Genetic basis of differential opsin gene expression in cichlid fishes.

Visual sensitivity can be tuned by differential expression of opsin genes. Among African cichlid fishes, seven cone opsin genes are expressed in different combinations to produce diverse visual sensitivities. To determine the genetic architecture controlling these adaptive differences, we analysed genetic crosses between species expressing different complements of opsin genes. Quantitative genetic analyses suggest that expression is controlled by only a few loci with correlations among some genes. Genetic mapping identifies clear evidence of trans-acting factors in two chromosomal regions that contribute to differences in opsin expression as well as one cis-regulatory region. Therefore, both cis and trans regulation are important. The simple genetic architecture suggested by these results may explain why opsin gene expression is evolutionarily labile, and why similar patterns of expression have evolved repeatedly in different lineages.

10-(Alkylamino)-4H-thieno[3,4-b][1,5]benzodiazepines. A novel class of potential neuroleptic agents.

An investigation of the structural requirements for CNS activity of the title compounds was undertaken. A synthesis of the precursor dihydro-10H-thieno[3,4-b][1,5]benzodiazepin-10-ones was achieved and three routes for their conversion to the title compounds were developed. The compounds were tested for neuroleptic activity by means of the blockade or d-amphetamine lethality in aggregated mice and/or effects on locomotor activity in rats. Antidepressant activity was examined using inhibition of tetrabenazine-induced depression in mice. Most of the compounds were found to be potent neuroleptic agents with several exhibiting additional antidepressant activity.

Thiophene systems. 5. Thieno[3,4-b][1,5]benzoxazepines, thieno[3,4-b][1,5]benzothiazepines, and thieno[3,4-b][1,4]benzodiazepines as potential central nervous system agents.

10-(Alkylamino)thieno[3,4-b][1,5]benzoxazepines (3) and 10-(alkylamino)thieno[3,4-b][1,5]benzothiazepines (4) were prepared by derivatization of the respective lactams (7 and 8) via phosphorus pentachloride and subsequent condensation with the appropriate alkylamines. 9-(Alkylamino)-4H-thieno[3,4-b][1,4]benzodiazepines (5) were prepared by titanium tetrachloride catalyzed condensation of the lactam 11 with alkylamines. 9-(Alkylamino)-4-methylthieno[3,4-b][1,4]benzodiazepines (6) were prepared by reductive alkylation of 5. The compounds were tested for potential neuroleptic activity by means of the blockade of d-amphetamine lethality in aggregated mice and/or effects on locomotor activity in rats. Antidepressant activity was examined using inhibition of tetrabenazine-induced depression in mice. Most of the title compounds 3-6 were found to have neuroleptic activity. In addition, introduction of a 3-chloro substituent in the oxygen and sulfur systems (3p and 4c), as well as introduction of an N-alkyl in the dinitrogen system (6), was found to produce antidepressant effects. Structure-activity relationships are discussed.

1-Aryl-3-azabicyclo[3.1.0]hexanes, a new series of nonnarcotic analgesic agents.

A series of 1-aryl-3-azabicyclo[3.1.0]hexanes was synthesized by hydride reduction of 1-arylcyclopropanedicarboximides. Hydroxyphenyl analogues 20, 22, and 24 were prepared by EtSNa--DMF ether cleavage of the corresponding methoxyphenyl analogues 2m, 2n, and 23, respectively, with the secondary amines 20 and 22 going through the N-formyl intermediate 19 and 21. The p-ethoxy analogue 26 was obtained by O-ethylation of 19, followed by base hydrolysis of the amide 25. The greatest analgesic potency in mouse writhing and rat paw-pain assays was observed for para-substituted compounds. Bicifadine, 1-(4-methylphenyl)-3-azabicyclo[3.1.0]hexane (2b), was the most potent member of the series and is presently undergoing clinical trials in man. Analgesic activity of 2b is limited to the (+) enantiomer 2v, which has the 1R,5S absolute configuration as determined by single-crystal X-ray analysis. The N-methyl analogue (27d) of 2b showed significant analgesic potency, whereas the N-allyl (27a), N-(cyclopropylmethyl) (27b), and N-(n-hexyl) (27c) analogues were inactive. Bicifadine (2b) showed a nonnarcotic profile different from analogous azabicycloalkane and 3-phenylpyrrolidine analgesics.

Use of granulocyte macrophage colony stimulating factor alone or in combination with interferon-alpha-2b for treatment of chronic hepatitis C.

BACKGROUND/AIMS: We have evaluated the effect of granulocyte macrophage colony-stimulating factor (GM-CSF) when utilized either alone or in combination with interferon for treatment of chronic hepatitis C virus (HCV). METHODS: A total of 71 patients with chronic HCV, elevated alanine aminotransferase and normal hepatic function were enrolled into these studies. Nineteen patients who had previously failed to achieve both biochemical and virologic response during interferon therapy were treated with increasing doses of GM-CSF alone (65-250 microg/m2 three times weekly) for 6 months. Another 52 patients who had not been previously treated with interferon entered a randomized controlled trial; 25 were treated with interferon alone (3 mU three times weekly) and 27 with a combination of interferon+GM-CSF (3 mU+250 microg/m2 three times weekly). All patients were treated for 6 months. Both groups were well matched for age, sex, race, serum alanine aminotransferase, HCV-RNA titer, liver histology score and cirrhosis. RESULTS: None of the patients treated with GM-CSF alone developed either biochemical or virologic response at any of the treatment dosages and mean HCV-RNA titer remained unchanged from baseline during this therapy. For patients in the randomized controlled trial, biochemical and virologic responses were observed in 54% of interferon-treated patients compared to 31.8% for those treated with interferon+GM-CSF. Long-term sustained virologic response was observed in only one patient in each group. No significant differences were observed in HCV-RNA titer during the course of treatment. GM-CSF treatment was associated with a significant increase in total white blood cell count and absolute eosinophil count, which peaked within the first month of therapy and declined spontaneously during the remaining 5 months. CONCLUSION: GM-CSF either alone or in combination with interferon does not appear to be effective for treatment of chronic HCV.

A randomized, controlled trial to determine whether continued ribavirin monotherapy in hepatitis C virus-infected patients who responded to interferon-ribavirin combination therapy will enhance sustained virologic response.

This study assessed the use of ribavirin monotherapy to enhance sustained virologic response in hepatitis C virus (HCV)-infected patients who achieved virologic response to interferon (IFN)-ribavirin combination therapy. Patients who had chronic HCV infection and prior relapse were retreated with IFN-ribavirin for 6 months. Patients with an end-of-treatment virologic response were assigned randomly to either stop use of both IFN and ribavirin or to continue use of ribavirin as monotherapy for an additional 6 months. HCV RNA became undetectable during treatment in 46 patients, who then entered the randomized trial. Sustained virologic response was observed in 13 of 26 patients who continued ribavirin monotherapy and in 15 of 20 patients who stopped use of both IFN and ribavirin (P, not significant). Sustained virologic response was significantly more common in patients with HCV genotype non-1 (75% vs. 56%) and in patients with a virus titer < 2 x 10(6) copies/mL (93% vs. 43%). The results indicate that continuing ribavirin monotherapy after achieving a virologic response does not improve sustained virologic response.

Improved sustained response following treatment of chronic hepatitis C by gradual reduction in the interferon dose.

Interferon (IFN) treatment of chronic hepatitis C virus (HCV) is associated with a high rate of relapse. IFN is thought to exert its effect against HCV via direct viral inhibition and immune stimulation. We have hypothesized that relapse following termination of therapy results from the sudden withdrawal of this immune modulatory effect and that gradual reduction in the IFN dose may decrease the incidence of relapse. One hundred six patients with chronic HCV were enrolled into this 24-month controlled, randomized prospective trial. All were treated with 5 mU of interferon-alpha-2b three times a week for 6 months. Patients who achieved biochemical response were randomized to either stop or taper IFN gradually at monthly intervals as follows; 3 mu, 2 mU, 1 mU, and 0.5 mU (all three times a week). 0.5 mU twice weekly and then once weekly. Liver histology was assessed by Knodell index and HCV RNA was measured by a quantitative polymerase chain reaction (PCR) assay. Of the 92 patients who completed the initial 6 months of IFN treatment, 47 (51%) achieved biochemical response. Twenty-one of these patients were randomized to stop IFN treatment and 25 to taper (1 drop-out). At randomization patients were well matched with respect to age, sex, race, serum alanine transaminase (ALT), and liver histology. Biochemical relapse was observed in 19 of 21 (91%) patients who stopped IFN treatment compared with only 60% who tapered IFN (P= .04). Virological relapse occurred in 90% of patients who stopped and only 48% of persons who tapered IFN therapy. At completion of the 24-month study patients who achieved long-term sustained biochemical response had a significantly lower mean Knodell score (3.5 vs. 6.5) and a significantly greater number were HCV RNA negative in serum (85% vs. 18%) compared with relapsers. We conclude that gradual reduction in IFN dose is associated with a significant higher rate of sustained response and clearance of HCV RNA from serum compared with abruptly stopping treatment. This in turn is associated with a significant improvement in hepatic histology supporting the premise that response to IFN therapy can prevent progression to cirrhosis.

Chronic infection with hepatitis C virus in patients with elevated or persistently normal serum alanine aminotransferase levels: comparison of hepatic histology and response to interferon therapy.

Ninety-five patients with chronic hepatitis C virus (HCV) infection, 35 with persistently normal serum alanine aminotransferase (ALT) levels, were randomized to treatment with daily interferon (IFN) for 3 months, followed by IFN 3 times weekly (TIW) for 12 months (group A) or TIW for 18 months (group B). Patients with elevated versus normal ALT levels had similar demographic and virologic characteristics but significantly (P<.05) more advanced liver histology (bridging fibrosis and cirrhosis, 37.9% vs. 11.4%). After 3 months of treatment, 38.3% of patients in group A were HCV RNA negative versus 18.8% in group B (P<.05). When the IFN dose was reduced from daily to TIW in group A, the percentage of patients who remained HCV RNA negative declined; sustained virologic response was similar in both groups (10.6% vs. 8.3%). Response to treatment was similar in patients with elevated or normal ALT levels. Persons with chronic HCV infection and persistently normal serum ALT levels have milder liver disease than, and respond to IFN therapy similarly to, persons with elevated ALT levels.

Treatment of chronic hepatitis C in patients who failed interferon monotherapy: effects of higher doses of interferon and ribavirin combination therapy. The Virginia Cooperative Hepatitis Treatment Group.

OBJECTIVE: The present study was designed to evaluate the effectiveness of interferon-ribavirin combination therapy for treatment of chronic hepatitis C virus (HCV) in patients who failed previous treatment with interferon monotherapy. METHODS: A total of 140 patients with well-documented chronic HCV who failed to achieve a virological (if HCV-RNA was assessed) or biochemical response (if HCV-RNA was not assessed) to interferon monotherapy, 3 mU three times weekly (TIW) for 3-18 months, were randomly assigned to one of three treatment groups. Group A patients were treated with 5 mU interferon TIW for 6 months. Ribavirin (1000-1200 mg daily) was added in those patients HCV-RNA positive at month 3. Group B patients were treated with 3 mU interferon TIW plus ribavirin (1000-1200 mg daily) for 6 months. The dose of interferon was increased to 5 mU TIW in those patients HCV-RNA positive at month 3. Group C patients were treated with 5 mU interferon TIW plus ribavirin (1000-1200 mg daily) for 6 months. Serum ALT and HCV-RNA were monitored during and after treatment for a total of 15 months. RESULTS: Seventeen percent of patients in group A became HCV-RNA negative by treatment month 3. Adding ribavirin resulted in one additional patient becoming HCV-RNA negative. However, none of the patients in this group achieved sustained virological response. Twenty-six percent of patients in group B became HCV-RNA negative by treatment month 3. Increasing the dose of interferon from 3 to 5 mU TIW increased virological response to 30%. However, sustained virological response was observed in only 14%. Thirty percent of patients in group C became HCV-RNA negative, but sustained virological response was observed in only 12%. Sustained virological response was found to be significantly greater in patients with a nontype I HCV genotype (p < 0.002) and in patients who had a decline in HCV-RNA titer to a value < 100,000 copies/ml during their previous course of interferon monotherapy (p < 0.0001). None of the 12 sustained responders were African Americans (p < 0.013). CONCLUSIONS: Retreatment of nonresponders with interferon-ribavirin combination therapy results in limited benefit; only 13% of patients achieved sustained virological response. Response was extremely poor in African Americans and those with HCV genotype 1.

Relationship between biochemical, virological, and histological response during interferon treatment of chronic hepatitis C.

The present study was conducted to evaluate the relationship between biochemical, virological, and histological response during the course of interferon therapy. Ninety consecutive patients with well-documented chronic hepatitis C virus (HCV) were treated with 5 MU of interferon alfa-2b three times weekly for 6 months. Liver biopsy was performed, and serum HCV RNA titer was measured before and at the completion of interferon treatment. Normalization of serum alanine transaminase (ALT) concentration (biochemical response) was observed in 50% of patients. In these patients, Knodell score declined significantly from 9.6 +/- 0.5 to 5.0 +/- 0.5 (P < .01), and 75% became HCV RNA negative. The remaining patients (50%) were biochemical nonresponders; mean Knodell score declined from 9.6 +/- 0.5 to 7.7 +/- 0.5 (P < .01), and 11% became HCV RNA negative. For both biochemical responders and nonresponders, the decline in Knodell score was confined to the components of hepatic inflammation (piecemeal necrosis + lobular + portal inflammation); no change in fibrosis was observed. Hepatic inflammation declined by 5 points or more in 69% of biochemical responders and 48% of biochemical nonresponders, and by at least 50% from pretreatment values in 74% and 38% of biochemical responders and biochemical nonresponders, respectively. For all patients (both biochemical responders and nonresponders) who remained viremic at the conclusion of interferon therapy, the reduction in hepatic inflammation was a linear function of the decline in HCV RNA titer. We conclude that more than one third of patients who had no biochemical response after 6 months of interferon therapy achieved a similar improvement in hepatic histology as was observed in patients with biochemical response. This improvement in hepatic histology appeared to correlate with a reduction in HCV RNA titer, especially in patients who remained viremic.

A randomized, controlled trial of maintenance interferon therapy for patients with chronic hepatitis C virus and persistent viremia.

BACKGROUND & AIMS: : At least half of patients with chronic hepatitis C virus (HCV) fail to respond to interferon or interferon/ribavirin therapy. Histological improvement is observed in some nonresponders. We conducted a randomized, controlled trial to determine if maintenance interferon therapy could prevent histological progression in this subset of nonresponders. METHODS: Fifty-three patients with chronic HCV were enrolled. All were HCV-RNA positive after 6 months of treatment with interferon alfa-2b but had a histological response. Twenty-seven of the patients were randomly assigned to continue interferon (3 MU 3 times weekly) for 24 months; 26 patients discontinued treatment and were observed prospectively. Alanine aminotransferase (ALT) level and HCV-RNA titer were monitored, and liver biopsy was repeated every 12 months. RESULTS: Before interferon therapy, the 2 groups were well matched for all demographic factors, serum ALT (94.0 +/- 15.6), log HCV-RNA titer (5. 85 +/- 0.15 copies/mL), histology score (9.5 +/- 0.2), and percentage with cirrhosis (25%). After 6 months of treatment, significant reductions (P < 0.05) in serum ALT level (62.6 +/- 9.6), log HCV-RNA titer (4.79 +/- 0.13 copies/mL), and hepatic inflammation (4.0 +/- 0.2) were observed. These improvements were maintained in the patients randomized to continue interferon. Stopping treatment was associated with an increase in serum ALT, log HCV-RNA, and hepatic inflammation back to baseline. After 30 months of treatment, mean fibrosis score declined from 2.5 to 1.7 and 80% of patients had histological improvement (P < 0.03). Discontinuation of interferon was associated with an increase in mean fibrosis score from 2.2 to 2.4 and worsening of hepatic histology in 30% of patients (P < 0.01). CONCLUSIONS: These data support the hypothesis that maintenance interferon may prevent histological progression of chronic HCV in patients who remain viremic.