External validation of Adjuvant! Online breast cancer prognosis tool. Prioritising recommendations for improvement

Background

Adjuvant! Online is a web-based application designed to provide 10 years survival probability of patients with breast cancer. Several predictors have not been assessed in the original Adjuvant! Online study. We provide the validation of Adjuvant! Online algorithm on two breast cancer datasets, and we determined whether the accuracy of Adjuvant! Online is improved with other well-known prognostic factors.

Patients and Methods

The French data set is composed of 456 women with early breast cancer. The Dutch data set is composed of 295 women less than 52 years of age. Agreement between observation and Adjuvant! Online prediction was checked, and logistic models were performed to estimate the prognostic information added by risk factors to Adjuvant! Online prediction.

Results

Adjuvant! Online prediction was overall well-calibrated in the French data set but failed in some subgroups of such high grade and HER2 positive patients. HER2 status, Mitotic Index and Ki67 added significant information to Adjuvant! Online prediction. In the Dutch data set, the overall 10-year survival was overestimated by Adjuvant! Online, particularly in patients less than 40 years old.

Conclusion

Adjuvant! Online needs to be updated to adjust overoptimistic results in young and high grade patients, and should consider new predictors such as Ki67, HER2 and Mitotic Index.     

Lack of association of CD36 SNPs with early onset obesity: a meta-analysis in 9,973 European subjects

A recent study suggested that four CD36 polymorphisms (namely rs3211867, rs3211883, rs3211908, and rs1527483) were associated with an increased risk of obesity, an increased BMI and percentage of body fat in European adolescents. We first attempted to confirm these results in three independent case-control genome-wide association studies (GWAS) data totaling 3,509 subjects of French and German origin, but we were unable to find any association of these variants with early onset obesity risk. We then genotyped the four CD36 single-nucleotide polymorphisms (SNPs) in a large population-based study of 4,667 Finnish subjects and we did not replicate any of the recently reported associations with BMI. By combining all available data in a meta-analysis (N = 9,973), we found no evidence for an association of the reported four variants in CD36 with increased obesity risk or increased BMI (0.07 ≤ P values ≤ 0.93). Finally, we assessed the contribution of the full CD36 locus gene variation to obesity risk in 3,509 subjects and we did not detect any significant association with obesity after correction for multiple testing. In summary, we were unable to confirm the recently reported association of variants in CD36 with early onset obesity in populations of European ancestry.     

Freezing tolerance of the European water frogs: the good, the bad, and the ugly

Survival and some physiological responses to freezing were investigated in three European water frogs (Rana lessonae, Rana ridibunda, and their hybridogen Rana esculenta). The three species exhibited different survival times during freezing (from 10 h for R. lessonae to 20 h for R. ridibunda). The time courses of percent water frozen were similar; however, because of the huge differences in body mass among species (from 10 g for Rana lessonae to nearly 100 g for Rana ridibunda), the ice mass accumulation rate varied markedly (from 0.75 +/- 0.12 to 1.43 +/- 0.11 g ice/h, respectively) and was lowest in the terrestrial hibernator Rana lessonae. The hybrid Rana esculenta exhibited an intermediate response between the two parental species; furthermore, within-species correlation existed between body mass and ice mass accumulation rates, suggesting the occurrence of subpopulations in this species (0.84 +/- 0.08 g ice/h for small R. esculenta and 1.78 +/- 0.09 g ice/h for large ones). Biochemical analyses showed accumulation of blood glucose and lactate, liver glucose (originating from glycogen), and liver alanine in Rana lessonae and Rana esculenta but not in Rana ridibunda in response to freezing. The variation of freeze tolerance between these three closely related species could bring understanding to the physiological processes involved in the evolution of freeze tolerance in vertebrates.     

Disruption of the langerin/CD207 gene abolishes Birbeck granules without a marked loss of Langerhans cell function

Langerin is a C-type lectin expressed by a subset of dendritic leukocytes, the Langerhans cells (LC). Langerin is a cell surface receptor that induces the formation of an LC-specific organelle, the Birbeck granule (BG). We generated a langerin(-/-) mouse on a C57BL/6 background which did not display any macroscopic aberrant development. In the absence of langerin, LC were detected in normal numbers in the epidermis but the cells lacked BG. LC of langerin(-/-) mice did not present other phenotypic alterations compared to wild-type littermates. Functionally, the langerin(-/-) LC were able to capture antigen, to migrate towards skin draining lymph nodes, and to undergo phenotypic maturation. In addition, langerin(-/-) mice were not impaired in their capacity to process native OVA protein for I-A(b)-restricted presentation to CD4(+) T lymphocytes or for H-2K(b)-restricted cross-presentation to CD8(+) T lymphocytes. langerin(-/-) mice inoculated with mannosylated or skin-tropic microorganisms did not display an altered pathogen susceptibility. Finally, chemical mutagenesis resulted in a similar rate of skin tumor development in langerin(-/-) and wild-type mice. Overall, our data indicate that langerin and BG are dispensable for a number of LC functions. The langerin(-/-) C57BL/6 mouse should be a valuable model for further functional exploration of langerin and the role of BG.     

Evolution of Plant-Like Crystalline Storage Polysaccharide in the Protozoan Parasite Toxoplasma gondii Argues for a Red Alga Ancestry

Single-celled apicomplexan parasites are known to cause major diseases in humans and animals including malaria, toxoplasmosis, and coccidiosis. The presence of apicoplasts with the remnant of a plastid-like DNA argues that these parasites evolved from photosynthetic ancestors possibly related to the dinoflagellates. Toxoplasma gondii displays amylopectin-like polymers within the cytoplasm of the dormant brain cysts. Here we report a detailed structural and comparative analysis of the Toxoplasma gondii, green alga Chlamydomonas reinhardtii, and dinoflagellate Crypthecodinium cohnii storage polysaccharides. We show Toxoplasma gondii amylopectin to be similar to the semicrystalline floridean starch accumulated by red algae. Unlike green plants or algae, the nuclear DNA sequences as well as biochemical and phylogenetic analysis argue that the Toxoplasma gondii amylopectin pathway has evolved from a totally different UDP-glucose-based metabolism similar to that of the floridean starch accumulating red alga Cyanidioschyzon merolae and, to a lesser extent, to those of glycogen storing animals or fungi. In both red algae and apicomplexan parasites, isoamylase and glucan–water dikinase sequences are proposed to explain the appearance of semicrystalline starch-like polymers. Our results have built a case for the separate evolution of semicrystalline storage polysaccharides upon acquisition of photosynthesis in eukaryotes.This article contains online-only supplementary material.Reviewing Editor:Dr. Patrick Keeling     

Flash photolysis using a light emitting diode: an efficient, compact, and affordable solution

Flash photolysis has become an essential technique for dynamic investigations of living cells and tissues. This approach offers several advantages for instantly changing the concentration of bioactive compounds outside and inside living cells with high spatial resolution. Light sources for photolysis need to deliver pulses of high intensity light in the near UV range (300-380 nm), to photoactivate a sufficient amount of molecules in a short time. UV lasers are often required as the light source, making flash photolysis a costly approach. Here we describe the use of a high power 365 nm light emitting diode (UV LED) coupled to an optical fiber to precisely deliver the light to the sample. The ability of the UV LED light source to photoactivate several caged compounds (CMNB-fluorescein, MNI-glutamate, NP-EGTA, DMNPE-ATP) as well as to evoke the associated cellular Ca(2+) responses is demonstrated in both neurons and astrocytes. This report shows that UV LEDs are an efficient light source for flash photolysis and represent an alternative to UV lasers for many applications. A compact, powerful, and low-cost system is described in detail.     

Mechanism of force generation of a viral DNA packaging motor

A large family of multimeric ATPases are involved in such diverse tasks as cell division, chromosome segregation, DNA recombination, strand separation, conjugation, and viral genome packaging. One such system is the Bacillus subtilis phage phi 29 DNA packaging motor, which generates large forces to compact its genome into a small protein capsid. Here we use optical tweezers to study, at the single-molecule level, the mechanism of force generation in this motor. We determine the kinetic parameters of the packaging motor and their dependence on external load to show that DNA translocation does not occur during ATP binding but is likely triggered by phosphate release. We also show that the motor subunits act in a coordinated, successive fashion with high processivity. Finally, we propose a minimal mechanochemical cycle of this DNA-translocating ATPase that rationalizes all of our findings.