A gene transfer strategy for making bone marrow cells resistant to trimetrexate

Trimetrexate (TMTX) is an anticancer drug with potential advantages over the more commonly used antifolate, methotrexate (MTX); however, its use has been limited by severe myelosuppression. Retroviral vectors containing mutant dihydrofolate reductase (DHFR) genes have been used to protect bone marrow cells from MTX, suggesting a similar approach could be used for TMTX. We first screened six variants of human DHFR to determine which allowed maximal TMTX resistance in fibroblasts. A variant enzyme containing a Leu-to-Tyr mutation in the 22nd codon (L22Y) was best, allowing a 100-fold increase in resistance over controls. Murine hematopoietic progenitor cells transduced with an L22Y- containing retroviral vector also showed high-level TMTX resistance in vitro. Mice reconstituted with L22Y-transduced bone marrow cells were challenged with a 5-day course of TMTX to determine whether hematopoiesis could be protected in vivo. Transfer of the L22Y vector resulted in consistent protection from TMTX-induced neutropenia and reticulocytopenia at levels that correlated with the proviral copy number in circulating leukocytes. We conclude that the L22Y vector is highly effective in protecting hematopoiesis from TMTX toxicity and may provide a means for increasing the therapeutic utility of TMTX in certain cancers.     

Commingling and segregation analyses: comparison of results from a simulation study of a quantitative trait

Commingling analysis is commonly used to provide preliminary evidence for a single genetic locus with a major effect on the quantitative trait of interest. In this paper, the effectiveness of commingling analysis as a screening technique to identify samples for segregation analysis is assessed by applying both commingling and segregation analyses to samples of simulated pedigree data in which a major locus is segregating in the presence of polygenes and an individual-specific environmental effect. Under the circumstances simulated here, there is evidence for a single locus from segregation analysis but not from commingling analysis in at least 20% of the samples. No more than 2% of the samples provided evidence for commingling but not for segregation of a single locus. Comparisons of the samples that give evidence for both commingling and segregation, evidence for one but not the other, and no evidence for either show that evidence for commingling depends on the distributional characteristics of the trait in the sample, while support for the single locus from segregation analysis depends on both the distributional characteristics as well as the transmission of the rarer allele from parents to offspring. Since lack of commingling does not rule out the existence of a single locus in the realistic situations considered here, commingling analysis has limited usefulness as a screening technique for the presence for a single locus. In contrast, evidence for commingling does suggest the possibility that a single locus has a major effect on the trait and commingling analysis can provide guidance in the choice of initial parameter estimates for segregation analysis.     

Small-bowel obstruction caused by intraluminal migration of prosthetic grafts

Two patients who had undergone prosthetic grafting in the venous system presented with small-bowel obstruction. The first had undergone mesocaval shunting with a Dacron graft and the second, replacement of the inferior vena cava with a polytetrafluoroethylene graft after an extensive resection of retroperitoneal tumor. In both instances the grafts were within the lumen of the small bowel, causing a mechanical obstruction. No evidence of retroperitoneal bleeding subsequent to detachment of the grafts was noted.     

Anticonvulsant effects of MK-801 and glycine on hippocampal afterdischarge

The effects of glycine and MK-801 on hippocampal seizure threshold and afterdischarge (AD) were determined in freely moving rats implanted with intracranial electrodes. The 0.25 mg/kg (ip) dose of MK-801 significantly reduced the primary hippocampal and cortical AD but induced neurological deficit in 4 of 16 rats. The 0.25 mg/kg MK-801 dose also significantly increased the seizure threshold as compared to the vehicle control treatment and reduced the rebound cortical AD as compared to the control (nondrug) seizure response. The 40 mmol/kg (po) glycine dose significantly reduced the rebound hippocampal and cortical AD without inducing neurological deficit. The 40 mmol/kg glycine dose did not significantly alter the response to the ineffective 0.125 mg/kg MK-801 dose. These results demonstrate the differential effects of MK-801 and glycine on primary and rebound hippocampal AD, respectively, which further establishes the independence of the hippocampal seizure AD parameters. The lack of interaction between MK-801 and glycine was unexpected considering that glycine potentiates MK-801 receptor binding as well as the activity of other anticonvulsants.     

Pulmonary arteriovenous malformations: techniques and long-term outcome of embolotherapy

Over a 10-year period, 276 pulmonary arteriovenous malformations (PAVMs) were occluded with balloon embolotherapy in 76 patients, 67 (88%) of whom had hereditary hemorrhagic telangiectasia. Eleven patients (14%) were discovered by means of family screening with measurement of arterial blood gases and chest radiography. Epistaxis, dyspnea, hemoptysis, and hemothorax occurred in 79%, 71%, 13%, and 9% of patients, respectively. Clinical histories of strokes and transient ischemic attacks were present in 18% and 37% of patients, respectively. Computed tomographic scans of 59 patients showed stroke in 36%. Sixty-five percent of PAVMs were located in the lower lobes, which correlated with the finding of more pronounced hypoxemia in the upright position. After embolotherapy, symptomatic hypoxemia was corrected, and serial values have remained constant for 5 years. Complications were minimal, and no patient required surgery. Balloon embolotherapy is effective long-term therapy for PAVMs, and family screening should be pursued because of the possibility of a higher frequency of paradoxical embolization (stroke) than previously recognized.