Evaluation of the developmental toxicity of lenalidomide in rabbits

BACKGROUND: Lenalidomide, a thalidomide analog, is indicated for treatment of patients with deletion-5q myelodysplastic syndromes or multiple myeloma. NZW rabbits were used because of sensitivity to thalidomide's teratogenicity. METHODS: Range-finding and pulse-dosing studies preceded a full developmental toxicity study in New Zealand white (NZW) rabbits (25/group) given lenalidomide (0, 3, 10, or 20 mg/kg/day) or thalidomide (180 mg/kg/day) by stomach tube on gestation days (GD) 7-19. Clinical signs, body weights, and feed consumption were recorded daily from GD 7. On GD 29, standard maternal necropsy, uterine content, and fetal evaluations were carried out. RESULTS: In all studies, thalidomide was selectively toxic to development. In the pulse-dosing study, lenalidomide did not affect development at 100 mg/kg/day. Increases in C(max) and AUC(0-24 hr) values for lenalidomide were slightly less than dose-proportional; lenalidomide occurred in the fetuses. At 10 and 20 mg/kg/day, lenalidomide was maternally toxic (reduced body weight gain and feed consumption; at 20 mg/kg/day, weight loss and one abortion). Developmental toxicity at 10 and 20 mg/kg/day included reduced fetal body weights and increased postimplantation losses and fetal variations (morbidity/purple-discolored skin, undeveloped intermediate lung lobe, irregular nasal-frontal suture, and delayed metacarpal ossification). Thalidomide selectively reduced fetal body weight, increased postimplantation loss and caused characteristic limb and other dysmorphology. CONCLUSIONS: The maternal and developmental NOAELs for lenalidomide are 3 mg/kg/day. Unlike thalidomide, lenalidomide affected embryo-fetal development only at maternally toxic dosages, confirming that structure-activity relationships may not predict maternal or developmental effects. No fetal malformations were attributable to lenalidomide.     

Comparative genomic analysis of Lactobacillus mucosae LM1 identifies potential niche-specific genes and pathways for gastrointestinal adaptation

Lactobacillus mucosae is currently of interest as putative probiotics due to their metabolic capabilities and ability to colonize host mucosal niches. L. mucosae LM1 has been studied in its functions in cell adhesion and pathogen inhibition, etc. It demonstrated unique abilities to use energy from carbohydrate and non-carbohydrate sources. Due to these functions, we report the first complete genome sequence of an L. mucosae strain, L. mucosae LM1. Analysis of the pan-genome in comparison with closely-related Lactobacillus species identified a complete glycogen metabolism pathway, as well as folate biosynthesis, complementing previous proteomic data on the LM1 strain. It also revealed common and unique niche-adaptation genes among the various L. mucosae strains. The aim of this study was to derive genomic information that would reveal the probable mechanisms underlying the probiotic effect of L. mucosae LM1, and provide a better understanding of the nature of L. mucosae sp.     

Nematode ascaroside enhances resistance in a broad spectrum of plant–pathogen systems

Recognition of specific molecule signatures of microbes, including pathogens, induces innate immune responses in plants, as well as in animals. Analogously, a nematode pheromone, the ascaroside ascr#18, induces hallmark plant defences including activation of (a) mitogen‐activated protein kinases, (b) salicylic acid‐ and jasmonic acid‐mediated defence signalling pathways and (c) defence gene expression and provides protection to a broad spectrum of pathogens. Ascr#18 is a member of an evolutionarily conserved family of nematode signalling molecules and is the major ascaroside secreted by plant–parasitic nematodes. Here, we report the effects of ascr#18 on resistance in four of the major economically important crops: maize, rice, wheat and soybean to some of their associated pathogens. Treatment with low nanomolar to low micromolar concentrations of ascr#18 provided from partial to strong protection in seven of eight plant–pathogen systems tested with viruses, bacteria, fungi, oomycetes and nematodes. This research may have potential to improve agricultural sustainability by reducing use of potentially harmful agrochemicals and enhance food security worldwide.     

Redefining the role of Ca2+-permeable channels in photoreceptor degeneration using diltiazem

Hereditary degeneration of photoreceptors has been linked to over-activation of Ca²⁺-permeable channels, excessive Ca²⁺-influx, and downstream activation of Ca²⁺-dependent calpain-type proteases. Unfortunately, after more than 20 years of pertinent research, unequivocal evidence proving significant and reproducible photoreceptor protection with Ca²⁺-channel blockers is still lacking. Here, we show that both D- and L-cis enantiomers of the anti-hypertensive drug diltiazem were very effective at blocking photoreceptor Ca²⁺-influx, most probably by blocking the pore of Ca²⁺-permeable channels. Yet, unexpectedly, this block neither reduced the activity of calpain-type proteases, nor did it result in photoreceptor protection. Remarkably, application of the L-cis enantiomer of diltiazem even led to a strong increase in photoreceptor cell death. These findings shed doubt on the previously proposed links between Ca²⁺ and retinal degeneration and are highly relevant for future therapy development as they may serve to refocus research efforts towards alternative, Ca²⁺-independent degenerative mechanisms.Subject terms: Cell death in the nervous system, Ion channels in the nervous system, Molecular neuroscience     

Biological mechanisms of aging predict age‐related disease co‐occurrence in patients

Genetic, environmental, and pharmacological interventions into the aging process can confer resistance to multiple age‐related diseases in laboratory animals, including rhesus monkeys. These findings imply that individual mechanisms of aging might contribute to the co‐occurrence of age‐related diseases in humans and could be targeted to prevent these conditions simultaneously. To address this question, we text mined 917,645 literature abstracts followed by manual curation and found strong, non‐random associations between age‐related diseases and aging mechanisms in humans, confirmed by gene set enrichment analysis of GWAS data. Integration of these associations with clinical data from 3.01 million patients showed that age‐related diseases associated with each of five aging mechanisms were more likely than chance to be present together in patients. Genetic evidence revealed that innate and adaptive immunity, the intrinsic apoptotic signaling pathway and activity of the ERK1/2 pathway were associated with multiple aging mechanisms and diverse age‐related diseases. Mechanisms of aging hence contribute both together and individually to age‐related disease co‐occurrence in humans and could potentially be targeted accordingly to prevent multimorbidity.     

MaGIC: a program to generate targeted marker sets for genome-wide association studies

High-throughput genotyping technologies such as DNA pooling and DNA microarrays mean that whole-genome screens are now practical for complex disease gene discovery using association studies. Because it is currently impractical to use all available markers, a subset is typically selected on the basis of required saturation density. Restricting markers to those within annotated genomic features of interest (e.g., genes or exons) or within feature-rich regions, reduces workload and cost while retaining much information. We have designed a program (MaGIC) that exploits genome assembly data to create lists of markers correlated with other genomic features. Marker lists are generated at a user-defined spacing and can target features with a user-defined density. Maps are in base pairs or linkage disequilibrium units (LDUs) as derived from the International HapMap data, which is useful for association studies and fine-mapping. Markers may be selected on the basis of heterozygosity and source database, and single nucleotide polymorphism (SNP) markers may additionally be selected on the basis of validation status. The import function means the method can be used for any genomic features such as housekeeping genes, long interspersed elements (LINES), or Alu repeats in humans, and is also functional for other species with equivalent data. The program and source code is freely available at http://cogent.iop.kcl.ac.uk/MaGIC.cogx.     

Stability and transformations of bis-PNA/DNA triplex structural isomers

Structurally isomeric complexes formed between homopyrimidine bis-PNAs (T(2)JT(2)JT(4)-linker-T(4)CT(2)CT(2)) and single- and double-stranded DNA targets were investigated. These complexes are triplexes designated S1, S2 and S3 in order of increased mobility by polyacrylamide gel electrophoresis. It is shown that the S3 isomer is formed only on double-stranded DNA and possesses highest stability. Isomers S2 and S1 are formed upon binding of bis-PNA to double-stranded as well as to single-stranded DNA. It was found that the stability of the isomer S1 increases dramatically in the presence of excess single-stranded oligonucleotide complementary to the bis-PNA. The structure of the stabilized S1 isomer is proposed to consist of two bis-PNA/DNA triplexes. The relationship between the yield of the isomer S1 formed on single-stranded DNA and the bis-PNA concentration was investigated and a kinetic model of the formation of S1 is presented.     

Development of definitive endoderm from embryonic stem cells in culture

The cellular and molecular events regulating the induction and tissue-specific differentiation of endoderm are central to our understanding of the development and function of many organ systems. To define and characterize key components in this process, we have investigated the potential of embryonic stem (ES) cells to generate endoderm following their differentiation to embryoid bodies (EBs) in culture. We found that endoderm can be induced in EBs, either by limited exposure to serum or by culturing in the presence of activin A (activin) under serum-free conditions. By using an ES cell line with the green fluorescent protein (GFP) cDNA targeted to the brachyury locus, we demonstrate that endoderm develops from a brachyury(+) population that also displays mesoderm potential. Transplantation of cells generated from activin-induced brachyury(+) cells to the kidney capsule of recipient mice resulted in the development of endoderm-derived structures. These findings demonstrate that ES cells can generate endoderm in culture and, as such, establish this differentiation system as a unique murine model for studying the development and specification of this germ layer.     

G1 cell cycle progression and the expression of G1 cyclins are regulated by PI3K/AKT/mTOR/p70S6K1 signaling in human ovarian cancer cells

Ovarian cancer is one of the most common cancers among women. Recent studies demonstrated that the gene encoding the p110alpha catalytic subunit of phosphatidylinositol 3-kinase (PI3K) is frequently amplified in ovarian cancer cells. PI3K is involved in multiple cellular functions, including proliferation, differentiation, antiapoptosis, tumorigenesis, and angiogenesis. In this study, we demonstrate that the inhibition of PI3K activity by LY-294002 inhibited ovarian cancer cell proliferation and induced G(1) cell cycle arrest. This effect was accompanied by the decreased expression of G(1)-associated proteins, including cyclin D1, cyclin-dependent kinase (CDK) 4, CDC25A, and retinoblastoma phosphorylation at Ser(780), Ser(795), and Ser(807/811). Expression of CDK6 and beta-actin was not affected by LY-294002. Expression of the cyclin kinase inhibitor p16(INK4a) was induced by the PI3K inhibitor, whereas steady-state levels of p21(CIP1/WAF1) were decreased in the same experiment. The inhibition of PI3K activity also inhibited the phosphorylation of AKT and p70S6K1, but not extracellular regulated kinase 1/2. The G(1) cell cycle arrest induced by LY-294002 was restored by the expression of active forms of AKT and p70S6K1 in the cells. Our study shows that PI3K transmits a mitogenic signal through AKT and mammalian target of rapamycin (mTOR) to p70S6K1. The mTOR inhibitor rapamycin had similar inhibitory effects on G(1) cell cycle progression and on the expression of cyclin D1, CDK4, CDC25A, and retinoblastoma phosphorylation. These results indicate that PI3K mediates G(1) progression and cyclin expression through activation of an AKT/mTOR/p70S6K1 signaling pathway in the ovarian cancer cells.