Nonhereditary p53 mutations in T-cell acute lymphoblastic leukemia are associated with the relapse phase

We have previously reported that greater than 60% of human leukemic T- cell lines possess mutations in the p53 tumor suppressor gene. To determine whether T-cell acute lymphoblastic leukemia (T-ALL) patient samples possess p53 mutations, we screened peripheral blood-and bone marrow-derived leukemia samples, taken at diagnosis and at relapse, for p53 mutations. Exons 4 through 9 and selected intron regions of the p53 gene were analyzed using polymerase chain reaction-single-strand conformation polymorphism and direct sequencing. p53 mutations were found in 0 of 15 T-ALL diagnosis samples, as compared with 10 of 36 (28%) T-ALL relapse samples. To determine whether p53 mutations play a role in the recurrence (relapse) of T-ALL, two special groups of T-ALL patients were studied: (1) a group of 8 relapse patients whose disease was refractory to chemotherapeutic treatment, and (2) a group of 6 "paired" T-ALL cell samples from patients for whom we possess both diagnosis and relapse samples. Three of 8 relapsed patients (37.5%) whose disease was refractory to the reinduction of remission by chemotherapy possessed missense mutations of the p53 gene. All 3 cases had mutations in exon 5. Among the paired samples, 3 of 6 patients harbored p53 mutations at disease recurrence, but possessed only wild- type p53 alleles at diagnosis. One case had mutation on exon 4, 1 case in exon 5, and 1 case in exon 8 with loss of heterozygosity. These data clearly indicate that recurrence of T-ALL is associated with missense mutations in p53. Our results indicate that (1) mutations of p53 do occur in T-ALL in vivo, and such mutations are associated with the relapse phase of the disease; and (2) p53 mutation is involved in the progression of T-ALL. This conclusion is supported by our observation that the introduction of T-ALL-derived mutant p53 expression constructs into T-ALL cell lines further increases their growth rate in culture, enhances cell cloning in methylcellulose, and increases tumor formation in nude mice.     

Differential modulation of proliferation, matrix metalloproteinase expression and invasion of human head and neck squamous carcinoma cells by c-erbB ligands

Evidence suggests that there is an association between the abnormal expression of members of the c-erbB receptor tyrosine kinase family and poor prognosis in head and neck squamous cell carcinomas (HNSCC). Until now, the relative contributions of different c-erbB ligands to HNSCC progression have not been clearly defined. In this paper we examined the effects of ligands with different c-erbB receptor specificities in terms of their stimulation of HNSCC proliferation, expression of matrix metalloproteinases (MMPs) and invasion. Heregulin-beta1 (HRG-β1; selective c-erbB3/B4 ligand) was found to stimulate proliferation in the majority of cell lines, whereas epidermal growth factor (EGF; EGFR ligand) and betacellulin (BTC; EGFR/B4 ligand) induced variable responses. All three ligands up-regulated multiple MMPs including collagenases, stromelysins, matrilysin and gelatinase B (MMP-9) but had minimal or no effects on gelatinase A (MMP-2), MT1-MMP and tissue inhibitors of MMPs (TIMPs). MMP-9 mRNA was induced to a higher level than other MMPs, although with slower kinetics. HRG-β1 was less active than EGF and BTC at the optimal concentration (relative potency of EGF:BTC:HRG = 3:4:1). In vitro invasion through Matrigel was also increased by all three ligands in proportion to their MMP up-regulation. A specific anti-EGFR monoclonal antibody (mAb ICR62) inhibited MMP up-regulation, migration and invasion induced by all three ligands, whereas an anti-c-erbB-2 mAb ICR12 inhibited mitogenic and motogenic responses following ligand stimulation but had no effect on MMP expression. These results suggest that c-erbB ligands may differentially potentiate the invasive phenotype of HNSCC via co-operative induction of cell proliferation, migration and proteolysis. The EGFR signalling pathway appears to be the dominant component controlling the proteolytic and invasive phenotype in HNSCC, whereas the c-erbB-2 signalling pathway is responsible, in part, for the mitogenic and motogenic effects of ligands. This revised version was published online in July 2006 with corrections to the Cover Date.     

Propofol hydroxylation by dog liver microsomes: assay development and dog breed differences.

Pharmacokinetic studies indicate that clearance of propofol, an anesthetic agent, is slower in greyhounds compared with other dog breeds. Biotransformation of propofol to 2,6-diisopropyl-1,4-quinol (4-hydroxypropofol) by cytochrome P-450 in the liver is proposed as a critical initial step in the elimination of this drug in dogs. Breed differences in the activity of this enzyme could therefore explain pharmacokinetic differences. An in vitro propofol hydroxylase assay was developed and then used to compare enzyme activities in liver microsomes from male greyhound, beagle, and mixed-breed dogs (five each). HPLC of incubate identified only one NADPH-dependent metabolite, which had a chromatographic retention time and UV absorbance, fluorescence, and mass spectra that were identical with authentic 4-hydroxypropofol standard. HPLC with fluorescence detection provided a highly sensitive quantitation method for 4-hydroxypropofol with a quantitation limit of 8 ng/ml using optimized excitation/emission wavelengths (288 nm/330 nm, respectively). Estimates of apparent K(m) and V(max) for propofol hydroxylation by microsomes from a male beagle dog were 7.3 microM and 3.8 nmol/mg/min, respectively. At a substrate concentration of 20 microM, propofol hydroxylase activity was significantly lower (p =.032) in greyhound microsomes (1.7 +/- 0.4 nmol/mg/min) compared with beagle microsomes (5.1 +/- 1.3 nmol/mg/min) but was not statistically different (p =.42) compared with mixed-breed microsomes (3.1 +/- 1.2 nmol/mg/min). These results indicate that there are breed differences in propofol hydroxylase activity and that deficient hydroxylation of propofol by one or more hepatic cytochrome P-450 isoforms may contribute to slow pharmacokinetic clearance of propofol by greyhounds.     

Stem bark alkaloids of Rauwolfia vomitoria

Forty-three indole alkaloids were isolated from 7 kg Rauwolfia vomitoria stem bark; 39 were identified and 2 partially characterised. At least 72 alkaloids classified into 19 types occur in the plant and the interrelationship of the alkaloids and their distribution throughout the plant has been discussed. The major alkaloids of the stems were heteroyohimbines (especially reserpiline) and N (a)-demethyldihydroindoles.     

Loss of sagittal plane correction after removal of spinal implants

STUDY DESIGN: A retrospective review of a clinical series was performed. OBJECTIVES: To evaluate the incidence of adult patients who experienced spinal collapse after spinal implant removal after a long spinal arthrodesis, and to assess the various factors that may influence the likelihood of collapse after implant removal. SUMMARY OF BACKGROUND DATA: Published reports describing the benefits or complications of spinal implant removal do not exist. Spinal implant removal, often considered a benign procedure, is even required by the Food and Drug Administration (FDA) for certain implants. METHODS: The medical records and radiographs of 116 consecutive adult patients with long posterior instrumented fusions (>5 segments) were reviewed. The information obtained included original diagnosis, patient age, number of previous surgeries before implant removal, levels of anterior and posterior fusion, time from fusion to implant removal, time from implant removal to failure, and reason for hardware removal. Radiographs also were assessed including scoliosis, lordosis, and kyphosis measurements before implant removal, after hardware removal, after failure, and after revision surgery. RESULTS: Of 116 patients, 14 underwent spinal implant removal. Most of these patients reported prominent implants either proximally in the thoracic spine or distally in the ilium (Galveston technique). Of these 14 patients, 4 experienced increased pain and collapse after implant removal despite thorough intraoperative explorations demonstrating solid fusion. CONCLUSIONS: Spinal implant removal after long posterior fusion in adults may lead to spinal collapse and further surgery. Removal of instrumentation should be avoided or should involve partial removal of the prominent implant.     

Validation of serotonin (5-hydroxtryptamine) as an in vitro substrate probe for human UDP-glucuronosyltransferase (UGT) 1A6.

Investigation of human UDP-glucuronosyltransferase (UGT) isoforms has been limited by a lack of specific substrate probes. In this study serotonin was evaluated for use as a probe substrate for human UGT1A6 using recombinant human UGTs and tissue microsomes. Of the 10 commercially available recombinant UGT isoforms, only UGT1A6 catalyzed serotonin glucuronidation. Serotonin-UGT activity at 40 mM serotonin concentration varied more than 40-fold among human livers (n = 54), ranging from 0.77 to 32.9 nmol/min/mg of protein with a median activity of 7.1 nmol/min/mg of protein. Serotonin-UGT activity kinetics of representative human liver microsomes (n = 7) and pooled human kidney, intestinal and lung microsomes and recombinant human UGT1A6 typically followed one enzyme Michaelis-Menten kinetics. Serotonin glucuronidation activity in these human liver microsomes had widely varying V(max) values ranging from 0.62 to 51.3 nmol/min/mg of protein but very similar apparent K(m) values ranging from 5.2 to 8.8 mM. Pooled human kidney, intestine, and lung microsomes had V(max) values (mean +/- standard error of the estimates) of 8.8 +/- 0.4, 0.22 +/- 0.00, and 0.03 +/- 0.00 nmol/min/mg of protein (respectively) and apparent K(m) values of 6.5 +/- 0.9, 12.4 +/- 2.0, and 4.9 +/- 3.3 mM (respectively). In comparison, recombinant UGT1A6 had a V(max) of 4.5 +/- 0.1 nmol/min/mg of protein and an apparent K(m) of 5.0 +/- 0.4 mM. A highly significant correlation was found between immunoreactive UGT1A6 protein content and serotonin-UGT activity measured at 4 mM serotonin concentration in human liver microsomes (R(s) = 0.769; P < 0.001) (n = 52). In conclusion, these results indicate that serotonin is a highly selective in vitro probe substrate for human UGT1A6.     

Communicating oesophageal duplication

Oesophageal duplications are rare congenital abnormalities. Most of them do not communicate with the oesophageal lumen. We present a very uncommon finding of communicating oesophageal duplication in which the connection between the oesophagus and its duplicate portion was demonstrated by CT.     

Squamous cell carcinoma of the penis: Multivariate analysis of prognostic factors and natural history in a monocentric study with a conservative policy

Background: Penile carcinoma is uncommon in Western countries. Here we report on a large series of patients with squamous cell carcinoma (SCC) of the penis, describing prognostic factors, survival and therapeutic results.Patients and methods: From 1973 to 1993, 102 patients with invasive SCC of the penis were treated at the Institut Gustave-Roussy. Precancerous lesions and conditions associated with penis cancer were analyzed retrospectively. Survival curves were estimated by the Kaplan–Meier method, and groups were compared for outcome by the log rank test for univariate comparisons and by Cox's proportional hazards model for multivariate analysis.Results: The median age at onset was 58 years. Sixty-nine patients presented with Jackson's stage I disease, 17 with stage II and 15 with stage III. The interval between the manifestation of symptoms and the diagnosis was more than a year in 13.7% of cases. Precancerous lesions were found in 17 (16.6%) patients, and a history of phimosis was noted in 25 (24.5%). In situ and invasive carcinoma were observed together in 17 (16.6%) cases and dysplasia was associated with invasive carcinoma in eight (7.8%) further cases.Conservative treatment was administered whenever feasible. Interstitial brachytherapy was performed alone or associated with limited surgery (local excision or circumcision) in 72 (70.6%) patients. Of the 28 patients with a local relapse, nine have died of their neoplasms (32%) compared to 21 of 28 patients with lymph node relapse (75%).The median follow-up was 111 months. Disease-free survival, disease-specific survival and overall survival were, respectively, 56%, 72% and 63% at five years and 42%, 66% and 50% at 10 years. Age (P = 0.01), the N status (P < 0.00001) or palpable nodes (P < 0.0038), corpus involvement (P = 0.006) and a verrucous histology (P = 0.038) had significant prognostic relevance for survival in the univariate analysis whereas the performance status, T status and Broders' grade did not. In the multivariate analysis only two parameters, involvement of the corpus (P < 0.0001) and palpable nodes (P = 0.009), were singled out as being independent variables influencing survival. A subgroup of nine patients with verrucous histologies were distinguished by their freedom from node involvement. These patients had an excellent prognosis: all are alive and disease-free. Penile integrity was preserved during follow-up in 54 patients (52.9%), 31 of whom are still alive. Of 72 patients treated by a conservative approach including brachytherapy, long-term penile integrity was maintained in 49 (68%).Conclusion: Corpus involvement and clinically palpable nodes are highly statistically significant independent factors influencing overall survival. Node relapses remain a major cause of death. Thus, better management of lymph nodes is essential for improving survival even when conservative therapy is used to treat the primary.