Application of an interleukin 2 slow delivery system to the immunotherapy of established murine colon 26 adenocarcinoma liver metastases.

We evaluated the antitumor effect of an interleukin 2 (IL-2) slow delivery system, the IL-2 minipellet, using a murine hepatic metastasis model. The IL-2 minipellet consists of atelocollagen derived from natural bovine skin together with 1 x 10(6) units of recombinant IL-2. Administration of the IL-2 minipellet was performed into the spleens of BALB/c mice after translocation of the spleens to the s.c. position. Administration produced detectable serum IL-2 levels for 72 h. The IL-2 minipellet was evaluated for its efficacy against hepatic metastases from colon 26 adenocarcinoma in the BALB/c mice. Both the administration of the IL-2 minipellet alone and its combination with the injection of 5 x 10(7) lymphokine-activated killer cells resulted in significant reductions of the number of metastatic nodules. Moreover, increased survival of mice bearing colon 26 adenocarcinoma was noted in these two treatment groups. To investigate the mechanism of the IL-2 minipellet activity, we tested the lytic potential of splenocytes obtained after administration of the IL-2 minipellet in a 51Cr release assay. Cytotoxicity against YAC-1 cells and colon 26 cells was significantly augmented on Day 2 after minipellet administration. These results demonstrated that local administration of the IL-2 minipellet into the hepatic circulation was extremely effective against metastatic liver cancer.

Impaired differentiation of endocrine and exocrine cells of the pancreas in transgenic mouse expressing the truncated type II activin receptor.

Activin A is expressed in endocrine precursor cells of the fetal pancreatic anlage. To determine the physiological significance of activins in the pancreas, a transgenic mouse line expressing the truncated type II activin receptor under the control of beta-actin promoter was developed. Histological analyses of the pancreas revealed that the pancreatic islets of the transgenic mouse were small in size and were located mainly along the pancreatic ducts. Immunoreactive insulin was detected in islets, some acinar cells, and in some epithelial cells in the duct. In addition, there were abnormal endocrine cells outside the islets. The shape and the size of the endocrine cells varied and some of them were larger than islets. These cells expressed immunoreactive insulin and glucagon. In the exocrine portion, there were morphologically abnormal exocrine cells, which did not form a typical acinar structure. The cells lacked spatial polarity characteristics of acinar cells but expressed immunoreactive amylase, which was distributed diffusely in the cytoplasm. Plasma glucose concentration was normal in the transgenic mouse before and after the administration of glucose. The insulin content of the pancreas in transgenic and normal mice was nearly identical. These results suggest that activins or related ligands regulate the differentiation of the pancreatic endocrine and exocrine cells.

Adenosine A1 antagonists. 3. Structure-activity relationships on amelioration against scopolamine- or N6-((R)-phenylisopropyl)adenosine-induced cognitive disturbance.

The effects of a variety of adenosine A1 and A2 antagonists on N6-((R)-phenylisopropyl)adenosine (R-PIA)- and scopolamine-induced amnesias were investigated in rodents in order to clarify the role of adenosine receptors in learning and memory. Some of the selective adenosine A1 antagonists exhibited antiamnesic activities at several doses where they did not induce an increase of spontaneous locomotion. These results suggest that the blockade of A1 receptors is more important than that of A2 receptors in learning and memory. Detailed studies of structure-activity relationships of adenosine A1 antagonists in two amnesia models demonstrated that there were three types of adenosine A1 antagonists: (A) Compounds 3-5 (8-substituted 1,3-dipropylxanthines) ameliorated the shortened latency in both models. (B) Compounds 7-11 (8-substituted 1,3-dialkylxanthines) and 19-21 (imidazo[2,1-i]purin-5(4H)-one derivatives) ameliorated the shortened latency in the (R)-PIA-induced amnesia model but not in the scopolamine-induced amnesia model. (C) Compounds 14-16 ameliorated the shortened latency in the scopolamine model but not in the (R)-PIA model. Aminophenethyl-substituted compounds C did not exhibit adenosine A1 antagonism in vivo presumably due to rapid metabolism. The dramatic change in the activities of A and B could not be explained by their simple pharmacokinetic differences because both types of compounds showed clear blockade of central adenosine A1 receptors in the (R)-PIA model. 8-(3-Dicyclopropylmethyl)-1,3-dipropylxanthine (5) (KF15372) was chosen for further studies and is currently under preclinical development as a cognition enhancer.

Adenosine A(2A) receptor-mediated modulation of GABA and glutamate release in the output regions of the basal ganglia in a rodent model of Parkinson's disease.

A target neuron of adenosine A(2A) receptor antagonists to exert anti-parkinsonian activities has been currently identified to be, at least in part, striatopallidal medium spiny neurons (MSNs). In the present study, we determine whether A(2A) receptor-mediated modulation is associated with changes in the release of GABA and glutamate in the substantia nigra pars reticulata (SNr), an output structure of the whole basal ganglia network, using in vivo microdialysis in a rat Parkinson's disease (PD) model. In 6-hydroxydopamine (OHDA)-lesioned rats compared with normal rats, basal extracellular GABA levels in the SNr show no change, whereas basal glutamate levels are significantly increased. Oral administration of the A(2A) receptor-selective antagonist (E-1,3-diethyl-8-(3,4-dimethoxystyryl)-7-methyl-3,7-dihydro-1-H-purine-2,6-dion (KW-6002) to 6-OHDA-lesioned rats at 1 mg/kg caused a marked and sustained increase of GABA and glutamate levels in the SNr. The increase of nigral glutamate by KW-6002 was abolished by a kainic acid-induced lesion of the globus pallidus (GP) or subthalamic nucleus (STN) in 6-OHDA-lesioned rats, whereas the increase of nigral GABA was completely blocked by the GP-lesion but only partially blocked by the STN-lesion. These results indicate that changes in neurotransmitter release in the SNr brought about by KW-6002 are largely attributable to blockade of A(2A) receptor-mediated modulation of striatopallidal MSNs. Thus, these actions of KW-6002 on striatopallidal MSNs may be the main mechanism for ameliorating PD by A(2A) antagonists.

Calcium as a second messenger of the action of transforming growth factor-beta on insulin secretion.

In MIN6 insulinoma cells, transforming growth factor-beta (TGF-beta) induced the oscillatory elevation of the cytoplasmic free calcium concentration, [Ca2+]c, in the presence of 5.5 mM glucose. The increase in [Ca2+]c induced by TGF-beta was totally dependent on calcium entry and attenuated by nifedipine or nickel chloride. In contrast, carbachol elevated [Ca2+]c in the presence of nickel chloride. When the plasma membrane was hyperpolarized by diazoxide, TGF-beta did not raise [Ca2+]c, whereas both carbachol and depolarizing concentration of potassium elevated [Ca2+]c under the same conditions. TGF-beta did not affect either the cellular cyclic AMP or inositol trisphosphate levels. In the presence of 5.5 mM glucose, TGF-beta induced a 3-fold increase in insulin secretion and the effect of TGF-beta was blocked by either nifedipine or nickel chloride. TGF-beta did not stimulate insulin secretion in the presence of 100 microM diazoxide, whereas both carbachol and 40 mM potassium chloride significantly increased insulin secretion. These results suggest that TGF-beta induces the oscillatory elevation of [Ca2+]c in MIN6 cells by stimulating calcium entry via voltage-dependent calcium channels. Calcium is an intracellular messenger of the action of TGF-beta on insulin secretion.

Actions of adenosine A2A receptor antagonist KW-6002 on drug-induced catalepsy and hypokinesia caused by reserpine or MPTP.

RATIONALE: Current treatment of Parkinson's disease (PD) is based on dopamine replacement therapy, but this leads to long term complications, including dyskinesia. Adenosine A2A receptors are particularly abundant in the striatum and would be a target for an alternative approach to the treatment of PD. OBJECTIVES: The purpose of this study is to examine the efficacy and potency of the novel selective adenosine A2A receptor antagonist (E)-1,3-diethyl-8-(3,4-dimethoxystyryl)-7-methyl-3,7-dhydro- 1H-purine-2,6- dione (KW-6002) in ameliorating the motor deficits in various mouse models of Parkinson's disease. METHODS: We evaluated the efficacy and potency of KW-6002 and other reference compounds in the selective adenosine A2A receptor agonist 2-[p-(2-carboxyethyl)phenethylamino]-5'-N-ethylcarboxamidoadenosin e (CGS 21680)-, haloperidol- or reserpine-induced catalepsy models. The effect of KW-6002 on reserpine or 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine hydrochloride(MPTP)-induced hypolocomotion was also examined. RESULTS: The ED50s of KW-6002 in the reversal of CGS21680-induced and reserpine-induced catalepsy were 0.05 mg/kg, PO and 0.26 mg/kg, PO, respectively. Compared to the ED50 of other adenosine antagonists and dopamine agonist drugs, KW-6002 is over 10 times as potent in these models. KW-6002 also ameliorated the hypolocomotion (minimum effective dose; 0.16 mg/kg) induced by nigral dopaminergic dysfunction with MPTP or reserpine treatment. Combined administrations of subthreshold doses of KW-6002 and L-dopa (50 mg/kg, PO) exerted prominent effects on haloperidol-induced and reserpine-induced catalepsy, suggesting that there may be a synergism between the adenosine A2A receptor antagonist KW-6002 and dopaminergic agents. CONCLUSIONS: To our knowledge, KW-6002 is the most potent and orally active adenosine A2A receptor antagonist in experimental models of Parkinson's disease, and may offer a new therapeutic approach to the treatment of Parkinson's disease.

KF17837: a novel selective adenosine A2A receptor antagonist with anticataleptic activity.

KF17837 is a novel selective adenosine A2A receptor antagonist. Oral administration of KF17837 (2.5, 10.0 and 30.0 mg/kg) significantly ameliorated the cataleptic responses induced by intracerebroventricular administration of an adenosine A2A receptor agonist, CGS 21680 (10 micrograms), in a dose-dependent manner. KF17837 also reduced the catalepsy induced by haloperidol (1 mg/kg i.p.) and by reserpine (5 mg/kg i.p.). These anticataleptic effects were exhibited dose dependently at doses from 0.625 and 2.5 mg/kg p.o., respectively. Moreover, KF17837 (0.625 mg/kg p.o.) potentiated the anticataleptic effects of a subthreshold dose of L-3,4-dihydroxyphenylalanine (L-DOPA; 25 mg/kg i.p.) plus benserazide (6.25 mg/kg i.p.). These results suggested that KF17837 is a centrally active adenosine A2A receptor antagonist and that the dopaminergic function of the nigrostriatal pathway is potentiated by adenosine A2A receptor antagonists. Furthermore, KF17837 may be a useful drug in the treatment of parkinsonism.

In vivo regulation of acetylcholine release via adenosine A1 receptor in rat cerebral cortex.

The roles of the endogenous adenosine on acetylcholine release via adenosine A1 receptor were investigated in rat cerebral cortex using brain microdialysis. Oral administration of KF15372 (8-dicyclopropylmethyl-1,3-dipropylxanthine), a novel selective adenosine A1 receptor antagonist, at doses of 1.25, 5, and 20 mg/kg, significantly increased the extracellular levels of acetylcholine in rat cerebral cortex. Selective A1 agonist N6-((R)-phenylisopropyl) adenosine (R-PIA) did not affect the extracellular level of acetylcholine by both oral (1.25 mg/kg) and intracortical administrations (0.3 microM) via dialysis probe. These results suggest that the extracellular level of acetylcholine is under tonic inhibitory control of endogenous adenosine via the A1 receptor.

Effects of serum fractions obtained from cancer patients by double-filtration plasmapheresis on tumor growth and metastasis in mice.

We administered serum fractions obtained from cancer patients by double-filtration plasmapheresis (DFPP) to cancer-bearing mice to examine the effects on tumor growth and metastasis. Fraction 1 (whole plasma), fraction 2 (a plasma fraction containing substances with higher particle size), fraction 3 (a plasma fraction containing substances with smaller particle size) and saline were administered intravenously to cancer-bearing mice for 10 days following the inoculation of tumor cells. The tumor growth and metastasis in mice administered fraction 2 was far more rapid than that in the control mice. On the other hand, tumor growth in mice administered fraction 3 was significantly delayed compared with that in mice injected with fraction 2. These results suggest that factors in the higher particle-size fraction of cancer patients' sera promote the growth and the metastasis of tumors in mice, and that DFPP, which remove these factors, is an effective therapy against cancer.

Artificial liver support for postoperative hepatic failure with anion exchange resin (BR-601).

An artificial liver support system for plasma exchange and plasma perfusion through BR-601 resin using a membrane separator was applied to 5 patients with postoperative liver failure. Percent absorption of total and direct bilirubin, and of bile acids were 77.1 +/- 6.4, 78.4 +/- 6.1, and 93.4 +/- 3.6%, respectively, when 250 ml of plasma was treated. Percent reductions in total and direct bilirubin, and in bile acids were 24.5 +/- 5.8, 25.5 +/- 5.8 and 30.9 +/- 8.5%, respectively. In contrast, percent reductions in total and direct bilirubin, and in bile acids by plasma exchange were 30.9 +/- 13.3, 34.5 +/- 12.5 and 24.2 +/- 8.5%, respectively. The coma grade was improved in 4 out of 5 cases, but unfortunately the patients did not recover. In conclusion, plasma perfusion through BR-601 resin is expected to play a promising role in artificial liver support systems because of its capacity to absorb bilirubin and bile acids.

Renal transplantation from HLA-haploidentical living-related donors: the effects of donor-specific blood transfusions and different immunosuppressive regimens.

One-hundred-nine HLA-haploidentical living related renal transplants have been retrospectively analysed to compare the effect of donor-specific blood transfusion (DST) and different immunosuppressive regimens on graft survival and acute rejection. The recipients were divided into four groups according to the immunosuppressive therapy. Group 1 (n = 44): conventional therapy with posttransplant azathioprine (AZP) + methylprednisolone (MP). Group 2 (n = 25): pretransplant DST + posttransplant AZP + MP. Group 3 (n = 12): triple-drug therapy with posttransplant AZP + MP + cyclosporine (CS). Group 4 (n = 25): pretransplant DST + posttransplant AZP + MP + CS. The five-year actuarial survival rates for groups 1, 2, 3 and 4 were 48%, 73%, 79%, and 89%, respectively. The graft survival rate in group 3 was significantly (p less than 0.01) better than that in group 1. The transfusion effect was reduced, and appears as a 10% improvement in the graft survival in the cyclosporin era compared with a 25% improvement at pre-cyclosporin era. Furthermore, the incidence of the first rejection episode was decreased in recipients that received DST. The present study revealed that DST, as pretransplant conditioning has a definite impact on rejection-free long-term graft survival in HLA-haploidentical living-related kidney recipients and the most favorable outcome in such patients could be achieved by DST pretreatment in conjunction with posttransplant triple-drug therapy including cyclosporine.

The impact of triple drug immunosuppression on clinical results of cadaveric kidney transplantation: a comparison of conventional immunosuppression.

A retrospective study was carried out in 110 cadaveric kidney transplant recipients to compare the effects of low doses of cyclosporine (CsA), azathioprine (AZP) and steroids (triple-drug therapy) with those of higher doses of steroids plus AZP (conventional immunosuppression). Graft survival rate in the triple-drug therapy was 77%, 69%, and 69% at 1, 3, and 5 years, respectively. This was significantly better than 48%, 34%, and 29% in conventional immunosuppression. The incidence of acute rejection episodes was significantly lower in the triple-drug therapy than in conventional immunosuppression (25% vs 58%). In conclusion, our study shows that triple-drug therapy using low-dose cyclosporine is the safest of the immunosuppressive regimens and provides a beneficial effect on the long-term survival of cadaveric kidney transplants.

Clinical application of cascade filtration combined with glassbead adsorbents in cancer patients.

Cascade filtration (CF) was applied to patients with advanced cancers. To selectively remove immunosuppressive factors with molecular weight lower than albumin, adsorbents combined with CF were examined. Among 11 kinds of adsorbents, glassbead adsorbents with pore size 100A were the most effective in adsorbing such substances. Clinically, a column packed with glassbead adsorbents was concomitantly used with CF. The second filtrate was passed through the column and returned to the patient. The removal rate of immunosuppressive substances with molecular weight lower than albumin was only 20% by CF alone, but more than 50% by CF combined with glassbead adsorbents. Clinical symptoms such as severe back pain and general malaise were drastically improved during and after therapy. These results suggest that CF combined with glassbead adsorbents is an effective adjunctive modality to cancer therapy.

[A case of recurrent gastric cancer successfully treated with TS-1].

A patient with recurrent gastric cancer which infiltrated the pelvic muscle after the treatment of paraaortic lymph node and ovarian metastases was successfully managed by a novel oral anticancer drug, TS-1. TS-1 was administered at a dose of 80 mg/day. One course consisted of two repetitions of consecutive administration of TS-1 for 14 days and withdrawal of TS-1 for 14 days. Adverse reactions were mild and the patient did not request hospitalization after two courses had been completed. Subjective symptoms such as difficulty in walking improved after one week and a partial response was obtained after 2 weeks of treatment. At the end of 4 courses we could remove an indwelling pyelocatheter for the ureter stricture. As of 14 months after the beginning of administration of TS-1, the patient is being treated as an outpatient and has attained a better QOL than before.

[Effect of Euro-Collins' or UW solution on the early graft function following cadaveric kidney transplantation].

From November 1985 to March 1990, 55 cadaveric kidney transplants were performed under cyclosporine therapy. All kidneys were harvested from non-heart beating donors and cold stored after being flushed with EC solution (Group I, n = 27) or UW solution (Group II, n = 28). Warm ischemic time (min) in groups I and II were 7.1 +/- 3.3 and 6.9 +/- 2.3, respectively. Cold ischemic times (hr) in groups I and II were 6.9 +/- 2.4 and 8.4 +/- 2.8, respectively. Mean numbers of days for postoperative dialysis were 14.0 +/- 7.9 in group I and 7.9 +/- 5.8 in group II (p less than 0.05). One-month creatinine (mg/dl) was 2.9 +/- 2.8 in group I and 1.75 +/- 1.0 in group II (NS). One-month graft survivals (%) in groups I and II were 81.4% and 92.8%, respectively. In conclusion, UW solution has provided beneficial effect of preservation on ischemic damaged kidney and appears to be method of choice in non-heart beating cadaveric kidney transplantation.