Focused library of phenyl-fused macrocyclic amidinoureas as antifungal agents.

The rise of antimicrobial-resistant phenotypes and the spread of the global pandemic of COVID-19 are worsening the outcomes of hospitalized patients for invasive fungal infections. Among them, candidiases are seriously worrying, especially since the currently available drug armamentarium is extremely limited. We recently reported a new class of macrocyclic amidinoureas bearing a guanidino tail as promising antifungal agents. Herein, we present the design and synthesis of a focused library of seven derivatives of macrocyclic amidinoureas, bearing a second phenyl ring fused with the core. Biological activity evaluation shows an interesting antifungal profile for some compounds, resulting to be active on a large panel of Candida spp. and C. neoformans. PAMPA experiments for representative compounds of the series revealed a low passive diffusion, suggesting a membrane-based mechanism of action or the involvement of active transport systems. Also, compounds were found not toxic at high concentrations, as assessed through MTT assays.

ABO incompatibility does not influence transfusion requirements in patients undergoing single-unit umbilical cord blood transplantation.

Hematopoietic stem cell transplantation is usually performed without considering the ABO compatibility between donor and recipient. There are few studies analyzing ABO matching impact on transfusion outcome of umbilical cord blood transplantation (UCBT) recipients. The aim of this study was to analyze factors influencing transfusion outcome, highlighting the ABO matching between donor and recipient. This study has reviewed data from 318 patients who underwent single unit UCBT at la Fe University Hospital from January 2000 to December 2014. There were no differences between RBC and platelet (PLT) requirements or RBC and PLT transfusion independence according to ABO matching between donor and recipient. RBC and PLT requirements were statistically correlated (ρ=0,841, P<0.001). A total of 170 and 188 patients achieved RBC and PLT independence, respectively, within 180 days after UCBT. Persistence of recipient isoagglutinins was detected in 6.8% of patients with major ABO incompatibility at median of 176 days (103-269) after UCBT. Autoimmune haemolytic anemia was diagnosed in 15 patients, 12 of them due to cold antibodies. In conclusion, ABO matching has not influenced transfusion requirements of patients undergoing UCBT.

Assessment of coastal marine pollution in Galicia (NW Iberian Peninsula); metal concentrations in seawater, sediments and mussels (Mytilus galloprovincialis) versus embryo-larval bioassays using Paracentrotus lividus and Ciona intestinalis.

Sediments from three Galician Rias were tested for toxicity using sea-urchin and ascidian sediment elutriate embryo-larval bioassays. Trace metal contents in seawater, sediments and mussels were also determined and subjected to multidimensional scaling methods which grouped stations according to chemical contamination. High metal contents were found in seawater, sediments and mussels from the Ria of Pontevedra, and moderate levels were detected in the Ria of Vigo and Ria of Arousa. The results revealed that samples assessed as toxic, according to the sea-urchin and ascidian embryo-larval bioassays, were among the most polluted by trace metals. A good agreement was reported between ordination plots resulting from applying multidimensional scaling to the chemical data, and the results of the biological endpoints tested.

Effect of humic acids on speciation and toxicity of copper to Paracentrotus lividus larvae in seawater.

The effects of humic acid (HA) on the toxicity of copper to sea urchin Paracentrotus lividus larvae were studied in chemically defined seawater. Square Wave Anodic Stripping Voltammetry (SWASV) was employed to study the complexation of copper in seawater medium. A simple complexation model assuming one ligand type and a 1:1 reaction stoichiometry successfully explained the inverse titration experiments. A conditional stability constant of 6.53+/-0.05 and a complexating capacity of 230+/-7 micromol Cu/g HA were obtained. Sea urchin bioassay tests with two endpoints, embryogenesis success and larval growth were carried out in order to study the toxicity of dissolved copper in both the presence and absence of HA. The toxicity data obtained fitted well into a logistic model, and the high sensitivity of both endpoints (EC(50) were 41.1 microg Cu/l and 32.9 microg Cu/l, respectively) encourages their use for biomonitoring. The HA had a clearly protective effect, reducing the toxicity of Cu to the sea urchin larvae. The labile copper, rather than the total copper concentrations, explained the toxicity of the Cu-HA solutions, and the Cu-HA complexes appeared as non-toxic forms. These results are in agreement with the Free Ion Activity Model, because the labile Cu concentrations in this buffered and chemically defined medium covary with the free ion activity of the Cu, validating the model to naturally occurring HA in the marine environment.

Successful treatment of severe intra-abdominal bleeding associated with disseminated intravascular coagulation using recombinant activated factor VII.

Recombinant activated factor VII (rFVIIa) is indicated mainly for the treatment of patients with haemophilia and inhibitors. However, little information is available on the use of rFVIIa in the treatment of the severe bleeding associated with disseminated intravascular coagulation (DIC). We report a pregnant woman with DIC, who developed severe intra-abdominal bleeding after caesarean section. Despite treatment with fresh-frozen plasma, fibrinogen, platelet transfusions and surgery, the abdominal bleeding persisted and intravenous treatment with rFVIIa was initiated. The response to treatment was rapid, with control of the bleeding and resolution of the coagulopathy. No side-effects related to rFVIIa were noted. This case suggests a potential role for rFVIIa in the treatment of severe and refractory bleeding associated with DIC.

Progression to acquired immunodeficiency syndrome in 94 human immunodeficiency virus-positive hemophiliacs with long-term follow-up.

BACKGROUND AND OBJECTIVES: Human immunodeficiency virus (HIV) infection was transmitted to many hemophilics treated with non-inactivated factor concentrates before 1986. The aim of this study was to know the long-term incidence of AIDS and risk factors for its development in HIV-infected hemophiliacs. DESIGN AND METHODS: This study was a retrospective analysis of 94 HIV-infected hemophilics. The cumulative incidence of AIDS during a follow-up of 16 years from seroconversion was determined by Kaplan-Meier analysis,and potential risk factors were also studied by multivariate analysis. RESULTS: The 16-year estimated incidence of AIDS was 38% (95%CI 27%-52%). The AIDS incidence was significantly higher in patients with hemophilia B (p <0.0001), older age at seroconversion (p=0.0004), lower CD4 counts at seroconversion (p=0.004), and lower concentrate consumption during follow-up (p=0.02), than it was in those patients without these characteristics. However, only hemophilia type and age at seroconversion remained significant in the multivariate analysis, with a relative risk of 0.06 (95%CI 0.02-0.20) for hemophilia A and 1.04(95%CI 1.01-1.06) for every year of increase in age at seroconversion. The severity of hemophilia, history of inhibitors and concentrate consumption before seroconversion were not significantly associated with AIDS development. INTERPRETATION AND CONCLUSIONS: A considerable proportion of HIV-infected hemophiliacs remained AIDS-free 16 years after seroconversion. The risk of AIDS was particularly high in patients with hemophilia B and for patients who were older at seroconversion.

Incidence of factor VIII inhibitors in severe haemophilia: the importance of patient age.

The potential effect of age at the start of replacement therapy on the development of factor VIII (FVIII) inhibitors was assessed in 62 severe (FVIII < 2 IU/dl) haemophilia A patients who started FVIII therapy at one of two haemophilia centres. Inhibitors were tested on an annual basis. Persistent or high-titre inhibitors were detected in 15 patients (24%). Kaplan-Meier cumulative incidence at 3 years from first FVIII exposure was 41% (95% CI 22-67%) in patients starting therapy before the age of 6 months, 29% (95% CI 13-57%) in patients starting therapy between 6 and 12 months of age, and 12% (95% CI 4-34%) in those starting therapy beyond 1 year of age (P = 0.03). By multivariate analysis, the influence of age was shown to be independent of other variables, including calendar year at the onset of therapy and baseline FVIII plasma levels. In conclusion, patient age at initial treatment appears to influence inhibitor formation. If confirmed, this finding would have a major impact on the management of haemophilia.

A comparison of FVII:C and FVIIa assays for the monitoring of recombinant factor VIIa treatment.

The use of recombinant factor VIIa (rFVIIa) is on the increase, not only to treat haemophilic patients with inhibitors, but also patients with other clotting disorders. However, the most appropriate method of monitoring this treatment remains a question that has yet to be resolved. We studied 24 plasma samples from patients receiving rFVIIa treatment (three had haemophilia A with inhibitors, and three a congenital FVII deficiency) and compared the results obtained from the FVII:C and FVIIa assays. Although a good correlation between the two methods was obtained (r = 0.91), the values of the FVII:C method were 1.63 higher than those of the FVIIa method, with a relatively wide margin in the interval of the FVII:C/FVIIa ratios obtained [95% confidence interval (CI) 1.38--1.88, range 0.68--3.68]. This interval became wider when we compared values of over 6 IU mL(-1), which led us to conclude that the two methods cannot be considered equivalent. As the FVIIa method specifically measures FVIIa, and FVII:C assay is known to have a wide interlaboratory variability, we believe that the FVIIa assay would be more suitable for the monitoring of rFVIIa treatment.

Q1311X: a novel nonsense mutation of putative ancient origin in the von Willebrand factor gene.

Type 3 von Willebrand disease, a recessive autosomally inherited bleeding disorder, refers to complete deficiency of von Willebrand factor (VWF). The novel Q1311X mutation was detected in the homozygous state in four Spanish patients from two apparently unrelated families of gypsy origin. The lack of specific amplification of platelet VWF cDNA from two of the patients indicates reduced levels of mutated gene expression. The similar haplotype linked to mutated alleles suggests a common origin. On the basis of the two instabilities observed and the estimated mutation rate of the microsatellites of intron 40 of the VWF gene, we can estimate that this mutation could have arisen about 2300 years ago.

Trasplante pulmonar y hemofilia. Consideraciones perioperatorias / Lung transplantation and hemophilia. Perioperative considerations

El trasplante pulmonar es una indicación terapéutica para pacientes seleccionados con enfermedad pulmonar terminal.Presentamos el caso clínico de un trasplante pulmonar unipulmonar izquierdo en una paciente de 16 años con hemofilia B y diagnóstico de fibrosis pulmonar idiopática, con antecedentes de desnutrición, osteoporosis, escoliosis severa, serología VHC positiva y neumotórax bilateral recidivante.La terapia sustitutiva con factor IX ultrapuro se inició en el momento en que se dispuso del pulmón donante y se mantuvo hasta 37 días postintervención. Los valores plasmáticos de factor IX se mantuvieron próximos al 100 por ciento durante el acto quirúrgico y postoperatorio inmediato, y por encima del 40 por ciento pasado éste, lo que permitió una correcta hemostasia durante todo el proceso, sin precisar la administración de hemoderivados.La evolución de la paciente fue favorable, con una estancia en la unidad de reanimación de 17 días, dándose el alta hospitalaria a los 40 días del trasplante.Se comentan las consideraciones de la hemofilia con respecto al trasplante pulmonar, y la influencia que tienen sobre éste la malnutrición, el tratamiento crónico esteroide y la osteoporosis (AU)

Haemophilia B in a female caused by skewed inactivation of the normal X-chromosome.

Hemophilia B (factor IX deficiency) is an X-linked recessive disorder with a prevalence of 1:30,000 male births, which rarely affects females. A missense mutation T38R (6488C>G) of the factor IX (FIX) gene was characterized in a young female with moderate-to-severe hemophilia B. She is heterozygous for this mutation, which she inherited from her carrier mother. Analysis of the methyl-sensitive HpaII sites in the first exon of the human androgen-receptor locus indicated a de novo skewed X-chromosomal inactivation. This indicates that the paternal X-chromosome carrying her normal FIX gene is the inactive one, which has led to the phenotypic expression of hemophilia B in this patient.

Estudio epidemiológico de la hemofilia en España / Epidemiological study of hemophilia in Spain

El objetivo de este estudio epidemiológico fue la caracterización de un grupo representativo de pacientes del territorio español afectos de hemofilia con relación a su situación diagnóstica, clínica, terapéutica y a las complicaciones de la enfermedad y de su tratamiento. Se llevó a cabo por medio de un cuestionario que se envió a todos los centros de tratamiento. Resultados: fueron remitidos 825 cuestionarios evaluables, lo que corresponde aproximadamente a un tercio de la población hemofílica española. El 88 por ciento de los hemofílicos padecían hemofilia A y el 12 por ciento hemofilia B. El 36 por ciento graves (nivel inferior al 2 por ciento). La gran mayoría de pacientes estaban siguiendo un programa de tratamiento a demanda (79 por ciento). Un reducido número seguía, en el momento del registro, un tratamiento de profilaxis secundaria (17 por ciento), y tan solo un 3,4 por ciento seguía un programa de profilaxis primaria. El consumo medio de factores por paciente durante el año 1995, fue de 45.945 UI, lo que correspondería a una media de 706 Ul/kg/año. El 30 por ciento de los pacientes presentaban una o más articulaciones con algún grado de afectación. Los pacientes graves presentaban un total de 627 articulaciones afectadas de media por paciente. El 7,3 por ciento del total presentaba o habían presentado inhibidor. El 27 por ciento tenían anticuerpos anti-VIH. Los anticuerpos de la hepatitis C estaban presentes en el 61 por ciento. El antígeno HBs fue positivo en el 4 por ciento. (AU)

Diagnosis of two related carriers of severe haemophilia B with no family history.

Haemophilia B is an X-linked disease affecting 1 in 30 000 males. Carrier diagnosis is usually carried out only in female relatives of haemophilic males, and the likelihood of discovering a carrier without a haemophilic male is very low. In this report we present the cases of two related women without a family history of haemophilia who were diagnosed as haemophilia B carriers. Following a minor haemorrhage in the proband, she and her mother were thought to be haemophilia B carriers because of a low factor IX level (16 and 23 IU dL-1, respectively; normal values >50 IU dL-1). The non-sense mutation C31118T, which is associated with severe haemophilia B, was detected in both women. This allowed us to diagnose them as being definite carriers of severe haemophilia B and give appropriate genetic counselling.

[Lung transplantation and hemophilia. Preoperative considerations]. / Trasplante pulmonar y hemofilia. Consideraciones perioperatorias.

Lung transplantation is indicated in certain patients with terminal pulmonary disease. We report a case in which a single lung (left) was transplanted to a 16-year-old girl with hemophilia B; she also suffered idiopathic pulmonary fibrosis and had a history of malnutrition, osteoporosis, severe scoliosis, hepatitis C positivity and recurrent bilateral pneumothorax. Treatment with pure factor IX was started the moment the donor lung was available and was continued for 37 days after surgery. Plasma levels of factor IX were kept at 100% during surgery and in the early postoperative period, and over 40% after that time. Correct hemostasis was thus achieved throughout the procedure, with no need for blood products. Patient outcome was satisfactory. The stay in the intensive care recovery ward was 17 days and discharge was 40 days after transplantation. We discuss aspects of hemophilia and lung transplantation, and the influence on malnutrition, chronic steroid treatment and osteoporosis.

Postoperative use of rFVIIa by continuous infusion in a haemophilic boy.

Continuous infusion of coagulation factor concentrates has proved to be safe and effective. Because rFVIIa (NovoSeven is a very expensive product and very frequent doses are needed, continuous infusion is expected to be highly cost-effective. The postoperative use of continuous infusion of rFVIIa in a haemophilic boy with a high titre FVIII inhibitor is reported. He presented with a large right knee haemarthrosis and was treated with intermittent doses of rFVIIa. After a transient improvement the haemarthrosis became worse and an open evacuation of the joint had to be made under treatment with bolus injections of rFVIIa for 3 days (120 microg kg(-1) every 2 h). A previous pharmacokinetic evaluation in this patient had showed that FVIIa recovery and half-life were less than expected. Continuous infusion of rFVIIa (20 microg kg(-1) h(-1)), with added low molecular heparin to prevent local thrombophlebitis, was started on the fourth postoperative day and maintained unchanged for 7 days. Four additional single bolus injections were given for early joint mobilization. The intervals between replacements of the pump syringes were progressively increased from 6 to 12 h and then up to 24 h. FVIIa plasma levels during continuous infusion ranged between 6.3 and 10.4 IU mL(-1). Although +FVIIa assays seemed to show good stability, we observed the formation of precipitates inside the syringes. The precipitates seemed to contain FVIIa. We concluded that FVIIa+ plasma levels of 6-10 IU mL(-1) were safe and effective to prevent postoperative haemorrhage in this patient. The addition of heparin to the rFVIIa concentrates, however, may cause precipitation and should be avoided. Individual pharmacokinetic evaluation may be useful to select the appropriate initial doses, especially in young patients.

Search for mutations in a segment of the exon 28 of the human von Willebrand factor gene: new mutations, R1315C and R1341W, associated with type 2M and 2B variants.

von Willebrand Disease (vWD) is the most frequently inherited bleeding disorder in humans, and is caused by a qualitative and/or quantitative abnormality of the von Willebrand factor (vWF). A large number of defects that cause qualitative variants have been located in the A1 domain of the vWF, which contains sites for interaction with platelet glycoprotein Ib (GPIb). We have developed a new approach to detect mutations based on DdeI digestion and single-strand conformation polymorphism analysis. A segment of 487 nucleotides, extending from intron 27 to codon 1368 of the pre-pro vWF was amplified from genomic DNA. The cleavage with DdeI yields two fragments of appropriate size for this kind of analysis and confirms that the gene, rather than the pseudogene, is being investigated. Six families with type 2B vWD, one type 2M vWD family, and one another type 2A vWD family were studied. After sequencing the fragments with an altered electrophoretic pattern, we found four mutations previously described--R1308C, V1316M, P1337L, and R1306W--in patients with 2B vWD. The last one arose de novo in the patient. In addition, two new candidate mutations were observed: R1315C and R1341W. The first one was associated to type 2M vWD, whereas the one second cosegregated with type 2B vWD. The fact that these new mutations were not found in 100 normal alleles screened further supports their causal relationship with the disease. These mutations, which induce either a gain or a loss of function, further show an important regulatory role of this region in the binding of vWF to GPIb and its implications in causing disease.

Zero incidence of inhibitor development in previously treated haemophilia A, HIV-negative patients upon exposure to a plasma-derived high-purity and double viral inactivated factor VIII concentrate.

Thirty-six haemophilia A, HIV-negative, previously treated patients were changed therapy to a highpurity and double-inactivated (solvent/detergent and dry-heating) previously unused factor VIII concentrate. The mean age of these patients was 27 years at the time of the change. Twenty-three patients were severe haemophiliacs (FVIII:C < 0.02 IU mL-1), seven moderate (FVIII:C between 0.02 and 0.05 IU mL-1) and six mild (FVIII:C > 0.05 IU mL-1). The mean follow-up with this single product was 16 months, with 82 accumulated exposure days and the mean consumption was 117,300 IU of FVIII corresponding to a mean of six batches per patient. No patient developed FVIII inhibitors (upper limit of the CI95: 7.98%), resulting in an incidence rate of 0/48 patient-years (upper limit of the CI95: 77/1000 patient-years). The change in therapy to this new factor VIII concentrate was not associated with the appearance of inhibitors.

Development of a factor VIII inhibitor in a newborn haemophiliac.

A major problem in the treatment of haemophilia A is the development of inhibitors (antibodies) against factor VIII. We report the case of a newborn male with no family history of haemophilia who developed an intracerebral haemorrhage. On day 10 post-delivery severe haemophilia A was diagnosed and treatment with recombinant FVIII (rFVIII) concentrate was started. Seventy-two hours later the presence of inhibitors was suspected because high doses of rFVIII were required to maintain therapeutic FVIII plasma levels. Days after, the inhibitor was detected. The quick detection of the inhibitor in this newborn haemophiliac allowed us to start the immunotolerance early, without interruption in the administration of rFVIII.

[Seroprevalence of hepatitis A in hemophiliacs]. / Seroprevalencia de la hepatitis A en hemofílicos.

PURPOSE: To study the seroprevalence of hepatitis A virus (HAV) infection in haemophiliacs treated with factor VIII/IX concentrates. PATIENTS AND METHODS: Anti-HAV IgG antibodies were tested in 133 haemophiliacs previously treated (20 of them only infused with virus-inactivated factor concentrates), 11 previously untreated haemophiliacs and 60 healthy individuals (> 25 yr. old). RESULTS: The overall anti-HAV prevalence was 43%. Anti-HAV was found in 2 (10%) of the patients treated only with virus-inactivated concentrates and in 55 (49%) of those who had received non-inactivated concentrates. The seroprevalence in the untreated haemophiliacs was 27% and 90% in the healthy control group. The anti-HAV seroprevalence showed a significant (p < 0.001) dependence on patient age, it being higher in patients aged > 25 (77%) than in those aged 10-25 (31%) and < 10 (4%). The seroprevalence of anti-HAV was lower in the treated haemophiliacs aged 25 or more than in the healthy individuals, although the difference did not reach statistical significance (p = 0.06). CONCLUSION: These results show that the seroprevalence of HAV infection in haemophiliacs is similar to that in the general population, and that there is not a significant excess of HAV infections amongst haemophiliacs with high exposure to coagulation factor concentrates.