A methylester of the glucuronide prodrug DOX-GA3 for improvement of tumor-selective chemotherapy

The glucuronide prodrug of doxorubicin, DOX-GA3, can be selectively activated in tumors by extracellular human beta-glucuronidase, resulting in a better therapeutic index than doxorubicin. DOX-GA3, however, is rapidly excreted by the kidney. We hypothesized that slow release of DOX-GA3 from its methylester, DOX-mGA3, by esterase activity in blood would result in improved circulation half-life (t(1/2)) of DOX-GA3. DOX-mGA3 was synthesized more efficiently with an overall yield of 60% as compared to 37% in the case of DOX-GA3. We showed that DOX-mGA3 was enzymatically converted to DOX-GA3 with a t(1/2) of approximately 0.5 min in mouse plasma to 2.5 h in human plasma, which was in agreement with differences in esterase activity between species. DOX-mGA3, similar to DOX-GA3, was at least 37-fold less potent than the parent drug doxorubicin in growth inhibition of four different human malignant cell lines in vitro. Incubation of OVCAR-3 cells with DOX-mGA3 in combination with an excess of human beta-glucuronidase (0.05 U mL(-1)) resulted in a similar growth inhibition to that of doxorubicin. Intravenous administration of DOX-mGA3 in FMa-bearing mice resulted in an area under the concentration versus time curve (AUC) of DOX-GA3 in tumor and most normal tissues that was 2.5- to 3-fold higher than after the same dose of DOX-GA3 itself. In tumor tissue, this was accompanied by a 2.7-fold increase in the AUC of doxorubicin from DOX-mGA3 than from DOX-GA3. In conclusion, an advantage of DOX-mGA3 over DOX-GA3 is that this prodrug can be produced with a higher yield. Another important advantage is the improved pharmacokinetics of the lipophilic DOX-mGA3 as compared to that of the hydrophilic DOX-GA3. This effect may even be more pronounced in man, because of the lower plasma esterase activity than measured in mice.     

Blood biochemistry and the risk of cancer

A longitudinal study based on a serum sample bank was carried out in Finland to find out the association between biochemical substances and the subsequent risk of cancer. The objective was to evaluate the consistency between means of individually estimated levels of these compounds and levels based on pooling. Levels of alpha-tocopherol, beta-carotene, retinol, retinol-binding protein, and ceruloplasmin were estimated by primary site and sex and partly by age and morphology. The concentrations in pooled samples were consistently lower than the averages of the individual samples. On the basis of individual samples, all the five biochemical compounds had a rather consistent protective effect on the risk of cancers at most primary sites. This protective effect disappeared in the pool analyses, and more than half of exposure contrasts showed an opposite sign. For ceruloplasmin, the effect of pooling was smaller but not negligible. The results of this study emphasize the demand to standardize the collecting, handling, and analysing of samples in serum banks. They are, furthermore, consistent with the hypothesis that pooling of biochemical samples affects the levels of the substances and may affect the conclusions of epidemiological studies on causes of diseases.     

Neurocognitive profiles in Salla disease

Salla disease, a free sialic acid storage disorder, is one of the 36 currently known disorders in Finland that form the Finnish disease heritage. Salla disease leads to learning disability* with a wide clinical variation. Two main categories of the disease have been classified: a conventional subtype and a severe subtype with more severe defects. We present detailed neurocognitive profiles of 41 Finnish patients with Salla disease (19 females, 22 males; age range 11mo to 63y, median 19y). The neurocognitive development of patients with Salla disease was assessed by psychological and neuropsychological testing. All patients were also examined by a paediatric neurologist and a speech therapist. The characteristic cognitive profile consisted of a lower non-verbal performance (mean developmental age 13mo) compared with linguistic skills (mean developmental age 17mo). In particular, spatial and visual-constructive impairments were typical of these patients. Tactile and visual discrimination of forms was poor. Tasks demanding hand-eye coordination, maintenance of visual attention, and those requiring short-term visual memory and executive skills were performed better. Receptive language skills were notably better compared with expressive speech. The patients' interactive and non-verbal communication skills were quite strong. Another typical pattern with Salla disease was severe motor disability. After the second decade of life, the decline in these skills was more pronounced than patients' cognitive deterioration. Our results indicate that even though there is a considerable variation in the clinical findings of patients with Salla disease, the characteristic neurocognitive profile of the disease can be outlined.     

Familial cryptic translocation (2;17) ascertained through recurrent spontaneous abortions

We report a young woman who presented with a reproductive history of three recurrent spontaneous abortions (RSA) and two neonatal deaths. Comparative genomic hybridization (CGH) was used to determine the chromosomal composition of the patient's last miscarriage. It showed the presence of monosomy for the distal end of chromosome 2 long arm (segment 2q37.2 to qter) and trisomy for the distal end of chromosome 17 long arm (segment 17q25 to qter). The mother was found to be a carrier for a cryptic translocation between chromosomes 2 and 17 long arms by fluorescence in situ hybridization using a subtelomeric probe for 17q. Retrospective CGH analysis on one baby who died neonatally showed that he had inherited the maternal translocation in the same unbalanced state as the last pregnancy loss. His detailed postmortem examination is reported.     

Effects of 4-tert-octylphenol, 4-tert-butylphenol, and diethylstilbestrol on prenatal testosterone surge in the rat

In the present study, we evaluated the effects that 4-tert-octylphenol (OP) and 4-tert-butylphenol (BP) had on the prenatal testicular testosterone surge at embryonic day (ED) 19.5 in the rat. In utero exposure to alkylphenols (0.1-100 mg/kg maternal weight) on EDs 13.5, 15.5, and 17.5 did not decrease testicular testosterone content, whereas exposure to diethylstilbestrol (DES) caused a significant depression in testosterone synthesis and secretion. The depression was maintained during ex vivo tissue culture. In order to elucidate the observed differences in the in vivo effects between alkylphenols and DES, the exposures were also carried out in tissue culture of intact ED 19.5 testes. Basal testosterone, progesterone, cAMP production and hCG-induced testosterone levels were determined during and after a 3-h culture period. DES (100 mg/l) did not alter testosterone production but caused a two-fold increase in progesterone. OP (10, 100, 500 mg/l) and BP (100 mg/l) significantly increased testosterone and progesterone levels by up to seven-fold. In the presence of BP 100 mg/l, however, the intratesticular testosterone content did not correlate with the significantly increased fraction of secreted, or leaked, testosterone. The latter was correlated with tissue damage observed at electron microscopic level. Consistent with this, BP 500 mg/l elevated testicular testosterone level slightly during the first hour in the culture but the level subsequently returned to the control value. At the electron microscopic level, alkylphenols caused most severe changes in Leydig cell membrane structures and lipid droplets. In the DES-treated testes, membrane vesicle formation around the lipid droplets and increased mitochondrial pleiomorphy were observed. Altogether, the present in vivo and in vitro analyses confirm different effects of alkylphenols and DES on fetal rat steroidogenesis and tissue structure.     

Regional cerebral glucose metabolism in monozygotic twins discordant for Alzheimer's disease

We investigated regional cerebral glucose metabolic rates (rCMRgluc) with positron emission tomography using 2-[18F]fluoro-2-deoxy-D-glucose (FDG) in 7 monozygotic twin pairs discordant for Alzheimer's disease (AD). Ten healthy volunteers with comparable mean age and educational level served as controls. In the hippocampus, the mean +/- SD rCMRgluc were 0.20 +/- 0.03 micromol/ml/min for the demented twins, 0.21 +/- 0.03 micromol/ml/min for their non-demented co-twins, and 0.23 +/- 0.02 micromol/ml/min for the controls. The mean hippocampal rCMRgluc was reduced in the demented twins (p = 0.006), compared with the controls. In the lateral temporal cortex, the mean +/- SD rCMRgluc were 0.27 +/- 0.05, 0.28 +/- 0.04, and 0.32 +/- 0.02 micromol/ml/min, respectively. These mean rates were reduced both in the demented (p = 0.02) and the non-demented (p = 0.01) twins, compared with the controls. In conclusion, in the demented twins, the reduction of rCMRgluc was detected in the hippocampus and lateral temporal cortex, i.e. the 2 brain areas which show early changes in pathological and imaging studies in AD. Their non-demented co-twins showed milder reductions, which may be an indication of genetic susceptibility for dementia, and an early sign of a dementing illness in them.     

Human pHyde is not a classical tumor suppressor gene in prostate cancer

A novel putative tumor suppressor gene, pHyde, was recently cloned from rat prostate. The rat gene has been shown to inhibit prostate cancer cell proliferation both in vitro and in vivo. However, the role of human pHyde in prostate cancer has not been studied before. Here, we analyzed human prostate cancer cell lines (LNCaP, DU145, PC-3, 22Rv1), xenografts (LuCaP 23.1, 35, 41, 49, 58, 69, 70 and 73) and clinical prostate carcinomas for genetic alterations and expression of pHyde. The expression of pHyde in normal human tissues as well as in prostate cancer was studied by Northern analysis and real-time quantitative RT-PCR. It was ubiquitously expressed in all normal tissues analyzed. Although, the expression was significantly (p=0.007) lower in poorly differentiated than in well and moderately differentiated carcinomas, there were no differences in the expression levels between benign prostate hyperplasia, untreated primary and recurrent hormone-refractory prostate carcinomas (p=0.607). Altogether, missense mutations were detected in 2 out of 68 samples studied ( approximately 3%) by denaturing high-performance liquid chromatography (DHPLC) and sequencing. One of the samples with the mutation also exhibited a loss of a gene copy by fluorescence in situ hybridization (FISH). This was the only sample that exhibited a genetic alteration in both alleles, suggesting that the human pHyde is not a classical prostate tumor suppressor gene. The reduced expression of the gene found in some tumors warrant further studies.     

Invasive group B streptococcal infections in Finland: a population-based study

We analyzed surveillance data on group B streptococcus (GBS) infection in Finland from 1995 to 2000 and reviewed neonatal cases of early-onset GBS infection in selected hospitals in 1999 to 2000. From 1995 to 2000, 853 cases were reported (annual incidence 2.2-3.0/100,000 population). We found 32-38 neonatal cases of early-onset GBS disease per year (annual incidence 0.6-0.7/1,000 live births). In five hospitals, 35% of 26 neonatal cases of early-onset GBS infection had at least one risk factor: prolonged rupture of membranes, preterm delivery, or intrapartum fever. Five of eight mothers screened for GBS were colonized. In one case, disease developed despite intrapartum chemoprophylaxis. Although the incidence of early-onset GBS disease in Finland is relatively low, some geographic variation exists, and current prevention practices are suboptimal. Establishing national guidelines to prevent perinatal GBS is likely to reduce the incidence of the disease.     

Effects of carriers and storage of formulation on the lung deposition of a hydrophobic and hydrophilic drug from a DPI

The effects of carriers, the drug:carrier ratio and a 1 month storage period of a formulation in permeable polystyrene tube at 40 degrees C/75% RH on the in vitro pulmonary deposition of model drugs from dry powder inhaler (DPI) were evaluated. Budesonide (hydrophobic) and salbutamol sulphate (hydrophilic) were used as model drugs. Mannitol and glucose were used as the carriers. In addition, lactose 110M was used as the carrier for budesonide. The novel multiple dose Taifun was used as a DPI; Taifun is a breath-actuated inhaler that contains the powder formulation in a reservoir chamber. The respirable fractions (RF%) values of the drugs were determined by the "Andersen" sampler. The RF% values of salbutamol sulphate increased with an increase in the drug:carrier ratio before storage, whereas the drug:carrier ratio did not affect the RF% values after storage. In the case of budesonide, the drug:carrier ratio did not affect the RF% values before storage, instead the RF% values of budesonide increased with an increase in the drug:carrier ratio after storage. The RF% values of salbutamol sulphate decreased after storage of the formulation, this was not dependent on the carrier and the drug:carrier ratio. However, with budesonide the effect of the storage on its RF% values was dependent on which carrier was used and also the drug:carrier ratio. Overall, storage had less effect on the RF% values of budesonide than those of salbutamol sulphate. The highest RF% values of budesonide were obtained when mannitol was used as the carrier. Furthermore, the RF% values of salbutamol sulphate tended to be higher when mannitol was used as the carrier instead of glucose.