Enhanced Risk Stratification for Children and Young Adults with B-Cell Acute Lymphoblastic Leukemia: A Children's Oncology Group Report.

Current strategies to treat pediatric acute lymphoblastic leukemia rely on risk stratification algorithms using categorical data. We investigated whether using continuous variables assigned different weights would improve risk stratification. We developed and validated a multivariable Cox model for relapse-free survival (RFS) using information from 21199 patients. We constructed risk groups by identifying cutoffs of the COG Prognostic Index (PICOG) that maximized discrimination of the predictive model. Patients with higher PICOG have higher predicted relapse risk. The PICOG reliably discriminates patients with low vs. high relapse risk. For those with moderate relapse risk using current COG risk classification, the PICOG identifies subgroups with varying 5-year RFS. Among current COG standard-risk average patients, PICOG identifies low and intermediate risk groups with 96% and 90% RFS, respectively. Similarly, amongst current COG high-risk patients, PICOG identifies four groups ranging from 96% to 66% RFS, providing additional discrimination for future treatment stratification. When coupled with traditional algorithms, the novel PICOG can more accurately risk stratify patients, identifying groups with better outcomes who may benefit from less intensive therapy, and those who have high relapse risk needing innovative approaches for cure.

Effects of Nordic walking in Alzheimer's disease: A single-blind randomized controlled clinical trial.

Non-pharmacological approaches, including exercise programs, have been proposed to improve cognitive function and behavioral symptoms, such as depression, agitation, or aggression, in the management of patients with Alzheimer's disease (AD). Indeed, physical inactivity is one of the main modifiable risk factors in patients with AD, as well as in the development of cardiovascular diseases and related pathologies. Although Nordic Walking (NW), a particular type of aerobic exercise, is known to benefit the health of aging populations, there is little evidence that patients with AD may benefit from this non-pharmacological treatment. In this context, we performed a pilot study in 30 patients with mild/moderate AD to evaluate whether NW influences different cognitive domains, including executive functions, visual-spatial abilities, and verbal episodic memory. To this aim, 15 patients (Control group, CG) underwent reality orientation therapy, music therapy, motor, proprioceptive and postural rehabilitation, and 15 patients (experimental group, EG) in addition to the activities performed by the CG also had the NW with a frequency of twice a week. Neuropsychological assessments and evaluations of daily activities and quality of life were performed at baseline and after 24 weeks. Twenty-two patients, including 13 in the CG and nine in the EG completed the activity program after 24 weeks. The EG showed a significant improvement in the Frontal Assessment Battery, Rey's auditory Verbal Learning Test Delayed Recall, Raven's Colored Progressive Matrices, and completion time for the Stroop Word-Color Interference test, compared to the CG. NW was able to improve cognitive domains like visual-spatial reasoning abilities, verbal episodic memory, selective attention, and processing speed in AD patients. These results, if confirmed by further studies with a larger number of patients and a longer training period, may prospect NW as a safe and likely useful strategy to slow down cognitive impairment in mild/moderate AD.

Molecular characterization of two sub-family specific monoclonal antibodies to meningococcal Factor H binding protein.

Factor H binding protein (FHbp) is a component of two licensed vaccines for prevention of sepsis and meningitis caused by serogroup B meningococci. FHbp binds human Factor H (FH), which contributes to evasion of host immunity and FHbp sequence variants can be classified into two sub-families. Antibodies against FHbp elicit complement-mediated killing and can inhibit recruitment of FH to the bacterial surface. We report epitope mapping studies of two murine IgG mAbs, designated JAR 31 and JAR 36, isolated from a mouse immunized with FHbp in sub-family A, which is present in ∼30-40% of invasive isolates. In the present study, we tested the reactivity of mAbs JAR 31 and JAR 36 with seven natural FHbp sequence variants from different phylogenic groups. We screened bacteriophage-displayed peptide libraries to identify amino acid residues contributing to the JAR 36 epitope. Based on the reactivities of mAbs JAR 31 and JAR 36 with the seven FHbp variants, and the frequent occurrences of aspartate (D) and lysine (K) residues in the JAR 36-bound phage peptides, we selected six residues in the carboxyl-terminal region of FHbp for replacement with alanine (A). The D201A and K203A substitutions respectively eliminated and decreased binding of mAbs JAR 31 and JAR 36 to FHbp. These substitutions did not affect binding of the control mAb JAR 33 or of human FH. JAR 31 or JAR 36 mediated cooperative complement-mediated bactericidal activity with other anti-FHbp mAbs. The identification of two amino acid residues involved in the epitopes recognized by these anti-FHbp mAbs may contribute to a more complete understanding of the spatial requirements for cooperative anti-FHbp mAb bactericidal activity.

Long-term results of the children's cancer group studies for childhood acute lymphoblastic leukemia 1983-2002: a Children's Oncology Group Report.

The Children's Cancer Group enrolled 13 298 young people age <21 years on 1 of 16 protocols between 1983 and 2002. Outcomes were examined in three time periods, 1983-1988, 1989-1995, 1996-2002. Over the three intervals, 10-year event-free survival (EFS) for Rome/National Cancer Institute standard risk (SR) and higher risk (HR) B-precursor patients was 68 and 58%, 77 and 63%, and 78 and 67%, respectively, whereas for SR and HR T-cell patients, EFS was 65 and 56%, 78 and 68%, and 70 and 72%, respectively. Five-year EFS for infants was 36, 38, and 43%, respectively. Seminal randomized studies led to a number of important findings. Stronger post-induction intensification improved outcome for both SR and HR patients. With improved systemic therapy, additional intrathecal (IT) methotrexate effectively replaced cranial radiation. For SR patients receiving three-drug induction, iso-toxic substitution of dexamethasone for prednisone improved EFS. Pegylated asparaginase safely and effectively replaced native asparaginase. Thus, rational therapy modifications yielded better outcomes for both SR and HR patients. These trials provide the platforms for current Children's Oncology Group trials.

Short communication: Effect of alphaS1-casein (CSN1S1) and kappa-casein (CSN3) genotypes on milk composition in Murciano-Granadina goats.

The effects of the caprine alpha(S1)-casein (CSN1S1) polymorphisms on milk quality have been widely demonstrated. However, much less is known about the consequences of the kappa-casein (CSN3) genotype on milk composition in goats. Moreover, the occurrence of interactions between CSN3 and CSN1S1 genotypes has not been investigated. In this study, an association analysis between CSN1S1 and CSN3 genotypes and milk quality traits was performed in 89 Murciano-Granadina goats. Total milk yield as well as total protein, fat, solids-not-fat, lactose, alpha(S1)-casein (CSN1S1), and alpha(S2)-casein (CSN1S2) contents were recorded every other month during a whole lactation (316 observations). Data analysis using a linear mixed model for repeated observations revealed no interaction between the CSN1S1 and CSN3 genotypes. With regard to the effect of the CSN3 locus, AB and BB genotypes were significantly associated with higher levels of total casein and protein content compared with the AA CSN3 genotype. In strong contrast with French breeds, the CSN1S1 genotype did not affect protein, casein, and fat concentrations in Murciano-Granadina goats. These results highlight the importance of taking into consideration the CSN3 genotype when performing selection for milk composition in dairy goats.

Selection in the making: a worldwide survey of haplotypic diversity around a causative mutation in porcine IGF2.

Domestic species allow us to study dramatic evolutionary changes at an accelerated rate due to the effectiveness of modern breeding techniques and the availability of breeds that have undergone distinct selection pressures. We present a worldwide survey of haplotype variability around a known causative mutation in porcine gene IGF2, which increases lean content. We genotyped 34 SNPs spanning 27 kb in 237 domestic pigs and 162 wild boars. Although the selective process had wiped out variability for at least 27 kb in the haplotypes carrying the mutation, there was no indication of an overall reduction in genetic variability of international vs. European local breeds; there was also no evidence of a reduction in variability caused by domestication. The haplotype structure and a plot of Tajima's D against the frequency of the causative mutation across breeds suggested a temporal pattern, where each breed corresponded to a different selective stage. This was observed comparing the haplotype neighbor-joining (NJ) trees of breeds that have undergone increasing selection pressures for leanness, e.g., European local breeds vs. Pietrain. These results anticipate that comparing current domestic breeds will decisively help to recover the genetic history of domestication and contemporary selective processes.

SNP-based markers for discriminating olive (Olea europaea L.) cultivars.

A set of 11 polymorphic markers (1 cleaved amplified polymorphic sequence (CAPS), 2 sequence-characterized amplified regions (SCARs), and 8 single-nucleotide polymorphism (SNP)-derived markers) was obtained for olive cultivar identification by comparing DNA sequences from different accessions. Marker development was more efficient, using sequences from the database rather than cloning arbitrary DNA fragments. Analyses of the sequences of 3 genes from 11 diverse cultivars revealed an SNP frequency of 1 per 190 base pairs in exons and 1 per 149 base pairs in introns. Most mutations were silent or had little perceptible effect on the polypeptide encoded. The higher incidence of transversions (55%) suggests that methylation is not the major driving force for DNA base changes. Evidence of linkage disequilibrium in 2 pairs of markers has been detected. The set of predominantly SNP-based markers was used to genotype 65 olive samples obtained from Europe and Australia, and was able clearly to discriminate 77% of the cultivars. Samples, putatively of the same cultivar but derived from different sources, were revealed as identical, demonstrating the utility of these markers as tools for resolving nomenclature issues. Genotyping data were used for constructing a dendrogram by UPGMA cluster analysis using the simple matching similarity coefficient. Relationships between cultivars are discussed in relation to the route of olive's spread.

Staged informed consent for a randomized clinical trial in childhood leukemia: impact on the consent process.

BACKGROUND: Children Cancer Group (CCG) 1991 is the first childhood acute lymphoblastic leukemia trial within CCG that allowed the utilization of a staged approach to the consent process. METHODS: One hundred and forty subjects participated in the Project on Informed Consent which compared the primary outcome measures in the consent process of patients enrolled in CCG-1991 with those enrolled in other CCG leukemia studies. RESULTS: The parents' trust scores were higher for the CCG-1991 compared with other protocols. Eighty percent of parents enrolled in CCG-1991 understood the distinction between the randomized clinical trial and the standard treatment arm, compared with 62.5% in the other studies, P = 0.05. Multiple other outcome measures suggested a positive impact from staged informed consent. CONCLUSIONS: Our results suggest that a consent process with a staged approach can help investigators obtain a more truly informed consent. Future research is needed to confirm the benefits of the staged approach to the informed consent process.

Characterization and genotyping of the caprine kappa-casein variants.

Kappa-Casein (kappa-CN) is the milk protein that determines the size and specific function of milk micelles, and its cleavage by chymosin is responsible for milk coagulation. We have previously detected and characterized four variants of the goat kappa-CN in Spanish, French, and Italian breeds by screening the major part of the coding region in exon 4. Here we have sequenced and analyzed the full coding region of the kappa-CN gene which includes exons 3 and 4. No additional mutations were found, with exception of a single nucleotide substitution in exon 3, which had no amino acid change. However, the analysis of the association between the different mutations resulted in two new variants designated kappa-CN F and G. The novel variants are present in the Italian breeds Teramana, Girgentana, and Sarda (variant F). A protocol for rapid simultaneous genotyping of all known kappa-CN variants using the primer extension method was described, and a total of 210 animals from nine European breeds were genotyped. Alleles A and B are the most frequent variants occurring in the majority of breeds with highest prevalence of the B variant, except for the Canaria breed where the A allele is more frequent. Sequence data suggest that the F variant is the original type of caprine kappa-CN, other alleles being derived from this type following two different trunks by successive mutations.

A first linkage map of olive (Olea europaea L.) cultivars using RAPD, AFLP, RFLP and SSR markers.

The first linkage map of the olive (Olea europaea L.) genome has been constructed using random amplified polymorphic DNA (RAPD) and amplified fragment length polymorphisms (AFLP) as dominant markers and a few restriction fragment length polymorphisms (RFLP) and simple-sequence repeats (SSR) as codominant markers. Ninety-five individuals of a cross progeny derived from two highly heterozygous olive cultivars, Leccino and Dolce Agogia, were used by applying the pseudo test-cross strategy. From 61 RAPD primers 279 markers were obtained - 158 were scored for Leccino and 121 for Dolce Agogia. Twenty-one AFLP primer combinations gave 304 useful markers - 160 heterozygous in Leccino and 144 heterozygous in Dolce Agogia. In the Leccino map 249 markers (110 RAPD, 127 AFLP, 8 RFLP and 3 SSR) were linked. This resulted in 22 major linkage groups and 17 minor groups with fewer than four markers. In the Dolce Agogia map, 236 markers (93 RAPD, 133 AFLP, 6 RFLP and 4 SSR) were linked; 27 major linkage groups and three minor groups were obtained. Codominant RFLPs and SSRs, as well as few RAPDs in heteroduplex configuration, were used to establish homologies between linkage groups of both parents. The total distance covered was 2,765 cM and 2,445 cM in the Leccino and Dolce Agogia maps, respectively. The mean map distance between adjacent markers was 13.2 cM in Leccino and 11.9 cM in Dolce Agogia, respectively. Both AFLP and RAPD markers were homogeneously distributed in all of the linkage groups reported. The stearoyl-ACP desaturase gene was mapped on linkage group 4 of cv. Leccino.

Short communication: characterization of a new genetic variant in the caprine kappa-casein gene.

A new polymorphism has been identified in the goat kappa-casein gene by evaluating genomic DNA from the Montefalcone breed in Italy. The polymorphic site consists of a single nucleotide substitution A to G at position 242 of the exon 4 and produces an amino acid substitution Asp/Gly. A polymerase chain reaction-restriction fragment length polymorphism protocol for rapid genotyping of the variant has been developed, using the HaeIII enzyme. Animals from Italian, Spanish, and French breeds have been analyzed to investigate the occurrence of the allele in other populations. The allele appears to be exclusive to the Montefalcone breed.

Structural and immunological similarities between high molecular weight zinc ion-dependent p-nitrophenylphosphatase and fructose-1,6-bisphosphate aldolase from bovine liver.

High molecular weight zinc ion-dependent acid p-nitrophenylphosphatase (HMW-ZnAPase) was purified from bovine liver to homogeneity as judged by native and sodium dodecyl sulfate polyacrylamide gel electrophoresis. The partial sequence of the purified enzyme electroblotted on PVDF membrane reveals a 95% sequence homology with human and bovine liver fructose-1,6-bisphosphate aldolase isozyme B (FALD B). FALD B was isolated from bovine liver using an affinity elution from phosphocellulose column. FALD B from bovine liver shows a native and subunit molecular weight that is indistinguishable from that of HMW-ZnAPase. In addition, an affinity purified antiserum raised in rabbits against purified HMW-ZnAPase cross-reacts with bovine liver FALD B and rabbit muscle isozymes. Despite these similarities, HMW-ZnAPase does not show FALD activity and bovine liver FALD does not display any zinc ion-p-nitrophenylphosphatase activity. These results suggested the existence of structural and immunological similarities between bovine liver HMW-ZnAPase and FALD B. Differences in some amino acid residues in enzyme activity indicate that they may be involved in different biochemical functions.

[Continence problems after radical prostatectomy: role of rehabilitation of the pelvic floor]. / Problemi di continenza dopo prostatectomia radicale: ruolo della rieducazione del piano perineale.

Continence mechanisms can be compromised after radical prostatectomy. Relatively low percentages of urinary incontinence are reported (2-15%). Perineal floor physiotherapy is considered an actual method of treatment of urinary incontinence in females. It is based on pelvic floor muscles exercises, biofeedback and functional electrical stimulation. The aim of physiotherapy is to improve pelvic floor muscles proprioception, to increase tone of levator ani and to favour automatization of these muscles in daily life. The reports in Literature on perineal floor physiotherapy in treating incontinence after radical prostatectomy are scarce. In this paper we present our experience about 9 patients with incontinence post radical prostatectomy (out of 74 patients operated on at our Institution). We obtained an improvement or a complete cure in 78% of the treated patients. We believed that pelvic floor physiotherapy can be considered a good and safe method of treatment of incontinence after radical prostatectomy, at least in less serious cases.

The chromosome complement of Olea europaea L.: characterization by differential staining of the chromatin and in-situ hybridization of highly repeated DNA sequences.

The chromosome complement of olive (Olea europaea L.) has been characterized by differential staining of the chromatin and chromosomal localization of highly repeated DNA sequences and ribosomal cistrons. DAPI staining produces different-sized positive bands in various locations on all the chromosomes. By combining this band pattern with the results obtained from cytological hybridization of OeTaq80, OeTaq178, and OeGEM86 DNA tandem repeats, most of the pairs can be distinguished from each other, in spite of the large number of chromosomes (2n = 46), their small size and similar morphology. Different tandem-repeated DNA sequences may be contained into single heterochromatic chromosome regions, even though there are regions where repeats of only one family are present. OeTaq80- and OeGEM86-related DNA sequences are rather specific to the heterochromatin at the chromosome ends, while most sequences related to the longer OeTaq178 probe are confined to interstitial heterochromatin. Some exceptions suggest that major chromosomal rearrangements occurred during genome evolution. Polymorphism, which may differentiate olive cultivars, was observed within chromosome pairs I, V, and VII.

Long-term treatment of refractory thrombocytopenia in a patient with Wiskott-Aldrich syndrome with vincristine, immunoglobulin, and methylprednisolone.

We report a child with Wiskott-Aldrich syndrome with severe, refractory, symptomatic thrombocytopenia who achieved an excellent response to combination therapy with vincristine 1.5 mg/m(2) x 1 day, intravenous immunoglobulin 1 g/kg x 3 days, and methylprednisolone 25 mg/kg x 3 days (VIM) for 7 years after failing multiple treatments. He did not have a histocompatible donor for bone marrow transplantation. When the patient ceased to respond to this regimen, he was rescued with pulse dexamethasone. Vincristine, immunoglobulin, and methylprednisolone might serve as a novel treatment option for the patient with refractory thrombocytopenia. Our patient had a sustained remission of symptomatic thrombocytopenia without toxicity. Furthermore, pulse dexamethasone might be an alternative treatment option to which patients with Wiskott-Aldrich syndrome may respond.

Stroke after zoster ophthalmicus in a 12-year-old girl with protein C deficiency.

A 12-year-old girl who had zoster ophthalmicus 10 months earlier presented with hemiparesis and corresponding basal ganglionic infarction related to middle cerebral artery branch thrombosis ipsilateral to the zoster. Hematologic evaluation disclosed protein C deficiency. This represents the first zoster-associated stroke reported in childhood associated with protein C deficiency, with extension of the latency period between zoster and infarction, previously reported to be 6 months.

Stable expression and purification of a secreted human recombinant prethrombin-2 and its activation to thrombin.

A human prothrombin cDNA has been engineered to obtain a cDNA coding for a secreted form of human prethrombin-2. The secreted prethrombin-2 has been produced in a mammalian expression system using DXB11 cells, a mutant strain of CHO cells in which the dihydrofolate reductase gene has been deleted, and an expression vector carrying the dihydrofolate reductase cDNA. Methotrexate-induced gene amplification favored an efficient production of the recombinant protein which accumulated in the culture medium of the DXB11 cells. Growth in suspension of the stable transformants in an airlift fermenter resulted in the production of 25 mg/L recombinant prethrombin-2. The recombinant protein was purified using single-step affinity chromatography on a recombinant-hirudin column and activated by agarose gel-immobilized ecarin. All purified recombinant prethrombin-2 was activated and the generated recombinant thrombin showed catalytic properties identical to those of plasma-derived alpha-thrombin. This expression system can be used to prepare mutants of prethrombin-2 for structure-function studies investigating thrombin interactions with substrate proteins, inhibitors, and cell membranes.

Interleukin-15 promotes angiogenesis in vivo.

IL-15, a cytokine with biological functions on cells of lymphoid lineage similar to those of IL-2, mediates its activities through the beta and gamma chains of the IL-2/15R and its own alpha chain. Unlike IL-2, IL-15 also binds to endothelial cells with high affinity. We report here that IL-15 is a stimulator of angiogenesis in vivo. When injected subcutaneously into nude mice, IL-15 consistently induced neovascularization of Matrigel plugs. Endothelial cells were found to express the IL-15R alpha chain and the IL-2/15R beta and common gamma chains. IL-15 induced the rapid tyrosine phosphorylation of proteins in endothelial cells, but did not stimulate endothelial cell proliferation in vitro. These findings document a previously unrecognized biological property of IL-15 and emphasize the role of IL-15 as an important mediator outside the immune system.

Mig, the monokine induced by interferon-gamma, promotes tumor necrosis in vivo.

Mig, the monokine induced by interferon-gamma, is a CXC chemokine active as a chemoattractant for activated T cells. Mig is related functionally to interferon-inducible protein 10 (IP-10), with which it shares a receptor, CXCR3. Previously, IP-10 was found to have antitumor activity in vivo. In the present study, murine Mig RNA was found to be expressed at higher levels in regressing Burkitt's lymphoma tumors established in nude mice compared with progressively growing tumors. Daily inoculations of purified recombinant human Mig into Burkitt's tumors growing subcutaneously in nude mice consistently caused tumor necrosis associated with extensive vascular damage. These effects were indistinguishable from those produced by intratumor inoculations of Burkitt's tumors with IP-10. These results support the notion that Mig, like IP-10, has antitumor activity in vivo.