Association of comorbid disease burden at diagnosis with higher tumor grade in men with non-metastatic prostate cancer.

BACKGROUND: While older age is associated with higher tumor grade, it is unknown whether comorbid disease burden has a similar, independent association. We sought to evaluate the impact of comorbid disease burden on tumor grade at diagnosis as indicated by biopsy Gleason score. METHODS: We conducted an observational cohort study of 1260 men newly diagnosed with non-metastatic prostate cancer from 1998 to 2004 at two Veterans Affairs Medical Centers. Multivariable ordinal and multinomial logistic regression were used to evaluate the association between Charlson Comorbidity Index score and biopsy Gleason score. RESULTS: Men with Charlson scores of 2 (odds ratio (OR) 1.8, P<0.001) and 3+ (OR 1.8, P<0.001) had significantly greater odds of higher Gleason scores, compared with men with Charlson scores of 0. In a multinomial logistic regression model predicting Gleason 7 vs ⩾6, only men with Charlson scores of 2 (OR 1.6, P=0.01) had greater odds of having a Gleason 7 tumor, compared with those with Charlson scores of 0. In a multinomial logistic regression model predicting Gleason 8-10 vs ⩽6, those with Charlson scores of 1 (OR 1.6, P=0.047), 2 (OR 2.8, P=0.01) and 3+ (OR 2.9, P=0.001) had higher odds of having a Gleason 8-10 tumor. CONCLUSIONS: Moderate-to-heavy comorbid disease burden at diagnosis may be associated with high tumor grade, independent of age, and is a stronger predictor of Gleason 8-10 than Gleason 7 disease.

Long-term outcome of detectable PSA levels after radical prostatectomy.

Detectable prostate-specific antigen levels (PSA) following radical prostatectomy (RP) are believed to represent treatment failure. In this retrospective review, we characterize long-term PSA outcomes following RP (n = 204) in a non-referral hospital performed between 1984 and 1994. With an average follow-up of 10 y, 90 (44%) patients developed a PSA recurrence: 15 (17%) died of prostate cancer despite hormonal intervention, 39 (43%) responded to hormonal therapy with stable remission and 36 (40%) were observed without intervention. Following RP many patients may have a detectable PSA that does not require treatment. PSA doubling time (< 12 months) was the best predictor of disease progression.

Laparoscopic radical prostatectomy: review and assessment of an emerging technique.

BACKGROUND: Several recent reports have affirmed the feasibility of the laparoscopic approach for radical prostatectomy. In this review, we discuss the morbidities associated with this technique and compare outcomes and convalescence with standard open radical prostatectomy. METHODS: We reviewed all currently published data on laparoscopic radical prostatectomy and our series of 45 robotic-assisted radical prostatectomies and compared them to several landmark series of open retropubic and perineal radical prostatectomies. RESULTS: Although the initial series reported long operating times, these times have been significantly reduced in more recent series. Data on blood loss, convalescence, impotence, and incontinence rates have also been promising. CONCLUSIONS: Although follow-up has been short thus far, laparoscopic radical prostatectomy has been shown to be similar to open radical prostatectomy in several areas.

Prostate needle biopsy infection after four or six dose ciprofloxacin.

OBJECTIVE: This retrospective analysis is to determine rates of clinical infection after prostate needle biopsy with four versus six doses of ciprofloxacin to previous literature. MATERIALS AND METHODS: Two groups were treated with pre and post biopsy 500 mg of ciprofloxacin twice daily by either six doses (n=337) or four doses (n=288) with the first dose given 24 or 12 hours prior to the procedure respectively. RESULTS: Six (0.96%) of the 625 patients had symptomatic urinary tract infections with a positive urinalysis and/or culture. One (0.3%) infection occurred among patients receiving six doses of ciprofloxacin, and five infections (1.7%), were identified among four dose patients. Two febrile episodes occurred in the four dose group, one requiring hospitalization. CONCLUSION: A low infection rate associated with prophylactic regimens. Six doses of ciprofloxacin appears more effective than four doses in reducing the clinical and febrile infection rate following ultrasound guided biopsy of the prostate. No obvious financial benefit was observed.

Simplified technique with short and long-term followup of conversion of an ileal conduit to an Indiana pouch.

PURPOSE: We report a simplified technique for converting an existing conduit to an Indiana pouch as well as short and long-term results. MATERIALS AND METHODS: From May 1988 to February 1998 we evaluated short and long-term outcome and complications in 23 patients 14 to 82 years old (average age 51.8) who underwent conversion of a conduit to an Indiana pouch. When no obstruction of the existing ureteroileal anastomoses was identified, the conduit was freed from the abdominal wall and surrounding bowel. The proximal conduit and ureteral anastomoses were not dissected. The conduit was opened along the antimesenteric wall proximal to the ureteral anastomoses and attached to 25 to 28 cm. of detubularized right colon as a refluxing Studer limb. The pouch was completed in the usual fashion and the stoma was matured at a virgin site. RESULTS: Surgical indications included stomal complications in 10 patients, an infected nonfunctioning kidney in 2 and patient preference in 11. There were no perioperative deaths although 3 patients died of cancer progression. Average operative time was 6.6 hours, estimated blood loss 518 cc and length of stay 7.8 days. Average followup after conversion was 4.7 years (range 0.2 to 11.0). Six late complications developed in 4 cases, including pyelonephritis in 2, severe pouchitis in 1, dehydration in 1 and stomal revision in 2. Renal function was well preserved with an average preoperative and postoperative creatinine of 0.91 and 1.14 mg./dl., respectively. CONCLUSIONS: This technique simplifies conversion and decreases bowel requirements. The low complication rate and stable serum creatinine support the finding that conversion of a conduit to an Indiana pouch is a safe, viable procedure.

On the evolution of Dopa decarboxylase (Ddc) and Drosophila systematics.

We have sequenced most of the coding region of the gene Dopa decarboxylase (Ddc) in 24 fruitfly species. The Ddc gene is quite informative about Drosophila phylogeny. Several outstanding issues in Drosophila phylogeny are resolved by analysis of the Ddc sequences alone or in combination with three other genes, Sod, Adh, and Gpdh. The three species groups, melanogaster, obscura, and willistoni, are each monophyletic and all three combined form a monophyletic group, which corresponds to the subgenus Sophophora. The Sophophora subgenus is the sister group to all other Drosophila subgenera (including some named genera, previously considered outside the Drosophila genus, namely, Scaptomyza and Zaprionus, which are therefore downgraded to the category of subgenus). The Hawaiian Drosophila and Scaptomyza are a monophyletic group, which is the sister clade to the virilis and repleta groups of the subgenus Drosophila. The subgenus Drosophila appears to be paraphyletic, although this is not definitely resolved. The two genera Scaptodrosophila and Chymomyza are older than the genus Drosophila. The data favor the hypothesis that Chymomyza is older than Scaptodrosophila, although this issue is not definitely resolved. Molecular evolution is erratic. The rates of nucleotide substitution in 3rd codon position relative to positions 1 + 2 vary from one species lineage to another and from gene to gene.

Erratic evolution of glycerol-3-phosphate dehydrogenase in Drosophila, Chymomyza, and Ceratitis.

We have studied the evolution of Gpdh in 18 fruitfly species by sequencing 1,077 nucleotides per species on average. The region sequenced includes four exons coding for 277 amino acids and three variable-length introns. Phylogenies derived by a variety of methods confirm that the nominal genus Zaprionus belongs within the genus Drosophila, whereas Scaptodrosophila and Chymomyza are outside. The rate of GPDH evolution is erratic. The rate of amino acid replacements in a lineage appears to be 1.0 x 10(-10)/site/year when Drosophila species are considered (diverged up to 55 million years ago), but becomes 2.3 x 10(-10) when they are compared to Chymomyza species (divergence around 60 My ago), and 4.6 x 10(-10) when species of those two genera are compared with the medfly Ceratitis capitata (divergence around 100 My ago). In order to account for these observations, the rate of amino acid replacement must have been 15 or more times greater in some lineages and at some times than in others. At the nucleotide level, however, Gpdh evolves in a fairly clockwise fashion.

Characterization of a Cu/Zn superoxide dismutase-encoding gene region in Drosophila willistoni.

A Cu/Zn superoxide dismutase-encoding gene (Sod) from Drosophila willistoni was cloned and sequenced. The gene shows a typical structure for a fruit-fly Sod gene, with a coding region of 462 bp in two exons separated by a 417-bp intron. Comparison of the Sod sequences from D. willistoni and D. melanogaster suggests that these species are only remotely related. Downstream from the Sod gene, there is an ORF on the opposite strand that putatively encodes the last exon of an unidentified gene. The polyadenylation signals of the two genes are separated by only 61 bp in D. willistoni, conforming to the common picture of compact dipteran genomes.

Phylogeny of Drosophila and related genera inferred from the nucleotide sequence of the Cu,Zn Sod gene.

The phylogeny and taxonomy of the drosophilids have been the subject of extensive investigations. Recently, Grimaldi (1990) has challenged some common conceptions, and several sets of molecular data have provided information not always compatible with other taxonomic knowledge or consistent with each other. We present the coding nucleotide sequence of the Cu,Zn superoxide dismutase gene (Sod) for 15 species, which include the medfly Ceratitis capitata (family Tephritidae), the genera Chymomyza and Zaprionus, and representatives of the subgenera Dorsilopha, Drosophila, Hirtodrosophila, Scaptodrosophila, and Sophophora. Phylogenetic analysis of the Sod sequences indicates that Scaptodrosophila and Chymomyza branched off the main lineage before the major Drosophila radiations. The presence of a second intron in Chymomyza and Scaptodrosophila (as well as in the medfly) confirms the early divergence of these two taxa. This second intron became deleted from the main lineage before the major Drosophila radiations. According to the Sod sequences, Sophophora (including the melanogaster, obscura, saltans, and willistoni species groups) is older than the subgenus Drosophila; a deep branch splits the willistoni and saltans groups from the melanogaster and obscura groups. The genus Zaprionus and the subgenera Dorsilopha and Hirtodrosophila appear as branches of a prolific "bush" that also embraces the numerous species of the subgenus Drosophila. The Sod results corroborate in many, but not all, respects Throckmorton's (King, R.C. (ed) Handbook of Genetics. Plenum Press, New York, pp. 421-469, 1975) phylogeny; are inconsistent in some important ways with Grimaldi's (Bull. Am. Museum Nat. Hist. 197: 1-139, 1990) cladistic analysis; and also are inconsistent with some inferences based on mitochondrial DNA data. The Sod results manifest how, in addition to the information derived from nucleotide sequences, structural features (i.e., the deletion of an intron) can help resolve phylogenetic issues.

Evidence for positive selection in the superoxide dismutase (Sod) region of Drosophila melanogaster.

DNA sequence variation in a 1410-bp region including the Cu,Zn Sod locus was examined in 41 homozygous lines of Drosophila melanogaster. Fourteen lines were from Barcelona, Spain, 25 were from California populations and the other two were from laboratory stocks. Two common electromorphs, SODS and SODF, are segregating in the populations. Our sample of 41 lines included 19 SodS and 22 SodF alleles (henceforward referred to as Slow and Fast alleles). All 19 Slow alleles were identical in sequence. Of the 22 Fast alleles sequenced, nine were identical in sequence and are referred to as the Fast A haplotypes. The Slow allele sequence differed from the Fast A haplotype at a single nucleotide site, the site that accounts for the amino acid difference between SODS and SODF. There were nine other haplotypes among the remaining 13 Fast alleles sequenced. The overall level of nucleotide diversity (pi) in this sample is not greatly different than that found at other loci in D. melanogaster. It is concluded that the Slow/Fast polymorphism is a recently arisen polymorphism, not an old balanced polymorphism. The large group of nearly identical haplotypes suggests that a recent mutation, at the Sod locus or tightly linked to it, has increased rapidly in frequency to around 50%, both in California and Spain. The application of a new statistical test demonstrates that the occurrence of such large numbers of haplotypes with so little variation among them is very unlikely under the usual equilibrium neutral model. We suggest that the high frequency of some haplotypes is due to natural selection at the Sod locus or at a tightly linked locus.

Genetic characterization of six parasitic protozoa: parity between random-primer DNA typing and multilocus enzyme electrophoresis.

We have assayed genetic polymorphisms in several species of parasitic protozoa by means of random amplified polymorphic DNA (RAPD). One goal was to ascertain the suitability of RAPD markers for investigating genetic and evolutionary problems, particularly in organisms, such as the parasitic protozoa, unsuitable for traditional methods of genetic analysis. Another goal was to test certain hypotheses concerning Trypanosoma cruzi, and other protozoa, that have been established by multilocus enzyme electrophoresis. The RAPD results corroborate the hypothesis that the population structure of T. cruzi is clonal and yield a phylogeny of the clonal lineages in agreement with the one obtained by enzyme electrophoresis. This parity between the two sets of results confirms that RAPD markers are reliable genetic markers. The RAPD markers are also suitable for reconstructing species phylogenies and as diagnostic characters of species and subspecific lineages. The number of DNA polymorphisms that can be detected by the RAPD method seems virtually unlimited, since the number of primers can be increased effectively at will. The RAPD method is well suited for investigating genetic and evolutionary questions in certain organisms, because it is cost effective and demands no previous genetic knowledge about the organism.

Structure and sequence of the Cu,Zn Sod gene in the Mediterranean fruit fly, Ceratitis capitata: intron insertion/deletion and evolution of the gene.

We have cloned a 4-kb region encompassing the Cu,Zn superoxide dismutase (Sod) gene from a genomic library of the Mediterranean fruit fly, Ceratitis capitata, using a cDNA probe from Drosophila melanogaster. The coding sequence of 462 bases is equally as long as that in Drosophila species. The rate of amino acid replacement over the past 100 million years is approximately the same in the Diptera and in mammals, thus excluding the hypothesis (proposed to account for an apparent acceleration in rate of evolution of Sod over geological time) that the evolution of the SOD protein is much higher in the mammals than in other organisms. The coding region is interrupted by two introns in Ceratitis, whereas only one occurs in Drosophila. Phylogenetic comparisons indicate that the second intron was present in the common dipteran ancestor, but was lost shortly after the divergence of the Drosophila lineage from other Diptera. Analysis of the exon/intron structure of Sod in various animal phyla, plants, and fungi indicates that intron insertions as well as deletions have occurred in the evolution of the Sod gene.

Structure and sequence of the Cu, Zn superoxide dismutase gene of Chymomyza amoena: phylogeny of the genus and codon-use evolution.

We have cloned and sequenced the Cu, Zn superoxide dismutase gene of Chymomyza amoena. The coding sequence has the same length as in Drosophila species and in Ceratitis capitata. There are two introns, located at the same sites as in Ceratitis. The second intron is absent in Drosophila: this places Chymomyza outside the Drosophila lineage, contrary to proposals based on anatomical and other evidence. The nucleotide or amino acid distances support a phylogeny in which Ceratitis first branches off the common stem, then Chymomyza splits before the divergence of the two major Drosophila subgenera. The estimated divergence times are 58 million for Chymomyza-Drosophila; 48 million years for the Drosophila subgenera. During the intervening 10 million years, the Drosophila lineage lost the second intron and evolved distinct codon-preferences: the G + C use in the third coding positions is increased by 69% in Drosophila relative to Chymomyza or Ceratitis.

A truncated P element is inserted in the transcribed region of the Cu,Zn SOD gene of an SOD "null" strain of Drosophila melanogaster.

The decreased Cu,Zn SOD activity (less than 5%) in a "null" SODCA1 Drosophila melanogaster strain isolated in our laboratory is due to the insertion of a truncated P element into the transcribed region of the Cu,Zn SOD gene. Using a cDNA Cu,Zn SOD probe from a wild type D. melanogaster (F allele) we isolated an EcoRI Cu,Zn SOD clone from an EMBL3 genomic library of the SODCA1 strain, subcloned it, restriction-mapped and partially sequenced it. The 2.5 kb clone consists of a wild-type 1.84 kb EcoRI fragment containing the Cu,Zn SOD gene previously isolated in our laboratory, with an insertion of 0.68 kb derived (by an internal deletion) from an autonomous, 2.9 kb P element. The insertion starts 21 bp upstream from the coding sequence and causes an 8 bp target site duplication characteristic of P elements. A point mutation in the second exon results in a replacement of Asn by Lys at position 96, confirming that the mature protein encoded by the SOCCA1 is the same one encoded by the S allele, commonly found in natural populations. The diminished expression of SODCA1 allele is most possibly due to a reduction of the rate of transcription attributable to the insertion of the P element.

A highly polymorphic locus very tightly linked to the Huntington's disease gene.

The genetic defect in Huntington's disease (HD), an inherited neuropsychiatric disorder of unknown etiology, has not been defined. The discovery of linkage between HD and the DNA marker D4S10(G8) raised the possibility of isolating the disease gene on the basis of its chromosomal location, in addition to providing a limited presymptomatic test for the late onset disorder. But it has been difficult to isolate other DNA markers nearer to the HD gene, and this has hampered attempts to identify the disease locus and limited the applicability and accuracy of predictive testing. Recently, several new DNA markers from the region of the genome near the HD gene have been isolated using a directed cloning strategy. We describe here the characterization of one of these new markers, D4S95, a highly polymorphic locus which displays no recombination with the HD gene in the families tested. The high degree of polymorphism at this locus and its proximity to the HD gene make it extremely useful for predictive testing and as a new starting point for attempts to clone the disease gene.

Isolation of DNA markers in the direction of the Huntington disease gene from the G8 locus.

To facilitate identification of additional DNA markers near and on opposite sides of the Huntington disease (HD) gene, we developed a panel of somatic-cell hybrids that allows accurate subregional mapping of DNA fragments in the distal portion of 4p. By means of the hybrid-cell mapping panel and a library of DNA fragments enriched for sequences from the terminal one-third of the short arm of chromosome 4, 105 DNA fragments were mapped to six different physical regions within 4p15-4pter. Four polymorphic DNA fragments of particular interest were identified, at least three of which are distal to the HD-linked D4S10 (G8) locus, a region of 4p previously devoid of DNA markers. Since the HD gene has also recently been shown to be distal to G8, these newly identified DNA markers are in the direction of the HD gene from G8, and one or more of them may be on the opposite side of HD from G8.

Deletion mapping of human chromosome 5 using chromosome-specific DNA probes.

A complete genomic DNA library was prepared from a Chinese hamster-human cell hybrid that contains human chromosome 5 as its only human DNA. Unique or low-copy DNA fragments, isolated form recombinant bacteriophage that contained human DNA inserts, were regionally mapped on chromosome 5 using Southern blot analysis of genomic DNA from a series of hybrid cell lines that were selected as having deletions of various portions of 5q. The chromosome 5-specific DNA library, together with a genetic selective procedure allowing the isolation of hybrid cell lines with deletions of virtually any portion of 5q, will provide a means to construct very accurate physical and recombinational maps of this human chromosome. This system represents an excellent opportunity to examine very precisely the relationship between physical and genetic distances for many loci along the length of this autosome.