Biomedical subjects
Publications and source records attributed to S Ally.
Bilateral ectopic pregnancy is a rare phenomenon. It has been 76 years since the first published report of a case of bilateral ectopic pregnancy. Since then about 200 cases have been reported in the literature; we report a case here.
Differential regulation of the regulatory subunits of cAMP-dependent protein kinase isozymes correlates with the growth inhibitory effect of site-selective 8-Cl-cAMP demonstrated in cancer cell lines (Ally, S., Tortora, G., Clair, T., Grieco, D., Merlo, G., Katsaros, D., Ogreid, D., Døskeland, S.O., Jahnsen, T., and Cho-Chung, Y.S. (1988) Proc. Natl. Acad. Sci. U. S. A. 85, 6319-6322). Such selective modulation of protein kinase isozyme regulatory subunits was also found in the 8-Cl-cAMP-induced inhibition of both transformation and transforming growth factor alpha (TGF alpha) production in Ki-ras-transformed rat kidney fibroblasts (Tortora, G., Ciardiello, F., Ally, S., Clair, T., Salomon, D. S., and Cho-Chung, Y. S. (1989) FEBS Lett. 242, 363-367). In this work, we have demonstrated that 8-Cl-cAMP antagonizes the TGF alpha effect in TGF alpha-transformed mouse mammary epithelial cells (NOG-8TFC17) at the level of gene expression for cAMP receptor protein isoforms, RI and RII (the regulatory subunits of protein kinase isozymes). Northern blot analysis demonstrated that in the transformed NOG-8TFC17 cells, compared with the nontransformed counterpart NOG-8 cells, the mRNA levels for the RI alpha cAMP receptor protein markedly increased, whereas the mRNA levels for the RII alpha and RII beta cAMP receptor proteins decreased. 8-Cl-cAMP, which induced growth inhibition and phenotypic reversion in NOG-8TFC17 cells, caused an inverse change in the mRNA patterns of the cAMP receptor proteins; RI alpha cAMP receptor mRNA sharply decreased to levels comparable with that of the nontransformed NOG-8 cells, whereas RII beta mRNA increased to a level even greater than that in the NOG-8 cells. In addition, one mRNA species of RII alpha increased, whereas the other RII alpha mRNA species decreased during the treatment. The mRNA level for the catalytic subunit of protein kinase, however, did not change during 8-Cl-cAMP treatment. In addition, 8-Cl-cAMP brought about a reduction in both TGF alpha mRNA and protein levels. These coordinated changes in the expression of the cAMP receptor proteins and TGF alpha were not observed during cis-hydroxyprolineor TGF beta-induced growth inhibition of the NOG-8TFC17 cells. Thus, the antagonistic effect of 8-Cl-cAMP toward TGF alpha-induced transformation involves modulation of the expression of a specific set of cellular genes.
Site-selective cyclic AMP (cAMP) analogues inhibit growth and induce changes in morphology in a spectrum of human cancer cell lines (D. Katsaros et al., FEBS Lett., 223:97, 1987). The cellular events underlying such effects of cAMP analogues include differential regulation of type I versus type II cAMP-dependent protein kinase isozymes (S. Ally et al., Proc. Natl. Acad. Sci. USA, 85: 6319, 1988). Infusion (i.p.) of 8-Cl-cAMP, the most potent site-selective cAMP analogue, for 7 days produced regression of LX-1 lung carcinoma in athymic mice in a dose-dependent manner. The tumor regression correlated with the changing levels of cAMP receptor proteins, RI alpha and RII beta, the regulatory subunits of cAMP-dependent protein kinase type I and type II, respectively. By photoaffinity labeling with 8-N3-[32P]cAMP and immunoblotting with a monospecific anti-RII antibody, RI alpha (Mr 49,000) and RII beta (Mr 51,000) were identified in the untreated control tumors. 8-Cl-cAMP treatment induced a rapid increase of both RI alpha and RII beta in tumor cytosols and translocation (within 1 h) of only RII beta from the cytosol to the nucleus. RII beta in both cytosols and nuclei remained elevated during 8-Cl-cAMP treatment, whereas RI alpha in the cytosols gradually decreased with time of treatment after its initial transient increase. Northern blot analyses demonstrated that the RII beta mRNA level increased within 6 h of 8-Cl-cAMP treatment and remained elevated during treatment, whereas the RI alpha mRNA level decreased to below that of the untreated control tumor level after its transient increase during 1-6 h of treatment. 8-Cl-cAMP treatment also caused a sharp decrease in both N-ras and c-myc mRNA levels. These results suggest that the fundamental basis for the antineoplastic activity of 8-Cl-cAMP may reside in the restoration of normal gene regulation in neoplasms in which cAMP receptor proteins play a role.
Control mechanisms of normal differentiation are disrupted in cancer cells but can be restored by treatment with site-selective cAMP analogs. The cellular events associated with such changes entail compartmental redistribution of the cAMP-dependent protein kinase type II regulatory subunit, RII beta. The results of this study indicate that the molecular mechanisms of action involve changes in specific DNA-binding activity of putative transcription factors. Gel retardation analyses revealed that nuclear extracts from cells of various human cancer cell lines [colon cancer (LS-174T), gastric cancer (TMK-1), and leukemia (K-562)] and rodent pheochromocytoma (PC12) show a concentration-dependent increase in binding activity to a synthetic DNA that contained the cAMP-responsive element 5'-TGACGTCA-3' after treatment with 8-Cl-cAMP. Such an increase in cAMP-responsive element binding activity was not observed in the 8-C1-cAMP-unresponsive MKN-1 gastric cancer cells. These findings indicate that the antitumor activity of site-selective cAMP analogs may reside in the induction of transcription factors that restore normal gene regulation in cancer cells.
A site-selective cAMP analog, 8-chloroadenosine 3',5'-cyclic monophosphate (8-Cl-cAMP), was demonstrated to be a potent inhibitor of both the monolayer and soft agar growth of normal rat kidney (NRK) fibroblasts that had been transformed with the v-Ki-ras oncogene or treated with transforming growth factor alpha (TGF alpha). The growth inhibition was dose dependent and reversible and was accompanied by reversion of the transformed phenotype, suppression of TGF alpha production, and a decrease in p21 ras protein levels. These effects of 8-Cl-cAMP were linked to the cAMP analog's selective modulation of the type I and type II cAMP-dependent protein kinase regulatory subunits, RI and RII, present in Ki-ras-transformed and TGF alpha-treated NRK cells.
Two classes (site 1- and site 2-selective) of cAMP analogs, which either alone or in combination demonstrate a preference for binding to type II rather than type I cAMP-dependent protein kinase isozyme, potently inhibit growth in a spectrum of human cancer cell lines in culture. Treatment of K-562 human leukemic cells for 3 days with 30 and 10 microM 8-chloroadenosine 3',5'-cyclic monophosphate (8-Cl-cAMP) (site 1-selective) resulted in 60% and 20% growth inhibition, respectively (with over 90% viability). N6-Benzyl-cAMP (site 2-selective) (30 microM) treatment resulted in 20% growth inhibition by day 3. When 8-Cl-cAMP (10 microM) and N6-benzyl-cAMP (30 microM) were both added, growth was almost completely arrested. The growth inhibition was accompanied by megakaryocytic differentiation in K-562 cells. The untreated control cells expressed little or no detectable levels of glycoprotein IIb-IIIa surface antigen complex. 8-Cl-cAMP (30 microM) treatment for 3 days substantially increased the antigen expression, while N6-benzyl-cAMP caused little or no change in the antigen expression. When cells were treated with 8-Cl-cAMP in combination with N6-benzyl-cAMP, antigen expression was synergistically enhanced, and cells demonstrated megakaryocyte morphology. By Northern blotting, we examined the mRNA levels of the type I and type II protein kinase regulatory subunits (RI alpha and RII beta), the catalytic subunit, and c-myc during 8-Cl-cAMP treatment. The steady-state level of RII beta cAMP receptor mRNA sharply increased within 1 hr of treatment and remained elevated for 3 days, while that of the RI alpha receptor markedly decreased to below control level within 6 hr and remained low during treatment. However, 8-Cl-cAMP did not affect the mRNA level of the catalytic subunit. 8-Cl-cAMP treatment also brought about a rapid decrease in c-myc mRNA. Thus, differential regulation of cAMP receptor genes is an early event in cAMP-induced differentiation and growth control of K-562 leukemia cells.
The physiologic role of cyclic adenosine monophosphate (cAMP) in the growth control of a spectrum of human cancer lines, including leukemic lines, and v-rasH oncogene-transformed NIH/3T3 cells is demonstrated by the use of site-selective cAMP analogs. These cAMP analogs, which can select either of the two known cAMP binding sites of the cAMP receptor protein, induce potent growth inhibition, phenotypic change, and differentiation (leukemic cells) of cancer cells at micromolar concentrations with no sign of cytotoxicity. The growth inhibition parallels selective modulation of cAMP-dependent protein kinase isozymes, type I versus type II, and suppression of cellular proto-oncogene expression. Site-selective cAMP analogs thus provide new biological tools for investigating cell proliferation and differentiation and also for the improved management of human cancers.
Cyclic AMP (cAMP) analogues that selectively bind to either one of the two binding sites of cAMP-dependent protein kinase demonstrate a potent inhibition of the growth stimulated by estrogen in MCF-7 human breast-cancer cells in culture. The site-selective analogues, which are more potent activators of protein kinase than the analogues studied earlier, exhibit growth inhibition at micromolar concentrations. Among the analogues tested, 8-Cl-cAMP (Site I-selective) and N6-benzyl-cAMP (Site 2-selective) are the 2 most potent inhibitors, causing 40-70% inhibition of the estrogen-stimulated growth at 10-20 microM concentrations with no sign of toxicity. 8-Cl-cAMP (1 microM) in combination with N6-benzyl-cAMP (0.5 microM) almost completely blocks estrogen-stimulated growth, demonstrating synergism between the Site 1- and Site 2-selective analogues. The growth inhibition parallels an increase in the R11 cAMP receptor protein with a decrease in the R1 receptor as well as reduction of c-myc and c-ras oncoproteins, whereas growth inhibition by tamoxifen does not affect the levels of the cAMP receptor proteins or the c-myc and c-ras protein levels. Site-selective cAMP analogues are antagonistic to estrogen stimulation of breast-cancer cell growth through a mechanism different from that of tamoxifen.
Our past studies on the mechanism of cyclic AMP (cAMP)-mediated control of tumor growth, using the experimental rat mammary tumor models as well as human breast cancer cell lines, indicated that the action of cAMP is mediated by the RII cAMP receptor protein, the regulatory subunit of cAMP-dependent protein kinase type II (Y. S. Cho-Chung, J. Cyclic Nucleotide Res., 6: 163, 1980). We now shown that the site-selective cAMP analogues, which are manyfold more active in binding to the cAMP receptor protein than previously studied analogues, demonstrate a potent growth inhibition of seven breast and three colon human cancer cell lines. The cAMP receptor protein has two different cAMP binding sites, and cAMP analogues that selectively bind to either one of the two binding sites are known as either site 1 selective (C-8 analogues) or site 2 selective (C-6 analogues). Nineteen site-selective analogues, C-6 and C-8 monosubstituted and C-6,-8 disubstituted, were tested for their growth regulatory effect. The majority of these analogues demonstrated an appreciable growth inhibition, with no sign of toxicity in all 10 cancer lines at micromolar concentrations. The three most potent inhibitors were 8-Cl-, N6-benzyl-, and N6-phenyl-8-thio-p-chlorophenyl-cAMP, demonstrating 50% growth inhibition at 5-25 microM concentrations (IC50). Furthermore, N6-analogues, in combination with halogen or thio derivatives of C-8 analogues, demonstrated synergistic enhancement of growth inhibition. The growth inhibition paralleled a change in cell morphology, an augmentation of the RII cAMP receptor protein, and a reduction in p21 ras protein. The growth inhibition by 8-Cl-cAMP was not due to its metabolite, 8-Cl-adenosine, since: (a) the growth inhibition by 8-Cl-cAMP was released upon cessation of treatment, whereas that by 8-Cl-adenosine was not released; (b) 8-Cl-cAMP treatment did not affect cell cycle progression, whereas 8-Cl-adenosine brought about G1 synchronization; (c) 8-Cl-cAMP treatment caused reduction of p21 ras protein, whereas 8-Cl-adenosine did not affect p21 levels; and (d) 8-Cl-adenosine was not detected in either cell extracts or medium from the cells treated with 8-Cl-cAMP for 48-72 h. Site-selective cAMP analogues thus provide a new physiological means to control the growth of breast and colon human cancer cells.
Differential expression of type I and type II cAMP-dependent protein kinase isozymes has been linked to growth regulation and differentiation. We examined the expression of protein kinase isozymes in the LS 174T human colon cancer cell line during 8-chloroadenosine 3',5'-cyclic monophosphate (8-Cl-cAMP)-induced growth inhibition. Two species of RII (the regulatory subunit of protein kinase type II) with apparent Mr 52,000 (RII52) and Mr 56,000 (RII56) and a single species of RI (the regulatory subunit of protein kinase type I) with Mr 48,000 were identified in the cancer cells. RI and both forms of RII were covalently labeled with 8-azidoadenosine 3',5'-cyclic [32P]monophosphate, and two anti-RII antibodies that exclusively recognize either RII52 or RII56 resolved two forms of the RII receptors. 8-Cl-cAMP treatment induced a decrease of RI and an increase of both RII52 and RII56 in the cytosols of cancer cells and rapid translocation (within 10 min) of RII52 from the cytosol to nucleus. 8-Cl-cAMP caused transcriptional activation of the RII52 receptor gene and inactivation of the RI receptor gene. It also exhibited high-affinity site-1-selective binding to the purified preparations of both RII receptor proteins. Thus, differential regulation of various forms of cAMP receptor proteins is involved in 8-Cl-cAMP-induced regulation of cancer cell growth, and nuclear translocation of RII52 receptor protein appears to be an early event in such differential regulation.
Site-selective cAMP analogs, depending on the position of their substituents on the adenine ring, selectively bind to either site 1 or site 2 of the known cAMP binding sites of protein kinase. Treatment of Harvey murine sarcoma virus-transformed NIH/3T3 cells with such site-selective analogs results in growth inhibition and phenotypic reversion, and the combination of a C-8 thio or halogen analog (site 1 selective) with an N6 analog (site 2 selective) produces a synergistic effect. We report here that the growth inhibitory effect of the analogs correlates with the nuclear translocation of the RII cAMP receptor protein, the regulatory subunit of protein kinase type II. The transformed NIH/3T3 cells contained no detectable level of RII in the nucleus, whereas nontransformed NIH/3T3 cells exhibited a high level of nuclear RII. Within 30 min after treatment of the transformed cells with the site-selective analogs, immunofluorescence against the RII protein markedly increased in the cell nucleus. The nuclear translocation of the RII cAMP receptor protein is an early event in the reverse transformation of the fibroblasts treated with site-selective cAMP analogs.
Site-selective cyclic AMP analogs bind to site 1 or site 2 of the known cAMP-binding sites depending on the position of substituents on the purine ring, either at C-2 and C-8 (site 1) or at C-6 (site 2). The growth inhibitory effect of such site-selective cAMP analogs used in this investigation with 15 human cancer cell lines surpassed that of analogs previously tested. The most potent analogs were 8-chloro, N6-benzyl and N6-phenyl-8-p-chlorophenylthio-cAMP. The combination of a C-8 with an N6 analog had synergistic effects. The 24 site-selective analogs tested produced growth inhibition ranging from 30 to 80% at micromolar concentrations with no sign of toxic effects. Growth inhibition was not due to a block in a specific phase of the cell cycle but paralleled a change in cell morphology, an increase of the RII cAMP receptor protein and a decrease of p21 ras protein. Since the adenosine counterpart of the 8-chloro analog produced G1 synchronization without affecting the RII and p21 ras protein levels, it is unlikely that an adenosine metabolite is involved in the analog effect. Site-selective cAMP analogs thus provide a new biological tool for control of cancer growth.
PURPOSE: This article describes an infant with a large abdominal mass and hypertension. PATIENT AND METHODS: A 5-month-old infant girl with diarrhea of 1 week's duration and a large right-sided abdominal mass was brought for treatment. Computed tomography of the abdomen revealed a large, generally homogeneous, hypodense mass, which compressed the right kidney, resulting in dilatation of the right renal collecting system. At surgery, the mass was adherent anteriorly to the transverse colon and attached by a stalk to the mesentery near the origin of the right colic artery. RESULTS: Examination of the mass showed an encapsulated lipoblastoma. Cytogenetic analysis revealed a 46,XX karyotype with a reciprocal translocation between chromosome 2 and chromosome 8 with breakpoints at q23 and q11.2, respectively. CONCLUSION: Lipoblastoma is a rapidly growing but benign tumor, which can cause severe medical problems by compressing major organs. Cytogenetic analysis can reveal translocations involving chromosome 8 band q11.2, which appears to be a specific chromosome marker for lipoblastoma.