A model of antimycin A binding based on structure-activity studies of synthetic antimycin A analogues.

The structural factors of antimycin A molecule required for inhibitory action were studied using newly synthesized antimycin A derivatives with bovine heart submitochondrial particles, in order to probe the interaction between antimycin A and its binding site. In particular, we focused upon the roles of the amide bond bridge, which connects the salicylic acid and dilactone ring moieties, and the 3-formylamino group in the salicylic acid moiety. The lack of formation of an intramolecular hydrogen-bond between phenolic OH and amide carbonyl groups resulted in a remarkable loss of the activity (by four orders of magnitude), indicating that this hydrogen-bond is essential for the inhibition. This result suggested that both the phenolic OH and the carbonyl groups form a hydrogen-bond with some residues at a fixed conformation. In addition, the inhibitory potency was remarkably decreased by N-methylation of the amide bond moiety, indicating that the NH group might function in hydrogen-bond interaction with the binding site. The N-methylation of 3-formylamino group also resulted in a decrease in the activity, probably due to a loss of the rotational freedom of this functional group. Molecular orbital calculation studies with respect to the conformation of the 3-formylamino group indicated that this group takes an active conformation when the formyl carbonyl projects to the opposite side of the phenolic OH group. Based upon a series of structure-activity studies of synthetic antimycin A analogues, we propose a tentative model for antimycin A binding in its binding cavity.

Expression of topoisomerase II alpha, Ki-67 and p53 in early stage laryngeal carcinomas not featuring vocal cord fixation.

AIM: To determine whether topoisomerase II alpha (topoIIa) expression is an additional prognostic marker for less advanced stage laryngeal cancers first treated without surgery. Ki-67 and p53 protein levels were also assessed for comparison. EXPERIMENTAL DESIGN: Formalin-fixed, paraffin-embedded tumor material from 63 cases of squamous cell carcinoma (SCC) of the larynx (glottis, stages 0,1,2) was immunohistochemically stained for topoIIa, Ki-67 (MIB-1) and p53 (DO-7) and the results were compared with clinicopathologic findings. RESULTS: There were 7 stage 0 (TisN0M0), 33 stage I (T1N0M0), and 23 stage II (T2N0M0) SCCs with the TNM classification. Significant differences between carcinomas and normal mucosa were found for the topoIIa-LI, Ki-67-LI, and topoIIa-to-Ki-67 ratio. Regarding histologic grade, a significant difference in topoIIa-to-Ki-67 ratio was evident between well or moderately and poorly differentiated lesions. There were 19 cases of recurrence and 44 cases of nonrecurrence, but no significant differences were found for either of the indices or their ratio. No significant variation with p53 positivity was evident with reference to histologic differentiation, T-factor, clinical course, or proliferation. CONCLUSIONS: The results demonstrate that the topoIIa-to-Ki-67 ratio is a more sensitive parameter reflecting proliferation, for histologic grading of less advanced laryngeal SCCs, than topoIIa- or Ki-67-LIs.

Xenotransplantation of canine tumors into severe combined immunodeficient (SCID) mice.

Canine tumors resected at surgery or autopsy were subcutaneously xenotransplanted into SCID mice. Thirty of the seventy-three tumors (41.1%) grew primarily in SCID mice. The primary take rate of mammary tumors and skin and subcutaneous tumors was 55.9% (19/34) and 28.6% (8/28), respectively. One thyroid carcinoma, one oral leiomyosarcoma and one osteosarcoma also underwent successful primary growth. Thirteen of the forty-one benign tumors (31.7%) and 17 of the thirty-two malignant ones (53.1%) were successfully transplantable. The histological features of the primary xenografts and their original tumors were similar to each other in all the cases. Three xenografts of the four malignant tumors with metastasis in the canine patients were shown to grow also in SCID mice and two mammary tumor xenografts out of the three metastasized to the lungs of SCID mice as well. All the 30 primary tumor xenografts grown in the first recipient were successfully transplantable to the second generation of the SCID mice. Xenotransplantation using SCID mice appears to be a valuable tool for investigating the biological characteristics of canine neoplasmas.

Epithelial inclusion cyst (epidermoid cyst) formation with epithelioid cell granuloma in an intrapancreatic accessory spleen.

The histologic characteristics of a case of epidermoid cyst formation with an epithelioid cell granuloma that developed in intrapancreatic accessory spleen were investigated, with the aim of clarifying its origin as well as etiologic factors. The patient, a 48-year-old male, was found to have a cystic lesion in the tail of the pancreas and renal cell carcinoma (clear cell carcinoma) during a medical check up. The pancreatic mass appeared to be a so-called "mucinous tumor" on imaging, and combined resection of the body and tail of the pancreas and the spleen was performed together with a left nephrectomy. The lesion in the tail of the pancreas was then demonstrated to have accessory splenic tissue with cyst formation in its central region. The cystic wall was covered with stratified squamous epithelium and ductal epithelium with squamous metaplasia, and pancreatic islet cells were evident at various sites within the accessory spleen. Formation of epithelioid cell granuloma tissue was observed around the cysts. The epithelium of the cyst was positive for CA19-9 and negative for antibodies against mesothelial cells, whereas the pancreatic island cells were positive for insulin. These results suggested that cystic epithelium in the accessory spleen could be derived from pancreatic duct epithelium. Frequent recurrence of chronic inflammation and epitheloid cell granuloma formation may have resulted from an aberration of the ectopic remaining pancreatic tissue in the spleen.

Cimetidine increases survival of colorectal cancer patients with high levels of sialyl Lewis-X and sialyl Lewis-A epitope expression on tumour cells.

Cimetidine has been shown to have beneficial effects in colorectal cancer patients. In this study, a total of 64 colorectal cancer patients who received curative operation were examined for the effects of cimetidine treatment on survival and recurrence. The cimetidine group was given 800 mg day(-1) of cimetidine orally together with 200 mg day(-1) of 5-fluorouracil, while the control group received 5-fluorouracil alone. The treatment was initiated 2 weeks after the operation and terminated after 1 year. Robust beneficial effects of cimetidine were noted: the 10-year survival rate of the cimetidine group was 84.6% whereas that of control group was 49.8% (P<0.0001). According to our previous observations that cimetidine blocked the expression of E-selectin on vascular endothelium and inhibited the adhesion of cancer cells to the endothelium, we have further stratified the patients according to the expression levels of sialyl Lewis antigens X (sL(x)) and A (sL(a)). We found that cimetidine treatment was particularly effective in patients whose tumour had higher sL(x) and sL(a) antigen levels. For example, the 10-year cumulative survival rate of the cimetidine group with higher CSLEX staining, recognizing sL(x), of tumours was 95.5%, whereas that of control group was 35.1% (P=0.0001). In contrast, in the group of patients with no or low levels CSLEX staining, cimetidine did not show significant beneficial effect (the 10-year survival rate of the cimetidine group was 70.0% and that of control group was 85.7% (P=n.s.)). These results clearly indicate that cimetidine treatment dramatically improved survival in colorectal cancer patients with tumour cells expressing high levels of sL(x) and sL(a).

Toward the antibody-catalyzed chemiluminescence. Design and synthesis of hapten.

Hapten 4 was synthesized to generate catalytic antibodies triggering chemiluminescence by catalyzing the decomposition of the 1,2-dioxetane 3. The hapten 4 was so designed as to elicit a negatively charged functional group in the antibody combining site to catalyze the beta-elimination of the protecting group in 3 as well as to lock the protecting group into an energetically favorable anti-periplanar conformation.