Renal clearance and protein binding of melphalan in patients with cancer

Summary The renal clearance of melphalan and the fraction unbound in plasma were determined after intravenous infusion of 5 mg/m² over 5 min in nine patients with cancer to obtain information regarding the mechanism of renal handling of melphalan. Four of the patients underwent bone marrow transplantation and also received an IV dose of 220 mg/m². Total melphalan clearance after the 5 mg/m² dose ranged from 66.0 to 272 ml/min per m²; the percentage of the dose excreted unchanged in urine, from 2.5% to 92.8%; renal clearance, from 4.1 to 188 ml/min per m²; the fraction unbound in plasma, from 0.0598 to 0.460; and t_1/2, from 39.4 to 84.3 min. Unbound melphalan clearance and renal clearance calculated from the unbound fraction in plasma for each patient ranged from 441 to 3356 ml/min per m² and 15 to 961 ml/min per m² respectively and were not related to serum albumin, serum creatinine or creatinine clearance. The percentage of the dose exctreted and melphalan renal clearance were not related to urine flow. There was evidence of active secretion of melphalan in the kidney an possible reabsorption. There were no significant paired differences in melphalan disposition between the high- and low-dose studies. Highly variable renal clearance involving active secretion may contribute in part to large interpatient differences in the total plasma clearance of melphalan in patients with cancer.This study was supported by a grant from The Queen Elizabeth Hospital Research Foundation     

The role of genetics in the development of asthma and atopy

PURPOSE OF REVIEW: The mapping of complex traits such as asthma and atopy is one of the most important and central areas of human genetics. This article will present an overview of the current status of genetic studies of asthma and atopy using genome screens and association studies that have occurred in the literature since January 2003. RECENT FINDINGS: Many regions of the genome have been found to have linkage with the phenotypes of asthma and atopy. Over 70 variants in candidate genes have been reported to be associated with these phenotypes. The main regions these variants have been found are on chromosomes 2q, 5q, 6p, 11q, 12q, 16q and 17q. Five potential asthma susceptibility genes or complexes have been identified using a positional approach. These are ADAM33, DPP10, PHF11 and SETDB2, GPRA and SPINK5. It is evident that environmental factors will influence the expression of genes and the ultimate clinical phenotype of asthma and atopy. SUMMARY: The development of asthma and atopy involves many genes and environmental factors. An understanding of their genetic basis has great implications for their management.     

Listeria monocytogenes intragastric and intraperitoneal approximate 50% lethal doses for mice are comparable, but death occurs earlier by intragastric feeding.

The intraperitoneal (i.p.) and intragastric (i.g.) mouse approximate 50% lethal dose values (ALD50S) were determined for 15 food and clinical isolates of Listeria monocytogenes. Although all strains gave i.g. ALD50S comparable to or less than their i.p. ALD50S, the i.g. feeding of most strains produced more deaths within the first 3 days of the 6-day test than did i.p. injection. ALD50S ranged from 50 to 4.4 x 10(5) cells with approximately 1-log 95% confidence intervals. Of five strains tested by suspension in milk or by growth in milk, none gave i.g. ALD50S that were lower than those of washed cells. Results with 10- to 21-g mice supported the use of 15-g mice for i.g. testing; 21-g mice were more resistant to i.g. infection. These results indicate that i.g. feeding permits an evaluation of the role of the carrier (such as milk) in the determination of listerial virulence, permits strain characterization by i.p. and i.g. ALD50S, and emphasizes a potentially more rapid infection when the bacterium is introduced i.g.     

A duplication/paracentric inversion associated with familial X-linked deafness (DFN3) suggests the presence of a regulatory element more than 400 kb upstream of the POU3F4 gene

X-linked deafness with stapes fixation (DFN3) is caused by mutationsin the POU3F4 gene at Xq21.1. By employing pulsed field gelelectrophoresis (PFGE) we identified a chromosomal aberrationin the DNA of a DFN3 patient who did not show alterations inthe open reading frame (ORF) of POU3F4. Southern blot analysisindicated that a DNA segment of 150 kb, located 170 kb proximalto the POU3F4 gene, was duplicated. Fluorescence in situ hybridization(FISH) analysis, PFGE, and detailed Southern analysis revealedthat this duplication is part of a more complex rearrangementincluding a paracentric inversion involving the Xq21.1 region,and presumably the Xq21.3 region. Since at least two DFN3-associatedminideletions are situated proximal to the duplicated segment,the inversion most likely disconnects the POU3F4 gene from aregulatory element which is located at a distance of at least400 kb upstream of the POU3F4 gene.     

Characterization of a subtype of primary osteoclastoma: Extracellular calcium but not calcitonin inhibits aggressive HLA-DR-positive osteoclastoma possessing ‘functional’ calcitonin receptors

We report here a case of primary osteoclastoma that despite possessing HLA-DR-positive status and ‘functional’ calcitonin receptors, exhibited aggressive in vitro and in vivo bone resorptive activity. In the osteoclast bone slice assay employing scanning electron microscopy, the giant cell-mediated bone resorption was uninhibited by salmon calcitonin (10 nM) and significantly inhibited by raised extracellular calcium (20 mM). In Fura-2AM based microspectrofluorimetric assays, the presence of the ‘functional’ calcitonin receptors was ascertained by a rise in intracellular calcium induced by calcitonin and high extracellular calcium. These findings provide evidence for a hitherto unrecognized subtype of giant cells that have HLA-DR-positive status, exhibit avid bone resorptive activity, but remain insensitive to calcitonin despite possessing calcitonin receptors.     

Structural diversity among anti-p-azophenylarsonate monoclonal antibodies from A/J mice: Comparison of Id and Id sequences

Amino acid sequence analyses were carried out on monoclonal anti-p-azophenylarsonate antibodies isolated from the ascites of mice carrying cell lines obtained from the fusion of A/J splenic lymphocytes with the myeloma cell line Sp2/0–Ag14. The partial primary structures of both heavy and light chains from seven idiotype negative hybridoma proteins are compared to those of six idiotype positive molecules. Amino-terminal amino acid sequences (40–47 residues) of heavy chains from molecules bearing the major cross-reacting idiotype, Id^CR, demonstrated 95% homology to each other. Similarly, aminoterminal sequences of Id^CR+light chains were homologous to each other. However, sequence variations were evident in individual antibodies in both framework and complementarity-determining regions, suggesting that a large family of molecules accounts for the major cross-reacting idiotype, as previously reported (Marshak-Rothstein et al., 1980b).

Heavy and light chains from seven Id^CR-negative monoclonal antibodies were subjected to amino-terminal (37–48 residues) amino acid sequence analysis. Four heavy chains were blocked to Edman degradation, but could be sequenced after enzymatic removal of the amino-terminal pyrrolidone carboxylic acid residue. In comparison with Id^CR-positive heavy chains, the Id^CR-negative heavy chains demonstrate greater diversity in both framework and complementarity-determining regions, with several different subgroups represented in contrast to the results from pooled serum Id^CR-positive antibodies (Capra et al., 1975). One of the seven Id^CR-negative light chains was blocked. The sequences of the remaining Id^CR-negative light chains exhibited marked variations in both framework and complementarity-determining regions, with different chain lengths in the first complementarity-determining region in several light chains.

Comparisons between the amino-terminal sequences of Id^CR-positive and Id^CR-negative monoclonal antibodies suggest that specific sequences in the first complementarity-determining regions of both heavy and light chains are not sufficient to account for the major cross-reacting idiotype. The structural basis for Id^CR in A/J mice is likely to be in other segments of the variable regions.     

Human herpes virus 6 and endogenous biotin in salivary glands.

AIMS: To detect the presence of human herpes virus 6 (HHV6) and endogenous biotin in paraffin wax embedded and frozen salivary glands. METHODS: Two stage indirect and streptavidin-biotin immunoperoxidase techniques were used to visualise the antigens. RESULTS: HHV6 could not be shown in any of the tissues. However, considerable endogenous biotin antigenicity was detected in the glandular elements of the paraffin wax embedded material. CONCLUSIONS: Results obtained with avidin-biotin detection systems should be interpreted with caution, especially when glandular epithelium is being stained. This may apply to both immunoperoxidase and in situ hybridisation techniques. The use of an anti-biotin antibody as a standard control should be considered.     

A comprehensive haplotype analysis of CYP19 and breast cancer risk: the Multiethnic Cohort

The CYP19 gene encodes for aromatase (P450arom), a key steroidogenic enzyme that catalyzes the final step of estrogen biosynthesis. Apart from rare mutations in CYP19 which result in severe phenotypes associated with estrogen insufficiency, little is known about whether common variation in CYP19 is associated with risk of hormone-related diseases. In this study, we employed a haplotype-based approach to search for common disease-associated variants in this candidate breast cancer susceptibility gene among African-American, Hawaiian, Japanese, Latina and White women in the Multiethnic Cohort Study (MEC). We utilized 74 densely spaced single-nucleotide polymorphisms (SNPs) (one every approximately 2.6 kb) spanning 189.4 kb of the CYP19 locus to characterize linkage disequilibrium (LD) and haplotype patterns among 69-70 individuals from each ethnic population. We detected four regions of strong LD (blocks 1-4) that were quite closely conserved across populations. Within each block there was a limited diversity of common haplotypes (5 to 10 with a frequency >/=5%) and most haplotypes were observed to be shared across populations. Twenty-five haplotype-tagging SNPs (htSNPs) were selected to predict the common haplotypes with high probability (average Rh2=0.92) and genotyped in a breast cancer case-control study in the MEC (cases, n=1355; controls, n=2580). We first performed global tests for differences in risk according to the common haplotypes and observed significant haplotype-effects in block 2 [P=0.01; haplotypes 2b (OR=1.23; 95% CI, 1.07-1.40), 2d (OR=1.28; 95% CI, 1.01-1.62)]. We also found a common long-range haplotype comprised of block-specific haplotypes 2b and 3c to be associated with increased risk of breast cancer (haplotype 2b-3c: OR=1.31; 95% CI, 1.11-1.54). Our findings suggest the hypothesis that women with the long-range CYP19 haplotype 2b-3c may be carriers of a predisposing breast cancer susceptibility allele.