CYP3A5 genotype and its impact on vincristine pharmacokinetics and development of neuropathy in Kenyan children with cancer.

BACKGROUND: Vincristine (VCR) is a critical part of treatment in pediatric malignancies and is associated with dose-dependent peripheral neuropathy (vincristine-induced peripheral neuropathy [VIPN]). Our previous findings show VCR metabolism is regulated by the CYP3A5 gene. Individuals who are low CYP3A5 expressers metabolize VCR slower and experience more severe VIPN as compared to high expressers. Preliminary observations suggest that Caucasians experience more severe VIPN as compared to nonCaucasians. PROCEDURE: Kenyan children with cancer who were undergoing treatment including VCR were recruited for a prospective cohort study. Patients received IV VCR 2 mg/m2 /dose with a maximum dose of 2.5 mg as part of standard treatment protocols. VCR pharmacokinetics (PK) sampling was collected via dried blood spot cards and genotyping was conducted for common functional variants in CYP3A5, multi-drug resistance 1 (MDR1), and microtubule-associated protein tau (MAPT). VIPN was assessed using five neuropathy tools. RESULTS: The majority of subjects (91%) were CYP3A5 high-expresser genotype. CYP3A5 low-expresser genotype subjects had a significantly higher dose and body surface area normalized area under the curve than CYP3A5 high-expresser genotype subjects (0.28 ± 0.15 hr·m2 /l vs. 0.15 ± 0.011 hr·m2 /l, P = 0.027). Regardless of which assessment tool was utilized, minimal neuropathy was detected in this cohort. There was no difference in the presence or severity of neuropathy assessed between CYP3A5 high- and low-expresser genotype groups. CONCLUSION: Genetic factors are associated with VCR PK. Due to the minimal neuropathy observed in this cohort, there was no demonstrable association between genetic factors or VCR PK with development of VIPN. Further studies are needed to determine the role of genetic factors in optimizing dosing of VCR for maximal benefit.

A novel role for an RNA polymerase III subunit POLR3G in regulating pluripotency in human embryonic stem cells.

The pluripotency of human embryonic stem cells (hESC) could have great potential for the development of cell replacement therapies. Previous studies have converged on the finding that OCT4, SOX2, and NANOG serve as key regulators in the maintenance of hESC. However, other signals that regulate hESC maintenance remain poorly studied. Here we describe a novel role of an RNA polymerase III (Pol III) subunit, POLR3G, in the maintenance of pluripotency in hESC. We demonstrate the presence of POLR3G in undifferentiated hESC, human induced pluripotent stem cells (hiPSC), and early mouse blastocysts. Downregulation of POLR3G is observed on differentiation of hESC and hiPSC, suggesting that POLR3G can be used as a molecular marker to readily identify undifferentiated pluripotent stem cells from their differentiated derivatives. Using an inducible shRNA lentiviral system, we found evidence that decreased levels of POLR3G result in loss of pluripotency and promote differentiation of hESC to all three germ layers but have no effect on cell apoptosis. On the other hand, overexpression of POLR3G has no effect on pluripotency and apoptosis in undifferentiated hESC. Interestingly, hESC expressing elevated levels of POLR3G are more resistant to differentiation. Furthermore, our experimental results show that POLR3G is a downstream target of OCT4 and NANOG, and our pharmacological study indicated that POLR3G expression can be readily regulated by the Erk1/2 signaling pathway. This study is the first to show an important role of POLR3G in the maintenance of hESC, suggesting a potential role of Pol III transcription in regulating hESC pluripotency.

Macromolecules involved in production and metabolism of beta-amyloid at the brain barriers.

One of the notable features of Alzheimer's disease (AD) is the overabundance of beta-amyloid peptides in brain fluids, leading to the formation and deposition of insoluble amyloid plaques. Previous work in this lab demonstrates that the normal choroid plexus, a primary component of the blood-cerebrospinal fluid barrier, has the capacity to remove beta-amyloid from the cerebrospinal fluid, potentially preventing the formation of beta-amyloid plaques. The purpose of this work was to determine whether the choroid plexus and/or the brain capillaries, a primary component of the blood-brain barrier, possessed the capacity to produce or degrade beta-amyloid peptides. Using quantitative real-time RT-PCR, immunodetection and enzyme activity assays, we demonstrated the presence in brain barriers of several key enzymes involved in beta-amyloid production, namely, amyloid precursor protein and beta-secretase, and in beta-amyloid metabolism and alternate processing, such as insulin degrading enzyme, endothelin-converting enzyme-1, neprilysin and alpha-secretase. Furthermore, beta-amyloid presence, in the absence of its application in culture media, was detected in an immortalized choroidal epithelial cell line, known as Z310 cells. The ability of the choroid plexus to produce and degrade beta-amyloid, in addition to its transport function, suggests a vital role of this tissue in maintaining beta-amyloid homeostasis. Disruption of this homeostasis due to aging, injury or toxicant exposure may contribute to accumulation of beta-amyloid peptides in the brain fluids, leading to AD.

Genomic scale profiling of nutrient and trace elements in Arabidopsis thaliana.

Understanding the functional connections between genes, proteins, metabolites and mineral ions is one of biology's greatest challenges in the postgenomic era. We describe here the use of mineral nutrient and trace element profiling as a tool to determine the biological significance of connections between a plant's genome and its elemental profile. Using inductively coupled plasma spectroscopy, we quantified 18 elements, including essential macro- and micronutrients and various nonessential elements, in shoots of 6,000 mutagenized M2 Arabidopsis thaliana plants. We isolated 51 mutants with altered elemental profiles. One mutant contains a deletion in FRD3, a gene known to control iron-deficiency responses in A. thaliana. Based on the frequency of elemental profile mutations, we estimate 2-4% of the A. thaliana genome is involved in regulating the plant's nutrient and trace element content. These results demonstrate the utility of elemental profiling as a useful functional genomics tool.

Parents' Analgesic Trade-Off Dilemmas: How Analgesic Knowledge Influences Their Decisions to Give Opioids.

OBJECTIVES: Ineffective analgesic decisions in the home may jeopardize the safety and comfort of children, yet little is known about factors influencing parental decisions. This study explored how parents' analgesic understanding influenced their hypothetical decisions to give opioids when faced with important trade-off dilemmas where pain and adverse drug event (ADE) symptoms were both present. METHODS: A total of 514 parents whose children required opioids after discharge completed surveys assessing their Gist ADE Understanding (ie, knowledge of opioid-related ADEs and their seriousness) and other comparative analgesic perceptions. Parents then made hypothetical decisions to give or withhold prescribed opioids to a postoperative child with varying pain levels and serious (oversedation) and nonserious (nausea) ADE symptoms. RESULTS: Gist ADE Understanding influenced decisions to withhold opioids for a child with nausea/vomiting (ß=0.85 [95% confidence interval (CI), 0.74-0.98]) or oversedation (ß=0.86 [95% CI, 0.77-0.97]), but not for one with no ADE (ß=1.00 [95% CI, 0.98-1.02]). However, while perceived higher seriousness of oversedation influenced withholding opioids when this ADE was present (mean difference=0.36 [95% CI, 0.11-0.61], P=0.005), knowledge that oversedation was possible did not by itself affect behavior (odds ratio=0.80 [95% CI, 0.50-1.29], P=0.362). DISCUSSION: These data suggest that gist understanding of ADE seriousness, not just its possible presence, is needed to facilitate safe analgesic decisions. Importantly, higher overall ADE understanding did not influence parents' opioid decisions in the presence of high pain and absence of ADEs. Thus, risk information about specific ADEs is unlikely to dissuade parents from efforts to manage pain but may improve their decisions if ADEs should occur.

Opioid-related adverse drug events: do parents recognize the signals?

OBJECTIVES: Evidence of unrelieved childhood pain, adverse drug events (ADE), and deaths suggest that parents may inadequately respond to pain and opioid-related ADE signals. This study examined parents' recognition and response to pain and ADE signals using both dynamic hypothetical scenarios and real at-home opioid decisions. MATERIALS AND METHODS: A total of 514 parents whose children required prescription opioids after discharge were surveyed. Parents made analgesic decisions for 4 hypothetical scenarios wherein the child's pain level and ADE symptoms were altered. After discharge, parents recorded their child's real pain levels, ADEs, and their analgesic decisions. Mixed-effects logistic regression examined the influence of pain and ADE signals on parents' opioid decisions. RESULTS: Pain intensity had a main effect on parents' hypothetical and real decisions to give opioids (P≤0.001). Nausea/vomiting influenced the decision to give the prescribed opioid dose (ß=-1.48 [95% confidence interval (CI): -1.78, -1.19], P<0.001) as did oversedation (ß=-1.02 [95% CI: -1.30, -0.75], P<0.001); however, parents were more likely to give the prescribed dose for oversedation than nausea/vomiting (odds ratio (OR) =1.53 [95% CI: 1.14, 2.05], P=0.005). Gastrointestinal effects were more likely to motivate a change in postdischarge opioid administration compared with other ADEs (OR=4.41 [95% CI: 1.91, 10.18], P<0.001), whereas sedation symptoms did not (OR=0.46 [95% CI: 0.21, 1.04], P=0.06). DISCUSSION: Findings demonstrated that many parents failed to withhold a prescribed opioid dose for oversedation, suggesting a lack of awareness regarding this potentially serious ADE. Strategies to improve parents' recognition of oversedation and its potential consequences are warranted to improve opioid safety.