Hepatitis B virus genotype E detected in Brazil in an African patient who is a frequent traveler

Genotype E of hepatitis B virus (HBV) has not been described in Brazil and is found mainly in Africa. Genotype A is the most prevalent in Brazil, and genotypes B, C, D, and F have already been reported. We report here an HBV genotype E-infected patient and some characterization of surface (S) protein, DNA polymerase (P) and precore/core (preC/C) coding regions based on the viral genome. The patient is a 31-year-old black man with chronic hepatitis B who was born and raised in Angola. He has been followed by a hepatologist in São Paulo, Brazil, since November 2003, and he is a frequent traveler to Latin America, Africa, and Europe. In 2003, he was diagnosed with HBV infection and started treatment with lamivudine with the later addition of adefovir dipivoxil. No known risk factor was identified. Serologically, he is HBsAg and anti-HBe positive, but HBeAg and anti-HBs negative. DNA sequence analysis of the S/P region confirmed that this patient is infected with genotype E, subtype ayw4. The preC/C region showed G1896A and G1899A mutations but no mutations in the basal core promoter. Nucleotide substitutions common in genotype E were also observed (C1772, T1858 and A1757). Although this is not an autochthonous case and there is no evidence of further spread, the description of this case in Brazil highlights the current risk of viral genotypes spreading with unprecedented speed due to constant travel around the world.

Hepatitis B virus genotype E detected in Brazil in an African patient who is a frequent traveler.

Genotype E of hepatitis B virus (HBV) has not been described in Brazil and is found mainly in Africa. Genotype A is the most prevalent in Brazil, and genotypes B, C, D, and F have already been reported. We report here an HBV genotype E-infected patient and some characterization of surface (S) protein, DNA polymerase (P) and precore/core (preC/C) coding regions based on the viral genome. The patient is a 31-year-old black man with chronic hepatitis B who was born and raised in Angola. He has been followed by a hepatologist in São Paulo, Brazil, since November 2003, and he is a frequent traveler to Latin America, Africa, and Europe. In 2003, he was diagnosed with HBV infection and started treatment with lamivudine with the later addition of adefovir dipivoxil. No known risk factor was identified. Serologically, he is HBsAg and anti-HBe positive, but HBeAg and anti-HBs negative. DNA sequence analysis of the S/P region confirmed that this patient is infected with genotype E, subtype ayw4. The preC/C region showed G1896A and G1899A mutations but no mutations in the basal core promoter. Nucleotide substitutions common in genotype E were also observed (C1772, T1858 and A1757). Although this is not an autochthonous case and there is no evidence of further spread, the description of this case in Brazil highlights the current risk of viral genotypes spreading with unprecedented speed due to constant travel around the world.

Effect of all-trans-retinoic acid on the hypercoagulable state of patients with breast cancer.

To evaluate whether all-trans-retinoic acid (ATRA) is able to modulate the hemostatic system in patients with solid tumors, we studied patients with locally advanced breast cancer who were enrolled in a Phase Ib study of ATRA +/- Tamoxifen (Tam). In this study, two groups of 15 patients/each were treated for 21 days before operation with ATRA at three doses (15, 45, or 75 mg/m(2)/day on alternate days) given alone (group 1) or in combination with Tam (group 2). One additional group received Tam alone. Plasma samples were evaluated for hypercoagulation markers (FVIIa, F1+2, TAT, D-dimer), fibrinolysis proteins (t-PA, PAI-1), and coagulation inhibitors (protein C, AT). At baseline, cancer patients had FVIIa, F1+2, TAT, and PAI-1 significantly greater than control subjects. During treatment, in the patients given ATRA alone, hypercoagulation markers appeared unmodified. Instead, subjects given Tam alone had a significant elevation of FVIIa, F1+2, and TAT versus baseline. However, in the ATRA + Tam groups, hypercoagulation markers were decreased compared with Tam alone. These results suggest that in selected conditions, pre-operative ATRA may modulate the hypercoagulable state of breast cancer patients.

Clinical activity and safety of combination immunotherapy with IFN-alpha 2a and Rituximab in patients with relapsed low grade non-Hodgkin's lymphoma.

BACKGROUND AND OBJECTIVES: To determine the clinical activity and safety of the combination immunotherapy of the chimeric anti-CD20 antibody, Rituximab, and Interferon (IFN)- alpha 2a DESIGN AND METHODS: Sixty-four patients with relapsed low-grade or follicular B-cell non Hodgkin's lymphoma received 4 infusion of Rituximab (375 mg/m(2) x dose) after priming and simultaneous treatment with IFN- alpha 2a. RESULTS: The overall response rate was 70% with 33% complete responses. Median for duration of response is 19 months, after a median follow-up of 22 months. By univariate analysis none of the most common prognostic factors predicted for response to therapy. After treatment 10 patients become bcl-2 negative in the bone marrow, but no correlation between molecular and clinical response was found. Fifty-three patients (83%) had drug related or unknown origin adverse events. The number of adverse events per patient varied from 1 to 21. Considering all 272 events, 231 (85%) were grade 1 or 2, 36 (13%) grade 3 and 5 (2%) grade 4. Twenty-three patients required reduction in the dose and/or short discontinuation of IFN treatment, either during priming or subsequent treatment. The most frequent adverse events were leukopenia, fever, neutropenia, hypotension and thrombocytopenia. INTERPRETATION AND CONCLUSIONS: this report shows that combination immunotherapy Rituximab + IFN- alpha 2a is active and relatively well tolerated. The overall response rate of 70% and the median duration remission of 19 months compare favorable with the results obtained with Rituximab alone in similar subset of patients. Randomized trials, investigating Rituximab versus combination immunotherapy are needed.

Treatment of B-cell non-Hodgkin's lymphoma with anti CD 20 monoclonal antibody Rituximab.

Rituximab is a chimeric anti CD-20 monoclonal antibody containing human IgG1 kappa constant regions, with murine variable regions. The anti-lymphoma effects of Rituximab are probably due to complement and antibody-dependent cell-mediated cytotoxicity, and induction of apoptosis. Phase II trials have demonstrated a strong activity of rituximab alone in indolent B non-Hodgkin lymphoma, especially in patients with follicular lymphoma. The most utilized dose-schedule is 375 mg/m(2) weekly x 4. The association with chemotherapy or with interferon-alpha increases Rituximab efficacy. More recently, Rituximab have showed activity also in diffuse large cell lymphoma, mantle cell lymphoma and in other B-malignancies. Good results have also been obtained utilizing Rituximab for in vivo purging. However, we are still far from having found a definite position for Rituximab in the treatment of lymphoproliferative disorders. The aim of future studies should be to develop new strategies that will hopefully produce the most effective Rituximab-based regimens in order to find the Rituximab key position in the treatment of B-malignancies

Biological activity of all-trans-retinoic acid with and without tamoxifen and alpha-interferon 2a in breast cancer patients.

In addition to suppressing breast cancer cell growth, retinoids potentiate growth inhibition in human breast cancer when tested in vitro and in vivo with tamoxifen and/or interferon. The purpose of this study was to ascertain the biologic effects of all-trans-retinoic acid (ATRA) administered alone and with tamoxifen +/- interferon and to identify the relationship between ATRA plasma concentrations and optimal biological dose (the lowest dose that produces a biological response). Three consecutive groups of 15 patients with locally advanced operable breast cancer were treated, in accordance with good clinical practice (GCP) requirements, with ATRA at 3 dose levels alone or with tamoxifen +/- alpha-interferon 2a at flat doses. After 3 weeks, the tumors were surgically removed. Biological parameters measured at the beginning (in biopsy tissue) and end (in surgical tissue) of the study were compared. The optimal biological dose for ATRA was 15 mg/m2/day. Treatments influenced tumor grade but not cell cycle kinetics (G0-G1 phase) or proliferation (Ki67 levels). ATRA induced progesterone receptors independent of dose level and co-administered drugs, but did not induce estrogen receptors when administered alone. Retinoic acid receptor (RAR)-alpha was not affected by treatment and RAR-alpha was moderately influenced whereas RAR-beta (concomitantly with transforming growth factor-beta) was induced in 33% of patients by ATRA alone. ATRA pharmacokinetics were dose- and time-dependent. Neither the ATRA + tamoxifen nor the ATRA + tamoxifen + interferon combinations potentiated the ATRA-induced biological changes. Future studies evaluating the role of RAR-beta as a biological marker of retinoid activity are warranted.

Successful in vivo purging of CD34-containing peripheral blood harvests in mantle cell and indolent lymphoma: evidence for a role of both chemotherapy and rituximab infusion.

Elimination of tumor cells ("purging") from hematopoietic stem cell products is a major goal of bone marrow-supported high-dose cancer chemotherapy. We developed an in vivo purging method capable of providing tumor-free stem cell products from most patients with mantle cell or follicular lymphoma and bone marrow involvement. In a prospective study, 15 patients with CD20(+) mantle cell or follicular lymphoma, bone marrow involvement, and polymerase chain reaction (PCR)-detectable molecular rearrangement received 2 cycles of intensive chemotherapy, each of which was followed by infusion of a growth factor and 2 doses of the anti-CD20 monoclonal antibody rituximab. The role of rituximab was established by comparison with 10 control patients prospectively treated with an identical chemotherapy regimen but no rituximab. The CD34(+) cells harvested from the patients who received both chemotherapy and rituximab were PCR-negative in 93% of cases (versus 40% of controls; P =.007). Aside from providing PCR-negative harvests, the chemoimmunotherapy treatment produced complete clinical and molecular remission in all 14 evaluable patients, including all 6 with mantle cell lymphoma (versus 70% of controls). In vivo purging of hematopoietic progenitor cells can be successfully accomplished in most patients with CD20(+) lymphoma, including mantle cell lymphoma. The results depended on the activity of both chemotherapy and rituximab infusion and provide the proof of principle that in vivo purging is feasible and possibly superior to currently available ex vivo techniques. The high short-term complete-response rate observed suggests the presence of a more-than-additive antilymphoma effect of the chemoimmunotherapy combination used.

Long term response to therapy of chronic anti-HBe-positive hepatitis B is poor independent of type and schedule of interferon.

OBJECTIVE: The response rate to alpha interferon (IFN) of chronic anti-HBe-positive hepatitis B is variable. We studied whether type, dose, and schedule of IFN, and type and frequency of posttreatment monitoring, influence the response rate. METHODS: Seventy-two consecutive anti-HBe-positive chronic hepatitis B patients (59 male and 13 female, median age 41 yr) stratified by sex and histology were randomly allocated to three treatment arms. Twenty-seven patients (A) received 10 million units alpha-N1 IFN i.m. t.w. for 24 wk (total dose: 720 million units); 21 (B) received 9 million units alpha-2a IFN i.m. t.w. for 4 wk, followed by 18 million units for 12 wk and 9 million units for 8 wk (972 million units); 24 (C) received 2 alpha-2a IFN courses (9 million units i.m. t.w. for 16 and 12 wk separated by a 6-month interval [756 million units]). Primary response was defined by normal ALT and serum HBV-DNA levels below 10 pg/ml at the end of therapy and sustained response by normal ALT (tested monthly), undetectable HBV-DNA and IgM anti-HBc (<7 I.U. Paul Ehrlich Institute) (tested every 3 months) during the posttreatment follow-up. RESULTS: At the end of treatment, 12, 8, and 13 patients from groups A, B, and C, respectively, were responders. At the 18-month follow-up, two patients in group A and only one in groups B and C maintained the response. Overall, after 34 months (median posttreatment follow-up), three patients were long term responders, whereas three showed a sustained remission after relapse. CONCLUSIONS: The rate of long term response to interferon of anti-HBe-positive chronic hepatitis B is poor, independent of IFN type, dose, or schedule; the more stringent the monitoring, the higher the relapse rate.

RARalpha antagonist Ro 41-5253 inhibits proliferation and induces apoptosis in breast-cancer cell lines.

Ro 41-5253 is a RARalpha-selective antagonist that binds RARalpha but does not induce transcriptional activation and does not influence RAR/RXR heterodimerization and DNA binding. This retinoid inhibits proliferation and induces apoptosis in MCF-7 and ZR-75.1 estrogen-receptor-positive breast-carcinoma cells in a dose-dependent way. The anti-proliferative effect is more evident in ZR-75.1 cells than in MCF-7 cells and is probably mediated by anti-AP1 activity, a mechanism known to be implied in the action of several retinoids. In the induction of apoptosis also ZR-75.1 cells are more sensitive to treatment with Ro 41-5253 than MCF-7 cells. In ZR-75.1 cells an apoptotic/hypodiploid DNA peak is already evident after 2 days of incubation, whereas in MCF-7 cells it appears only after 4 days. The highest percentage of apoptotic cells, for both cell lines, is reached after 6 days of treatment. The apoptosis pathway is p53-independent and bcl-2 downregulation seems to be correlated with an increase in TGF-beta1 protein. The MDA-MB-231 estrogen-receptor-negative cell line is poorly responsive to Ro 41-5253 treatment, both in terms of proliferation inhibition and apoptosis induction. Ro 41-5253 has proliferation-inhibiting and apoptosis-inducing properties that are not mediated by transcriptional activation from retinoic-acid response elements. This retinoid antagonist seems to be a compound that exerts an anti-tumor activity but does not induce the toxic side effects of retinoids and might, therefore, be considered as a candidate for cancer therapy.

5-Fluorouracil plus interferon alpha-2a compared to 5-fluorouracil alone in the treatment of advanced colon carcinoma: a multicentric randomized study.

Biochemical modulation is one of the most interesting fields in cancer chemotherapy. Interferon-alpha (IFNalpha) is a cytokine that is able to influence the pharmacodynamics of 5-fluorouracil (5FU) through a number of mechanisms. With the aim of confirming some data emerging from the literature, we initiated a multicentric randomized study comparing the combination of 5FU and IFNalpha-2a with 5FU alone in the treatment of advanced or metastatic colon cancer. A group of 205 colon cancer patients (104 in the 5FU arm and 101 in the 5FU + IFNapha-2a arm) were included in the final intention-to-treat analysis. Rectal cancers were not considered eligible. All patients had measurable disease, were aged 75 years or less, had a Karnofsky index of at least 60 and had good bone marrow, renal, liver and cardiac functions. No previous chemo-immunotherapy was allowed. The treatment was 750 mg/m2 5FU (4 h i.v. infusion) on days 1 5 and then i.v. bolus weekly, starting from day 12, with or without IFNalpha-2a given s.c. three times weekly (starting dose 3 x 10(6) IU rising to 9 x 10(6) IU, if tolerated). Patients were treated until progression or, if responsive, for a maximum of 48 weeks and then observed for a period of 2 years. The primary end-point of the study was objective clinical response (OR); secondary parameters were time to progression, overall survival, and time to death after progression. WHO criteria were used for both clinical response and toxicity measurements. Dose reduction was planned a priori in the event of significant toxicity due to 5FU, IFNalpha-2a or both. Association between primary and secondary end-points and treatment was studied by univariate and multivariate analysis. Altogether, 47 patients achieved a documented response. A 25% OR was observed in the combination arm while a 21% OR was seen in the 5FU arm; this difference is not statistically significant (P = 0.6). Patients with a small tumour burden (below 5 cm2) showed a higher probability of response in both arms. Patients in the experimental arm had a higher but not statistically significant cumulative progression-free probability. Median survival was 47.1 weeks overall, while it was 43.7 and 48.5 weeks in the control and experimental arms, respectively. The combination was clearly more toxic than 5FU alone, leukopenia being the most frequent side-effect in the experimental arm and nausea and vomiting in the control arm. In conclusion these results are quite disappointing and 5FU + IFNalpha-2a can not be considered a standard treatment for advanced colon cancer.

All-trans retinoic acid (ATRA) in patients with chronic myeloid leukemia in the chronic phase.

Since in vitro observations indicated that all-trans retinoic acid (ATRA), especially in combination with IFNalpha, can exert significant suppressive effects on Ph+ cells, we investigated the effects and the pharmacokinetic profile of ATRA in a selected cohort of patients with Ph+ chronic myeloid leukemia (CML) in chronic phase. Eighteen patients were treated with ATRA at a dose of 80 mg/m2/day (p.o.), divided into two equal doses after meals, for 7 consecutive days every other week for a maximum of 12 courses (1 course = 1 week on and 1 week off). Pharmacokinetic profiles of ATRA were evaluated during intermittent therapy on days 1 and 7 of course 1; on day 1 of course 2; on day 1 of course 6. Out of the 18 patients treated with ATRA, 11 (61%) went off study before the sixth course of treatment because of progressive hyperleukocytosis (seven cases), or thrombocytosis (one case), or refusal (three cases). Seven (39%) patients completed the first six courses (12 weeks) of treatment with ATRA and two of them (11%) maintained a white blood cell (WBC) <10 x 10[9]/l which was induced by the pretreatment with hydroxyurea. One patient completed the 12th course of ATRA maintaining WBC <10 x 10(9)/l, platelets <500 x 10(9)/l and spleen not palpable. The treatment with ATRA was well tolerated and only one patient discontinued the therapy because of non-hematological side-effects. The area under the concentration-time curve (AUC) decreased significantly (P< 0.001) during the first week of therapy. By adopting an intermittent dosing regimen, 1 week on/ 1 week off (1 course), at the start of courses 2 and 6, we obtained the ATRA AUCs equivalent to the ones achieved on day 1 of course 1. In conclusion, our results showed that ATRA alone appeared to be unable to control the WBC expansion in the CML patients in chronic phase. Moreover, it did not induce any remarkable cytoreductive effects on the platelet count and on the hemoglobin level. The major interest of ATRA would be in combination with other therapies. If ATRA was given in combination with IFNalpha or other agents, dose reduction of these would not be planned. On the basis of the pharmacokinetic profile, ATRA should be administered intermittently rather than continuously.

Time-Dependent Kinetics of Tretinoin in Chronic Myelogenous Leukaemia during Intermittent Dose Scheduling: 1 Week On/1 Week Off.

OBJECTIVE: This study investigated the pharmacokinetics of tretinoin during alternating cycles of 1 week of tretinoin treatment and 1 week drug-free in patients with Ph1+ chronic myelogenous leukaemia (CML) in the chronic phase. PATIENTS: Eighteen patients with CML were treated with tretinoin 80 mg/m(2)/day (in two divided doses) for 7 consecutive days every other week (one cycle = 1 week on/1 week off). RESULTS: Body systemic exposure to tretinoin as determined by the area under the plasma concentration-time curve (AUC) decreased significantly during the first week of drug administration, from (mean +/- SD) 678.3 +/- 498.1 to 258.7 +/- 272.4 microg/L.h. In about 40% of the patients the decline in plasma concentrations was >/=80%, while 17% of the population did not experience any decline. On day 7 of cycle 1, the mean apparent oral clearance (CL/F) was 2.6 times the corresponding value on day 1. After 1 week without tretinoin, the mean AUC on day 1 of cycle 2 was lower (down 15%) but not statistically different from the corresponding value observed on day 1 of cycle 1; 62% of patients showed an increase in the AUC, which was 40% higher than the corresponding value on day 7 of cycle 1. On day 1 of cycle 6, the AUC and CL/F of tretinoin during a dosage interval were not statistically different from those observed on day 1 of cycle 1 and cycle 2. On all occasions the peak plasma concentration (C(max)) was strongly correlated to the corresponding AUC. No significant change in the time to observed C(max) (t(max)) and in the elimination half-life (t((1/2))) was observed during the whole study. These results confirmed that the metabolism of tretinoin is rapidly up-regulated in CML patients, with significant declines in plasma drug exposure during the first week of drug administration. After tretinoin was discontinued, a return to the noninduced state followed a mean time-cycle similar to the induction. The strong decrease in the apparent oral drug clearance and the absence of significant variations in the drug half-life demonstrated that the presystemic extraction of tretinoin is the main cause of the marked decline in plasma drug exposure. CONCLUSION: The favourable pharmacokinetic profile of tretinoin obtained by an intermittent regimen, 1 week on/1 week off therapy (vs continuous administration), suggests that such a therapeutic schedule is the most appropriate for the assessment of clinical efficacy in those pathologies in which its use is suitable.

Synergistic effect between doxorubicin and a low dose of all-trans-retinoic acid in MCF-7 breast cancer cell line.

The effect of doxorubicin (DOX) used in combination with a low dose (10(-7) M) of all-trans-retinoic acid (tRA) was tested on MCF-7 breast carcinoma cell line. Both drugs are able to inhibit cell proliferation in these cells in a dose-dependent way. The combined treatment with DOX and tRA was more effective in inhibiting cell growth than each of the two compounds alone. This was evidenced in the following experimental conditions: pre-treatments with tRA, for 72 h or 18 h, before DOX incubation; post-treatment with tRA for 18 h after DOX incubation. A consistent synergism was reached by 72 h pre-treatment with tRA and also with brief pre- and post-treatments, but only if tRA was also present during DOX incubation (co-treatments). The mechanisms involved in this interaction between chemotherapeutics and differentiating agents are as yet unclear and should be evaluated further.

Effects of all-trans-retinoic acid and 13-cis-retinoic acid on breast-cancer cell lines: growth inhibition and apoptosis induction.

Interest has been increasingly focused on all-trans-retinoic acid (tRA) and 13-cis-retinoic acid (13cRA) in cancer chemoprevention and treatment. We have examined the in vitro effects of these 2 retinoic acids (RAs) on human breast-cancer cell lines MCF-7 and ZR-75.1 (both estrogen-receptor-positive, ER+) and MDA-MB-231 (estrogen-receptor-negative, ER-), in terms of inhibition of proliferation and induction of apoptosis. Both retinoic acids exerted an evident dose-dependent growth inhibition, although in the ER- cell line the anti-proliferative effect was obtained only with the highest concentration used; the anti-proliferative activity of tRA was more evident than 13cRA on all 3 tested cell lines. tRA and 13cRA induced apoptosis in MCF-7 and MDA-MB-231 cell lines, but not in ZR-75.1. The apoptotic phenomenon was clearly time-dependent, and in our experience it was not related to the arrest in a specific phase of cell cycle. After treatment with RAs the levels of bcl-2 were reduced in MCF-7, while in ZR-75.1 and in MDA-MB-231 no treatment-related modifications were observed. An analysis of estrogen-receptor status, used as a marker of differentiation, demonstrated that after treatment with RAs the levels of estrogen receptor (ER) decreased in ZR-75.1 only. Our study indicates that the anti-proliferative effects of RAs are sustained by induction of apoptosis in MCF-7 and MDA-MB-231 cells, while in ZR-75.1 cells an induction of differentiation without apoptosis was the prevalent mechanism of growth inhibition. Our results encourage further studies on in vivo effects of these retinoids in breast cancer.

All-trans retinoic acid in hematological malignancies, an update. GER (Gruppo Ematologico Retinoidi).

BACKGROUND AND OBJECTIVE: During the past ten years, the study of retinoids has undergone a total transformation. The Italian Society of Experimental Hematology decided to discuss these advances at a meeting in Florence on April 18, 1996. INFORMATION SOURCES: The material examined in the present review includes articles and abstracts published in journals covered by the Science Citation Index and Medline. In addition, all the authors of the present article have been actively working in the field of retinoids and have contributed several papers. Summaries of their oral presentations at the Florence meeting are reported in the Appendix to this review article. STATE OF ART AND PERSPECTIVES: One of the most important advances has been the elucidation of new molecular mechanisms of control of gene expression by retinoids. A number of new retinoids have been synthesized by chemists, some of which are being screened for potential clinical use, and a few have already had a tremendous impact on clinical practice. The most important achievements have been obtained in acute promyelocytic leukemia. In 1988 a Chinese group working in Shanghai showed that using all-trans retinoic acid (ATRA) alone 94% of acute promyelocytic leukemic patients obtained complete remission through differentiation of the leukemic clone. This result transformed a dream into reality and allowed researchers to move from laboratory experience to clinical applications of this differentiating therapy. Expanding the spectrum of hematological malignancies that may respond to ATRA remains a challenge; however, several results show some activity of retinoids alone or in combination with other drugs in juvenile chronic myeloid leukemia (CML), myelodysplastic syndrome, cutaneous T-cell lymphoma and CML. Particularly interesting are the studies that explored the potential clinical synergism of ATRA-based combination therapies with growth factors, other differentiating agents such as vitamin D3, immunomodulators like interferons, or chemotherapeutic agents, in particular Ara-C, all of which show promising in vitro effects when used in combination with retinoids.

Associations of retinoids, tamoxifen and alpha-interferon 2a in human breast cancer.

We evaluated in vitro the potentiating and/or synergistic antitumor effects among retinoids (all-trans-retinoic acid, tRA, and 13-cis-retinoic acid, 13cRA), alpha-interferon 2a (alpha-IFN 2a) and tamoxifen (TAM) on both estrogen receptor positive (ER(+)) and negative (ER(-)) human breast cancer cell lines. In our experimental model, the three studied agents showed antiproliferative activity in ER(+) cell lines MCF-7 and ZR-75.1, while alpha-IFN 2a was the most effective drug in the ER(-) cell line MDA-MB-231. Retinoids and TAM exerted a strong apoptotic effect in MCF-7 cells, while such an effect was obtained in MDA-MB-231 cells by alpha-IFN 2a. The tested combinations displayed different effects in the different evaluated cell lines: i) in MCF-7 cells tRA + TAM showed additive activity, both tRA + alpha-IFN 2a and TAM + alpha-IFN 2a association displayed a synergistic effect, and a further potentiation of the antiproliferative action was detected with the triple combination; ii) in ZR-75.1 cell line an additive activity was showed by tRA + TAM and TAM + alpha-IFN 2a, while tRA + alpha-IFN 2a produced synergistic action; iii) in MDA-MB-231 cell line only alpha-IFN 2a displayed a strong antiproliferative effect, and no significant potentiation was exerted by any drug association. The feasibility and activity of such combinations have been tested in two pilot clinical trials on patients with metastatic breast cancer: both the tested associations were tolerable, with good treatment compliance and low toxicity. The different antiproliferative and apoptotic effects observed in vitro on apparently similar breast cancer cell lines prompted us to a further investigation of the mutual biological modulations of these drug combinations, in view of a better selection of patients who might potentially benefit from these treatments.

Recombinant alpha 2a interferon and polycythemia vera: clinical results and biological evaluation by means of Fourier-transform infrared microspectroscopy.

Polycythemia vera (PV) is a chronic myeloproliferative disease. The use of recombinant alpha 2a Interferon (IFN) therapy in this disease is a novel approach. We applied Fourier-transform infrared microspectroscopy (FT-IR-M) to investigate the behavior and therapeutic responsiveness of PV patients treated with IFN. A spectroscopic parameter (A1/A2) was used, corresponding to the ratio of the integrated areas of the bands at 1080 cm-1 and at 1540 cm-1 due to nucleic acids and proteic components, respectively, calculated on the spectra of single megakaryocytes (MKs). In previous studies, we have pointed out that MKs in PV have a surprisingly strong myeloproliferative impulse when compared to MKs from other chronic myeloproliferative diseases. Nine patients out of the 11 studied exhibited a satisfactory responsiveness to the IFN treatment. Ten patients were evaluated by the A1/A2 parameter. In 8 of these, a good agreement was seen between this parameter and the laboratory data commonly used for the assessment of this disease. The infrared parameter, which we propose, proves to be an original, reliable method for the evaluation of recombinant alpha 2a IFN responsiveness in this disease.

Precore mutant hepatitis B virus and outcome of chronic infection and hepatitis in hepatitis B e antigen-positive children.

Mutant hepatitis B virus (HBV), responsible for the lack of hepatitis B virus "e" antigen (HBeAg) secretion because of a translational stop codon at nucleotide 1896 of the HBV-DNA precore region (HBeAg minus HBV), has been detected worldwide in acute and chronic HBV infections and diseases. HBeAg minus HBV appears to condition the outcome of infection and to be involved in the pathogenesis of hepatitis B. We investigated the mutant prevalence and its clinical implications in 30 hepatitis B surface antigen/HBeAg-positive children (17 treated with interferon) with chronic hepatitis B. Wild-type and HBeAg minus HBV were characterized by quantitative oligohybridization assays in sera from 29 children followed up for a mean of 33 mo (12 mo to 9 y). At admission, 18 children (62%) circulated wild-type HBV alone; mutant HBV became detectable in two of them during the follow-up before HBeAg/anti-HBe seroconversion. Wild-type and HBeAg minus HBV were detected in 10 children (34.5%); mutant HBV levels were lower than 20% of total viremia in four of them and higher in six. Serum HBV-DNA from one child did not hybridize with our probes. HBeAg minus HBV was associated with older age (p < 0.009) and higher histologic activity (p < 0.069). HBeAg/anti-HBe seroconversion occurred independently from HBeAg minus HBV detection; it was observed in six (37.5%) of 16 children with wild-type HBV alone and in four (33.3%) of 12 children with mixed viremia.(ABSTRACT TRUNCATED AT 250 WORDS)

Continuous versus intermittent therapy for chronic hepatitis C with recombinant interferon alfa-2a.

BACKGROUND/AIMS: Prolonged interferon administration to patients with chronic hepatitis C, although increasing the sustained response rate, is poorly accepted and may favor drug resistance. A pulse-treatment schedule would be preferred for compliance and costs. METHODS: One hundred thirty-five patients with chronic hepatitis C received 6 MU units of interferon alfa-2a, three times weekly, continuously for 9 months (group 1: 66 patients) or for two 3-month cycles, separated by 6 months pause (group 2: 69 patients). RESULTS: At the end of therapy, 25 of 54 patients of group 1 (46.3%) and 28 of 60 of group 2 (46.7%) had normal serum aminotransferase levels. Six months after the end of treatment, sustained responders were still similar in the two groups (11 or 16.7% vs. 7 or 10.1%; NS). A loss of response before the end of therapy was seen in 10 patients of group 1 and 6 of group 2; interferon-neutralizing antibodies developed in 1 of 7 and 6 of 6 of such patients, respectively. CONCLUSIONS: The intermittent administration of interferon alfa-2a to patients with chronic hepatitis C shows a sustained response rate comparable with that achieved with continuous treatment at the same dosage. Hepatitis breakthroughs during pulse therapy appeared to be limited to interferon neutralizing antibodies, whereas a prolonged, continuous treatment is more likely to induce other forms of interferon resistance.

Synergistic effects of alpha interferon and 1,25 dihydroxyvitamin D3: preliminary evidence suggesting that interferon induces expression of the vitamin receptor.

BACKGROUND: To active metabolite of vitamin D3-1,25(OH)2D3-is a well-known differentiation inducer. The addition of this metabolite to sensitive cell cultures inhibits proliferation and induces monocytic-macrophagic differentiation. Alpha interferon may also inhibit proliferation and increase the expression of some surface antigens in some neoplastic cells. In the present report, we describe the synergistic activity of these two drugs on U-937 and on cultured cells from a leukemic patient. METHODS: Proliferation was studied by 3H-thymidine incorporation; differentiation markers were evaluated immunologically by monoclonal antibodies and by cytochemical tests. Phagocytosis and NBT reduction test were also performed in order to confirm the differentiating properties of these drugs. Finally, the expression of the 1,25(OH)2D3 receptor was evaluated by immunochemical methods. RESULTS: After culturing these cells for 72 hours in the presence of 1,25(OH)2D3, cell proliferation was reduced and the expression of some phenotypic and functional markers suggested monocytic-macrophagic differentiation. Alpha interferon and 1,25(OH)2D3 synergistically inhibit the proliferation of U-937 cells. Alpha interferon increased the expression of the 1,25(OH)2D3 receptor in U-937 cells. CONCLUSIONS: The reported results confirm the synergistic activity of INF and 1,25(OH)2D3 on cell proliferation in monoblastic cells. The possible role of the increased expression of the vitamin receptor in cells cultured in the presence of INF is discussed.