In vitro and in vivo effects of cisplatin and etoposide in combination on small cell lung cancer cell lines

The effects of cisplatin (CDDP) and etoposide (ETP) in combination were evaluated in vitro and in vivo using small cell lung cancer cell lines. The combination effects in vitro were investigated using isobologram analysis. Used together, CDDP and ETP showed a synergistic effect against cell growth on only 1 cell line (SBC-3), additive effects on 6 (SBC-2, SBC-5, Lu130, Lu134AH, Lu135T and H69) and an antagonistic effect on 1 (SBC-1). In the in vivo experiment, nude mice were inoculated with SBC-1, SBC-3 and SBC-5 cells. Two or 5 mg/kg CDDP and 10 or 30 mg/kg ETP were administered intraperitoneally alone and simultaneously in combination to nude mice. The in vivo effects of the combination were determined by comparing the observed growth ratio in mice treated with the combination with the expected value of this ratio calculated based on the assumption that the effects of the drugs were simply additive. According to this definition, synergistic effects were observed against all 3 tumors. Thus, the in vivo and in vitro effects differed. The toxicity of the combination therapy, which was analyzed by estimating the body weight change of mice, was no higher than that of CDDP or ETP alone. These results suggest that the excellent clinical effects of CDDP and ETP combination therapy may be attributable not to drug interaction at the cellular level but to the feasibility of combined use of them at full doses without overlapping side effects.     

Interaction of combinations of drugs, chemosensitizers, and peptides with the P-glycoprotein multidrug transporter

P-Glycoprotein functions as an ATP-driven efflux pump for hydrophobic natural products and peptides, and gives rise to resistance to multiple chemotherapeutic drugs. The inhibition of colchicine transport via P-glycoprotein by various compounds was determined in a plasma membrane vesicle model system. A chemotherapeutic drug (vinblastine) and several chemosensitizers (verapamil, reserpine, cyclosporin A) and hydrophobic peptides (N-acetyl-leucyl-leucyl-methioninal, leupeptin, pepstatin A, valinomycin) were examined, both as individual species and as combinations of compounds. The median effect analysis was used to determine the concentration of each combination required to produce a median effect, Dm, as well as the sigmoidicity of the concentration-effect plot, m. The combination of cyclosporin A and verapamil was the only one established to be mutually nonexclusive, whereas several mutually exclusive pairs of compounds were identified. The combination index, CI, was calculated for several combinations of drugs, chemosensitizers, and peptides, and used to ascertain whether effects were synergistic, antagonistic, or additive. Some combinations (vinblastine/verapamil; verapamil/valinomycin) showed antagonism over the entire concentration range. Other combinations (valinomycin/N-acetyl-leucyl-leucyl-methioninal; cyclosporin A/verapamil) displayed both synergism and antagonism over different regions of the CI plot. Many combinations of compounds displayed additive interactions over most of the CI plot. The median effect analysis may be helpful in identifying potentially useful additive or synergistic combinations of compounds for reversal of Pgp-mediated drug resistance.     

Efficient designs for studying synergistic drug combinations

Distinguishing between pharmacologically additive and synergistic drug combinations requires experimental designs and statistical analyses that often require appreciable numbers of animals and much experimenter time. The current study employed a design in which individual dose-effect data from each drug were translated into theoretically additive total dose combinations, in a fixed drug proportion, in order to produce a composite additive dose-effect relation that could be compared with that of an actual mixture having the same proportion. Results from this approach, using a combination of intrathecal doses of morphine and clonidine, were virtually identical to those using isobolographic analysis of the same data set. Both analyses showed significant synergism for this combination and, in each method, it was not necessary to constrain the drug regression lines to parallelism. In contrast to the isobole approach, the use of the composite additive dose-effect relation also allows observation of the interaction over a range of effects while reducing the size of the data sets needed.     

Adenovirus-mediated p53 gene therapy and paclitaxel have synergistic efficacy in models of human head and neck, ovarian, prostate, and breast cancer

Synergy (or antagonism) between two chemical agents is an in vitro empirical phenomenon, in which the observed effect of the combination is more (or less) than what would be predicted from the effects of each agent working alone. Although mathematical synergy is not directly provable in the clinical setting, it does predict a favorable outcome when the two therapeutics are combined in vivo and strongly suggests the presence of in vivo synergy. In contrast, overt antagonism warns of future problems. Sophisticated three-dimensional statistical modeling was used to evaluate the presence of synergistic, additive, or antagonistic efficacy between adenovirus (Ad)-mediated p53 gene therapy (p53 Ad) and paclitaxel (Taxol) in a panel of human tumor cell lines. Cells were either pretreated with paclitaxel 24 h before p53 Ad or treated with both agents simultaneously. Cell proliferation was measured 3 days later. Paclitaxel had synergistic or additive efficacy with p53 gene therapy. In no case was the interaction antagonistic. Cell cycle analysis demonstrated that p53 Ad arrested cells in G0/G1 prior to apoptotic cell death, whereas paclitaxel arrested cells in G2-M prior to apoptotic cell death. When combined, the relative concentration of each agent determined the dominant cellular response. These results are consistent with the previously reported cell cycle effects of p53 or paclitaxel, respectively; however, these data fail to explain the observed drug synergy. We found that low concentrations of paclitaxel (1-14 nM) increased the number of cells transduced by recombinant Ad 3-35% in a dose-dependent manner, which is one possible mechanism for the observed synergy. Of particular note, the concentrations of paclitaxel responsible for increased Ad transduction were lower than the concentrations required for microtubule condensation. The efficacy of combination therapy was also evaluated in vivo. In the p53null SK-OV-3 xenograft model of ovarian cancer, a dosing schedule of p53 Ad that, by itself, had a relatively minimal effect on tumor burden (16%) caused a much greater decrease in tumor burden (55%) when combined with paclitaxel. Greater combined efficacy was also observed in the p53mut DU-145 prostate, p53mut MDA-MB-468 breast, and p53mut MDA-MB-231 breast cancer xenograft models in vivo. In summary, p53 Ad for cancer shows enhanced efficacy when combined with paclitaxel. This combination is recommended for clinical cancer trials.     

Confusion of concepts in mixture toxicology

Regulatory limit values are generally set for single compounds. However, humans are exposed both simultaneously and sequentially to a wide variety of compounds. Some concepts on mixture toxicology are discussed in this introduction to the European Conference on Combination Toxicology. Studies on mixtures are often accompanied by statements about the type of combined action, which can be, for example, additive, synergistic or antagonistic. Unfortunately, comparison of results is hardly possible for various reasons. First, the terminology for indicating combined action is far from consistent. Bearing this in mind, researchers should be explicit in the definitions of terms. Secondly, depending on the model, different conclusions may be drawn from the same results. It is therefore important to provide clear definitions of the null hypothesis. Thirdly, adequate statistical methods should be used for testing the null hypothesis. In the past, many mixtures studies either used no statistics or used statistics incorrectly. Last, but not least, the study should be designed in such a way that it should be possible to obtain clear answers. In this introduction, it is stressed that environmental toxicologists should focus on the low-dose region of the dose-effect curves. It appears that interactions are less plausible at low doses. Dose additivity, however, cannot be excluded.     

Primary cutaneous cryptococcosis as the presenting manifestation of AIDS

Presented is a case of primary cutaneous cryptococcosis, presenting as a papular facial lesion with subsequent ulceration, as the initial manifestation of AIDS. This unusual condition must be considered in the workup of unexplained or therapeutically unresponsive skin lesions. If it is diagnosed in an ostensibly "healthy" individual, evaluation for evidence of immunosuppression, and its underlying cause, must be undertaken.     

Synergistic antiproliferative activity of tamoxifen and docetaxel on three oestrogen receptor-negative cancer cell lines is mediated by the induction of apoptosis

The taxanes are a promising family of anti-tumour drugs that block cell cycle replication by interfering with the microtubule network. The clinical use of these drugs involves some problems related to their low solubility and occurrence of resistance, which is mainly dependent on the multidrug-resistant (MDR) phenotype. To investigate the possible interaction between docetaxel and tamoxifen (TAM), three oestrogen receptor-negative cancer cell lines, MDR- MDA-MB 231, MDR + CEM-VBLr and MCF-7 ADRr, were used. In all three cell lines, the combination of docetaxel and TAM was more effective in terms of growth inhibition than single drug exposure. Isobolic analysis confirmed the presence of synergism in all cell lines when docetaxel was used at 0.2 microM and TAM at a dose equal to or higher than 1 microM. Flow cytometric DNA analysis performed on the three cell lines showed that TAM was able to increase the G2/M blocking activity of docetaxel. This blocking activity was followed by an increased flow cytometric DNA fragmentation suggestive of the presence of apoptosis, which was confirmed by DNA gel fragmentation and morphological analysis. While an antagonistic effect on P-glycoprotein (P-gp) activity may contribute to the synergistic effect of tamoxifen and docetaxel on CEM-VBLr and MCF-7 ADRr, other mechanisms must be involved, as the synergistic effect is also apparent with a P-gp-negative cell line.     

Potencies of oral contraceptives

Oral contraceptives are combinations of estrogens and progestogens or, in the case of the mini-pills, progestogens alone. With specific test procedures in laboratory animals or human subjects, it is possible to assign potency evaluations to the components relative to the progestational, estrogenic, or antiestrogenic activities of the progestogen or to the estrogenic potencies of the estrogenic component. It might even be possible to quantify the synergistic effects of the estrogen on the progestational agent. Unfortunately, however, it is impossible now to amalgamate such assay results into single estimates of the potencies of the combinations (either the combination products per se or the combination tablets of sequential products). For example, an over-all estrogenic potency of a combination preparation would involve the integration of contributions form the estrogen itself plus the estrogenic products of metabolism of the progestogen minus the antagonistic effect of the progestational agent, if any. These factors cannot now be quantified independently, much less merged into a single figure of clinical significance. Further, even if it were possible to produce such an estimate, it is unlikely that the evaluation would be meaningful in relation to any putative side effect or adverse reaction, i.e., the alleged thrombogenic effects of oral contraceptives cannot currently be related directly to any measure of potency that will allow prediction of these clinical conditions from laboratory models. Any evaluation of the potential of a given contraceptive to produce a specific side effect will depend upon data generated with specific regard to that adverse reaction and the individual product in question.     

The Effect of the Herbicide Mecoprop on Heliscus lugdunensis and Its Influence on the Preferential Feeding of Gammarus Pseudolimnaeus

Ribonucleotide reductase, a key enzyme in deoxyribonucleotide synthesis, is an important target for cancer chemotherapy. Drugs that inhibit its individual components may act synergistically to block DNA synthesis. Prior work has established that gallium inhibits the R2 subunit of ribonucleotide reductase. We show that gallium acts synergistically with the ribonucleotide reductase inhibitors gemcitabine and hydroxyurea to inhibit the proliferation of CCRF-CEM cells. In contrast, combinations of gallium with the ribonucleotide reductase inhibitors amidox, didox, or trimidox produced antagonistic effects on cell growth. Spectroscopy analysis revealed that as a result of their metal-binding properties, amidox, didox and trimidox formed complexes with gallium, thus negating potential synergistic actions. Our results have important implications in the design of clinical trials using these ribonucleotide reductase inhibitors in combination.     

Synergistic effect of the Bacillus thuringiensis toxins CryIAa and CryIAc on the gypsy moth, Lymantria dispar

The insecticidal activity of toxins CryIAa, CryIAb, and CryIAc against Lymantria dispar (gypsy moth) and Bombyx mori (silkworm) was examined by force-feeding bioassays. Toxin CryIAa exhibited higher toxicity than toxins CryIAb and CryIAc for L. dispar and B. mori. To evaluate possible synergism among these toxins, bioassays were performed with mixtures of CryIAa and CryIAb, CryIAb and CryIAc, and CryIAa and CryIAc. Expected toxicity was calculated from the activity of each individual toxin and its proportion in the mixture by using the equation described by Tabashnik (B. E. Tabashnik, Appl. Environ. Microbiol. 58:3343-3346, 1992). Observed 50% growth-inhibitory doses were calculated from mixing experiments by probit analysis. In L. dispar bioassays, synergism was observed with a mixture of CryIAa and CryIAc while a mixture of CryIAa and CryIAb exhibited an antagonistic effect. No synergistic effect on B. mori was observed with any toxin combination. Voltage clamping assays of isolated L. dispar midguts also demonstrated that the mixture of CryIAa and CryIAc induced a greater slope of inhibition of short circuit current than did other toxin combinations.     

Regulation of Na+ channel beta 1 and beta 2 subunit mRNA levels in cultured rat astrocytes

This work describes the molecular mechanism of fatty acid and hormonal modulation of retinoid X receptor (RXR alpha) in rat liver. We examined the effects of different fatty acids (myristic-, stearic-, linolenic-, oleic-, arachidonic- and tetradecylthioacetic acid (TTA)) and the synthetic glucocorticoid dexamethasone on RXR alpha mRNA and protein steady-state levels in hepatoma cells and cultured hepatocytes. Fatty acids induced the RXR alpha gene expression where TTA showed the most inductive effect (three-fold induction). Dexamethasone alone resulted in a stronger induction (up to seven-fold in hepatocytes), and in combination with fatty acids, an additive or synergistic effect was observed. The RXR alpha protein level in cultured hepatocytes showed a similar pattern of regulation, with a slight inductive effect of fatty acids and an additive or synergistic effect was observed in combination with dexamethasone. Our results indicate that the RXR alpha gene expression is under distinct regulation by fatty acids and dexamethasone acid which strongly suggests a coupling with the lipid metabolizing system and the retinoid signaling pathway.     

A queue-series model for reaction time, with discrete-stage and continuous-flow models as special cases.

Presents a new reaction time (RT) model that includes both sequential-stage (discrete) and overlapping-stage (continuous-flow) models as special cases. In the new model, task performance is carried out by a series of distinct processing stages, each of which functions as a queue. A stimulus conveys 1 or more distinct components of information (e.g., features), and each stage can begin processing as soon as it receives 1 component from its predecessor. If a stimulus activates only 1 component, successive stages operate in strict sequence; if it activates multiple components, successive stages operate with temporal overlap. Within this class of models, experimental factors affecting different processing stages always have additive effects on RT with sequential stages but rarely do so with overlapping stages. Within this class of models, then, observations of factor additivity support discrete-stage models. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Heme oxygenase and carbon monoxide: regulatory roles in islet hormone release: a biochemical, immunohistochemical, and confocal microscopic study

Two randomized, double-blind, parallel-group single-dose 2 x 2 factorial analgesic studies compared a single-dose or a 2-tablet dose of a combination of 7.5 mg hydrocodone bitartrate with 200 mg ibuprofen with each constituent alone and with a placebo in women with moderate or severe postoperative pain from abdominal or gynecologic surgery. A nurse-observer recorded patient reports of pain intensity and pain relief periodically for 8 hours. In both studies, the combination was significantly superior to placebo for sum of the pain intensity differences (SPID), total pain relief (TOTPAR), peak pain intensity difference (PID) and pain relief, global evaluation, and time to remedication. The combination was likewise significantly superior to both hydrocodone and ibuprofen for most of these summary measures of analgesia. In a factorial analysis, both the hydrocodone and ibuprofen effects were significant for most summary measures of analgesia, whereas results of the interaction contrast were consistent with the concept that the analgesic effect of the combination represents the additive analgesia of its 2 constituents.     

Dose-response characteristics of cholesterol-lowering drug therapies: implications for treatment

PURPOSE: To develop an optimal treatment strategy that reduces low-density lipoprotein (LDL) cholesterol levels and improves adherence to therapy by reviewing clinical trials that define the dose-response characteristics for 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors (statins), bile acid sequestrants, and niacin. DATA SOURCES: Data were obtained from a MEDLINE search of the English-language literature published from 1975 through November 1995 and from an extensive bibliography review. STUDY SELECTION: Controlled, clinical trials were reviewed if they evaluated 1) the effectiveness and toxicity of one LDL cholesterol-lowering agent (statins, bile acid sequestrants, or niacin, at two or more doses) or 2) monotherapy with two LDL cholesterol-lowering agents at defined doses used alone and in combination. Studies that had fewer than 10 patients in a treatment group or that selected patients on the basis of previous response to therapy were not included. DATA EXTRACTION: Trials were reviewed for overall methodology, inclusion and exclusion criteria, sources of bias, and outcomes. DATA SYNTHESIS: Dose-response relations for bile acid sequestrants and statins are nonlinear, and most of their LDL cholesterol-lowering effects can be obtained with lower doses. The few dose-response studies of niacin that have been done suggest that most of niacin's high-density lipoprotein cholesterol-increasing effect can also be achieved with relatively low doses, but higher doses are needed to substantially reduce LDL cholesterol levels. If bile acid sequestrants or niacin are added to statin therapy, the effect of combined therapy on LDL cholesterol levels is additive. CONCLUSION: The nonlinear dose-response relation of statins, bile acid sequestrants, and niacin and their additive LDL cholesterol-lowering effect when used together suggest a strategy for treating hypercholesterolemia that may optimize effectiveness while minimizing adverse effects and cost.     

Age-specific inbreeding depression and components of genetic variance in relation to the evolution of senescence

Two major theories of the evolution of senescence (mutation accumulation and antagonistic pleiotropy) make different predictions about the relationships between age, inbreeding effects, and the magnitude of genetic variance components of life-history components. We show that, under mutation accumulation, inbreeding decline and three major components of genetic variance are expected to increase with age in randomly mating populations. Under the simplest version of the antagonistic pleiotropy model, no changes in the severity of inbreeding decline, dominance variance, or the genetic variance of chromosomal homozygotes are expected, but additive genetic variance may increase with age. Age-specific survival rates and mating success were measured on virgin males, using lines extracted from a population of Drosophila melanogaster. For both traits, inbreeding decline and several components of genetic variance increase with age. The results are consistent with the mutation accumulation model, but can only be explained by antagonistic pleiotropy if there is a general tendency for an increase with age in the size of allelic effects on these life-history traits.     

Synergistic enhancement of small and large intestinal carcinogenesis by combined treatment of rats with five heterocyclic amines in a medium-term multi-organ bioassay

The carcinogenic potential of five heterocyclic amines in combination was analyzed using a medium-term multi-organ bioassay. Male F344 rats were initially treated with five known carcinogens (diethylnitrosamine, N-methyl-N-nitrosourea, N-butyl-N-(4-hydroxybutyl)-nitrosamine, 1,2-dimethylhydrazine and 2,2'-dihydroxy-di-n-propylnitrosamine) over a 4 week period to induce preneoplastic changes in a variety of organs (wide spectrum initiation) and then given the five heterocyclic amines, all having the intestines as a target of their carcinogenicity, individually or in combination in the diet for a further 24 weeks. In the small and large intestines, simultaneous administration of five heterocyclic amines at doses 1/5 or 1/25 of those used in reported carcinogenicity studies resulted in higher incidences and multiplicities of adenocarcinomas than expected from the five individual effects, although the differences were not statistically significant. A synergistic effect based on the additive model was most evident (P < 0.141) with multiplicity data for carcinoma in the small intestine at the 1/25 dose. A similar trend was observed for Zymbal gland (P < 0.077), but not other carcinoma induction. Thus the results suggested that synergism depends on the carcinogenic organotropism of individual agents as well as the doses applied in combination.     

The validity of melatonin as an oncostatic agent

The validity of melatonin as a prominent, naturally occurring oncostatic agent is examined in terms of its putative oncostatic mechanism of action, the correlation between melatonin levels and neoplastic activity, and the outcome of therapeutically administered melatonin in clinical trials. Melatonin's mechanism of action is summarized in a brief analysis of its actions at the cellular level, its antioxidative functions, and its indirect immunostimulatory effects. The difficulties of interpreting melatonin levels as a diagnostic or prognostic aid in cancer is illustrated by referral to breast cancer, the most frequently studied neoplasm in trials regarding melatonin. Trials in which melatonin was used therapeutically are reviewed, i.e., early studies using melatonin alone, trials of melatonin in combination with interleukin-2, and controlled studies comparing routine therapy to therapy in combination with melatonin. A table compiling the studies in which melatonin was used in the treatment of cancer in humans is presented according to the type of neoplasm. Melatonin's suitability in combination chemotherapy, where it augments the anticancer effect of other chemotherapeutic drugs while decreasing some of the toxic side effects, is described. Based on the evidence derived from melatonin's antiproliferative, antioxidative, and immunostimulatory mechanisms of action, from its abnormal levels in cancer patients and from clinical trials in which melatonin was administered, it is concluded that melatonin could indeed be considered a physiological anticancer substance. Further well-controlled trials should, however, be performed in order to find the link between its observed effects and the underlying mechanisms of action and to define its significance as a therapeutic oncostatic agent.     

Suppression of TNF-alpha expression, inhibition of Th1 activity, and amelioration of collagen-induced arthritis by rolipram

Rolipram is a type IV phosphodiesterase inhibitor that suppresses inflammation and TNF-alpha production. As anti-TNF-alpha therapy is effective in rheumatoid arthritis, we investigated the effect of rolipram on collagen-induced arthritis (CIA), a murine model of rheumatoid arthritis. Rolipram was administered after the onset of clinical arthritis at doses of 0.5, 3, 5, or 10 mg/kg twice daily, with a dose-dependent therapeutic effect on clinical severity and joint erosion. Immunohistochemical analysis of joints of rolipram-treated mice revealed 67% reduction in TNF-alpha-expressing cells compared with control arthritic mice. In vitro studies using bone marrow-derived macrophages confirmed that rolipram directly suppressed TNF-alpha and IL-12 production following stimulation with IFN-gamma and LPS. The effect of rolipram on T cell activity was studied by measuring Th1/Th2 cytokine production by collagen-stimulated draining lymph node cells from arthritic mice treated in vivo with rolipram. Rolipram reduced IFN-gamma production and increased IL-10, indicating that rolipram down-regulated the ongoing Th1 response to type II collagen. Finally, the effect on CIA of combination therapy was studied using rolipram plus either anti-TNF-alpha or anti-CD4 mAbs. Rolipram plus anti-TNF-alpha was not therapeutically additive, whereas rolipram plus anti-CD4 mAb was clearly additive. This result indicates that the therapeutic effects of rolipram overlap with TNF-alpha blockade, but are complementary to anti-CD4 treatment. It is therefore proposed that a major mechanism of action of rolipram in CIA is suppression of TNF-alpha activity. These findings suggest that type IV phosphodiesterase inhibitors may be effective in pathologic conditions, such as RA, with overexpression of TNF-alpha.     

Interactions of erythropoietin, granulocyte colony-stimulating factor, stem cell factor, and interleukin-11 on murine hematopoiesis during simultaneous administration

We investigated how in vivo effects of single hematopoietic cytokines change if given in combination for a prolonged time. Mice were treated with every combination of recombinant human (rh) erythropoietin (EPO), rh granulocyte colony-stimulating factor (G-CSF), recombinant rat (rr) stem cell factor (SCF), and rh interleukin (IL)-11 by continuous infusion over 7 days (full factorial design with three dose levels for each cytokine). Burst-forming unit-erythroid (BFU-E), colony-forming unit-erythroid (CFU-E), and colony-forming unit-granulocyte-macrophage (CFU-GM) were determined in bone marrow and spleen, reticulocytes, hematocrit, granulocytes, and thrombocytes in the peripheral blood. An analysis of variance (ANOVA) and multiple comparison of means was used to evaluate the data. For several cell types, cytokine effects superimposed in an additive way if combined. However, in a large number of circumstances, nonadditive pairwise interactions were found. They differed in type and magnitude involving high-dose saturation, high-dose antagonistic effects, and even effect reversals (qualitative interactions). Hence, in general, it was not possible to foresee the combination effects on the basis of existing knowledge of single effects. On the other hand, the cytokine network was robust and no system hazards were observed under multiple cytokine combinations. The results illustrate that the cytokine network has nonlinear dynamic properties in vivo with dose-response characteristics of one cytokine being continuously modified by other cytokines.     

Evaluation of the efficacy and tolerability of a low-dose combination of isradipine and spirapril in the first-line treatment of mild to moderate essential hypertension

To investigate the concept of initiating therapy with low doses of a calcium antagonist and an ACE inhibitor, a fixed combination of isradipine 2.5 mg plus the ACE inhibitor spirapril 3 mg was compared with its components, with the full-dose monotherapies (isradipine 5 mg or spirapril 6 mg), and with placebo. After a 2-week wash out phase in pretreated patients and a subsequent 2-week placebo period, 405 patients with a diastolic blood pressure (DBP) between 100 and 114 mmHg were randomly allocated to 12-week once-daily double-blind treatment in one of the six treatment arms. In patients whose blood pressure was not normalized (defined as DBP< or =90 mmHg) after 6 weeks of treatment, the dosage of either medication was doubled or, in the placebo group, was switched to the fixed combination. After week 6, the mean reductions from baseline in sitting systolic/diastolic blood pressure 24 hours after dosing (trough) for the fixed combination or the monotherapies isradipine 5 mg, isradipine 2.5 mg, spirapril 6 mg, spirapril 3 mg, and placebo were 10.4/8.7, 10.0/9.4, 6.5/6.7, 10.0/8.3, 7.0/5.8, and 2.2/4.7 mmHg, respectively. The blood pressure changes obtained with the low-dose fixed combination were essentially identical to those observed with the full-dose monotherapies, thus showing an additive effect of low-dose isradipine and spirapril. In terms of tolerability, the lowest rate of any adverse events was found in the combination group. In this group, typical adverse events of calcium antagonists, such as headache, flushing, ankle edema, or palpitations, were observed only in 5%, 2%, 1%, and 0%, respectively, dry cough, considered typical for ACE inhibitors, was observed in only 1% of the combination group. In conclusion, the low-dose components isradipine 2.5 mg and spirapril 3 mg were shown to have an additive effect when combined, exerting a blood pressure-lowering effect comparable with the full doses and a trend to a better tolerability profile in comparison with the standard doses. Thus, low-dose combination therapy with these drugs appears to be a rational alternative to conventional monotherapy in the first-line treatment of hypertension.