Cystic fibrosis detection in high-risk Egyptian children and CFTR mutation analysis.

BACKGROUND: Knowledge about Cystic Fibrosis (CF) in Egypt is very limited. The objective of this study was to screen for CF in Egyptian children with suggestive clinical features and to identify causative genetic mutations. METHODS: Sixty-one patients from the Chest Unit, Cairo University Children's Hospital, Egypt, were included. Subjects presented with persistent or recurrent respiratory symptoms, failure to thrive, diarrhea and/or steatorrhea and unexplained persistent jaundice. Patients were screened using the CF Indicatortrade mark sweat test system (PolyChrome Medical, Inc., Brooklyn Center, MN). A quantitative sweat testing was conducted on 10 of the 12 positive patients. Seven probands and one sibling underwent molecular analysis by direct DNA sequencing of the coding region and of the intronic sequences adjacent to the 27 exons of the CFTR gene. RESULTS: Of 61 patients, 12 (20%) had positive sweat chloride screening. Ten of the 12 patients underwent quantitative sweat testing and were positive. Eight CFTR sequence changes were identified in seven affected probands and two were confirmed in one sibling by direct DNA sequencing. CONCLUSION: The study results suggest that CF is more common in Egypt than previously anticipated. Larger studies are warranted to identify the incidence, molecular basis and clinical pattern of CF in the Egyptian population.     

δ-aminolevulinic acid dehydrase (porphobilinogen synthase) in two families with inherited enzyme deficiency

The inheritance of a deficient delta-aminolevulinic acid dehydrase (ALA-D; synonym: porphobilinogen synthase; EC 4.2.1.24) was studied in blood samples of two families over three generations. The propositus in each family was a young male acute hepatic porphyria patient with an almost complete ALA-D deficiency in the homozygous state (ALA-D activity less than 2% of controls). Heterozygotes are clinically non-affected (mean ALA-D 36% of controls). The mode of transmission could be traced by enzyme activity and electrophoretic polymorphism studies. Heterozygotes are detected by the demonstration of enzyme activity in the gel. The notation D was used for the gene expressing the defective enzyme. The "phenotype" D-1 was observed in six, the "phenotype" D-2 in three of all heterozygotes studied. These results are compatible with a single normal allele in heterozygotes responsible for enzyme activity. Quantitative assays and the segregation pattern in both families suggest a 3-allele-system for the inheritance of ALA-D deficiency.     

Hereditary uroporphyrinogen-decarboxylase deficiency predisposing porphyria cutanea tarda (chronic hepatic porphyria) in females after oral contraceptive medication

Summary Porphyria cutanea tarda (PCT) was diagnosed in 27 women aged 23–48 years (mean, 35 years) who had been under oral-hormonal-contraceptive medication for 1–18 years, in 3 women under substitutional estrogen treatment in the menopause, and in 2 men aged 65 and 76 years after estrogen treatment of prostatic carcinoma. In all patients, total urinary porphyrin excretion was elevated, with an average uro-and heptacarboxyporphyrin predominance of 88%, thus proving PCT. On the patients, 84% showed a significant decrease of erythrocyte uroporphyrinogen-decarboxylase (UD; EC 4.1.1.37) activity to 50% of control levels suggesting a hereditary predisposition for the development of a chronic hepatic porphyria. Estrogens and alcohol are capable of reducing hepatic UD activity. Women with hereditary red cell UD deficiency may be regarded as predisposed to PCT when under estrogen intake, especially in combination with the potentiating influence of alcohol and chronic liver disease. Normal erythrocyte UD values in patients with additive alcohol consumption may implicate a stronger inhibitory effect for alcohol on UD, suggesting a merely toxic form of chronic hepatic porphyria.     

Intraplacental Choriocarcinoma Associated with Viable Pregnancy: Pathologic Features and Implications for the Mother and Infant

Choriocarcinoma arising in the placenta, or intraplacental choriocarcinoma, has seldom been reported, particularly in the absence of maternal metastases. Reluctance to diagnose choriocarcinoma in the presence of chorionic villi can delay diagnosis; however, timely diagnosis of choriocarcinoma is prognostically important, both for the mother and infant. We report the clinicopathologic findings in five mothers and infants in whom choriocarcinoma was identified in the placenta. None of the mothers had a history of gestational trophoblastic disease in previous pregnancies. Three placentas were similar with a single small lesion grossly suggesting a small infarct; microscopically these consisted of infarcted areas surrounded by choriocarcinoma. These three mothers were unusual in that none had metastatic choriocarcinoma; two were treated with chemotherapy and remained disease-free; the third was lost to follow-up shortly following delivery. The remaining two mothers had known pulmonary metastases at time of delivery. One of these latter two placentas contained a large marginal lesion microscopically identified as choriocarcinoma. The fifth placenta had rare microscopic foci of choriocarcinoma, and sheets of necrotic choriocarcinoma were identified in “blood clot” submitted with the placenta. In four of the five cases the choriocarcinoma appeared to be arising from otherwise normal chorionic villi, and in no case was there invasion of the villous stroma. All of the infants survived, and none had evidence of choriocarcinoma. These cases support the concept that choriocarcinoma associated with otherwise normal pregnancy arises in the placenta and may be more common than reported. Received August 11, 1997; accepted December 8, 1997.     

Metabolic activation of fluoropyrrolidine dipeptidyl peptidase-IV inhibitors by rat liver microsomes.

The current study evaluated the potential for two dipeptidyl peptidase-IV (DPP-IV) inhibitor analogs (1S)-1-(trans-4-([(4-trifluoromethoxyphenyl)sulfonyl]amino)cyclohexyl)-2-[(3S)-3-fluoropyrrolidin-1-yl]-2-oxoethanaminium chloride and (1S)-1-(trans-4-([(2,4-difluorophenyl)sulfonyl]amino)cyclohexyl)-2-[(3S)-3-fluoropyrrolidin-1-yl]-2-oxoethanaminium chloride (MRL-A and MRL-B), containing a fluoropyrrolidine moiety in the structure, to undergo metabolic activation. The irreversible binding of these tritium-labeled compounds to rat liver microsomal protein was time- and NADPH-dependent and was attenuated by the addition of reduced glutathione (GSH) or N-acetylcysteine (NAC) to the incubation, indicating that chemically reactive intermediates were formed and trapped by these nucleophiles. Mass spectrometric analyses and further trapping experiments with semicarbazide indicated that the fluoropyrrolidine ring had undergone sequential oxidation and defluorination events resulting in the formation of GSH or NAC conjugates of the pyrrolidine moiety. The bioactivation of MRL-A was catalyzed primarily by rat recombinant CYP3A1 and CYP3A2. Pretreatment of rats with prototypic CYP3A1 and 3A2 inducers (pregnenolone-16alpha-carbonitrile and dexamethasone) enhanced the extent of bioactivation which, in turn, led to a higher degree of in vitro irreversible binding to microsomal proteins (5- and 9-fold increase, respectively). Herein, we describe studies that demonstrate that the fluoropyrrolidine ring is prone to metabolic activation and that GSH or NAC can trap the reactive intermediates to form adducts that provide insight into the mechanisms of bioactivation.     

In vitro metabolism of MK-0767 [(+/-)-5-[(2,4-dioxothiazolidin-5-yl)methyl]-2-methoxy-N-[[(4-trifluoromethyl)-phenyl] methyl]benzamide], a peroxisome proliferator-activated receptor alpha/gamma agonist. II. Identification of metabolites by liquid chromatography-tandem mass spectrometry.

The in vitro metabolism of MK-0767 [(+/-)-5-[(2,4-dioxothiazolidin-5-yl) methyl]-2-methoxy-N-[[(4-trifluoromethyl)-phenyl] methyl]benzamide], a novel 2,4-thiazolidinedione (TZD)-containing peroxisome proliferator-activated receptor alpha/gamma agonist, was studied in rat, dog, monkey, and human liver microsomes and hepatocytes, as well as in recombinant human CYP3A4-containing microsomes. Twenty-two metabolites (some at trace levels) were detected by liquid chromatography-tandem mass spectrometry analysis. All appeared to be phase I metabolites except for a glucuronide conjugate of a hydroxylated metabolite that was detected at trace levels. A constant neutral loss scan experiment performed on a triple quadrupole mass spectrometer proved to be very useful for resolving the metabolites from endogenous compounds. It was observed that the initial site of metabolism of MK-0767 was at the TZD ring leading to two major metabolites, namely the 5-hydroxy-TZD metabolite (M24) and the mercapto metabolite (M22). The latter was formed via the cleavage of the TZD ring with the elimination of the carbonyl adjacent to the sulfur atom. The structure of M24 was established by accurate mass measurements and NMR analysis. This hydroxy-TZD metabolite might represent an important precursor for a group of metabolites formed by TZD ring opening and subsequent loss of the sulfur moiety. The mercapto metabolite, on the other hand, is probably the key precursor for the TZD ring-opened metabolites with retention of the sulfur, even though the detailed mechanism of the ring scission remains to be characterized. From these studies, it was concluded that the TZD ring was the major site of metabolism of MK-0767. All the metabolites produced in vitro from human preparations were detected in the corresponding preparations from the nonclinical species.