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BH3 mimetics, including the BCL2/BCLXL/BCLw inhibitor navitoclax and MCL1 inhibitors S64315 and tapotoclax, have undergone clinical testing for a variety of neoplasms. Because of toxicities, including thrombocytopenia after BCLXLinhibition as well as hematopoietic, hepatic and possible cardiac toxicities after MCL1 inhibition, there is substantial interest in finding agents that can safely sensitize neoplastic cells to these BH3 mimetics. Building on the observation that BH3 mimetic monotherapy induces AMP kinase (AMPK) activation in multiple acute leukemia cell lines, we report that the AMPK inhibitors (AMPKis) dorsomorphin and BAY-3827 sensitize these cells to navitoclax or MCL1 inhibitors. Cell fractionation and phosphoproteomic analyses suggest that sensitization by dorsomorphin involves dephosphorylation of the proapoptotic BCL2 family member BAD at Ser75 and Ser99, leading BAD to translocate to mitochondria and inhibit BCLXL. Consistent with these results, BAD knockout or mutation to BAD S75E/S99E abolishes the sensitizing effects of dorsomorphin. Conversely, dorsomorphin synergizes with navitoclax or the MCL1 inhibitor S63845 to induce cell death in primary acute leukemia samples ex vivo and increases the antitumor effects of navitoclax or S63845 in several xenograft models in vivo with little or no increase in toxicity in normal tissues. These results suggest that AMPK inhibition can sensitize acute leukemia to multiple BH3 mimetics, potentially allowing administration of lower doses while inducing similar antineoplastic effects.

The authors are experienced in the design, installation, testing, safety of personnel, and commissioning of shore power projects, “connection/disconnection of ships to shore power during berthing at ports.” Herein, the authors provide their views of some of the issues that may require first-hand input to the industry professionals involved in these types of projects. Technical references and recommendations included in the article should enhance the reader’s knowledge of shore power projects mandated in numerous regions around the world at larger ports to enhance safety of operators, minimize air pollution in the vicinity of ports for health benefits, and produce a cleaner environment for the pubic.

How BAK and BAX induce mitochondrial outer membrane (MOM) permeabilization (MOMP) during apoptosis is incompletely understood. Here we have used molecular dynamics simulations, surface plasmon resonance, and assays for membrane permeabilization in vitro and in vivo to assess the structure and function of selected BAK subdomains and their derivatives. Results of these studies demonstrate that BAK helical regions α5 and α6 bind the MOM lipid cardiolipin. While individual peptides corresponding to these helical regions lack the full biological activity of BAK, tandem peptides corresponding to α4–α5, α5–α6, or α6–α7/8 can localize exogenous proteins to mitochondria, permeabilize liposomes composed of MOM lipids, and cause MOMP in the absence of the remainder of the BAK protein. Importantly, the ability of these tandem helices to induce MOMP under cell-free conditions is diminished by mutations that disrupt the U-shaped helix-turn-helix structure of the tandem peptides or decrease their lipid binding. Likewise, BAK-induced apoptosis in intact cells is diminished by CLS1gene interruption, which decreases mitochondrial cardiolipin content, or by BAKmutations that disrupt the U-shaped tandem peptide structure or diminish lipid binding. Collectively, these results suggest that BAK structural rearrangements during apoptosis might mobilize helices involved in specific protein-lipid interactions that are critical for MOMP.

We present a highly elastic strain gauge based on capacitive sensing of parallel, carbon nanotube-based percolation electrodes separated by a dielectric elastomer. The fabrication, relying on vacuum filtration of single-walled carbon nanotubes and hydrophobic patterning of silicone, is both rapid and inexpensive. We demonstrate reliable, linear performance over thousands of cycles at up to 100% strain with less than 3% variability and the highest reported gauge factor for a device of this class (0.99). We further demonstrate use of this sensor in a robotics context to transduce joint angles.

The BCL2 family of proteins regulates cellular life and death decisions. Among BCL2 family members, BH3-only proteins have critical roles by neutralizing antiapoptotic family members, as well as directly activating BAX and BAK. Despite widespread occurrence of BH3-only protein upregulation in response to various stresses, this process is rarely quantified. Moreover, it is unclear whether all BH3-only proteins are equipotent at inducing cell death. Here we show that BH3-only proteins increase as much as 15- to 20-fold after various treatments and define a parameter, termed BH3-only tolerance, which measures how many copies of a particular BH3-only protein can be expressed before the majority of cells in a population undergo apoptosis. We not only assess the relative contributions of anti- and proapoptotic BCL2 family members to BH3-only tolerance, but also illustrate how the study of this parameter can be used to understand cellular sensitivity to anticancer drugs and new combinations. These observations provide a new quantitative framework for assessing apoptotic susceptibility under various conditions.

Congenital heart disease (CHD) affects up to 1% of live births. Although a genetic etiology is indicated by an increased recurrence risk, sporadic occurrence suggests that CHD genetics is complex. Here, we show that hypoplastic left heart syndrome (HLHS), a severe CHD, is multigenic and genetically heterogeneous. Using mouse forward genetics, we report what is, to our knowledge, the first isolation of HLHS mutant mice and identification of genes causing HLHS. Mutations from seven HLHS mouse lines showed multigenic enrichment in ten human chromosome regions linked to HLHS. Mutations in Sap130 and Pcdha9, genes not previously associated with CHD, were validated by CRISPR–Cas9 genome editing in mice as being digenic causes of HLHS. We also identified one subject with HLHS with SAP130 and PCDHA13 mutations. Mouse and zebrafish modeling showed that Sap130 mediates left ventricular hypoplasia, whereas Pcdha9 increases penetrance of aortic valve abnormalities, both signature HLHS defects. These findings show that HLHS can arise genetically in a combinatorial fashion, thus providing a new paradigm for the complex genetics of CHD.

Many ecological processes depend on the regular rise and fall of water levels (WLs), and artificial manipulations to WL regimes can impair important ecosystem services. Previous research has suggested that differences in WL between late summer and early spring may alter the suitability of shoals used by Walleyes Sander vitreusfor spawning. Other species, such as the Yellow Perch Perca flavescens, are unlikely to be affected in the same way by WL fluctuations because their spawning requirements are quite different. We used 11–23 years of data from six northern Minnesota lakes to assess the effects of WL fluctuations on the abundances of young-of-the-year (age-0) Walleyes and Yellow Perch. In two lakes (Rainy Lake and Lake Kabetogama), a change in WL management occurred in 2000, after which these lakes saw increased age-0 Walleye abundance, while the other study lakes experienced decreases or no change. Rainy Lake and Lake Kabetogama also had increases in age-0 Yellow Perch, but another study lake did also. We used partial least-squares regression to assess whether WL metrics were associated with variation in age-0 Walleye and Yellow Perch abundances, but WL metrics were seldom associated with age-0 abundance for either species. Our analysis suggested a potential influence of WL regulation on age-0 Walleye abundance, but we found no evidence that early spring access to spawning shoals was the mechanism by which this occurred. Received August 19, 2015; accepted July 15, 2016 Published online November 10, 2016

AbstractMany ecological processes depend on the regular rise and fall of water levels (WLs), and artificial manipulations to WL regimes can impair important ecosystem services. Previous research has suggested that differences in WL between late summer and early spring may alter the suitability of shoals used by Walleyes Sander vitreusfor spawning. Other species, such as the Yellow Perch Perca flavescens, are unlikely to be affected in the same way by WL fluctuations because their spawning requirements are quite different. We used 11–23 years of data from six northern Minnesota lakes to assess the effects of WL fluctuations on the abundances of young-of-the-year (age-0) Walleyes and Yellow Perch. In two lakes (Rainy Lake and Lake Kabetogama), a change in WL management occurred in 2000, after which these lakes saw increased age-0 Walleye abundance, while the other study lakes experienced decreases or no change. Rainy Lake and Lake Kabetogama also had increases in age-0 Yellow Perch, but another study lake did also. We used partial least-squares regression to assess whether WL metrics were associated with variation in age-0 Walleye and Yellow Perch abundances, but WL metrics were seldom associated with age-0 abundance for either species. Our analysis suggested a potential influence of WL regulation on age-0 Walleye abundance, but we found no evidence that early spring access to spawning shoals was the mechanism by which this occurred.Received August 19, 2015; accepted July 15, 2016 Published online November 10, 2016

Approximately one-third of all mammalian genes are essential for life. Phenotypes resulting from knockouts of these genes in mice have provided tremendous insight into gene function and congenital disorders. As part of the International Mouse Phenotyping Consortium effort to generate and phenotypically characterize 5,000 knockout mouse lines, here we identify 410 lethal genes during the production of the first 1,751 unique gene knockouts. Using a standardized phenotyping platform that incorporates high-resolution 3D imaging, we identify phenotypes at multiple time points for previously uncharacterized genes and additional phenotypes for genes with previously reported mutant phenotypes. Unexpectedly, our analysis reveals that incomplete penetrance and variable expressivity are common even on a defined genetic background. In addition, we show that human disease genes are enriched for essential genes, thus providing a dataset that facilitates the prioritization and validation of mutations identified in clinical sequencing efforts.

Objective:To determine the effects of motivational interviewing (MI) counselling versus a minimal intervention control on the enrollment of smokers in the Minnesota QUITPLAN Helpline.Study design:Two-group randomized controlled trial of 235 community-dwelling adult smokers. Participants were randomly assigned to receive either a three-session phone based motivational interviewing intervention (n= 118) or the control (n= 117) consisting of mailed printed materials about the Minnesota QUITPLAN helpline services.Data collection:Participants completed demographic and smoking-related survey questionnaire at baseline (by mail) and at weeks 4 and 26 (by phone) follow-up. Quitline enrollment status data was provided by the Minnesota QUITPLAN Helpline.Principal findings:At week 4 follow-up, a higher proportion of participants in the MI group (22.3%) had enroled in the Minnesota QUITPLAN Helpline compared to those in the control arm (13.6%; p= 0.098). At week 26 follow-up, enrollment rates were similar for MI (28.0%) and control (26.5%) arms. Of those who enroled in the Helpline, participants in the MI group reported completing more Helpline sessions than those in control group (4.9 vs. 3.2; p= 0.087).There was no significant interaction between readiness to quit and intervention for the outcome of enrollment in the Helpline.Conclusions:A minimal intensity control such as mailing printed materials resulted in quitline enrollment rates similar to a more resource intensive intervention like motivational interviewing and several folds higher than the current state or national averages. We recommend that health plans should consider mailing smoking cessation promotional messages to encourage smokers to enrol in quitlines.

The mammalian target of rapamycin (mTOR), a kinase that regulates proliferation and apoptosis, has been extensively evaluated as a therapeutic target in multiple malignancies. Rapamycin analogs, which partially inhibit mTOR complex 1 (mTORC1), exhibit immunosuppressive and limited antitumor activity, but sometimes activate survival pathways through feedback mechanisms involving mTORC2. Thus, attention has turned to agents targeting both mTOR complexes by binding the mTOR active site. Here we show that disruption of either mTOR-containing complex is toxic to acute lymphocytic leukemia (ALL) cells and identify 2 previously unrecognized pathways leading to this cell death. Inhibition of mTORC1-mediated 4EBP1 phosphorylation leads to decreased expression of c-MYC and subsequent upregulation of the proapoptotic BCL2 family member PUMA, whereas inhibition of mTORC2 results in nuclear factor-κB–mediated expression of the Early Growth Response 1 (EGR1) gene, which encodes a transcription factor that binds and transactivates the proapoptotic BCL2L11 locus encoding BIM. Importantly, 1 or both pathways contribute to death of malignant lymphoid cells after treatment with dual mTORC1/mTORC2 inhibitors. Collectively, these observations not only provide new insight into the survival roles of mTOR in lymphoid malignancies, but also identify alterations that potentially modulate the action of mTOR dual inhibitors in ALL.

The mammalian target of rapamycin (mTOR), a kinase that regulates proliferation and apoptosis, has been extensively evaluated as a therapeutic target in multiple malignancies. Rapamycin analogs, which partially inhibit mTOR complex 1 (mTORC1), exhibit immunosuppressive and limited antitumor activity, but sometimes activate survival pathways through feedback mechanisms involving mTORC2. Thus, attention has turned to agents targeting both mTOR complexes by binding the mTOR active site. Here we show that disruption of either mTOR-containing complex is toxic to acute lymphocytic leukemia (ALL) cells and identify 2 previously unrecognized pathways leading to this cell death. Inhibition of mTORC1-mediated 4EBP1 phosphorylation leads to decreased expression of c-MYC and subsequent upregulation of the proapoptotic BCL2 family member PUMA, whereas inhibition of mTORC2 results in nuclear factor-κB–mediated expression of the Early Growth Response 1(EGR1) gene, which encodes a transcription factor that binds and transactivates the proapoptotic BCL2L11locus encoding BIM. Importantly, 1 or both pathways contribute to death of malignant lymphoid cells after treatment with dual mTORC1/mTORC2 inhibitors. Collectively, these observations not only provide new insight into the survival roles of mTOR in lymphoid malignancies, but also identify alterations that potentially modulate the action of mTOR dual inhibitors in ALL.

Signaling through the phosphatidylinositol-3 kinase (PI3K)/Akt pathway, which is aberrantly activated in >50% of carcinomas, inhibits apoptosis and contributes to drug resistance. Accordingly, several Akt inhibitors are currently undergoing preclinical or early clinical testing. To examine the effect of Akt inhibition on the activity of multiple widely used classes of antineoplastic agents, human cancer cell lines were treated with the Akt inhibitor A-443654 [(2S)-1-(1H-indol-3-yl)-3-[5-(3-methyl-2H-indazol-5-yl)pyridin-3-yl]oxypropan-2-amine; ATP-competitive] or MK-2206 (8-[4-(1-aminocyclobutyl)phenyl]-9-phenyl-2H-[1,2,4]triazolo[3,4-f][1,6]naphthyridin-3-one;dihydrochloride; allosteric inhibitor) or with small interfering RNA (siRNA) targeting phosphoinositide-dependent kinase 1 (PDK1) along with cisplatin, melphalan, camptothecin, or etoposide and assayed for colony formation. Surprisingly different results were observed when Akt inhibitors were combined with different drugs. Synergistic effects were observed in multiple cell lines independent of PI3K pathway status when A-443654 or MK-2206 was combined with the DNA cross-linking agents cisplatin or melphalan. In contrast, effects of the Akt inhibitors in combination with camptothecin or etoposide were more complicated. In HCT116 and DLD1 cells, which harbor activating PI3KCAmutations, A-443654 over a broad concentration range enhanced the effects of camptothecin or etoposide. In contrast, in cell lines lacking activating PI3KCAmutations, partial inhibition of Akt signaling synergized with camptothecin or etoposide, but higher A-443654 or MK-2206 concentrations (>80% inhibition of Akt signaling) or PDK1 siRNA antagonized the topoisomerase poisons by diminishing DNA synthesis, a process that contributes to effective DNA damage and killing by these agents. These results indicate that the effects of combining inhibitors of the PI3K/Akt pathway with certain classes of chemotherapeutic agents might be more complicated than previously recognized.

Canonical Wnt signaling supports the pluripotency of embryonic stem cells (ESCs) but also promotes differentiation of early mammalian cell lineages. To explain these paradoxical observations, we explored the gene regulatory networks at play. Canonical Wnt signaling is intertwined with the pluripotency network comprising Nanog, Oct4, and Sox2 in mouse ESCs. In defined media supporting the derivation and propagation of ESCs, Tcf3 and β‐catenin interact with Oct4; Tcf3 binds to Sox motif within Oct‐Sox composite motifs that are also bound by Oct4‐Sox2 complexes. Furthermore, canonical Wnt signaling upregulates the activity of the Pou5f1distal enhancer via the Sox motif in ESCs. When viewed in the context of published studies on Tcf3 and β‐catenin mutants, our findings suggest Tcf3 counters pluripotency by competition with Sox2 at these sites, and Tcf3 inhibition is blocked by β‐catenin entry into this complex. Wnt pathway stimulation also triggers β‐catenin association at regulatory elements with classic Lef/Tcf motifs associated with differentiation programs. The failure to activate these targets in the presence of a mitogen‐activated protein kinase kinase (MEK)/extracellular signal‐regulated kinase (ERK) inhibitor essential for ESC culture suggests MEK/ERK signaling and canonical Wnt signaling combine to promote ESC differentiation. StemCells2013;31:2667–2679

Analysis of large-scale human genomic data has yielded unexplained mutations known to cause severe disease in healthy individuals. Here, we report the unexpected recovery of a rare dominant lethal mutation in TPM1, a sarcomeric actin-binding protein, in eight individuals with large atrial septal defect (ASD) in a five-generation pedigree. Mice with Tpm1mutation exhibit early embryonic lethality with disrupted myofibril assembly and no heartbeat. However, patient-induced pluripotent-stem-cell-derived cardiomyocytes show normal beating with mild myofilament defect, indicating disease suppression. A variant in TLN2, another myofilament actin-binding protein, is identified as a candidate suppressor. Mouse CRISPR knock-in (KI) of both the TLN2and TPM1variants rescues heart beating, with near-term fetuses exhibiting large ASD. Thus, the role of TPM1in ASD pathogenesis unfolds with suppression of its embryonic lethality by protective TLN2variant. These findings provide evidence that genetic resiliency can arise with genetic suppression of a deleterious mutation.

We describe five patients with HIV-2 infection (four antiretroviral-experienced and one antiretroviral-naive) treated with a regimen containing raltegravir. All responded to treatment as demonstrated by viral load and CD4+T-cell count monitoring. Our series confirms the clinical effectiveness of raltegravir in HIV-2-infected patients when given with other antiretrovirals to which the virus is susceptible.

Palm sized legged robots show promise for military and civilian applications, including exploration of hazardous or difficult to reach places, search and rescue, espionage, and battlefield reconnaissance. However, they also face many technical obstacles, including- but not limited to- actuator performance, weight constraints, processing power, and power density. This paper presents an overview of several robots from the Biomimetic Millisystems Laboratory at UC Berkeley, including the OctoRoACH, a steerable, running legged robot capable of basic navigation and equipped with a camera and active tail; CLASH, a dynamic climbing robot; and BOLT, a hybrid crawling and flying robot. The paper also discusses, and presents some preliminary solutions to, the technical obstacles listed above plus issues such as robustness to unstructured environments, limited sensing and communication bandwidths, and system integration.

Many of the antiretrovirals used against HIV-1 are either ineffective or less effective in HIV-2 infection. There is in vitroevidence of the potency of maraviroc and several investigational agents against HIV-2. We conclude that, whilst specific boosted protease inhibitors combined with nucleoside analogues should still be considered the mainstays of HIV-2 treatment, maraviroc, T-1249, TAK-779 and AMD3100, as well as raltegravir, could contribute to regimens for treatment-experienced individuals. Factors bearing on the use and timing of these alternative agents are discussed.

Despite much interest in amniote systematics, the origin of turtles remains elusive. Traditional morphological phylogenetic analyses place turtles outside Diapsida—amniotes whose ancestor had two fenestrae in the temporal region of the skull (among the living forms the tuatara, lizards, birds and crocodilians)—and allied with some unfenestrate-skulled (anapsid) taxa. Nonetheless, some morphological analyses place turtles within Diapsida, allied with Lepidosauria (tuatara and lizards). Most molecular studies agree that turtles are diapsids, but rather than allying them with lepidosaurs, instead place turtles near or within Archosauria (crocodilians and birds). Thus, three basic phylogenetic positions for turtles with respect to extant Diapsida are currently debated: (i) sister to Diapsida, (ii) sister to Lepidosauria, or (iii) sister to, or within, Archosauria. Interestingly, although these three alternatives are consistent with a single unrooted four-taxon tree for extant reptiles, they differ with respect to the position of the root. Here, we apply a novel molecular dataset, the presence versus absence of specific microRNAs, to the problem of the phylogenetic position of turtles and the root of the reptilian tree, and find that this dataset unambiguously supports a turtle + lepidosaur group. We find that turtles and lizards share four unique miRNA gene families that are not found in any other organisms' genome or small RNA library, and no miRNAs are found in all diapsids but not turtles, or in turtles and archosaurs but not in lizards. The concordance between our result and some morphological analyses suggests that there have been numerous morphological convergences and reversals in reptile phylogeny, including the loss of temporal fenestrae.

The mammalian target of rapamycin (mTOR) plays crucial roles in proliferative and antiapoptotic signaling in lymphoid malignancies. Rapamycin analogs, which are allosteric mTOR complex 1 (mTORC1) inhibitors, are active in mantle cell lymphoma and other lymphoid neoplasms, but responses are usually partial and short-lived. In the present study we compared the effects of rapamycin with the dual mTORC1/mTORC2 inhibitor OSI-027 in cell lines and clinical samples representing divers lymphoid malignancies. In contrast to rapamycin, OSI-027 markedly diminished proliferation and induced apoptosis in a variety of lymphoid cell lines and clinical samples, including specimens of B-cell acute lymphocytic leukemia (ALL), mantle cell lymphoma, marginal zone lymphoma and Sezary syndrome. Additional analysis demonstrated that OSI-027–induced apoptosis depended on transcriptional activation of the PUMA and BIM genes. Overexpression of Bcl-2, which neutralizes Puma and Bim, or loss of procaspase 9 diminished OSI-027–induced apoptosis in vitro. Moreover, OSI-027 inhibited phosphorylation of mTORC1 and mTORC2 substrates, up-regulated Puma, and induced regressions in Jeko xenografts. Collectively, these results not only identify a pathway that is critical for the cytotoxicity of dual mTORC1/mTORC2 inhibitors, but also suggest that simultaneously targeting mTORC1 and mTORC2 might be an effective anti-lymphoma strategy in vivo.

The mammalian target of rapamycin (mTOR) plays crucial roles in proliferative and antiapoptotic signaling in lymphoid malignancies. Rapamycin analogs, which are allosteric mTOR complex 1 (mTORC1) inhibitors, are active in mantle cell lymphoma and other lymphoid neoplasms, but responses are usually partial and short-lived. In the present study we compared the effects of rapamycin with the dual mTORC1/mTORC2 inhibitor OSI-027 in cell lines and clinical samples representing divers lymphoid malignancies. In contrast to rapamycin, OSI-027 markedly diminished proliferation and induced apoptosis in a variety of lymphoid cell lines and clinical samples, including specimens of B-cell acute lymphocytic leukemia (ALL), mantle cell lymphoma, marginal zone lymphoma and Sezary syndrome. Additional analysis demonstrated that OSI-027–induced apoptosis depended on transcriptional activation of the PUMA and BIM genes. Overexpression of Bcl-2, which neutralizes Puma and Bim, or loss of procaspase 9 diminished OSI-027–induced apoptosis in vitro. Moreover, OSI-027 inhibited phosphorylation of mTORC1 and mTORC2 substrates, up-regulated Puma, and induced regressions in Jeko xenografts. Collectively, these results not only identify a pathway that is critical for the cytotoxicity of dual mTORC1/mTORC2 inhibitors, but also suggest that simultaneously targeting mTORC1 and mTORC2 might be an effective anti-lymphoma strategy in vivo.

A phase 2 study of the oral farnesyltransferase inhibitor tipifarnib was conducted in 93 adult patients with relapsed or refractory lymphoma. Patients received tipifarnib 300 mg twice daily on days 1-21 of each 28-day cycle. The median number of prior therapies was 5 (range, 1-17). For the aggressive B-cell, indolent B-cell, and T-cell and Hodgkin lymphoma (HL/T) groups, the response rates were 17% (7/42), 7% (1/15), and 31% (11/36), respectively. Of the 19 responders, 7 were diffuse large B-cell non-Hodgkin lymphoma (NHL), 7 T-cell NHL, 1 follicular grade 2, and 4 HL. The median response duration for the 19 responders was 7.2 months (mean, 15.8 months; range, 1.8-62), and 5 patients in the HL/T group are still receiving treatment at 29-64+ months. The grade 3/4 toxicities observed were fatigue and reversible myelosuppression. Correlative studies suggest that Bim and Bcl-2 should be examined as potential predictors of response in future studies. These results indicate that tipifarnib has activity in lymphoma, particularly in heavily pretreated HL/T types, with little activity in follicular NHL. In view of its excellent toxicity profile and novel mechanism of action, further studies in combination with other agents appear warranted. This trial is registered at www.clinicaltrials.govas #NCT00082888.

The mechanism of cytotoxicity of farnesyltransferase inhibitors is incompletely understood and seems to vary depending on the cell type. To identify potential determinants of sensitivity or resistance for study in the accompanying clinical trial (Witzig et al, page 4882), we examined the mechanism of cytotoxicity of tipifarnib in human lymphoid cell lines. Based on initial experiments showing that Jurkat variants lacking Fas-associated death domain or procaspase-8 undergo tipifarnib-induced apoptosis, whereas cells lacking caspase-9 or overexpressing Bcl-2 do not, we examined changes in Bcl-2 family members. Tipifarnib caused dose-dependent up-regulation of Bim in lymphoid cell lines (Jurkat, Molt3, H9, DoHH2, and RL) that undergo tipifarnib-induced apoptosis but not in lines (SKW6.4 and Hs445) that resist tipifarnib-induced apoptosis. Further analysis demonstrated that increased Bim levels reflect inhibition of signaling from c-Raf to MEK1/2 and ERK1/2. Additional experiments showed that down-regulation of the Ras guanine nucleotide exchange factor RasGRP1 diminished tipifarnib sensitivity, suggesting that H-Ras or N-Ras is a critical farnesylation target upstream of c-Raf in lymphoid cells. These results not only trace a pathway through c-Raf to Bim that contributes to tipifarnib cytotoxicity in human lymphoid cells but also identify potential determinants of sensitivity to this agent.

A phase 2 study of the oral farnesyltransferase inhibitor tipifarnib was conducted in 93 adult patients with relapsed or refractory lymphoma. Patients received tipifarnib 300 mg twice daily on days 1-21 of each 28-day cycle. The median number of prior therapies was 5 (range, 1-17). For the aggressive B-cell, indolent B-cell, and T-cell and Hodgkin lymphoma (HL/T) groups, the response rates were 17% (7/42), 7% (1/15), and 31% (11/36), respectively. Of the 19 responders, 7 were diffuse large B-cell non-Hodgkin lymphoma (NHL), 7 T-cell NHL, 1 follicular grade 2, and 4 HL. The median response duration for the 19 responders was 7.2 months (mean, 15.8 months; range, 1.8-62), and 5 patients in the HL/T group are still receiving treatment at 29-64+ months. The grade 3/4 toxicities observed were fatigue and reversible myelosuppression. Correlative studies suggest that Bim and Bcl-2 should be examined as potential predictors of response in future studies. These results indicate that tipifarnib has activity in lymphoma, particularly in heavily pretreated HL/T types, with little activity in follicular NHL. In view of its excellent toxicity profile and novel mechanism of action, further studies in combination with other agents appear warranted. This trial is registered at www.clinicaltrials.gov as #NCT00082888.

The mechanism of cytotoxicity of farnesyltransferase inhibitors is incompletely understood and seems to vary depending on the cell type. To identify potential determinants of sensitivity or resistance for study in the accompanying clinical trial (Witzig et al, page 4882), we examined the mechanism of cytotoxicity of tipifarnib in human lymphoid cell lines. Based on initial experiments showing that Jurkat variants lacking Fas-associated death domain or procaspase-8 undergo tipifarnib-induced apoptosis, whereas cells lacking caspase-9 or overexpressing Bcl-2 do not, we examined changes in Bcl-2 family members. Tipifarnib caused dose-dependent up-regulation of Bim in lymphoid cell lines (Jurkat, Molt3, H9, DoHH2, and RL) that undergo tipifarnib-induced apoptosis but not in lines (SKW6.4 and Hs445) that resist tipifarnib-induced apoptosis. Further analysis demonstrated that increased Bim levels reflect inhibition of signaling from c-Raf to MEK1/2 and ERK1/2. Additional experiments showed that down-regulation of the Ras guanine nucleotide exchange factor RasGRP1 diminished tipifarnib sensitivity, suggesting that H-Ras or N-Ras is a critical farnesylation target upstream of c-Raf in lymphoid cells. These results not only trace a pathway through c-Raf to Bim that contributes to tipifarnib cytotoxicity in human lymphoid cells but also identify potential determinants of sensitivity to this agent.

OBJECTIVE To assess the cross-sectional association of thiazolidinediones with diabetic macular edema (DME). METHODS The cross-sectional association of DME and visual acuity with thiazolidinediones was examined by means of baseline fundus photographs and visual acuity measurements from the Action to Control Cardiovascular Risk in Diabetes (ACCORD) trial. Visual acuity was assessed in 9690 participants in the ACCORD trial, and 3473 of these participants had fundus photographs that were centrally read in a standardized fashion by masked graders to assess DME and retinopathy from October 23, 2003, to March 10, 2006. RESULTS Among the subsample, 695 (20.0%) people had used thiazolidinediones, whereas 217 (6.2%) people had DME. Thiazolidinedione use was not associated with DME in unadjusted (odds ratio [OR], 1.01; 95% confidence interval [CI], 0.71-1.44; P = .95) and adjusted (OR, 0.97; 95% CI, 0.67-1.40; P = .86) analyses. Significant associations with DME were found for retinopathy severity (P < .001) and age (OR, 0.97; 95% CI, 0.952-0.997; P = .03) but not for hemoglobin A1c (P = .06), duration of diabetes (P = .65), sex (P = .72), and ethnicity (P = .20). Thiazolidinedione use was associated with slightly greater visual acuity (0.79 letter; 95% CI, 0.20-1.38; P = .009) of uncertain clinical significance. CONCLUSIONS In a cross-sectional analysis of data from the largest study to date, no association was observed between thiazolidinedione exposure and DME in patients with type 2 diabetes; however, we cannot exclude a modest protective or harmful association. TRIAL REGISTRATION clinicaltrials.gov Identifier: NCT00542178Arch Ophthalmol. 2010;128(3):312-318--

ABSTRACTDrug design, antiretroviral therapy (ART), and drug resistance studies have focused almost exclusively on human immunodeficiency virus type 1 (HIV-1), resulting in limited information for patients infected with HIV-2 and for those dually infected with HIV-1 and HIV-2. In this study, 20 patients, 12 infected with HIV-2 and 8 dually infected with HIV-1 and HIV-2, all treated with zidovudine (ZDV), lamivudine (3TC), and lopinavir-ritonavir (LPV/r), were followed up longitudinally for about 3 years. For 19/20 patients, viral loads were reduced to undetectable levels; the patient whose viral load remained detectable reported adverse effects associated with LPV/r that had caused him to stop taking all the drugs. HIV-2 strains containing mutations in both the protease and the reverse transcriptase gene that may confer drug resistance were observed in two patients with viral rebound, as early as 130 days (4.3 months) after the initiation of therapy. We conclude that the combination of ZDV, 3TC, and LPV/r is able to provide efficient and durable suppression of HIV-1 and HIV-2 for as long as 3 years in HIV-2-infected and dually infected patients. However, the emergence of HIV-1 and HIV-2 strains containing drug-resistant mutations can compromise the efficacy of this highly active ART.

Low phytic acid(lpa) crops have reductions in the amount of seed phytic acid (myo‐inositol hexakisphosphate, InsP6) and increases in inorganic phosphorus (Pi) with little change to the amount of total seed P. In this study, four barley lpagenotypes (lpa1–2, lpa2–1, lpa3–1, and M955), backcross wild‐type (wt) sib‐selections, and original parental line ‘Harrington’ were grown in the field over 2 yr. Developing seed was harvested once a week for 3 wk and then again at physiological maturity, and the seeds assayed for levels of total P, Pi, and phytic acid P. Total phosphorus concentration showed no consistent differences between the lpagenotypes and Harrington. Inorganic P declined during development in the wtgenotypes; however in lpagenotypes, inorganic P declined during the first few weeks of development, and then increased from 24 d to maturity. Phytic acid concentration increased steadily during development for the wtlines and barley lpa1–1 and lpa2–1, although the increase was much slower in the lpalines. The lpa3–1 and M955 had very little InsP6accumulation until later in development, with little to no increase in the amount of phytic acid in mature seed of M955 compared to the developing seeds of lpa3–1. This information is useful in understanding timing of phosphorus accumulation in seeds, as well as the nature of the low phytic acidmutation in seed development.

How complex body plans evolved in animals such as fruit flies and vertebrates, as compared to the relatively simple jellyfish and sponges, is not known, given the similarity of developmental genetic repertoires shared by all these taxa. Here, we show that a core set of 18 microRNAs (miRNAs), non‐coding RNA molecules that negatively regulate the expression of protein‐coding genes, are found only in protostomes and deuterostomes and not in sponges or cnidarians. Because many of these miRNAs are expressed in specific tissues and/or organs, miRNA‐mediated regulation could have played a fundamental evolutionary role in the origins of organs such as brain and heart—structures not found in cnidarians or sponges—and thus contributed greatly to the evolution of complex body plans. Furthermore, the continuous acquisition and fixation of miRNAs in various animal groups strongly correlates both with the hierarchy of metazoan relationships and with the non‐random origination of metazoan morphological innovations through geologic time. J. Exp. Zool. (Mol. Dev. Evol.) 306B: 2006. © 2006 Wiley‐Liss, Inc.

While the highly consistent gene order and axial colinear patterns of expression seem to be a feature of vertebrate hox gene clusters, this pattern may be less well conserved across the rest of the bilaterians. We report the first deuterostome instance of an intact hox cluster with a unique gene order where the paralog groups are not expressed in a sequential manner. The finished sequence from BAC clones from the genome of the sea urchin, Strongylocentrotus purpuratus, reveals a gene order wherein the anterior genes (Hox1, Hox2 and Hox3) lie nearest the posterior genes in the cluster such that the most 3′ gene is Hox5. (The gene order is 5′‐Hox1, 2, 3, 11/13c, 11/13b, 11/13a, 9/10, 8, 7, 6, 5‐3′.) The finished sequence result is corroborated by restriction mapping evidence and BAC‐end scaffold analyses. Comparisons with a putative ancestral deuterostome Hox gene cluster suggest that the rearrangements leading to the sea urchin gene order were many and complex. J. Exp. Zool. (Mol. Dev. Evol.) 304B, 2005. © 2005 Wiley‐Liss, Inc.

Numerous studies indicate a gap between evidence-based recommendations for care and clinical outcomes. Barriers reported by physicians include inefficiencies in data gathering. Information technology provides a potent solution to this problem. Diabetes registries are increasingly being utilized as a critical feature for population-based disease management. As an increasing number of organizations realize the need to choose among a plethora of different diabetes registry systems, we seek to identify important features in successful programs and address several of the critical features essential to an effective diabetes registry. A diabetes registry can be used to target high-risk patients, send reminders to patients, and provide physicians with feedback on their care, all important aspects of an optimal diabetes registry that can lead to significant improvements in clinical outcomes. As the healthcare system moves toward wider adoption of diabetes registries, key features of diabetes registry use are discussed.

Land application of manure can increase P transfer in runoff, although the risk depends in part on the characteristics of the manure. We assessed this for calcareous soils using manures from swine (Sus domesticus) fed one of five barley varieties (Hordeum vulgareL.), including four low phytate mutants and a normal variety, to produce manures with a range of total P (6.8–4.9 g P kg−1), water‐soluble P (4.3–8.0 g P kg−1), total N/P ratios (2.5:1–5.5:1), and total C/P ratios (31:1–67:1). Two experiments were conducted. First, manures were incorporated into three soils on a N (150 mg N kg−1soil) or P (27.5 mg P kg−1soil) basis three times during a 7‐wk incubation. Second, 10 additional soils were incubated for 2 wk following a single P‐based manure application (82.5 mg P kg−1soil). Water and NaHCO3(Olsen) extractable P were determined at regular intervals, with microbial P determined by fumigation–extraction after each incubation. For N‐based application (i.e., variable P amendment), extractable P increased with total P applied. For P‐based applications, the increase in soil P was more closely correlated to microbial P concentration than manure P composition or soil properties. These results suggest that stimulation of the microbial biomass by added organic C is important in determining soil P solubility following manure application.

About 75% of the total P in conventional soybean [Glycine max(L.) Merr.] seed is phytate P, which cannot be readily digested by nonruminant livestock, such as swine and poultry. The phytate P in soybean lines homozygous for the recessive alleles pha1and pha2is reduced to about 25% of the total P. The objective of this study was to determine the influence of low phytate (LP) on agronomic and seed traits of soybean. Three populations were developed by crossing three cultivars with normal phytate (NP) to the LP line CX1834‐1‐6. From each population, 10 LP and 10 NP lines were selected and grown in replicated tests at three Iowa environments during 2003. The mean total P of the LP and NP lines was not significantly different, but the mean phytate P, inorganic P, and other P were significantly different for the two types of lines in the three populations. The mean seedling emergence of the LP lines was 45% compared with 68% for the NP lines. The mean differences between the LP and NP lines for the other agronomic and seed traits were not significant in one or more of the populations. On the basis of these results, reduced seedling emergence will be a major factor to consider in the development of commercially viable cultivars with the pha1pha1pha2pha2genotype for LP.

In the present study, we report the finding of high concentrations of D-Asp (D-aspartate) in the retina of the cephalopods Sepia officinalis, Loligo vulgaris and Octopus vulgaris. D-Asp increases in concentration in the retina and optic lobes as the animal develops. In neonatal S. officinalis, the concentration of D-Asp in the retina is 1.8±0.2 μmol/g of tissue, and in the optic lobes it is 5.5±0.4 μmol/g of tissue. In adult animals, D-Asp is found at a concentration of 3.5±0.4 μmol/g in retina and 16.2±1.5 μmol/g in optic lobes (1.9-fold increased in the retina, and 2.9-fold increased in the optic lobes). In the retina and optic lobes of S. officinalis, the concentration of D-Asp, L-Asp (L-aspartate) and L-Glu (L-glutamate) is significantly influenced by the light/dark environment. In adult animals left in the dark, these three amino acids fall significantly in concentration in both retina (approx. 25% less) and optic lobes (approx. 20% less) compared with the control animals (animals left in a diurnal/nocturnal physiological cycle). The reduction in concentration is in all cases statistically significant (P=0.01–0.05). Experiments conducted in S. officinalis by using D-[2,3-3H]Asp have shown that D-Asp is synthesized in the optic lobes and is then transported actively into the retina. D-aspartate racemase, an enzyme which converts L-Asp into D-Asp, is also present in these tissues, and it is significantly decreased in concentration in animals left for 5 days in the dark compared with control animals. Our hypothesis is that the dicarboxylic amino acids, D-Asp, L-Asp and L-Glu, play important roles in vision.