The impact of genetic markers on selection

Genetic marker technologies, such as marker-assisted selection, parentage identification, and gene introgression can be applied to livestock selection programs. Highly saturated genetic maps are now available for cattle, swine, and sheep to provide the genetic framework for developing MAS programs. These programs rely on three phases for commercialization of the technology: the detection phase, in which quantitative trait loci are located and their effects on the phenotype measured; the evaluation phase, in which the markers are evaluated in commercial populations; and the implementation phase, in which markers are combined with phenotypic and pedigree information in genetic evaluation for predicting the genetic merit of individuals within the population. Predicting the economic impact of genetic technologies is a complex process that requires quantitative prediction and economic analysis. Evaluating the impact of these benefits across an industry can be achieved through a process in which gains from implementation of a genetic technology are assessed at the individual, enterprise, and industry levels. A pattern of annual benefits and costs can be predicted using gene flows that can be evaluated by conventional economic analysis.     

Inhibition of tumor cell growth by RTP/rit42 and its responsiveness to p53 and DNA damage

Through a differential screening technique, we have identified a cDNA clone with differential expression in normal versus tumor cells. This clone, designated rit42 (reduced in tumor, 42 kDa), was previously isolated as a homocysteine-inducible gene in human endothelial cells (RTP), and the same or a highly related androgen-responsive gene in mouse has also been identified. Both Northern blot analysis and in situ hybridization demonstrated a significantly diminished expression in tumor cells, including those derived from breast and prostate when compared with normal cells. It was shown that RTP/rit42 mRNA cycles with cell division, peaking at G1 and G2-M, with lower expression in S phase. The biphasic expression of RTP/rit42 mRNA was absent in tumor cells. Introduction of rit42 cDNA into human cancer cells reduced cell growth both in vitro and in nude mice. Moreover, analysis of a tetracycline-regulated p53-inducible system in null-p53 cell lines showed that RTP/rit42 mRNA expression increased concomitantly with p53 expression and followed a similar time course. In addition, DNA-damaging agents induced RTP/rit42 expression in a p53-dependent manner but independent of a p53-mediated G1 arrest. Immunofluorescence analysis of a FLAG epitope-tagged RTP/rit42 protein revealed a cytoplasmic localization pattern with redistribution to the nucleus upon DNA damage. We have localized RTP/rit42 to human chromosome 8q24.3. Taken together, these results are consistent with a growth inhibitory role for RTP/rit42, and its down-regulation may contribute to the tumor malignant phenotype.     

Biochemical and immunochemical characterization of Brassica juncea glyoxalase I

A homogenous preparation of glyoxalase I (S-lactoylglutathione-lyase, EC 4.4.1.5) was obtained from Brassica juncea seedlings. The enzyme is a heterodimer with 27,000 and 29,000 M(r) subunits and native M(r) of 56,000. The circular dichroic spectra of the protein showed characteristics of a distinctly helical protein, and magnesium affected the secondary structure. It is a zinc metalloenzyme. Amino acid modification studies suggested the involvement of histidine residues in catalysis. Apo-glyoxalase I was reactivated by divalent cations Mn2+ (0.5 Mm) > Mg2+ (5 Mm) > Zn2+ (0.05 Mm) and Ca2+ (0.01 Mm). Monospecific, polyclonal anti-glyoxalase I antibodies were raised, which showed its presence in seeds, roots, hypocotyl, cotyledon and different flower parts. They showed varied degree of cross reactivity with the extracts from various plants, yeast, bacteria and animal system.     

The effects of reverse micelles on the reaction mechanism of alpha-chymotrypsin

Reverse micelles were employed to test the accuracy of the widely accepted mechanism for alpha-chymotrypsin in a highly structured aqueous system similar to intracellular conditions. Results yielded from spectrophotometrical assays of the alpha-chymotrypsin catalyzed hydrolysis of both p-nitrophenyl acetate (p-NPA) and p-nitrophenyl trimethylacetate (p-NPTA) were kinetically analyzed to determine constants typical of the proposed mechanistic model. This was accomplished through the establishment of a control, i.e. the well studied buffer system, for comparison between the reverse micellular environment and a bulk aqueous solution. Control group results yielded kinetic constants in favor of the proposed mechanism (Km = 1.55 x 10(-5) +/- 1.40 x 10(-6) M for p-NPA and a Km = 4.97 x 10(-6) +/- 2.29 x 10(-7) M, Km(app) = 4.92 x 10(-6) +/- 2.33 x 10(-8) M, k2 = 4.34 x 10(-3) +/- 1.31 x 10(-3), k(cat) = 1.96 x 10(-3) +/- 2.47 x 10(-4), and Ks = 1.60 x 10(-5) +/- 4.61 x 10(-6) M for p-NPTA). In contrast, similar reactions of the enzyme in a reverse micellular system produced kinetic constants atypical to that representative of the textbook mechanism. (Km = 1.59 x 10(-4) +/- 2.70 x 10(-5) M, Ks = -8.67 x 10(-5) +/- 4.46 x 10(-5) M and Km(app) = -4.80 x 10(-5) +/- 7.05 x 10(-5) M for p-NPA and Km = 1.95 x 10(-4) +/- 9.28 x 10(-5) M, Km(app) = -1.79 x 10(-4) +/- 2.36 x 10(-5) M, and Ks = -3.95 x 10(-4) +/- 1.18 x 10(-4) M for p-NPTA). In addition to negative kinetic constants, alpha-chymotrypsin seemed to display characteristics indicative of super-activity and a hysteretic response. Overall, the widely accepted mechanism for alpha-chymotrypsin appeared to fail within the confines of reverse micelles, due to the direct influence of the system's highly structured form.     

Phase I trial of intravenous carboplatin and cyclosporin A in refractory gynecologic cancer patients

Our objective was to determine the maximum tolerated dose of cyclosporin A (CsA) delivered as a loading dose (LD) and continuous i.v. infusion (CI) in combination with carboplatin in patients with refractory gynecologic cancers. Twenty-nine heavily pretreated patients (25 ovarian epithelial, 2 cervical, and 2 endometrial carcinomas) received 113 cycles of CsA and carboplatin from September 1989 to September 1991. Twenty-four of these 29 carcinomas were strictly defined to be platinum resistant. CsA was administered as a LD escalated from 6 to 10 mg/kg followed by a 24-h CI from 2.5 to 14.5 mg/kg/day. Carboplatin was targeted to an area under the time versus concentration curve (AUC) of 6 mg/ml x min and was not dose escalated. Whole-blood CsA concentrations (fluorescence polarization immunoassay) at the maximum tolerated dose (10 mg/kg LD, 14.5 mg/kg/day CI) ranged from 2.4 to 3.0 microgram/ml over 12 h. Estimated median carboplatin AUC, based on calculated carboplatin clearance, was 7.9 mg/ml x min. The dose-limiting toxicity of the combination of CsA and carboplatin was grade 4 thrombocytopenia. Grade 3 or 4 thrombocytopenia occurred in 35% of the patients, which could be explained by the effects of carboplatin (AUC of 6 mg/ml x min) alone. Overall, neutropenia occurred in 24% of the patients and anemia in 17% of the patients. Grade 3 or 4 nausea or vomiting was noted in 10 and 14% of the patients, respectively. Grade 3 hypertension during CsA administration occurred in 14% of the patients. No grade 3 or 4 nephrotoxicity was seen in this trial. Three objective responses were noted: one complete response (11 months) and one partial response (5 months), both in potentially platinum-sensitive patients with platinum-free intervals of only 9 months each. One platinum-resistant patient had a partial response for 21 months. Five additional patients experienced >75% reduction of CA-125 or a return to a normal CA-125 titer. We concluded that whole-blood CsA concentrations of >3.0 microgram/ml (as seen when CsA is used as a modulator of multidrug resistance) were not achievable in this combination with carboplatin in this population of heavily pretreated gynecologic cancer patients. However, because CsA is used in this trial as a chemosensitizer in platinum-sensitive tumors and as a chemomodulator of platinum resistance, we targeted a CsA concentration of >1.0 microgram/ml, which was achieved. The CsA dose recommended for a Phase II trial of this combination is 10 mg/kg LD and 11.6 mg/kg/day CI, which results in blood CsA concentrations ranging from 1.2 to 1.3 microgram/ml over 12 h. Responses in this population of refractory gynecologic cancer patients are unusual, and these encouraging results form the basis for a Phase II trial of this combination.     

Cytogenetic abnormalities in pediatric myelodysplastic syndrome: a report of three cases

Three consecutive cases of pediatric myelodysplastic syndrome (MDS) diagnosed over a three-year period in Queen Mary Hospital, Hong Kong, were described. Depending on the classification system used, they comprised two cases of chronic myelomonocytic leukemia (CMMoL) of which one can be reclassified as juvenile chronic myeloid leukemia (JCML) and one cases of refractory anemia with excess of blasts (RAEB) or an alternative diagnosis of atypical CML. Cytogenetic abnormalities were detected in all of them on examination of bone marrow cells. Of the two CMMoL, one had monosomy 21, whereas the other had hypodiploidy. The patient with RAEB had a complex karyotype of 46,X,del(X)(q24),t(1;7) (p22;q32),add(15)(q26)(8). The balanced translocation (1;7) seen in this patient was exceedingly rare and, to the best of our knowledge, was reported only twice in the literature. The karyotypic abnormalities that we saw in our patients were not well recognized in pediatric MDS. This report emphasizes the importance of cytogenetic study in children suspected of suffering from MDS, which remains a rare disorder of childhood, and a need to rationalize current classification schemes.