Distribution of estramustine in the BT4C rat glioma model

 Estramustine (EaM), a carbamate ester of 17β-estradiol and nor-nitrogen mustard, is a cytotoxic compound with antitumoral effect in malignant glioma in vitro and in vivo . However, knowledge of the pharmacokinetics of EaM in experimental glioma is limited. The objective of this study was therefore to investigate further the distribution of EaM in the BT4C rat glioma model. Assessment of EaM uptake and distribution was performed by quantitative whole-body autoradiography. In addition, the uptake of EaM and its metabolites estromustine (EoM), estradiol, and estrone were analyzed by gas chromatography. EaM was taken up from the circulation and was found to be the main product in glioma tissue. Whole-body autoradiography after [¹⁴C]-EaM administration revealed a strong ¹⁴C label simultaneously in tumor and normal brain tissue at 0.5 h after drug administration. In tumor tissue, sustained high levels of ¹⁴C label were detected at 12 h after drug administration. In contrast to the tumor, radioactivity in normal brain tissue rapidly leveled off, indicating a retention of radioactivity in the tumor. The tumor/brain radioactivity ratio reached a peak of 4.5 at 12 h after drug administration. High levels of ¹⁴C label were also found in pulmonary tissue. By gas chromatography, EoM was found to be the main metabolite in plasma. However, EaM reached higher levels in tumor tissue, with the mean tumor/plasma ratio being 11.7 as compared with 2.0 for EoM. Only low plasma levels of the estrogen metabolites were detected. In conclusion, EaM is taken up in the BT4C rat glioma tissue and is retained in the tumor as compared with normal brain tissue and plasma. EaM showed a greater selectivity for tumor tissue, exhibiting a high tumor/plasma ratio as compared with EoM. The distribution pattern after administration of EaM, as evaluated by both whole-body autoradiography and gas chromatography, supports the earlier suggestion that the uptake is related to a protein with EaM-binding characteristics. Received: 12 January 1997 / Accepted: 28 August 1997     

Routine application of a novel MLPA-based first-line screening test uncovers clinically relevant copy number aberrations in haematological malignancies undetectable by conventional cytogenetics

Objective

The presence of numerical and/or structural chromosomal abnormalities is a frequent finding in clonal hematopoietic malignant disease, typically diagnosed through routine karyotyping and/or fluorescent in situ hybridization (FISH) analysis. Recently, the application of array comparative genomic hybridization (aCGH) has uncovered many new cryptic genomic copy number imbalances, most of which are now recognized as clinically useful markers of haematological malignancies. In view of the limitations of both FISH and aCGH techniques, in terms of their routine application as a first line screening test, we designed a new multiple ligation-dependent probe amplification (MLPA) probemix for use in addition to classic karyotype analysis.

Methods

A novel MLPA probemix was developed to interrogate copy number changes involving chromosomal regions: 2p23-24 (MYCN, ALK), 5q32-34 (MIR145A, EBF1, MIR146A), 6q21-27, 7p12.2 (IKZF1), 7q21-36, 8q24.21 (MYC), 9p24 (JAK2 V617F point mutation), 9p21.3 (CDKN2A/2B), 9p13.2 (PAX5), 10q23 (PTEN), 11q22.3 (ATM), 12p13.2 (ETV6), 13q14 (RB1, MIR15A, DLEU2, DLEU1), 17p13.1 (TP53), and 21q22.1 (RUNX1/AML1) and was applied to DNA extracted from 313 consecutive bone marrow patient samples, referred for routine karyotype analysis.

Results

More than half of the samples originated from newly investigated patients. We discovered clinically relevant genomic aberrations, involving a total of 24 patients (8%) all with a normal karyotype, which would have remained undiagnosed.

Discussion

Our data clearly indicate that routine application of this MLPA screening panel, as an adjunct to karyotype analysis, provides a sensitive, robust, rapid and low-cost approach for uncovering clinically important genomic abnormalities, which would have otherwise remained undetected.     

Enamel Matrix Derivative in Intrabony Defects: Prognostic Parameters of Clinical and Radiographic Treatment Outcomes

Background: The purpose of this retrospective case series study is to identify possible preoperative parameters that could predict postoperative probing depth (PD), clinical attachment level (CAL) gain, or radiographic defect resolution in intrabony defects treated with enamel matrix derivative (EMD). Methods: Sixty‐one chronic periodontitis patients, each contributing a 2‐ or 3‐wall intrabony defect treated with EMD, were included. Clinical parameters recorded included the following: PD; CAL; gingival margin position; supracrestal soft tissue (SST); surgical distances of cemento‐enamel junction (CEJ) to bone crest (CEJ‐BC), CEJ to base of the defect (CEJ‐BD), and BC to BD (BC‐BD); and depth of 2‐ and 3‐wall components. Radiographic parameters recorded included the following: CEJ‐BC, CEJ‐BD, BC‐BD distances, and radiographic defect angle. Postoperative assessments were performed at 12 months. Results: The probability of postoperative PD >4 mm increased 1.6‐fold (odds ratio [OR] = 1.6; 95% confidence interval [CI] = 1.2 to 2.3) with each 1‐mm baseline PD increase. Baseline PD and surgical CEJ‐BD were statistically significant predictors of CAL gain; the greater the baseline PD (OR = 0.5; 95% CI = 0.3 to 0.8) and bone loss (OR = 0.6; 95% CI = 0.3 to 0.9), the less likely that postoperative CAL gain was ≤3 mm. Smoking and SST were significantly associated with defect resolution; failure to achieve ≥65% defect resolution was six‐fold greater for smokers (OR = 6.5; 95% CI = 1.7 to 24.5) and almost double (OR = 1.7; 95% CI = 1.1 to 2.8) for each millimeter of SST increase. Conclusion: In EMD‐treated intrabony defects, baseline PD predicts both CAL gain and postoperative PD. Smoking and SST are predictors of defect resolution.     

Hypermethylation of CpG islands in the promoter region of the p15INK4B gene in childhood acute leukaemia

It has been reported that the cyclin-dependent kinase inhibitor (CDKI) gene p15INK4B is frequently inactivated by genetic alterations and may be responsible for various malignant tumours. Another way of inactivation of this CDKI is by hypermethylation of 5'CpG islands in the promoter region of the p15INK4B gene and this inactivation seems to be a frequent event in various haematological malignancies. In the present study, we investigated the methylation status of the p151NK4B gene to clarify its role in the pathogenesis of childhood acute myeloid (AML) and acute lymphoblastic leukaemia (ALL). The study included 23 cases of B-cell origin ALL, 13 cases of T-cell origin ALL, 32 cases of AML, and 10 apparently healthy controls. Hypermethylation was studied by methylation-specific polymerase chain reaction. Hypermethylation of the p15INK4B gene was more frequent in cases with T-cell origin ALL (46.2%), but similar among children with B-cell origin ALL (13.0%) and AML (18.8%). Hypermethylation of p15INK4B may be involved in the pathogenesis of T-cell origin ALL, but not in that of AML or B-cell origin ALL.