PET imaging of oncogene overexpression using 64Cu-vasoactive intestinal peptide (VIP) analog: comparison with 99mTc-VIP analog.

The purpose of this study was to assess the feasibility of PET imaging of oncogene VPAC1 receptors overexpressed in human breast cancer cells. METHODS: Vasoactive intestinal peptide (VIP) analog (TP3982) was synthesized to harbor a carboxy-terminus lysine (Lys) residue separated from VIP-asparagine (Asn(28)) by 4-aminobutyric acid (Aba) as a spacer. Lys was derivatized with diaminopropionic acid coupled to a pair of dibenzoylthioglycolic acid residues as protecting groups. The analog was labeled with (64)Cu at pH 9 ((64)Cu-TP3982) and (99m)Tc at pH 12 ((99m)Tc-TP3982). (99m)Tc-TP3982 and VIP derivatized with Aba-GAGG and labeled with (99m)Tc ((99m)Tc-TP3654) were used as reference agents. Smooth muscle relaxivity assays performed with each derivative and compared with unaltered VIP(28) demonstrated functional integrity. In vitro stability of (64)Cu-TP3982 was determined by challenging the complex with 100-mol excess of diethylenetriaminepentaacetic acid (DTPA), human serum albumin (HSA), and cysteine. In vivo stability was determined in urine and serum for up to 24 h. The mass of the Cu-TP3982 complex was determined by mass spectrometry. Human T47D breast tumor xenografts were grown in athymic nude mice. Planar scintigraphic imaging was performed at 4 and 24 h after the intravenous administration of (99m)Tc-TP3982 and (99m)Tc-TP3654 and PET imaging was performed using a small animal MOSAIC PET scanner, also at 4 and 24 h after injection of (64)Cu-TP3982. Tissue-distribution studies were also performed. In a separate experiment, receptors were blocked by intravenous injection of authentic VIP(28) 30 min before the administration of (64)Cu-TP3982 and tissue distribution was examined. RESULTS: (64)Cu-TP3982 labeling yields were 98% +/- 1.2% and those for (99m)Tc-TP3982 and (99m)Tc-TP3654 were 98.2% +/- 1.1% and 97% +/- 1.6%, respectively. The biologic activity of both VIP analogs was uncompromised. When (64)Cu-TP3982 was challenged with 100-mol excess of DTPA, HSA, or cysteine, >98% radioactivity remained as (64)Cu-TP3982. In vivo, >98% of (64)Cu circulating in plasma remained as (64)Cu-TP3982. Of the (64)Cu excreted in urine 4, 20, and 24 h after injection, >98%, 89.9% +/- 0.9%, and 85% +/- 3%, respectively, were bound to TP3982. The mass of Cu-TP3982 as determined by surface-enhanced laser desorption/ionization time of flight (SELDI-TOF) was 4,049.7 Da. Four hours after receptor blocking with VIP(28), there was a significant reduction in uptake of all tissues except in the liver. With (64)Cu-TP3982, the 4-h postinjection tumor uptake was 10.8 +/- 2.1 %ID/g versus 0.5 +/- 0.02 %ID/g and 0.24 +/- 0.08 %ID/g for (99m)Tc-TP3982 and (99m)Tc-TP3654, respectively. Twenty-four hours after injection, the corresponding numbers were 17 +/- 0.7 %ID/g, 0.77 +/- 0.1 %ID/g, and 0.23 +/- 0.1 %ID/g. The severalfold greater uptake (21.2-74) of (64)Cu-TP3982 is attributable to the in vivo stability of the agent. CONCLUSION: The results suggest that the uncompromised biologic activity and the significantly greater tumor uptake of (64)Cu-TP3982, combined with the high sensitivity and enhanced resolution of PET imaging, make (64)Cu-TP3982 highly desirable for further studies in PET imaging of oncogene receptors overexpressed in breast and other types of cancers.     

Scale-up of Adhesive Transdermal Drug Delivery Systems

Pharmaceutical Research -     

Protection against 7,12-dimethylbenzanthracene-induced tumour initiation by protein A in mouse skin.

Protein A is an immunostimulating glycoprotein obtained from Staphylococcus aureus Cowan I. Its antitumour activity is proven in various tumour models. Its ability to provide protection against tumour initiation by the chemical carcinogen 7,12-dimethylbenzanthracene (DMBA) has been investigated in the present study using a mouse skin model of two-stage carcinogenesis. Protein A was administered intraperitoneally (1 microgram/animal 20 g body wt.) twice a week for 2 weeks, prior to initiation by DMBA. The promotion was performed by twice weekly applications of 12-O-tetradecanoyl phorbol-13-acetate (TPA) (3 or 5 micrograms/animal in 100 microliters acetone). Protein A provided significant protection to animals from DMBA-induced tumour initiation as was observed by the decrease in cumulative number of tumours, percent of animals developing tumours, number of tumours per animal and rate of tumour growth. Our data indicate that protein A has anticarcinogenic properties.     

Effects of MR imaging on murine natural killer cell cytotoxicity

To determine the effect of MR imaging on the immune system, 21 male C57BL/6 X DBA/2 F1 mice were exposed to MR imaging at a field strength of 0.15 T for 2 hr. Another nine mice (controls) were sham exposed for the same amount of time. Mice were sacrificed and their spleens removed 24, 72, and 144 hr after the exposure (MR or sham). Spleen cell suspensions were passed over nylon wool columns and then used as effector cells in a short-term natural killer cell cytotoxicity assay with 51Cr-labeled YAC-1 cells as target cells. The results showed no evidence of decreased cytotoxicity due to exposure to MR. On the contrary, at all three times after exposure and for all target-to-effector cell ratios, mean cytotoxicity was greater for MR-exposed groups than for sham-exposed groups. The results show that MR exposure has no adverse effect on the immune system, as evidenced by natural killer cell activity.     

Does cardiac SPECT using attenuation and scatter correction accurately predict coronary artery disease in a minority women population?

BACKGROUND: Myocardial perfusion imaging (MPI) is a well-established diagnostic method for evaluation and risk stratification of coronary artery disease (CAD).We undertook this study to validate both the positive predictive value (when compared to cardiac catheterization) and the prognosis afforded by MPI in a group of minority women patients. MATERIAL/METHODS: The database of our Nuclear Imaging and Catheterization Laboratory was retrospectively queried for consecutive minority (African-American, Hispanic and Asian) women patients who underwent MPI and cardiac catheterization within 90 days of each other. Patients with recent revascularization were excluded. Attenuation/scatter correction was utilized in the final interpretation of the study. RESULTS: Of the 54 women patients who underwent MPI, 7 underwent exercise stress testing, 26 had stress testing with adenosine, 18 with dipyridamole and 3 with dobutamine. Eighteen patients (53%) had same number of vessels predicted by MPI and coronary angiography (7 patients with triple vessel disease, 7 with 2-vessel disease and 4 with single vessel disease). Five (3 with intermediate and 2 with high risk scans) out of the 54 patients (9.3%) were dead at 2 years. The sensitivity, specificity and positive predictive value of MPI as compared to angiography were 87.2%, 26.7%, 75.6% and 44.4% respectively. CONCLUSIONS: The sensitivity of MPI in this group of patients is comparable to the general population though the specificity is lower in spite of using attenuation and scatter correction. Low risk perfusion scan signifies favorable prognosis at 2 years with regards to mortality.     

Revisiting the biofragmentable anastomotic ring: Is it safe in colonic surgery?

INTRODUCTION: The use of the biofragmentable anastomotic ring (BAR) has been reported in the literature with good results. Our purpose in this review was to document the clinical outcomes after gastrointestinal anastomoses performed with use of the BAR. METHODS: Data were gathered systematically through chart review with the help of data collection forms from 159 patients who underwent 173 intestinal anastomoses performed with use of the BAR between 1992 and 1999. Of the 165 patients who had anastomoses (6 had 2 anastomoses constructed on separate occasions and were considered separately), 23 (13.9%) had surgery with anastomosis under emergency conditions, and 44 (26.7%) were steroid-dependent patients. The indications for surgery were malignant disease in 63 (38.2%) patients, inflammatory bowel disease in 54 (32.7%) patients, diverticular disease in 13 (7.9%) patients and other conditions in 35 (21.2%) patients. RESULTS: A clinical anastomotic leak developed in the first 2 weeks after surgery in 7 (4.2%) patients, 6 of whom required reoperation. All recovered well, withno deaths related to use of the BAR. Early small-bowel obstruction developed in 13 patients (7.9%), none of whom required reoperation. The average postoperative length of hospital stay was 9.0 days, the average time to pass the first flatus was 3.2 days, and the average time to begin oral fluid intake was 3.3 days. The rate of leakage at the anastomosis in our series was comparable to that found in randomized trials with the BAR (2.0%-4.4%) and as reported with hand-sewn and stapled anastomoses (1.9%-8.2%). CONCLUSIONS: Our data indicate that use of the BAR is safe and effective in both elective and emergent surgery. The rate of leakage is comparable to that reported in the literature when a BAR is used.     

Impact of nutritional status on peritonitis in CAPD patients.

OBJECTIVE: To determine the impact of nutritional status on peritonitis in patients on continuous ambulatory peritoneal dialysis (CAPD) in a developing country. METHODS: 56 patients with end-stage renal disease on CAPD were randomly selected for this study. These patients were assessed for nutritional status and peritonitis episodes. Nutritional parameters were assessed by anthropometry, diet, body mass index (BMI), Nutritional Risk Index (NRI), serum albumin level, and Subjective Global Assessment (SGA). Based on SGA, patients were categorized into either group 1 (malnutrition, n = 31) or group 2 (normal nutritional status, n = 25). Peritonitis was considered the primary outcome and was compared between the two groups. RESULTS: Demographic profiles, Kt/V, creatinine clearance, and mean follow-up of the two groups were similar. Number of peritonitis episodes was significantly higher in patients with malnutrition (25/31) compared to patients with normal nutritional status (4/25) (p = 0.001). Mean peritonitis rate per patient per year was also significantly higher in patients with malnutrition (0.99 +/- 1.07) compared to patients with normal nutritional status (0.18 +/- 0.42) (p = 0.007). On univariate analysis, malnutrition based on SGA (p = 0.009), NRI (p = 0.02), serum albumin level (p = 0.005), and calorie intake (p = 0.006) was a significant predictor of peritonitis. On multivariate Cox regression analysis, only SGA (p = 0.001, odds ratio 0.08, 95% confidence interval 0.02-0.36) was found to be a significant predictor of peritonitis. On general linear model, the observed power of prediction of peritonitis was 0.96 based on SGA. On Kaplan-Meier survival analysis, peritonitis-free survival in patients with normal nutrition (42 months) was significantly higher compared to patients with malnutrition (21 months) based on SGA (log rank p = 0.003). CONCLUSION: We conclude that peritonitis rate is high in patients with malnutrition and that malnutrition indices, especially SGA, can predict the peritonitis rate in CAPD patients.     

Three-dimensional structure of a membrane-containing virus.

The structure of Sindbis virus was determined by electron cryomicroscopy. The virion contains two icosahedral shells of viral-encoded proteins separated by a membrane bilayer of cellular origin. The three-dimensional structure of the ice-embedded intact Sindbis virus, reconstructed from electron images, unambiguously shows that proteins in both shells are arranged with the same icosahedral lattice of triangulation number T = 4. These studies also provide structural evidence of contact between the glycoprotein and the nucleocapsid protein across the membrane bilayer. The structural organization of Sindbis virus has profound implications for the morphogenesis of the alphaviruses. The observed interactions confirm stoichiometric and specific protein associations that may be crucial for virion stability and predict a mechanism for assembly.