OBJECTIVE

This study evaluated the frequency and clinical significance of abnormal Thyroid Stimulating Hormone (TSH) and Free Thyroxine Index (FTI) in patients with bipolar I disorder treated primarily with lithium.

METHODS

We evaluated the medical records of 143 participants in the Pittsburgh study of Maintenance Therapies in Bipolar Disorder who did not have a thyroid abnormality at entry.

RESULTS

Thirty-six percent of the 143 patients developed abnormal TSH and/or FTI values. Thirty-eight percent of the 135 patients who received lithium developed abnormal TSH and/or FTI, spent significantly longer time in the acute treatment phase (t = -3.6, df = 133, p = .0004), and had significantly higher mean Hamilton Scale for Depression scores over the course of the maintenance phase (t = -2.3, df = 71.6, p = .03). Time on lithium and development of abnormal TSH and/or FTI were positively correlated (r = .25, p = .004).

CONCLUSIONS

Thyroid dysfunction can be frequent in patients exposed to lithium treatment for bipolar I disorder; it also appears to be correlated with a slower response to acute treatment, and may be related to poorer quality of long-term remission. A prospective study is needed to confirm our findings and determine whether more aggressive thyroid replacement can optimize thyroid function to facilitate clinical recovery.

BACKGROUND

Supported in part by National Institute of Mental Health Grants MH 029618 (Drs. Frank and Fagiolini) and MH 030915 (Drs. Kupfer and Fagiolini), and the Bosin Memorial Fund of The Pittsburgh Foundation (Drs. Fagiolini, Kupfer, Cook, Scott, Novick and Frank). Dr. Fagiolini is on the advisory board and a consultant to Pfizer Inc, and Bristol Myers Squibb, and is on the speaker bureau of Bristol Myers Squibb, Eli Lilly Italy, Pfizer Inc, and Shire. Dr. Frank is on the advisory board of Pfizer Inc. and Eli Lilly & Company, and is a consultant to Pfizer Italia and Servier Amerique. Dr. Kupfer is on the advisory board of Pfizer, Inc., Forest Pharmaceuticals, Inc., and Solvay-Wyeth Pharmaceuticals, and is a consultant to Servier Amerique.

BACKGROUND

Pulmonary vein reconnection (PVR) still determines recurrences of atrial fibrillation after contact force (CF)-guided pulmonary vein isolation. We studied whether acute PVR (adenosine and waiting time) and late PVR (at repeat) are explained by incomplete transmurality and contiguity within the deployed radiofrequency circle.

RESULTS

We analyzed 42 CF-guided ipsilateral pulmonary vein isolation procedures. For each radiofrequency tag within the circle, we collected data reflecting lesion depth (time of application, power, impedance drop [Δ-Imp], CF, force-time integral [FTI], and ablation index [AI]) and contiguity (automated interlesion distance [ILD]). Ablation line contiguity index (ALCI) was developed as a novel automated algorithm combining depth and contiguity into one single criterion. Each circle was subdivided into 10 segments. For each segment, we determined its weakest link by annotating timemin, powermin, Δ-Impmin, CFmin, FTImin, AImin, ILDmax, and ALCImin. Compared with segments without PVR (n=758), PVR segments (n=44) were characterized by lower Δ-Impmin (4.8 versus 7.4 Ω), CFmin (8.5 versus 11.8 g), FTImin (351 versus 473 gs), AImin (367 versus 408 arbitrary unit [au]), and higher ILDmax (6.8 versus 5.5 mm). ALCImin was significantly lower in segments with PVR (74% versus 104%; P<0.001) and was associated with the highest accuracy to predict durable segments (area under the curve=0.73).

CONCLUSIONS

In CF-guided pulmonary vein isolation, PVR is explained by lack of both lesion depth and contiguity within the deployed radiofrequency circle. ALCI, a novel measure combing contiguity and depth, is the most accurate predictor for durable segments. By avoiding weak links in the ablation chain, ALCI-guided ablation is expected to improve success rate of point-by-point radiofrequency ablation.

OBJECTIVE

Severe iodine deficiency disorders (IDDs) may have been eradicated in many parts of the world, but milder forms still exist and may escape detection. We evaluated the impact of pregnancy on the maternal and fetal thyroid axis in Hong Kong, a coastal city in southern China with borderline iodine intake.

METHODS

A prospective study performed in a maternity hospital.

METHODS

Two hundred and thirty pregnant women were prospectively studied and their neonates assessed at birth.

METHODS

Urine iodine concentration, thyroid function tests and thyroid volume (TV) by ultrasound were determined in the mothers during pregnancy and up to 3 months postpartum and in the neonates.

RESULTS

Increased urinary iodine concentration was seen from first trimester onwards and the proportion of women having urine iodine concentration of < 0.4 micromol/l decreased from 11.3% in the first trimester to 4.7% in the third trimester. There was progressive reduction in circulating fT4 and fT3 concentrations and free thyroxine index (FTI) with increasing gestation and the percentage of women having subnormal levels at term were 53.2%, 61.1% and 4.8%, respectively. The serum TSH concentration during pregnancy doubled towards term. In the first trimester, multiparous women had significantly larger TV than the nulliparous women (P < 0.001). By the third trimester, TV had increased by 30% (range 3-230%) so that the goitre incidence was 14.1%, 21.8%, 25.9% during the three trimesters of pregnancy, and 24.3% and 21.9% at 6 weeks and 3 months postpartum (ANOVA, P < 0.05). The change in thyroid volume during pregnancy correlated positively with the change in thyroglobulin (r = 0.225, P < 0.002) and negatively with urinary iodine concentration (r = - 0.149, P < 0.02). Fourteen women with excessive thyroidal stimulation in the second trimester (defined as those with thyroglobulin (Tg) concentrations in the highest tertile and FTI in the lowest tertile) were found to have lower urine iodine concentrations and larger TV (both P < 0.005) throughout pregnancy, and their neonates had higher cord TSH (P < 0.05), Tg (P < 0.05) and slightly larger TV (P = 0.06) as compared to the findings in 216 pregnant women without evidence of thyroid stimulation. Seven neonates (50%) born to these women had subnormal fT4 levels at birth.

CONCLUSIONS

In a borderline iodine sufficient area, pregnancy posed an important stress resulting in higher rates of maternal goitrogenesis as well as neonatal hypothyroxinaemia and hyperthyro- trophinaemia. An adequate iodization program is necessary to eliminate iodine deficiency disorders during pregnancy.

Protein farnesylation, catalyzed by protein farnesyltransferase, plays important roles in the membrane association and protein-protein interaction of a number of eukaryotic proteins. Recent development of farnesyltransferase inhibitors (FTIs) has led to further insight into the biological significance of farnesylation in cancer cells. A number of reports point to the dramatic effects FTIs exert on cancer cells. In addition to inhibiting anchorage-independent growth, FTIs cause changes in the cell cycle either at the G1/S or at the G2/M phase. Furthermore, induction of apoptosis by FTIs has been reported. FTIs also affects the actin cytoskeleton and cell morphology. This review summarizes these reports and discusses implications for farnesylated proteins responsible for these FTI effects.

OBJECTIVE

To compare the fatal toxicities of antidepressant drugs in England, Scotland and Wales 1985-1989.

METHODS

Epidemiological retrospective study using Department of Health prescription data and mortality data from the Office of Population Censuses and Surveys, and the Registrar General for Scotland, for the years 1974-1989. The fatal toxicity index (FTI) of groups of drugs and individual drugs was compared with the FTI for all antidepressant drugs for the years 1985-1989.

RESULTS

Of 3,604 single antidepressant deaths between 1975 and 1989, the majority (70.95%) were from amitriptyline or dothiepin. The mean FTI for all drugs for the years 1985-1989 was 35.6; the FTIs for dothiepin, amitriptyline, nortriptyline and tranylcypromine were significantly higher than the mean of all, while those for clomipramine, lofepramine, fluvoxamine, trimipramine, maprotiline, trazodone, mianserin, protriptyline, isocarboxazid and phenelzine were lower. The FTI for the older tricyclic drugs was higher at 43.03 (p < 0.001). The FTI for the monoamine oxidase inhibitors, of 27.03 (p = 0.045), and for all drugs introduced after 1973, of 5.32 (p < 0.001), were each significantly lower than the mean of all drugs.

CONCLUSIONS

Overdose deaths from antidepressants have not decreased over the last 15 years. A trend away from prescribing drugs with a higher fatal toxicity index in favour of those with a lower index, would reduce the number of deaths from antidepressant poisoning.

We investigated for the first time the ability of farnesyltransferase inhibitors (FTI) to radiosensitize human glioma. For this, human glioma cell lines were treated with the specific FTI, R115777, 48 hr prior to a 2Gy irradiation. The treatment with R115777 decreased by 45% the SF2 value of the more radioresistant glioma cell lines (SF763 and U87) without any significant effect on the radioresistance of the radiosensitive ones (SF767 and U251-MG). This radiosensitizer effect was due to the induction of post-mitotic necrotic cell death. We then tested the hypothesis that wild-type Ras or RhoB, which has been proposed as potential FTI target, could control the glioma radioresistance. For this, we expressed inducible dominant negative forms of Ras (RasN17) and RhoB (RhoBN19) in radioresistant U87 glioma cell line and analyzed the survival after irradiation of the obtained clones. While blocking Ras pathways by expression of RasN17 did not affect the SF2 value of the U87 glioma cell line, the expression of RhoBN19 dramatically reduced the cell survival after irradiation of these cells. Taken together, these data demonstrated that RhoB, but not Ras, is implicated in glioma radioresistance. Furthermore, the R115777 differential radiosensitizer effect underlines the potential therapeutic interest of using this drug as a radiosensitizer of human glioma.

Statins, which inhibit 3-hydroxy-3-methylglutaryl-CoA reductase and thus the synthesis of cholesterol, are remarkably effective in the treatment of cardiovascular disease. In addition to their favorable effect on lipid profile, these drugs may also prevent the proliferation of vascular smooth muscle that is characteristic of atherosclerosis. We hypothesize that statins prevent the post-translational prenylation, and thus inhibit the function, of critical small GTPases in vascular smooth muscle cells. We have therefore assayed the effect of lovastatin on both the growth of A10 vascular smooth muscle cells and the status of their Ras and RhoB proteins. We find that < or =1 microM lovastatin potently inhibits the proliferation of A10 cultures, and higher concentrations >> or =3 microM) induce apoptosis. We have also tested the effect of 3-allylfarnesol (3-alFOH), an inhibitor of farnesyl transferase (FTI). The data show that although>> or =10 microM 3-alFOH is required for a cytostatic effect, the action of 3 microM 3-alFOH can be greatly potentiated by even nanomolar levels of lovastatin. We also find that lovastatin and 3-alFOH exhibit synergism to cause the up-regulation and relocalization of RhoB from the membrane to cytosolic compartments. This relocalization of RhoB, which is presumed to reflect an inhibition of its prenylation, correlates with the proapoptotic activities of combined 3-alFOH and lovastatin treatment. These data suggest that RhoB may be a valuable pharmacological target in cardiovascular disease, and that combinations of statins and certain FTIs may be of value in treatment of disorders that are characterized by excess cell proliferation.

BACKGROUND

Bronchial fibroblasts contribute to airway remodelling, including airway wall fibrosis. Transforming growth factor (TGF)-β1 plays a major role in this process. We previously revealed the importance of the mevalonate cascade in the fibrotic response of human airway smooth muscle cells. We now investigate mevalonate cascade-associated signaling in TGFβ1-induced fibronectin expression by bronchial fibroblasts from non-asthmatic and asthmatic subjects.

METHODS

We used simvastatin (1-15 μM) to inhibit 3-hydroxy-3-methlyglutaryl-coenzyme A (HMG-CoA) reductase which converts HMG-CoA to mevalonate. Selective inhibitors of geranylgeranyl transferase-1 (GGT1; GGTI-286, 10 μM) and farnesyl transferase (FT; FTI-277, 10 μM) were used to determine whether GGT1 and FT contribute to TGFβ1-induced fibronectin expression. In addition, we studied the effects of co-incubation with simvastatin and mevalonate (1 mM), geranylgeranylpyrophosphate (30 μM) or farnesylpyrophosphate (30 μM).

RESULTS

Immunoblotting revealed concentration-dependent simvastatin inhibition of TGFβ1 (2.5 ng/ml, 48 h)-induced fibronectin. This was prevented by exogenous mevalonate, or isoprenoids (geranylgeranylpyrophosphate or farnesylpyrophosphate). The effects of simvastatin were mimicked by GGTI-286, but not FTI-277, suggesting fundamental involvement of GGT1 in TGFβ1-induced signaling. Asthmatic fibroblasts exhibited greater TGFβ1-induced fibronectin expression compared to non-asthmatic cells; this enhanced response was effectively reduced by simvastatin.

CONCLUSIONS

We conclude that TGFβ1-induced fibronectin expression in airway fibroblasts relies on activity of GGT1 and availability of isoprenoids. Our results suggest that targeting regulators of isoprenoid-dependent signaling holds promise for treating airway wall fibrosis.

Resistance of glioblastoma multiforme to radiotherapy poses a major clinical challenge. Farnesyltransferase inhibitors (FTI), such as R115777, have potential to increase radiotherapeutic benefit in this disease, although their mechanism of action is unclear. In our study with eight glioblastoma multiforme cell lines, the most sensitive ones underwent cell cycle arrest in response to FTI treatment. Radiosensitization by FTIs, however, seemed to involve other pathways. If R115777 treatment was initiated < 6 hours before irradiation, all eight glioblastoma multiforme lines were radiosensitized. However, if the time between drug and radiation was extended to 24 hours, cells harboring wild type but not mutated p53 were able to counteract drug-induced radiosensitization. The involvement of the p53/p21 pathway in the development of resistance was confirmed by showing that U87 cells transfected with human papillomavirus E6 to block p53 or interfering RNA to inhibit p21 stayed radiosensitive for 24 hours after drug treatment. The time dependency of R115777-induced radiosensitization suggested that the initial FTI target for early radiosensitization was short-lived, and that a p21-directed pathway restored resistance. Consideration of prenylated molecules that could potentially be involved led us to consider HDJ-2, a co-chaperone of heat shock protein 70. This hypothesis was strengthened by finding that cellular radiosensitivity was increased by genetic inhibition of HDJ-2, whereas overexpression conferred radioresistance. Importantly, irradiation of cells caused HDJ-2 to migrate from the cytoplasm to the nucleus, and this migration was inhibited by prior FTI treatment. These results have clinical relevance in that they help explain the variability in responses to FTIs that occurs following radiotherapy and elucidate some of the reasons for the complexity underlying FTI-induced radiosensitization.

Previous studies have demonstrated that astrocytomas express elevated levels of activated Ras.GTP despite the absence of activating Ras mutations. Farnesyl transferase inhibitors (FTIs) exert their antitumor effect in part through inhibition of Ras-mediated signaling. SCH66336 is a potent FTI presently undergoing clinical trials in patients with solid tumors. We evaluated the efficacy of SCH66336 against a panel of eight human astrocytoma cell lines and three human astrocytoma explant xenograft models in NOD-SCID mice. SCH66336 demonstrated variable antiproliferative effects against the cell lines, with IC(50) ranging from 0.6 microM to 32.3 microM. Two of the three human glioblastoma multiforme (GBM) xenografts demonstrated substantial growth inhibition in response to SCH66336, with up to 69% growth inhibition after 21 days of treatment. Drug efficacy could be accurately predicted using a combination of the H-, K-, and N-isotype-specific Ras.GTP levels. These data indicate that the absence of Ras mutations does not preclude chemotherapeutic efficacy by FTIs, that Ras is likely a major target of FTIs regardless of Ras mutational status, and that isotype-specific Ras.GTP levels are a promising marker of drug efficacy.

The fact that proteins such as Ras, Rac and RhoA require farnesylation or geranylgeranylation to induce malignant transformation prompted many investigators to develop farnesyltransferase (FTase) and geranylgeranyltransferase I (GGTase I) inhibitors (FTIs and GGTIs, respectively) as novel anticancer drugs. Although FTIs have been shown to antagonise oncogenic signalling, reverse malignant transformation, inhibit human tumour growth in nude mice and induce tumour regression in transgenic mice without any signs of toxicity, their mechanism of action is not known. This review will focus on important mechanistic issues as well as bench to bedside translational issues. These will include the relevance to cancer therapy of the alternative geranylgeranylation of K-Ras when FTase is inhibited; a thorough discussion about evidence for and against the involvement of inhibition of prenylation of Ras and RhoB in the mechanism of FTIs' antitumour activity as well as effects of FTIs and GGTIs on the cell cycle machinery and the dynamics of bipolar spindle formation and chromosome alignment during mitosis. Bench to bedside issues relating to the design of hypothesis-driven clinical trials with biochemical correlates for proof-of-concept in man will also be discussed. This will include Phase I issues such as determining maximally tolerated dose (MTD) versus effective biological dose (EBD), as well as whether Phase II trials are still needed for clinical evaluations of anti-signalling agents. Other questions that will be addressed include: what levels of inhibition of FTase activity are required for tumour response in Phase II clinical evaluations? What FTase substrates are most relevant as biochemical correlates? Are signalling pathways such as H-Ras/PI3K/Akt and K-Ras/Raf/MEK/Erk significant biological readouts? Does Ras mutation status predict response? What are appropriate clinical end-points for FTI Phase II trials? For this latter important question, time to tumour progression, median survival, percentage of patients that progress, clinical benefits and improvement in quality of life will all be discussed.

Muscle contractions induced by functional electrical stimulation (FES) tend to result in rapid muscle fatigue, which greatly limits activities such as FES-assisted standing and walking. It was hypothesized that muscle fatigue caused by FES could be reduced by randomly modulating parameters of the electrical stimulus. Seven paraplegic subjects participated in this study. While subjects were seated, FES was applied to quadriceps and tibialis anterior muscles bilaterally using surface electrodes. The isometric force was measured, and the time for the force to drop by 3 dB (fatigue time) and the normalized force-time integral (FTI) were determined. Four different modes of FES were applied in random order: constant stimulation, randomized frequency (mean 40 Hz), randomized current amplitude, and randomized pulsewidth (mean 250 micros). In randomized trials, stimulation parameters were stochastically modulated every 100 ms in a range of +/-15% using a uniform probability distribution. There was no significant difference between the fatigue time measurements for the four modes of stimulation. There was also no significant difference in the FTI measurements. Therefore, our particular method of stochastic modulation of the stimulation parameters, which involved moderate (15%) variations updated every 100 ms and centered around 40 Hz, appeared to have no effect on muscle fatigue. There was a strong correlation between maximum force measurements and stimulation order, which was not apparent in the fatigue time or FTI measurements. It was concluded that a 10-min rest period between stimulation trials was insufficient to allow full recovery of muscle strength.

Isoprenoid inhibitors are being evaluated as agents for the treatment of cancer. Their antitumor activity is attributed to inhibition of post-translational modification of Ras, which is crucial for its translocation and attachment to the plasma membrane, and ultimate involvement in signal transduction. However, whether blocking of Ras is solely responsible for the observed antitumor activity is unresolved. In this report, we propose an alternate mechanism. Using breast tumor models, we show that agents possessing a lactone moiety, including statins (such as lovastatin) and the isoprenoid inhibitors (such as FTI-277 and GGTI-298), mediate their cell cycle inhibitory activities by blocking the chymotrypsin activity of the proteasome in vitro. This results in the accumulation of cyclin-dependent kinase inhibitors p21 and p27 with subsequent G(1) arrest. Cells devoid of p21 were refractory to the growth-inhibitory activity of lovastatin, FTI-277, and GGTI-298. However, in these p21 null cells, isoprenylation of key substrates of farnesyl transferase (such as Ras) and of geranylgeranyl transferase (such as RAP-1) were inhibited by FTI-277 and GGTI-298, respectively, suggesting that although both these isoprenoid inhibitors reached and inhibited their intended targets, inhibition of the isoprenylation of Ras and RAP-1A are not sufficient to mediate G(1) arrest. We also show that the cell cycle effects can be attributed to the functional lactone moiety of the aforementioned agents. Collectively, our data suggest that FTI and GGTI and other agents containing an active lactone moiety mediate G(1) arrest via inhibition of the proteasome and up-regulation of p21, independent of the inhibition of isoprenylation of Ras or RAP-1.

The study aimed to characterize curcumin (CCM) (fluorescent yellow curry pigment) labeling of neuronal fibrillar tau inclusions (FTIs) in representative cases of 3 main tauopathies: Alzheimer disease (AD), progressive supranuclear palsy, and Pick disease. After identification of FTIs in hematoxylin and eosin-stained brain sections, sequential labeling and signal colocalization image analysis were used to compare CCM with thioflavine S (ThS), monoclonal antibody AT8 immunofluorescence, and Gallyas silver staining by visualizing the same FTIs. Curcumin preference for specific tau isoforms was tested with 3-repeat tau and 4-repeat tau isoform-specific immunofluorescence. Curcumin proved highly comparable to ThS and Gallyas staining in its detection of FTIs. When comparing CCM with AT8, ThS, and Gallyas staining in AD and progressive supranuclear palsy, 3 types of neuronal tau deposits were observed: nonfibrillar intracellular material labeled only with AT8, fibrillar intracellular inclusions labeled by all the methods, and fibrillar extracellular FTIs labeled with CCM, ThS, and Gallyas staining but not with AT8. Although CCM labeling overlapped with both 3-repeat tau and 4-repeat tau in AD, it did not label 3-repeat tau FTIs in Pick disease probably because of their different ultrastructural characteristics. In summary, CCM fluorescence reliably detected neuronal FTIs in AD and progressive supranuclear palsy and surpassed AT8 immunolabeling in visualizing later stages of FTIs, including ghost tangles. These results provide the basis for potential future applications of CCM binding of tau aggregates in diagnostic pathology and in vivo.

OBJECTIVE

Prenylation is essential for membrane localization and participation of proteins in various signaling pathways. The following study was conducted to examine the importance of protein farnesylation and geranylgeranylation for the regulation of lymphoma cell survival and proliferation.

METHODS

Lymphoma cells were treated with the beta-hydroxy-beta-methylglutaryl-CoA reductase inhibitor lovastatin, which inhibits protein farnesylation and geranylgeranylation by the depletion of intracellular pools of farnesylpyrophosphate and geranylgeranylpyrophosphate. In addition, farnesyl transferase and geranylgeranyl transferase activities were specifically inhibited by FTI-277 and GGTI-298, respectively.

RESULTS

Only inhibition of geranylgeranylation by lovastatin led to reduction of cell viability in lymphoma cell lines and purified tumor cells from lymphoma patients in a time- and dose-dependent way. Reduction in the number of viable cells was mediated by both induction of apoptosis and inhibition of proliferation. In addition, GGTI-298 was more effective in induction of apoptosis and inhibition of proliferation than FTI-277. Apoptosis induced by inhibition of protein geranylgeranylation was associated with a reduction of Mcl-1 protein levels, collapse of the mitochondrial transmembrane potential, and caspase-3 activation. Inhibition of proliferation resulted from the induction of G(1) arrest. Furthermore, lovastatin at low concentrations sensitized lymphoma cells to dexamethasone, including cells resistant to this drug.

CONCLUSIONS

These results indicate that protein geranylgeranylation is critical for the regulation of lymphoma tumor cell survival and proliferation and that pharmacological agents such as lovastatin or geranylgeranyl transferase inhibitors, alone or in combination with other drugs, may be useful in the treatment of lymphoma.

Daily vaginal douching with polyvinylpyrrolidone-iodine in 12 euthyroid volunteers for 14 days resulted in a significant increase in serum total iodine concentration and urine iodine excretion. The increase in serum total iodine was associated with a marked decrease in 24-hour iodine 123 uptake by the thyroid and a small but significant increase in serum thyrotropin (TSH) concentration. However, values for serum TSH never rose above the normal range. No significant changes in serum thyroxine (T4), free T4 index (FTI), or triiodothyronine concentrations were observed, although serum T4 and FTI did decrease slightly during treatment. The findings suggest that iodine is absorbed across the vaginal mucosa and that the subsequent increase in serum total iodine does induce subtle increases in serum TSH concentration. There was no evidence, however, of overt hypothyroidism in these euthyroid women.

Endotoxemia plays an important role in the pathogenesis of sepsis and is accompanied by dysregulated apoptosis of immune and non-immune cells. Treatment with statins reduces mortality in rodent models of sepsis and endotoxemia. Inhibition of protein isoprenylation, including farnesylation, has been proposed as a mechanism to mediate the lipid-lowering-independent effects of statins. Nonetheless, the effects of the inhibition of isoprenylation have not yet been studied. To investigate the role of farnesylation, we evaluated the effects of farnesyltransferase inhibitor and statin on survival following lipopolysaccharide (LPS) challenge in mice. Both simvastatin (2mg/kg BW) and FTI-277 (20mg/kg BW) treatment improved survival by twofold after LPS injection, as compared with vehicle alone (p<0.01). LPS-induced cleavage (activation) of caspase-3, an indicator of apoptotic change, and increased protein expression of proapoptotic molecules, Bax and Bim, and activation of c-Jun NH(2)-terminal kinase (JNK/SAPK) in the liver and spleen were attenuated by both simvastatin and FTI-277. These results demonstrate that farnesyltransferase inhibitor as well as statin significantly reduced LPS-induced mortality in mice. Our findings also suggest that inhibition of protein farnesylation may contribute to the lipid-lowering-independent protective effects of statins in endotoxemia, and that protein farnesylation may play a role in LPS-induced stress response, including JNK/SAPK activation, and apoptotic change. Our data argue that farnesyltransferase may be a potential molecular target for treating patients with endotoxemia.

Cardiac atrial and ventricular parameters were determined by Doppler two-dimensional echocardiography at rest and exercise in 8 patients with subclinical hypothyroidism (SCH) (6 women and 2 men; age range: 28-48 years) before and 3 months after achievement of a euthyroid state with incremental adjustment of L-thyroxine therapy. None of the patients had known heart disease. At 3 months of L-thyroxine therapy, TSH levels decreased from 14.8 +/- 9.4 mIU/L to 3.0 +/- 1.5 mIU/L and FTI increased from 7.1 +/- 1.8 to 8.1 +/- 1.9. The cardiac studies were performed at rest, and during incremental exercise load (50, 100, 150 W workload) on a Quinton exercise bicycle. No significant differences were found between the subclinical hypothyroid and euthyroid states in systolic blood pressure at rest (104.8 +/- 12.3 vs 105 +/- 10.1 mm Hg) and exercise (158 +/- 24.9 vs 158.5 +/- 20.9 mm Hg) or diastolic blood pressure at rest (70 +/- 4.7 vs 69 +/- 5.7 mm Hg) and exercise (86 +/- 11.4 vs 89.2 +/- 7.3 mm Hg). All echocardiographic atrial and ventricular parameters were similar before and during L-thyroxine therapy with the exception of a small but significant change in left ventricular diastolic dimension (4.5 +/- 0.3 vs 4.8 +/- 0.4 cm; p < 0.05). All Doppler parameters were not significantly affected by L-thyroxine therapy with the exception of preejection period at stage III exercise (51 +/- 17 vs 39 +/- 13 msec; p < 0.05). Preejection period at other stages of exercise showed trends toward similar differences between subclinical hypothyroidism and euthyroidism, but the differences were not statistically significant. We conclude that the cardiac structure and function overall remains for practical purposes normal in subclinical hypothyroidism. However, the latter may be responsible for a mild prolongation of the preejection period during exercise and a slightly smaller left ventricular diastolic dimension at rest, changes that may not be of clinical significance in patients without underlying heart disease.

OBJECTIVE

To investigate the effect of endogenous hormone levels on central abdominal fat during the menopausal transition in a population-based cohort of Australian-born women.

METHODS

Prospective observational study.

METHODS

Population-based sample. Body composition was assessed in the Royal Melbourne Hospital, and interviews were conducted at the patient's home.

METHODS

One hundred two women from the Melbourne Women's Midlife Health Project. Data, physical measures, and blood were obtained by interview when the longitudinal study commenced (baseline) and at the time of the total body scan approximately 5 years later. Body composition was measured using dual-energy X-ray absorptiometry.

METHODS

None.

METHODS

Total body fat and central abdominal fat.

RESULTS

The 102 women were either premenopausal or in the early menopausal transition at baseline. At the time of their dual-energy X-ray absorptiometry scan, 31 were in the early menopausal transition, 22 were in the late menopausal transition, and 49 were postmenopausal. Multiple regression analysis found that total percentage of body fat was associated with weight measures, whereas central abdominal fat was also positively associated with baseline free T index (FTI) and with the increase in FTI since baseline.

CONCLUSIONS

The major hormonal change associated with central adiposity during the menopausal transition is the increase in the FTI. This effect is significant even after allowing for baseline and final weight.

Genetic mutations that lead to an accumulation of farnesyl-prelamin A cause progeroid syndromes, including Hutchinson-Gilford progeria syndrome. It seemed possible that the farnesylated form of prelamin A might be toxic to mammalian cells, accounting for all the disease phenotypes that are characteristic of progeria. This concept led to the hypothesis that protein farnesyltransferase inhibitors (FTIs) might ameliorate the disease phenotypes of progeria in mouse models. Thus far, two different mouse models of progeria have been examined. In both models, FTIs improved progeria-like disease phenotypes. Here, prelamin A post-translational processing is discussed and several mutations underlying human progeroid syndromes are described. In addition, recent data showing that FTIs ameliorate disease phenotypes in a pair of mouse models of progeria are discussed.

Recent genetic investigations have established that RhoB gain-of-function is sufficient to mediate the antitransforming effects of farnesyltransferase inhibitors (FTIs) in H-Ras-transformed fibroblast systems. In this study, we addressed the breadth and mechanism of RhoB action in epithelial cells transformed by oncoproteins which are themselves insensitive to FTI inactivation. Rat intestinal epithelial (RIE) cells transformed by activated K-Ras or Rac1 were highly sensitive to FTI-induced actin reorganization and growth inhibition, despite the inability of FTI to block prenylation of either K-Ras or Rac1. Ectopic expression of the geranylgeranylated RhoB isoform elicited in cells by FTI treatment phenocopied these effects. Analysis of RhoB effector domain mutants pointed to a role for PRK, a Rho effector kinase implicated in the physiological function of RhoB in intracellular receptor trafficking, and these findings were supported further by experiments in a fibroblast system. We propose that FTIs recruit the antioncogenic RhoB protein in the guise of RhoB-GG to interfere with signaling by pro-oncogenic Rho proteins, possibly by sequestering common exchange factors or effectors such as PRK that are important for cell transformation.

The ras mutation, which is observed in 20-30% of human nonsmall cell lung cancers (NSCLCs), is one of common genetic alterations and has been proposed to be a prognostic factor in lung cancer. Oncogene ras appears to be essential for tumor progression and maintenance. Several therapeutic agents have been developed to inhibit ras, such as FTIs and antisense oligonucleotides. A new tool for blocking oncogenes in cancer cells has emerged with the discovery that RNA interference can specifically silence expression of endogenous human genes. In the current study, we used small interfering RNA (siRNA) directed against mutant K-ras to determine the anti-tumor effects of decreasing the levels of this protein in lung cancer cell lines. Adenovirus-mediated siRNA (AdH1/siK-ras(V12)) against K-ras(V12) markedly decreased K-ras(V12) gene expression and inhibited cellular proliferation of NSCLC H441 cells that express the relevant mutation (K-ras codon 12 GGT ->> GTT), but produced minimal growth inhibition on NSCLC H1650 cells that lack the relevant mutation. Pretreatment with AdH1/siK-ras(V12) completely abrogated subcutaneous engraftment of H441 cells, as compared with a 100% tumor take in animals that received control vector-treated tumor cells. The in vivo effect of AdH1/siK-ras(V12) treatment was further examined by intratumoral injections after tumor induction. Pre-existing tumor growth was reduced by 45% by a single intratumoral injection. Three or five repeat injections resulted in complete tumor regression in eight of ten nude mice. Further, 23.12% of AdH1/siK-ras(V12) treated H441 cells underwent apoptosis, as compared with 6.13%, and 8.27% in untreated and control vector-treated cells, respectively. These results indicate that adenovirus-mediated siRNA can specifically and efficiently target factors whose expression is altered in malignancy and may have the potential as a therapeutic modality to treat human lung cancer.

To understand the role of BRAF dysfunction in the carcinogenesis and progression/development of colorectal tumors, the authors investigated genetic alterations in the BRAF gene in human colorectal neoplasms as well as the effects of an RAS inhibitor in BRAF-mutant cells. Seven colon cancer cell lines and 116 colorectal tumors (34 adenomas and 82 adenocarcinomas) were analyzed. Genetic alterations in the BRAF and K-ras genes were examined using polymerase chain reaction-single strand conformation polymorphism and direct sequencing analyses. The growth-inhibitory and apoptosis-inducing effects of the FTI-277 RAS inhibitor in colon cancer cell lines were analyzed as well. An immunohistochemical study was also performed to investigate the correlations between the clinicopathologic parameters involved in the Ki-67 labeling index and the number of apoptotic bodies in tumor cells. FTI-277 did not suppress the proliferation of BRAF-mutant cells (WiDr and TCO), but remarkably inhibited the growth of K-ras mutant cells (LoVo). Interestingly, LoVo cells underwent apoptosis by FTI-277 in a dose-dependent manner, whereas WiDr cells were resistant to this agent. In tumor samples, BRAF mutations were found in 1 (3.0%) of 33 adenomas and 6 (7.2%) of 83 adenocarcinomas. No tumor exhibited mutations in both the BRAF and K-ras genes. Neither BRAF nor K-ras mutations correlated with the Ki-67 labeling index immunohistochemically. However, the number of apoptotic bodies was significantly decreased in the BRAF-mutant tumors. Mutation in the BRAF gene may contribute to colorectal carcinogenesis by upregulating the antiapoptotic role of the RAS/RAF/MEK/ERK pathway.

Farnesyltransferase inhibitors (FTIs) target and inhibit the peptide prenylating enzyme farnesyltransferase. This new class of signal transduction inhibitors is being tested clinically in diverse malignancies, with encouraging results in hematololgic malignancies and breast cancer in particuarl. Critical questions have yet to be answered, for example, optimal dose and schedule, disease subgroups most likely to respond, and appropriate combinations with standard cytotoxics and new biologics. Gene profiling studies of malignant target cells obtained during FTI clinical trials will help to identify patients who are likely to respond to FTIs and to develop mechanisms for overcoming FTI resistance. Clinical trials and correlative laboratory studies in progress and under development will define the optimal roles of FTIs in cancer patients.