals and decreased cell viability in GNAQ mutant uveal melanoma cells. CONCLUSION: AuNPs may in future be developed to serve as sensors for mutations of vital importance. The new release system for siRNA-AuNP improves previous systems, which conceivably will be useful for future therapeutic gene regulatory approaches.

Setting, Structure, and Timing of the Preparticipation Examination: The Wilderness Adventure Consultation.

Patients pursue wilderness experiences throughout the entire life cycle, and while outdoor pursuits are relatively safe, injuries do occur. Many of these adverse events can be anticipated, identified, and prevented through a wilderness preparticipation examination (PPE). To accomplish this, it is incumbent on the physician to assess the extrinsic and intrinsic factors faced by the patient and attempt to correct them to ensure an enjoyable experience in the outdoors. This article outlines the goals of the PPE along with identification of various risk factors that can influence a trip. Most injuries and rescues occur from underestimating the risks from extrinsic, environmental factors, and/or overestimating one's intrinsic skills. By matching the patient's fitness and skill level to the environment, the physician can help reduce the risk of serious injury.

Setting, Structure, and Timing of the Preparticipation Examination: The Wilderness Adventure Consultation.

Patients pursue wilderness experiences throughout the entire life cycle, and while outdoor pursuits are relatively safe, injuries do occur. Many of these adverse events can be anticipated, identified, and prevented through a wilderness preparticipation examination (PPE). To accomplish this, it is incumbent on the physician to assess the extrinsic and intrinsic factors faced by the patient and attempt to correct them to ensure an enjoyable experience in the outdoors. This article outlines the goals of the PPE along with identification of various risk factors that can influence a trip. Most injuries and rescues occur from underestimating the risks from extrinsic, environmental factors, and/or overestimating one's intrinsic skills. By matching the patient's fitness and skill level to the environment, the physician can help reduce the risk of serious injury.

Doping control for the team physician: a review of drug testing procedures in sport.

Drug testing is now ubiquitous in sport, and it often falls to the team physician to perform a variety of roles including interpreting test results, designing drug-testing programs, acting as medical review officer, and providing therapeutic use exemptions, education, and counseling. Proper understanding of current testing methods for drugs such as anabolic-androgenic steroids, erythropoietin, and growth hormone is essential if the team physician is going to assume these positions. This article outlines the basics of athletic drug testing from the collection process through the interpretation of results to assist the team physician in this field.

Mild traumatic brain injury in major and Minor League Baseball players.

BACKGROUND: Although mild traumatic brain injury (MTBI) is not as common in professional baseball as in collision sports, it does occur and frequently results in significant loss of time away from the sport. To date, no study has investigated MTBI among an entire cohort of professional baseball players. PURPOSE: To investigate MTBIs in major and minor league baseball players to determine the most common mechanisms of injury, activity at time of injury, position, level of play, and time lost, as well as ultimately inform prevention efforts. A secondary objective was to document the association between MTBI and return to play using several different measures. STUDY DESIGN: Descriptive epidemiologic study. METHODS: Data were captured from a newly implemented league-wide injury surveillance system that records injuries among all professional baseball players as entered by certified athletic trainers and physicians. The MTBIs were identified with respect to level of play, activity, field location, and mechanism of injury. Time loss was assessed by 3 measures of return to play, and MTBI game rates were reported as injuries per 1000 athlete-exposures. Data were combined over the 2011-2012 seasons for analysis, and results were presented separately for minor and major league players. Chi-square tests were used to test the hypothesis of equal proportions between the various categories of MTBI injury characteristics. RESULTS: There were 41 reported MTBIs in the major leagues and 266 in the minor leagues over the 2-year period under study. The overall MTBI game rate across both major and minor league ball clubs was 0.42 per 1000 athlete-exposures. The median time lost was 9 days. Mild traumatic brain injury accounted for 1% of all injuries resulting in time lost from play. For MTBIs that occurred while fielding, catchers were significantly overrepresented. No differences were noted among the 3 measures of time lost. CONCLUSION: Mild traumatic brain injury is an important problem in professional baseball players, especially for catchers. This study provides a foundation for future inquiry to reduce the incidence of MTBI in those positions at greatest risk and to provide a baseline as rules and equipment evolve.

Performance-enhancing drugs: understanding the risks.

To help clinicians understand the risks associated with performance-enhancing drugs, this overview covers prohibited lists of substances and methods, therapeutic use exemptions, the legitimate indications and adverse effects, including for megadose and polypharmacy doping of stimulants, anabolic steroids, erythropoiesis-stimulating agents, and growth hormone and ways in which physicians or patients risk committing anti-doping rule violations inadvertently.

Performance-enhancing drug use.

According to Gary Green, MD, everyone who deals with athletes is a "stakeholder" in the issue of performance-enhancing drugs and can influence athletes in a positive or negative role. In this issue of ORTHOPEDICS, Dr Green shares his thoughts on testing, prevention, and education of performance-enhancing drug use.

Court of Dreams.

This article was written in May 1992, after Magic Johnson announced he had tested positive for the human immunodeficiency virus that causes AIDS and was retiring from the Los Angeles Lakers. Since then, Johnson played in the 1992 Summer Olympic Games and, after a brief return to the Lakers, permanently retired from the NBA.

Detection of recombinant human erythropoietin in urine by isoelectric focusing.

BACKGROUND: Doping with erythropoietic proteins such as recombinant human erythropoietin (rHuEPO) and darbepoetin alfa is a serious issue in sport. There is little information on the time course of detection of rHuEPO in urine and on methods to evaluate electrophoresis-based data. METHODS: We used a recently described isoelectric focusing method for detecting rHuEPO and endogenous EPO in urine obtained from individuals treated with placebo or epoetin alfa. The latter was administered subcutaneously at 50 IU/kg on days 0, 2, 4, 7, 9, 11, 14, 16, and 18. Blood and urine samples were collected during the morning of study days -3, 0, 2, 4, 7, 9, 11, 14, 16, and 18 and on days 2, 3, 4, and 7 postadministration. We developed visual and numerical (two-band ratio) techniques to evaluate the electropherograms for the presence of rHuEPO. RESULTS: Compared with the placebo group, the epoetin alfa-treated group responded with increases in hematocrit, reticulocytes, macrocytes, serum EPO, and serum soluble transferrin receptor. The electropherograms showed that the pattern of bands arising from urinary rHuEPO is different from that of endogenous urinary EPO. Both the two-band ratio and the visual technique detected rHuEPO in all 14 epoetin alfa-treated individuals 3 days after the last dose. On the 7th day after the last dose, both techniques detected rHuEPO in approximately one-half of the participants. rHuEPO was not detected in the placebo-treated individuals. CONCLUSIONS: The isoelectric focusing method detects rHuEPO in most urine samples collected 3 days after nine doses of epoetin alfa. The numerical two-band ratio was equivalent to a visual method for detecting rHuEPO in urine.