Agonist-induced GTPgamma35S binding mediated by human 5-HT(2C) receptors expressed in human embryonic kidney 293 cells

The 5-HT(2C) receptor as heterologously expressed in various mammalian cells mediates inositol 1,4,5-triphosphate (IP(3)) signal by activating G(q/11) subtypes of G proteins, but minimally promotes agonist-induced GTPgamma35S binding in membranes due to slow GTP turnover rates of the G proteins. Here we discovered robust (over 200%) agonist-induced GTPgamma35S binding mediated by the human receptor expressed in human embryonic kidney (HEK) 293 cells, and investigated its pharmacology. Agonists concentration-dependently increased GTPgamma35S binding in isolated membranes, which was competitively blocked by antagonists. Intrinsic efficacies of agonists from GTPgamma35S binding were comparable to those from IP(3) measurement. Pertussis toxin treatment largely blocked serotonin-induced GTPgamma35S binding, serotonin high affinity sites by 70% without altering the total binding sites, and reduced IP(3) release by 40%. GTPgamma35S-bound Galpha subunits from serotonin-activated membranes were mainly Galpha(i), judging from immobilization studies with various Galpha-specific antibodies. Inhibition of forskolin-stimulated cAMP formation, however, was not observed. Apparently, the 5-HT(2C) receptor-mediated GTPgamma35S binding is a unique phenotype observed in HEK293 cells, reflecting its coupling to pertussis toxin-sensitive G(i) subtypes, which contribute to the IP(3) signal, along with pertussis toxin-insensitive G(q/11) subtypes.     

Combination chemotherapy with cisplatin, bleomycin, and methotrexate in patients with advanced head and neck cancer

Twenty-six patients with advanced head and neck cancer, 22 of whom had failed prior surgery and/or radiotherapy, were treated with a combination regimen of cis-diamminedichloroplatinum II, bleomycin, and methotrexate. There were no complete responses. Nine patients achieved a partial response (35%). Three of the four (75%) patients without prior therapy achieved a partial response while only 6 of the 22 patients (27%) with prior surgery and/or radiation therapy obtained a partial response. The median response duration was 3 months. Patients with a partial response did not live significantly longer than those who did not live significantly longer than those who did not respond. Toxic reactions were frequent: there were three episodes of sepsis secondary to leukopenia and two cases of bleomycin pulmonary toxicity. Mucositis was noted in 40% and nausea and vomiting in 60% of patients. We conclude that this triple-drug regimen has little value in the treatment of patients with advanced squamous cell carcinoma of the head and neck who have failed prior surgery and radiotherapy.     

Effects of a high-selenium yeast supplement on celecoxib plasma levels: a randomized phase II trial.

A combination of celecoxib and selenium was used in a randomized double-blind Phase II trial as a preliminary study to a multicenter Phase III colorectal cancer chemoprevention trial using these two agents together. The purpose of this trial was to determine whether high-selenium baker's yeast [(Saccharomyces cerevisiae) 200 microg once daily] in combination with celecoxib (400 mg once daily) altered the steady-state plasma concentration of celecoxib or produced clinically significant toxicities. Seventy-three healthy subjects (ages 40-75 years) were recruited to the 6-week study from the general local population and were randomized to either the celecoxib plus selenized baker's yeast group or the celecoxib plus placebo group after a 2-week run in period of celecoxib only. Blood samples were taken at baseline (to document that there was no evidence of celecoxib intake), after the 2-week run-in period on celecoxib to verify steady-state blood levels of this agent, and at end of study (4 weeks postrandomization). Toxicities were monitored at 2 weeks after initiation of celecoxib, at 4 weeks after initiation, and at the end of the study. Blood level concentrations of celecoxib did not differ between the two groups as determined by high-performance liquid chromatography analysis nor were there significant differences in blood chemistry values between the two groups. Subjects' self-report of general physical toxicities was uncommon and limited to National Cancer Institute toxicity grade 2 or less; however, 2 female participants (3%) were removed from the study medications because of grade 2 edema and significant weight gain after 2 and 2.5 weeks of celecoxib administration. In conclusion, high-selenium yeast and celecoxib can be taken at the described doses with minimum short-term negative effects. In future Phase III chemoprevention trials of celecoxib, weight gain should be carefully monitored, and participants should be made aware of this potential side effect before study entry.     

Gemcitabine and ISIS-2503 for patients with locally advanced or metastatic pancreatic adenocarcinoma: a North Central Cancer Treatment Group phase II trial.

PURPOSE: Gemcitabine remains the standard therapy for metastatic pancreatic adenocarcinoma (ACA), but has limited activity. ISIS-2503 is an antisense compound directed against H-ras with preclinical activity against pancreatic ACA in tumor models. The combination of ISIS-2503 and gemcitabine has been evaluated in a prior phase I study. METHODS: Patients with metastatic or locally advanced pancreatic ACA not amenable to surgery or local radiation received gemcitabine 1,000 mg/m(2) intravenously over 30 minutes on days 1 and 8 and ISIS-2503 6 mg/kg/d as a continuous intravenous infusion over 14 days of an every-3-weeks cycle. Responses were monitored by radiologic imaging every 6 weeks. RESULTS: Forty-eight eligible patients were enrolled, 43 with metastatic disease. Median follow-up was 12.6 months (range, 2.2 to 16.8 months) for living patients. A median of four cycles of treatment was given (range, 1 to 18 cycles). All patients were assessable for response and toxicity. The 6-month survival percentage was 57.5% (95% CI, 44.9% to 73.5%) and the median survival was 6.6 months. The response rate was 10.4% (one complete response, four partial responses). Clinically significant toxicity was limited except for one fatal pulmonary embolism. CONCLUSION: This study shows a promising response rate to the combination of gemcitabine and ISIS-2503 in patients with pancreatic ACA. The observed 6-month survival rate in these patients met our protocol-defined criteria for success. This regimen is tolerable, but is of unclear benefit. Additional studies evaluating the role of gemcitabine and ISIS-2503 in the treatment of pancreatic ACA should be considered.     

A randomized controlled trial of fluorouracil plus leucovorin, irinotecan, and oxaliplatin combinations in patients with previously untreated metastatic colorectal cancer.

PURPOSE: Three agents with differing mechanisms of action are available for treatment of advanced colorectal cancer: fluorouracil, irinotecan, and oxaliplatin. In this study, we compared the activity and toxicity of three different two-drug combinations in patients with metastatic colorectal cancer who had not been treated previously for advanced disease. PATIENTS AND METHODS: Patients were concurrently randomly assigned to receive irinotecan and bolus fluorouracil plus leucovorin (IFL, control combination), oxaliplatin and infused fluorouracil plus leucovorin (FOLFOX), or irinotecan and oxaliplatin (IROX). The primary end point was time to progression, with secondary end points of response rate, survival time, and toxicity. RESULTS: A total of 795 patients were randomly assigned between May 1999 and April 2001. A median time to progression of 8.7 months, response rate of 45%, and median survival time of 19.5 months were observed for FOLFOX. These results were significantly superior to those observed for IFL for all end points (6.9 months, 31%, and 15.0 months, respectively) or for IROX (6.5 months, 35%, and 17.4 months, respectively) for time to progression and response. The FOLFOX regimen had significantly lower rates of severe nausea, vomiting, diarrhea, febrile neutropenia, and dehydration. Sensory neuropathy and neutropenia were more common with the regimens containing oxaliplatin. CONCLUSION: The FOLFOX regimen of oxaliplatin and infused fluorouracil plus leucovorin was active and comparatively safe. It should be considered as a standard therapy for patients with advanced colorectal cancer.     

Smoking exposure as a risk factor for prevalent and recurrent colorectal adenomas.

Colorectal adenomatous polyps are considered to be the precursor lesion of colorectal cancer (1-3). Greater understanding of the association between smoking and adenoma development enable better detection and prevention of colorectal cancer. This study was conducted in men and women, ages 40-80, participating in a randomized trial testing the effects of wheat bran fiber supplement on adenoma recurrence. First, we investigated smoking exposure (status, cigarettes/day, and years of smoking) and colorectal adenoma characteristics (location, histology, size, and multiplicity) at baseline colonoscopy (n = 1429). Second, we evaluated smoking exposure and adenoma recurrence (n = 1304). The prevalence of distal versus proximal adenomas was greater for < or =30 cigarettes/day [odds ratio (OR), 1.48; 95% CI, 1.02-2.16] and 15 to <25 years of smoking (OR, 1.95; 95% CI, 1.23-3.09) compared with never smokers. Tubular versus villous histology prevalence was increased for > or =30 cigarettes and > or =35 years of smoking (OR, 1.74; 95% CI, 1.21-2.49 and OR, 1.74; 95% CI, 1.24-2.45, respectively) compared with never-smokers. Years of smoking increased prevalence of multiple versus single adenomas, whereas cigarettes/day and years of smoking were associated with large adenomas (> or =1 cm) prevalence as compared with small lesions (< or =0.5 cm). Greater than 35 years of smoking was significantly associated with an increased risk of adenoma recurrence (OR, 1.42; 95% CI, 1.01-1.98). These results suggest that the association between smoking and adenoma prevalence varies by the characteristic of the lesion. Furthermore, the association between smoking and adenoma recurrence is modest and was only significant after a long duration of exposure. Additional investigations that characterize the genetic changes in specific subgroups of prevalent and recurrent adenomas associated with smoking exposure are needed.     

Total but not resting energy expenditure is increased in infants with ventricular septal defects

OBJECTIVE: The purpose of this study was to determine the effect of left-to-right shunting on the resting energy expenditure (REE), total energy expenditure (TEE), and energy intake in a group of 3- to 5-month-old infants with moderate to large unrepaired ventricular septal defects (VSDs) compared with age-matched, healthy infants. METHODS: Eight infants with VSDs and 10 healthy controls between 3 to 5 months of age participated in the study. Indirect calorimetry was used to measure REE and the doubly-labeled water method was used to measure TEE and energy intake. An echocardiogram and anthropometric measurements were performed on all study participants. Daily urine samples were collected at home for 7 days. Samples were analyzed by isotope ratio mass spectrometry. Data were compared using analysis of variance. RESULTS: No significant differences were found in REE (VSD, 42.2 +/- 8.7 kcal/kg/d; control, 43.9 +/- 14.1 kcal/kg/d) or energy intake (VSD, 90.8 +/- 19.9 kcal/kg/d; control, 87.1 +/- 11.7 kcal/kg/d) between the groups. The percent total body water was significantly higher in the VSD infants and the percent fat mass was significantly lower. TEE was 40% higher in the VSD group (VSD, 87.6 +/- 10.8 kcal/kg/d; control, 61.9 +/- 10.3 kcal/kg/d). The difference between TEE and REE, reflecting the energy of activity, was 2.5 times greater in the VSD group. CONCLUSIONS: REE and energy intake are virtually identical between the two groups. Despite this, infants with VSDs have substantially higher TEE than age-matched healthy infants. The large difference between TEE and REE in VSD infants suggests a substantially elevated energy cost of physical activity in these infants. These results demonstrate that, although infants with VSDs may match the energy intake of healthy infants, they are unable to meet their increased energy demands, resulting in growth retardation.     

Isolation and characterization of propagable cell lines (HUNC) from the androgen-sensitive Dunning R3327H rat prostatic adenocarcinoma

The Dunning H rat prostate tumor (R3327H) is a widely used experimental model of human prostatic adenocarcinoma (CaP). The Dunning H tumor has been characterized as androgen-sensitive, androgen-receptor (AR) positive, prostate-specific antigen and prostatic acid phosphatase (PAP) positive. To date, the tumor has been maintained by serial passage in vivo because of the lack of an in vitro cell line that retains the characteristics of the in vivo tumor. The objective of the present study was to establish a propagable cell line from R3327H adenocarcinoma that maintained androgen sensitivity and expression of AR, PSA and PAP. Tissue harvested from an in vivo R3327H tumor was dissociated with collagenase and placed into Richter's improved media (with supplements). A cytokeratin-positive epithelial cell line (HUNC- E) and a vimentin-positive stromal cell line (HUNC-S) were generated from the primary culture, subcultured continuously for >300 days, and passaged >50 times. Survival of the HUNC-E cell line in vitro depended on several media supplements, including nicotinamide, insulin, transferrin, selenium and epidermal growth factor (EGF). HUNC-E cells expressed AR and produced PSA and PAP throughout the culture period, as confirmed by immunocytochemistry and Western blot analyses. Addition of 14 nM testosterone (T) or dihydrotestosterone (DHT) to HUNC-E cells, stimulated DNA synthesis as well as anchorage-independent growth and PSA production, which demonstrated the androgen-sensitive nature of the cells in vitro. When HUNC-E and HUNC-S cells were combined in a 3:1 ratio and introduced subcutaneously into syngeneic male hosts, tumors formed in 2/3 animals with an average latency of 7 months. RT-PCR and immunocytochemical characterization of the HUNC cell lines revealed that the cells expressed several growth factors and their cognate receptors, including HGF, TGF-alpha and the TGF-betas, indicating the establishment of potential autocrine loops in the neoplastic cells. The HUNC-E and HUNC-S CaP cell lines, which retain the characteristics of the epithelial and stromal components of the in vivo R3327H tumor, will allow a more thorough and informative molecular and biological analysis of prostatic adenocarcinoma.      

STRESS FRACTURES: EFFECT OF PRIOR PHYSICAL ACTIVITY,SPORTS PARTICIPATION AND MILITARY TRAINING

A study was carried out to find out the effects of prior physical activity, sports participation and prior military training on the incidence of stress fractures among Gentlemen Cadets (GC''s) undergoing military training at Indian Military Academy (IMA). One thousand and fourteen GC''s were followed up for a period of 12 weeks. Thirty-seven GC''s developed stress fractures during the study period. The incidence of stress fractures was significantly higher in GC''s without any prior military training (p=0.0009). They were compared with 100 healthy controls drawn from the study population to study the influence of the other mentioned factors. There was no significant association between prior physical activity and stress fractures (OR=0.74, 95% CL=0.26 to 2.05, p=0.688). There was also no significant relationship between sports participation and stress fractures (OR=0.79. 95% CCL=0.35 to 1.81, p=0.684).KEY WORDS: Risk factors, Stress fractures