Effects of antiandrogens on growth of androgen-dependent mouse mammary tumor (Shionogi carcinoma 115) in vivo and in vitro

Binding affinities of modified steroidal anthrasteroids, 3 beta-hydroxy-3a beta,6-dimethyl-2,3,3a,4,5,8,9,10,10a beta,11,11a beta, 11b alpha-dodecahydro-1H-cyclopenta[a]anthracene-8-one (1) and 3a beta,6-dimethyl-2,3,3a,4,5,8,9,10,10a beta,11,11a beta,11b alpha-dodecahydro-1H-cyclopenta[a]anthracene-3,8-dione (2), the steroid oxendolone and the nonsteroid AA560, for the androgen receptor (AR) of Shionogi carcinoma 115 (SC115) and their effects on the growth of SC115 were investigated in vivo and in vitro. The inhibitory effects of these compounds on testosterone 5 alpha-reductase of SC115 tissues were also measured. The relative binding affinities of these compounds were 3.17-0.03% of that of dihydrotestosterone, and their rank order was (1) greater than AA560 greater than oxendolone much greater than (2). In the presence of 10(-9) M testosterone, anthrasteroids and AA560 inhibited the growth of SC115 cells at 10(-7) M in a serum-free medium, but oxendolone did not. In the absence of testosterone, (1), (2) and oxendolone promoted cell growth at 10(-6), 10(-7) and 10(-7) M, respectively. However, AA560 nearly completely blocked cell growth at 10(-5) M. At a 2 mg daily dose for 13 days, (1) and AA560 powerfully inhibited tumor growth in castrated DS mice treated with testosterone propionate but oxendolone had almost no effect. Anthrasteroids and oxendolone showed weak but significant agonistic activity in vivo. Anthrasteroids markedly inhibited 5 alpha-reductase activity of SC115, oxendolone weakly and AA560 not at all. The remarkable antiandrogenic activities of (1) and AA560 may partially result from their higher affinities for the AR of SC115 but other yet unknown mechanisms may also contribute to these activities.     

Rejection of B16 melanoma induced by expression of a transfected major histocompatibility complex class I gene.

Transfection of a functional major histocompatibility complex class I gene into certain tumor cells, induced by oncogenic viruses or chemical carcinogens, can effectively abrogate their tumorigenic activity. Since experimentally induced tumors possess strong tumor-specific transplantation antigens, expression of cell surface class I antigens may present the tumor cells to appropriate immune effector cells. Most spontaneously arising tumors do not possess tumor-specific transplantation antigens, and their tumorigenicity may not be affected by the expression of a transfected class I gene. We demonstrate that the poorly immunogenic B16-BL6 melanoma can be rendered nontumorigenic in syngeneic mice by the expression of the class I H-2K antigen but not the class II I-A antigen. Furthermore, the poorly tumorigenic, class I-expressing B16-BL6-transfected cells can effectively immunize syngeneic C57BL/6 mice against the highly tumorigenic, class I-deficient B16-BL6 parental cells. Our success in experimentally manipulating the tumorigenicity of a spontaneously derived neoplasm offers hope for a potential modality for the effective treatment of human cancer.     

Studies on peptides CLVIII. Model experiments for the synthesis of open-chain unsymmetrical cystine-peptides

Treatment of a mixture of Cys(R)(O) and Cys(R) with an acid was found to generate cystine in fairly good yields, when suitable R, R, and an acid were selected. An unsymmetrical cystine peptide was prepared by treatment of a mixture of Z(OMe)-Cys(R) (0)-Ala-NH₂ (R=Acm or MBzl) and Z(OMe)-Cys(MBzl)-Gly-OBzl with TFA or 1 M TFMSA/TFA.³ Oxytocin was obtained in an excellent yield by TFA treatment of the protected peptide containing Cys(Acm)(0) and Cys(MBzl). Thus, formation of the disulfide bond was found feasible at the position of Cys(R) (0).The following abbreviations are used Boc t-butyloxycarbonyl - Z(OMe) p-methoxybenzyloxycarbonyl - MBzl p-methoxybenzyl - Acm acetamidomethyl - Bzl benzyl - Ad l-adamantyl - tBu t-butyl - TFA trifluoroacetic acid - TFMSA trifluoromethanesulfonic acid - TMSOTf trimethylsilyl trifluoromethane sulfonate     

Fermentation of 2-methoxyethanol by Acetobacterium malicum sp. nov. and Pelobacter venetianus

Anaerobic bacteria degrading 2-methoxyethanol were enriched from freshwater sediments, and three strains were isolated in pure culture. Two of them were Grampositive non-spore-forming rods and grew strictly anaerobically by acetogenic fermentation. Optimal growth occurred at 30°C, initial pH 7.5–8.0. 2-Methoxyethanol and 2-ethoxyethanol were fermented to acetate and corresponding alcohols. Hydrogen plus carbon dioxide, formate, acetoin, l-malate, lactate, pyruvate, fructose, and methoxyl groups of 3,4,5-trimethoxybenzoate and 3,4,5-trimethoxycinnamate were fermented to acetate. 1,2-Propanediol was fermented to acetate, propionate, and propanol. Strain MuME1 was described as a new species, Actetobacterium malicum. It had a DNA base composition of 44.1 mol% guanine plus cytosine. The third strain, which was identified as Pelobacter venetianus, fermented 2-methoxyethanol to methanol, ethanol, and acetate.     

Inhibition by Cyclic AMP of Phorbol Ester-Potentiated Norepinephrine Release from Guinea Pig Brain Cortical Synaptosomes

The involvement of Ca2+/phospholipid-dependent protein kinase (protein kinase C, PKC) and cyclic AMP-dependent protein kinase in the K+-evoked release of norepinephrine (NE) was studied using guinea pig brain cortical synaptosomes preloaded with [3H]NE. 12-O-Tetradecanoylphorbol-13-acetate (TPA), a potent activator of PKC, enhanced the K+-evoked release of [3H]NE, in a concentration-dependent manner, but with no effect on the spontaneous outflow and uptake of [3H]NE in the synaptosomes. The apparent affinity of the evoked release for added calcium but not the maximally evoked release was increased by TPA (10(-7) M). Inhibitors of PKC, polymyxin B, and a more potent inhibitor, staurosporine, counteracted the TPA-induced potentiation of the evoked release. Both forskolin and dibutyryl cyclic AMP (DBcAMP) enhanced the evoked release, but reduced the TPA-potentiated NE release. A novel inhibitor of cyclic AMP-dependent protein kinase, KT5720, blocked both the forskolin-induced increase in the evoked release and its inhibition of TPA-induced potentiation in the evoked release, thereby suggesting that forskolin or DBcAMP counteracts the Ca2+-dependent release of NE by activating cyclic AMP-dependent protein kinase. These results suggest that the activation of PKC potentiates the evoked release of NE and that the activation of cyclic AMP-dependent protein kinase acts negatively on the PKC-activated exocytotic neurotransmitter release process in brain synaptosomes of the guinea pig.     

gamma-Aminobutyric acid in synovial membrane of rat knee joint

gamma-Aminobutyric acid (GABA) content was measured, and the release of GABA was studied in the synovial membrane of the rat knee joint. GABA content of the synovial membrane was 20.1 nmol/g tissue. Ten days after unilateral dissection of the sciatic nerve, femoral nerve or both nerves, the GABA contents of the ipsilateral membrane were 13.8, 14.6 and 7.8 nmol/g tissue, respectively. High K+ evoked the Ca2+-dependent release of [3H] GABA from the synovial membranes of intact rats preloaded with [3H] GABA, but did not evoke release from the membrane ipsilateral to the dissection of both sciatic and femoral nerves. Evoked release of [3H] GABA was obtained in the synovial membrane preloaded with [3H] GABA in the presence of beta-alanine, but not in the presence of 2,4-L-diaminobutyric acid. These results indicate that GABA is present in the neuronal elements of the synovial membrane of the rat knee joint.     

Anti-idiotypic antibodies against UV-induced tumor-specific CTL clones. Preparation in syngeneic combination

In this study, we first established several CTL clones of (BALB/c x C57BL/6)F1 origin that were specific for either syngeneic UV female 1 or UV male 1 fibrosarcoma cell lines. All the CTL clones had Thy-1+ Lyt-2+ L3T4- phenotypes and showed Kd restriction when lysing the corresponding target cells. Sera obtained from syngeneic animals immunized with three CTL clones, 10B-5 for UV female 1, and CTL9 and CTL10 for UV male 1, showed specific inhibition of target cell lysis with the corresponding CTL clones. The inhibitory activities were found in sera of the majority of immunized animals. Because the inhibitory activity resides in protein A-binding fraction, mAb were produced by hybridizing spleen cells of hyperimmune animals. N1-56 was thus obtained from a mouse immunized with 10B-5 CTL clone reactive with UV female 1. N1-56 was clonotype specific, reacting with 10B-5 but not with other CTL lines or leukemia cell lines. No N1-56+ cells were detectable in thymocytes, lymph node cells, or spleen cells of either naive or UV female 1-immune CB6F1 mice. Immunoprecipitation showed that N1-56 reacts with 90,000 Mr molecules on 10B-5 CTL clone under nonreducing conditions and 45,000 Mr molecules under reducing conditions, indicating its reactivities with idiotypic determinants of TCR on the CTL clone. N1-56 inhibited lytic activity of 10B-5, but neither N1-56 nor alpha-10B-5 hyperimmune serum inhibited that of alpha-UV female 1 mixed lymphocyte tumor cell culture cells. N1-56 induced proliferation of 10B-5 without addition of Ag.     

An assessment of nasal functions in control of breathing

Breathing pattern and steady-state CO2 ventilatory response during mouth breathing were compared with those during nose breathing in nine healthy adults. In addition, the effect of warming and humidification of the inspired air on the ventilatory response was observed during breathing through a mouthpiece. We found the following. 1) Dead space and airway resistance were significantly greater during nose than during mouth breathing. 2) The slope of CO2 ventilatory responses did not differ appreciably during the two types of breathing, but CO2 occlusion pressure response was significantly enhanced during nose breathing. 3) Inhalation of warm and humid air through a mouthpiece significantly depressed CO2 ventilation and occlusion pressure responses. These results fit our observation that end-tidal PCO2 was significantly higher during nose than during mouth breathing. It is suggested that a loss of nasal functions, such as during nasal obstruction, may result in lowering of CO2, fostering apneic spells during sleep.