Biomedical subjects

Sandra Franco

Publications and source records attributed to Sandra Franco.

13 recordsLinked to original sources

PubMed2006

Phase I trial and antitumor effects of BZL101 for patients with advanced breast cancer.

BACKGROUND: Botanical therapies are often used by breast cancer patients yet few clinical trials have evaluated their safety and efficacy. We studied mechanisms of activity and performed a phase I clinical trial in patients with advanced breast cancer to evaluate BZL101, an aqueous extract from Scutellaria barbata. METHODS: Preclinical studies were conducted in vitro to characterize cell death induced by BZL101. In a phase I trial, eligible patients had histologically confirmed, measurable metastatic breast cancer. Treatment consisted of 350 ml per day of oral BZL101, administered as sole cancer therapy until disease progression, toxicity or personal preference to discontinue. Primary endpoints were safety, toxicity and tumor response. RESULTS: BZL101 extract induced strong growth inhibition and apoptosis of breast cancer cell lines. In the phase I trial, 21 patients received BZL101. Mean age was 54 years (30-77) and mean number of prior treatments for metastatic disease was 3.9 (0-10). There were no grade III or IV adverse events (AEs). The most frequently reported BZL101-related grade I and II AEs included: nausea (38%), diarrhea (24%), headache (19%) flatulence (14%), vomiting (10%), constipation (10%), and fatigue (10%). Sixteen patients were evaluable for response. Four patients had stable disease (SD) for >90 days (25%) and 3/16 had SD for >180 days (19%). Five patients had objective tumor regression, one of which was 1 mm short of a PR based on RECIST criteria. CONCLUSIONS: BZL 101 inhibits breast cancer cell lines by inducing apoptosis. In a phase I clinical trial, BZL101 was safe and had a favorable toxicity profile. BZL101 demonstrated encouraging clinical activity in this heavily pretreated population.

Administration, Oral↗

PubMed2006

Mutational patterns associated with the 69 insertion complex in multi-drug-resistant HIV-1 reverse transcriptase that confer increased excision activity and high-level resistance to zidovudine.

Human immunodeficiency virus type 1 (HIV-1) strains having dipeptide insertions in the fingers subdomain and other drug resistance-related mutations scattered throughout their reverse transcriptase (RT)-coding region show high-level resistance to zidovudine (AZT) and other nucleoside analogues. Those phenotypic effects have been correlated with their increased ATP-dependent phosphorolytic activity on chain-terminated primers. Mutations T69S and T215Y and a dipeptide insertion (i.e. Ser-Ser) between positions 69 and 70 are required to achieve low-level resistance to thymidine analogues. However, additional amino acid substitutions are necessary to achieve the high-level phenotypic resistance to AZT shown by clinical HIV isolates carrying a dipeptide insertion in their RT-coding region. In order to identify those mutations that contribute to resistance in the sequence context of an insertion-containing RT derived from an HIV clinical isolate (designated as SS RT), we expressed and purified a series of chimeric enzymes containing portions of the wild-type or SS RT sequences. ATP-mediated excision activity measurements using AZT- and stavudine (d4T)-terminated primers and phenotypic assays showed that molecular determinants of high-level resistance to AZT were located in the fingers subdomain of the polymerase. Further studies, using recombinant RTs obtained by site-directed mutagenesis, revealed that M41L, A62V and in a lesser extent K70R, were the key mutations that together with T69S, T215Y and the dipeptide insertion conferred high levels of ATP-dependent phosphorolytic activity on AZT and d4T-terminated primers. Excision activity correlated well with AZT susceptibility measurements, and was consistent with phenotypic resistance to d4T. Structural analysis of the location of the implicated amino acid substitutions revealed a coordinated effect of M41L and A62V on the positioning of the beta3-beta4 hairpin loop, which plays a key role in the resistance mechanism.

Adenosine Triphosphate↗

PubMed2006

Complete nucleotide sequence of genotype 4 hepatitis C viruses isolated from patients co-infected with human immunodeficiency virus type 1.

The hepatitis C virus (HCV) genotype 4 is spreading among southern European intravenous drug users, who are frequently co-infected with human immunodeficiency virus type 1 (HIV-1). Response to interferon (IFN) alpha-based therapies in HIV-1 positive patients co-infected with HCV genotype 4 is poor, similar to that obtained for HCV genotype 1 and much lower than for HCV genotypes 2 and 3. The lack of sequence data related to HCV of genotype 4 prompted us to sequence the complete genome of two genotype 4 variants isolated from two HIV-1 co-infected patients (24 and 25). Our aim was to investigate the evolutionary relationships of the former variants with other genotypes and/or genotype 4 subtypes. Sequence alignments and phylogenetic analysis from genomic regions 5'NC, core-E1 and NS5B revealed that the variants isolated from patients 24 and 25 (both subtyped 4c/4d by INNO-LIPA II HCV) belong to subtypes 4d and 4a, respectively. When looking at the complete genome sequence one of the variants showed a new genotype 4 subtype. Interestingly, sequence length differences in the interferon sensitivity determining region coding regions were observed when compared with sequences from other genotypes. Similarly, when the catalytic efficiency of the NS3/4 protease from patients 24 and 25 samples were determined, they displayed 70.6+/-7.7 and 23.5+/-3.4%, respectively, of the activity shown by genotype 1 NS3/4 proteases. Overall, pairwise comparison and phylogenetic analysis of nucleotide sequences of the complete genome or the different protein-encoding regions showed that genotype 4 sequences were more closely related to genotype 1 sequences. The description of new HCV genome variants may help our understanding of the HCV biology as well as the role of different genotypes in HCV treatment and therapy response.

Amino Acid Sequence↗

PubMed2005

Dynamics of hepatitis C virus NS5A quasispecies during interferon and ribavirin therapy in responder and non-responder patients with genotype 1b chronic hepatitis C.

The quasispecies nature of hepatitis C virus (HCV) may have important implications concerning resistance to antiviral agents. To determine whether HCV NS5A quasispecies composition and dynamics are related to responsiveness to combined interferon (IFN) and ribavirin therapy, extensive sequence analyses of cloned RT-PCR amplification products of HCV-1b NS5A quasispecies of sequential isolates from 15 treated (nine sustained responders and six non-responders) and three untreated patients were performed. Accumulation of mutations in NS5A during therapy was relatively frequent in the V3 domain, but unusual elsewhere. Amino acid changes were the result of the imposition of minor variants that were already present before treatment and always occurred within the first week of therapy. Before treatment, the complexity and diversity of quasispecies were lower in isolates from responders than in those from non-responders, particularly in the V3 domain, where differences in nucleotide entropy (0.35 vs 0.64, P=0.003), genetic distance (0.0145 vs 0.0302, P=0.05) and non-synonymous substitutions (0.0102 vs 0.0203, P=0.036) were statistically significant. These differences became more apparent during treatment, because complexity and diversity remained stable or tended to increase in non-responders, whereas they tended to decrease in responders. These observations suggest that the composition and dynamics of HCV NS5A quasispecies, particularly in the V3 domain, may play a role in the response to combined IFN/ribavirin therapy.

Amino Acid Sequence↗

PubMed2004

Relative replication fitness of multi-nucleoside analogue-resistant HIV-1 strains bearing a dipeptide insertion in the fingers subdomain of the reverse transcriptase and mutations at codons 67 and 215.

A two-serine insertion at position 69 (i69SS) of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) appears to be critical to enhance multi-nucleoside RT inhibitor resistance (MNR) in the sequence context of multiple zidovudine (AZT) resistance mutations (i.e., M41L, L210W, T215Y). In this study, we measured the replication capacity relative to the wild-type (WT) HIV-1 of a series of recombinant viruses carrying the i69SS in the background of a clinical isolate with MNR in which we introduced mutations D67N, Y215T, Y215S, or Y215N. In vitro measurements included replication kinetics and growth competition assays at different multiplicities of infection (MOI). While the addition of D67N had a minor effect on replication capacity, the reversion of Tyr-215 to Thr, Ser, or Asn was sufficient to increase the virus ability to replicate in a drug-free environment. The same genotypic changes at position 215 rendered the MNR virus susceptible to AZT and stavudine. Interestingly, the presence of the insertion together with mutation T215Y in an otherwise WT sequence background was not sufficient to confer high-level resistance to AZT, although its replication capacity was clearly impaired. Therefore, the RT residue 215 plays a critical role in both replication capacity and drug resistance of multidrug-resistant viruses containing the i69SS.

Adult↗

PubMed2004

Molecular determinants of multi-nucleoside analogue resistance in HIV-1 reverse transcriptases containing a dipeptide insertion in the fingers subdomain: effect of mutations D67N and T215Y on removal of thymidine nucleotide analogues from blocked DNA primers.

Human immunodeficiency virus type 1 isolates having dipeptide insertions in the fingers subdomain of the reverse transcriptase (RT) show high level resistance to 3 '-azido-3 '-deoxythymidine (AZT) and other nucleoside analogues. Insertions are usually associated with thymidine analogue resistance mutations, such as T215Y. The resistance phenotype correlates with increased ATP-dependent phosphorolytic activity, which facilitates removal of thymidine analogues from inhibitor-terminated primers. In this report, we show that substituting Thr, Ser, or Asn for Tyr-215 in a multidrug-resistant RT, bearing a Ser-Ser insertion between codons 69 and 70, leads to AZT and stavudine resensitization through the loss of the ATP-mediated removal activity. The mutation D67N, which is rarely found in insertion-containing strains, had no effect on excision and a minor influence on resistance. Substituting Tyr-215 had a larger effect than deleting the dipeptide insertion. The presence of both the insertion and mutation T215Y in the wild-type BH10 RT conferred significant ATP-mediated removal activity and moderate resistance to AZT. However, resistance levels and unblocking activities were lower than those observed with the multidrug-resistant enzyme. Removal reactions can be inhibited by the next complementary dNTP. Both Tyr-215 and the dipeptide insertion affect RT-DNA.DNA-dNTP ternary complex formation, an effect that was not detected in the presence of foscarnet. Based on crystal structures of binary and ternary complexes of HIV-1 RT, we propose that Tyr-215 exerts its action by facilitating a proper orientation of the pyrophosphate donor molecule, whereas the effects on dNTP binding are indirect and could be related to significant conformational changes occurring during polymerization.

Adenosine Triphosphate↗

PubMed2004

Early HCV dynamics on Peg-interferon and ribavirin in HIV/HCV co-infection: indications for the investigation of new treatment approaches.

OBJECTIVES: To describe the 28-day hepatitis C virus (HCV) kinetics under Pegylated-interferon (Peg-IFN) + ribavirin (RBV) therapy in HIV/HCV co-infected patients. To evaluate the predictive value of early virological response (EVR) of achieving a sustained virological response (SVR). To investigate the baseline mutations in the interferon sensitivity determining region (ISDR)2209-2248 in the non-structural 5A protein of HCV according to genotype. METHODS: Open, prospective trial including 28 co-infected patients with directly observed treatment with Peg-IFN + RBV. We assessed the predictive values of EVR (> or = 2 log10 of HCV decay or a negative qualitative test) at days 1, 7, 28 and in week 12 of the SVR. RESULTS: The SVR in an intention-to-treat analysis was 28.6% (genotype 1, 1/13; genotype 3, 6/10; genotype 4, 1/5). Patients who reached SVR presented a significantly faster HCV plasma viral load reduction compared to non-responders from the first 24 h [-1.06 log10 (interquartile range, -1.7 to -0.4) versus -0.05 log10 (interquartile range, -0.4 to +0.14) respectively; P = 0.002]. The median HCV viral load at week 12 was significantly different from that at baseline in responder and transient responders but not in non-responder patients. The positive predictive value was 100% within the first month and the best negative predictive value was 92% and 88.8% at weeks 4 and 12 respectively. The only genotype 1 responder patient had eight mutations in ISDR2209-2248. CONCLUSIONS: A very early HCV viral decay is observed in responder patients. An early virological response assessment at week 4 and 12 might be a useful tool in the clinical management of the co-infected population.

Adult↗

PubMed2004

Response to fulvestrant in heavily pretreated postmenopausal women: a single-center experience.

Fulvestrant ('Faslodex') is a new estrogen receptor (ER) antagonist that has no agonist effects. It binds, blocks and accelerates degradation of the ER, leading to a complete abrogation of estrogen-sensitive gene transcription. In postmenopausal women with advanced breast cancer progressing on prior endocrine therapy, fulvestrant is at least as effective as the third-generation aromatase inhibitor (AI) anastrozole. In this single-center experience, 42 postmenopausal patients with metastatic breast cancer who had been heavily pretreated with prior endocrine therapy and chemotherapy were treated with fulvestrant. Prior endocrine therapies included selective ER modulators (including tamoxifen and toremifene), AIs, megestrol acetate, and high-dose estrogens. In total, eight patients (19%) achieved stable disease (SD) for > or =24 weeks, including two patients with SD for 2 years and one with SD for 14 months. Fulvestrant was well tolerated with the majority of adverse events related to the site of metastatic disease. These data demonstrate that fulvestrant is a well tolerated and effective endocrine therapy for postmenopausal women with metastatic breast cancer who have been heavily pretreated with prior therapies. The novel mechanism of action of fulvestrant reduces the likelihood of cross-resistance with other endocrine therapies and therefore this agent may be active in patients who have proved to be resistant to treatments such as tamoxifen or AIs. The use of fulvestrant earlier in the sequence of endocrine treatments may achieve better responses than observed in this heavily pretreated patient population.

Adult↗

PubMed2003

Characterization and evolution of NS5A quasispecies of hepatitis C virus genotype 1b in patients with different stages of liver disease.

The quasispecies nature of hepatitis C virus (HCV) is thought to play a central role in modulating viral functions. Recent work has linked NS5A protein with viral replication, resistance to interferon (IFN), and control of cellular growth, probably through the interaction of its protein kinase R (double stranded RNA-activated protein kinase, PKR) binding domain (PKR-bd) with cellular PKR, but knowledge of how PKR-bd viral population evolves during disease progression is limited. Since we have previously described an association between amino acid composition of the PKR-bd and the presence of HCC, in this report we further investigated the dynamic behavior of viral population parameters by sequencing an average of 20 clones per sample in 27 samples from 19 untreated patients with different degrees of liver disease, 8 of whom were followed over time. Viral population parameters varied widely from patient to patient, but no differences were observed in the complexity, diversity, types of nucleotide changes, or evolutionary pattern of the quasispecies according to the stage of liver disease. In five samples, we detected "quasispecies-tails"; that is, clones whose minimum genetic distance to the remaining clones of their own quasispecies were higher than the maximum genetic distance found between any other two clones of the same sample. In summary, independent of the degree of liver disease, or the mutations detected within the consensus sequence of the PKR-bd, the NS5A of HCV presents a flexible and variable quasispecies structure that remains largely stable during the natural course of an HCV infection, highlighting the central role of NS5A protein in viral life cycle.

Amino Acid Sequence↗

PubMed2003

Influence of human immunodeficiency virus type 1 subtype on mother-to-child transmission.

The present study was designed to assess whether the subtype of human immunodeficiency virus type 1 (HIV-1) could affect the rate of HIV-1 mother-to-child transmission in a cohort of 31 HIV-1-seropositive pregnant Tanzanian women. In order to assign a subtype to the samples analysed, nucleotide sequencing of the HIV-1 long terminal repeat U3 and C2V3C3 envelope regions was performed from the sera of these 31 pregnant women. Except in three cases, amplification of both regions was achieved in all samples. Subtypes A (n=13, 46 %), C (n=6, 21 %) and D (n=2, 7 %), as well as a number (25 %) of A/C, C/A, D/A and C/D recombinant forms (n=3, 2, 1 and 1, respectively), were identified. Of the 31 HIV-1 seropositive pregnant women analysed, eight (26 %) transmitted HIV-1 to their infants. Among the eight transmitter mothers, four (4 of 13, 31 %) were infected with HIV-1 subtype A, one (1 of 6, 17 %) with HIV-1 subtype C, none (0 of 2, 0 %) with HIV-1 subtype D and three (3 of 7, 43 %) with HIV-1 subtype recombinant A/C. These findings show no significant differences in the mother-to-child transmissibility of HIV-1 subtypes A, C and D and detected recombinants forms.

Cohort Studies↗

PubMed2002

Outbreak of nosocomial hepatitis C virus infection resolved by genetic analysis of HCV RNA.

In July 2000, symptomatic acute hepatitis C was diagnosed in five patients who had attended the emergency room of a municipal hospital on the same day, about 6 weeks before. Investigation of the remaining 65 patients visited at the emergency room on that day disclosed that 8 patients had a positive anti-hepatitis C virus (anti-HCV) test and 4 of them had biochemical evidence of acute anicteric hepatitis. HCV RNA was detected in 12 of the 13 anti-HCV-positive patients. Phylogenetic analysis of the nonstructural 5A (NS5A) and E2 regions showed that 10 patients, including all 9 with acute hepatitis, were infected with a closely related HCV strain, while the remaining 2 patients harbored unrelated strains. Flushing of intravenous catheters with heparin retrieved from a multidose heparin solution in saline was carried out for all the patients involved in the hepatitis outbreak but in only 1 of 23 (4%) matched controls recruited among HCV-noninfected patients attending the emergency room on the same day, and this was the only significant difference concerning risk factors for HCV infection between patients and controls. Thus, accidental contamination of a multidose heparin solution with blood from an unrecognized HCV carrier was identified as the source of this nosocomial outbreak of hepatitis C.

Adult↗

PubMedDate not supplied

Measuring corneal thickness with a rotary scanning system.

PURPOSE: Recently the authors presented a technique that allows measurement of the corneal thickness along a vertical meridian from optical sections obtained using a slit-lamp microscope. We present a new system that allows rotary scanning of the cornea and consequent measurement of the corneal thickness along any meridian. METHOD: The corneal thickness along two perpendicular meridians was measured from optical sections obtained with a new rotary scanning system. To obtain the optical sections, the cornea is illuminated with a light beam that is previously expanded in a fan by a small cylindrical lens rotated on a plane-containing axis to explore the cornea. The light diffused by both corneal surfaces is collected by two video cameras placed at an angle with the light beam. The axes of the two cameras define, with the visual axis, two planes normal to each other. After image acquisition, a processing algorithm is applied in order to compute the corneal thickness along the two meridians. RESULTS: Some preliminary results for the vertical and the horizontal meridians of one eye are shown, as well as all the intermediate computations. CONCLUSIONS: With this system, we expect to be able to measure corneal topography and thickness along an arbitrary meridian. The rotary scanning of the cornea is mechanically simple and will eventually allow automated scanning.

Cornea↗

PubMedDate not supplied

Corneal thickness and elevation maps computed from optical rotary scans.

PURPOSE: Recently, the authors presented a technique that allows corneal thickness measurements along any meridian from optical sections obtained using a rotary scanning system. This paper presents three-dimensional mapping of the corneal thickness and topography of both corneal surfaces, obtained with the rotary system. METHOD: Corneal thickness and topography are computed from optical sections obtained by illumination with a collimated beam expanded in a fan by a small cylindrical lens. This lens is provided with motor driven rotation to perform automated rotary scanning of the whole cornea. Two cameras are used to capture the images of the optical sections. RESULTS: With this system, it is possible to obtain measurements of corneal thickness, as well as corneal topography. Corneal thickness and elevation maps are shown. CONCLUSIONS: Although still under development, this new optical system allows measurement of the thickness of the whole cornea as well as topographical mapping of both corneal surfaces.

Corneal Topography↗