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[Classification of malignant lymphomas--current status and clinical implications].

New techniques such as cytogenetics, molecular biology and immunology have provided new insights into the pathogenesis and development of malignant lymphomas. By this way they provided a new basis for a classification of lymphoid malignancies which is increasingly oriented on biology and pathogenesis. This process resulted in the 'Revised European-American Lymphoma' (R.E.A.L.) classification, which was recently proposed by an international group of pathologists. This classification comprises a preliminary end point of a continuing process which will require a reevaluation and adaptation to new knowledge in the future again. In order to facilitate the clinical use of this histopathologically and biologically oriented classification, a proposal for a clinical grouping was developed, which divides the R.E.A.L. lymphoma subtypes into the following categories: I. Indolent lymphomas (low risk group). II. Aggressive lymphomas (intermediate risk group). III. Very aggressive lymphomas (high risk group). IV. Morbus Hodgkin. This internationally developed consensus of a histopathologic classification and a clinical grouping is a substantial step forward towards an international cooperation in the management of malignant lymphomas. It is background for cooperative studies and justifies the hope that further advances will be achieved in the treatment of malignant lymphomas and that more effective strategies can be developed in the near future.

Cytogenetics↗

Genetic diversity and high proportion of intersubtype recombinants among HIV type 1-infected pregnant women in Kisumu, western Kenya.

The high genetic diversity of HIV-1 continues to complicate effective vaccine development. To better understand the extent of genetic diversity, intersubtype recombinants and their relative contribution to the HIV epidemic in Kenya, we undertook a detailed molecular epidemiological investigation on HIV-1-infected women attending an antenatal clinic in Kisumu, Kenya. Analysis of gag-p24 region from 460 specimens indicated that 310 (67.4%) were A, 94 (20.4%) were D, 28 (6.1%) were C, 9 (2.0%) were A2, 8 (1.7%) were G, and 11 (2.4%) were unclassifiable. Analysis of the env -gp41 region revealed that 326 (70.9%) were A, 85 (18.5%) D, 26 (5.7%) C, 9 (2.0%) each of A2 and G, 4(0.9%) unclassifiable, and 1 (0.2%) CRF02_AG. Parallel analyses of the gag-p24 and env-gp41 regions indicated that 344 (74.8%) were concordant subtypes, while the remaining 116 (25.2%) were discordant subtypes. The most common discordant subtypes were D/A (40, 8.7%), A/D (27, 5.9%), C/A (11, 2.4%), and A/C (8, 1.7%). Further analysis of a 2.1-kb fragment spanning the gag-pol region from 38 selected specimens revealed that 19 were intersubtype recombinants and majority of them were unique recombinant forms. Distribution of concordant and discordant subtypes remained fairly stable over the 4-year period (1996-2000) studied. Comparison of amino acid sequences of gag-p24 and env-gp41 regions with the subtype A consensus sequence or Kenyan candidate vaccine antigen (HIVA) revealed minor variations in the immunodominant epitopes. These data provide further evidence of high genetic diversity, with subtype A as the predominant subtype and a high proportion of intersubtype recombinants in Kenya.

AIDS Serodiagnosis↗

Desmocollins form a distinct subset of the cadherin family of cell adhesion molecules.

The desmosomal adhesive core is formed by four major components: desmoglein (Mr, 165,000), desmocollins I and II (Mr, 120,000 and 110,000, respectively), and a Mr 22,000 protein. Here, we report the cloning and sequencing of cDNAs encoding a bovine desmocollin. The open reading frame found in the longest cDNA, 5 kilobases, contains a region encoding a protein of 839 amino acids. The features of the deduced amino acid sequence imply that the mature 707-amino acid desmocollin is a type I transmembrane protein that is produced by proteolytic cleavage of an 810-amino acid precursor. The ectodomain of desmocollin contains repeats that show extensive sequence similarity to members of the cadherin family of calcium-dependent cell adhesion molecules. A comparison of the amino acid sequences of desmocollin, desmoglein, and the cadherins shows that although these intercellular junctional adhesion molecules share a consensus sequence in their adhesive domains that defines them as a family, several features, including the divergence in the sequence of their cytoplasmic tails, divide them into three distinct subtypes.

Amino Acid Sequence↗

Characterization and purification of H1TF2, a novel CCAAT-binding protein that interacts with a histone H1 subtype-specific consensus element.

Definition of mechanisms regulating human histone H1 gene transcription during the cell cycle requires the isolation and biochemical characterization of protein factors which interact with specific promoter elements. Two distinct binding activities have been identified in nuclear extracts from HeLa cells and mapped within a 180-base-pair (bp) region of a cell cycle-regulated H1 gene promoter. H1TF1 bound to an H1-specific A + C-rich sequence (AC box), 100 bp upstream of the cap site; H1TF2 interacted with the H1 subtype-specific consensus element and was dependent on the presence of an intact CCAAT box for binding. H1TF2 was purified through a combination of ion-exchange and oligonucleotide affinity chromatographies. Analysis of purified fractions by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and UV crosslinking showed that H1TF2 was a single polypeptide of 47 kilodaltons. This factor was distinct from previously characterized CCAAT-binding proteins in both molecular size and binding properties. Fractions containing H1TF2 activity activated transcription in vitro only if programmed with an H1 DNA template carrying an intact H1TF2-binding site.

Binding Sites↗

Hodgkin's lymphoma: the pathologist's viewpoint.

Despite its well known histological and clinical features, Hodgkin's lymphoma (HL) has recently been the object of intense research activity, leading to a better understanding of its phenotype, molecular characteristics, histogenesis, and possible mechanisms of lymphomagenesis. There is complete consensus on the B cell derivation of the tumour in most cases, and on the relevance of Epstein-Barr virus infection and defective cytokinesis in at least a proportion of patients. The REAL/WHO classification recognises a basic distinction between lymphocyte predominance HL (LP-HL) and classic HL (CHL), reflecting the differences in clinical presentation and behaviour, morphology, phenotype, and molecular features. CHL has been classified into four subtypes: lymphocyte rich, nodular sclerosing, with mixed cellularity, and lymphocyte depleted. The borders between CHL and anaplastic large cell lymphoma have become sharper, whereas those between LP-HL and T cell rich B cell lymphoma remain ill defined. Treatments adjusted to the pathobiological characteristics of the tumour in at risk patients have been proposed and are on the way to being applied.

Diagnosis, Differential↗

Partial nucleotide sequencing of six subtype 2c hepatitis C viruses detected in Italy.

The great majority of 121 hepatitis C virus (HCV) isolates obtained from 117 Italian patients with community-acquired infection could readily be typed by genotype-specific PCR. Subtype 1b was dominant (74 isolates); subtypes 2b, 2a, and 1a followed, with 19, 14, and 8 isolates, respectively. The six isolates that remained untyped by this method were classified as subtype 2c on the basis of sequence analysis of PCR amplicors obtained from the core and NS5 genes. These findings indicate that HCV subtype 2c has a relatively high prevalence in Italy. Sequencing the core region from positions 160 to 259 is sufficient to distinguish subtype 2c from other known HCV genotypes.

Adolescent↗

Peptide/benzodiazepine hybrids as ligands of CCK(A) and CCK(B) receptors.

The (neuro)hormones gastrin and cholecystokinin (CCK) share a common C-terminal tetrapeptide amide sequence that has been recognized as the message portion while the N-terminal extensions are responsible for the CCK(A) and CCK(B) receptor subtype selectivity and avidity. 1,4-Benzodiazepine derivatives are potent and selective antagonists of these receptors, and according to comparative molecular field analysis, the structures of these nonpeptidic compounds could well mimic the message sequence of the peptide agonists at least in terms of spatial array of the aromatic residues. Docking of a larger series of low molecular weight nonpeptide antagonists to a homology modeling derived CCK(B) receptor structure revealed a consensus binding mode that is further validated by data from site-directed mutagenesis studies of the receptors. Whether this putative binding pocket of the nonpeptide antagonists is identical to that of the message portion of the peptide agonists, or whether it is distinct and spatially separated, or overlapping, but with distinct interaction sites, is still object of debate. Using a 1,4-benzodiazepine core amino-functionalized at the C3 position, related tryptophanyl derivatives were synthesized as mimics of the tetrapeptide and subsequently extended N-terminally with gastrin and CCK address sequences. All hybrid constructs were recognized as antagonists by the CCK(A) and CCK(B) receptors, but their address portions were incapable of enhancing in significant manner selectivity and avidity. Consequently, the binding of the peptide/benzodiazepine hybrids has to be dictated mainly by the benzodiazepine moiety, which apparently prevents optimal interactions of the address peptides with extracellular receptor subdomains. These findings would strongly support the view of distinct binding sites for the message portion of the peptide agonists and the benzodiazepine-based nonpeptide antagonists.

Benzodiazepines↗

Evolution of the human immunodeficiency virus type 1 subtype-specific V3 domain is confined to a sequence space with a fixed distance to the subtype consensus.

Human immunodeficiency virus type 1 (HIV-1) strains can be separated into genetic subtypes based on phylogenetic analysis of the envelope gene. Once it had been shown that population-wide intrasubtype genetic variation of HIV-1 strains increases in the course of the AIDS epidemic, it remained uncertain whether HIV-1 subtypes are phenotypic entities spreading as distinct virus populations. To examine this, we applied Eigen's concepts of sequence geometry and fitness topography to the analysis of intrasubtype evolution of the gp120 V3 domain of HIV-1 subtypes A, B, C, and D in the course of the global AIDS epidemic. We observed that despite the high evolution rate of HIV-1, the nonsynonymous distances to the subtype consensus of sequences obtained early in the epidemic are similar to those obtained more than 10 years later, in contrast to the synonymous distances, which increased steadily over time. For HIV-1 subtype B, we observed that the evolution rate of the individual sequences is independent of their distance from the subtype B consensus, but for the individual sequences most distant from the consensus evolution away from the consensus is constrained. As a result, individual HIV-1 genomes fluctuate within a sequence space with fixed distance to the subtype consensus. Our findings suggest that the evolution of the V3 domain of HIV-1 subtypes A, B, C, and D is confined to an area in sequence space within a fixed distance to the consensus of a respective subtype. This in turn indicates that each HIV-1 subtype is a distinct viral quasispecies that is well adapted to the present environment, able to maintain its identity in the V3 region over time, and unlikely to merge during progression of the AIDS epidemic.

Consensus Sequence↗

In vivo genetic variability of the HIV-1 vif gene.

The human immunodeficiency virus type 1 (HIV-1) vif gene encodes a 23-kDa protein (viral infectivity factor) whose exact mechanism of action is not entirely clear. Vif is believed to be highly conserved among different HIV-1 strains. We have analyzed the proviral vif sequences of 61 peripheral blood mononuclear cell samples from HIV-1 positive patients by direct solid phase sequencing and temperature gradient gel electrophoresis of polymerase chain reaction products. Inter- and intraindividual sequence variations, conserved motifs, and prevalent vif subtypes were investigated. The consensus proviral vif DNA sequence of the 61 samples as well as the consensus sequence of the 61 deduced vif amino acid sequences were found to be less conserved than previously thought. The vif proviral sequences were 58% conserved, with the 5' end of the vif gene being the most conserved region (84%). Of the vif amino acids only 45% were absolutely conserved in the 61 samples, i.e., the absolutely conserved and as such possibly functionally important domains of the vif protein comprised less than half of the vif amino acids. In two-thirds of the variable positions residues belonging to different amino acid groups were found. In individual patients the prevalent vif sequence changed with the course of disease, but the differences found in two serial samples of a patient were < or = 10%. Phylogenetic analysis revealed that one African vif subtype had been introduced in the investigated population.

Amino Acid Sequence↗

Structural requirements for and consequences of an antiviral porphyrin binding to the V3 loop of the human immunodeficiency virus (HIV-1) envelope glycoprotein gp120.

Several porphyrin derivatives were reported to have anti-HIV-1 activity. Among them, meso-teta(4-carboxyphenyl)porphine (MTCPP) and other carboxyphenyl derivatives were the most potent inhibitors (EC50 <0.7 mu M). MTCPP bound to the HIV-1 envelope glycoprotein gp120 and to full-length V3 loop peptides corresponding to several HIV-1 isolates but not to other peptides from gp120 + gp41. However, it remained possible that MTCPP bound to regions on gp120 which cannot be mimicked by peptides. Further characterization of the binding domain for MTCPP is important for understanding the antiviral activity of porphyrins and for the design of anti-HIV-1 drugs interfering with functions of the virus envelope. Results presented here show that: (i) deletion of the V3 loop from the gp120 sequence resulted in drastically diminished MTCPP binding, suggesting that the V3 loop is the dominant if not the only target site on gp120; (ii) this site was only partially mimicked by full-length V3 loop peptides; (iii) MTCPP binding to the gp120 V3 loop elicited allosteric effects resulting in decreased accessibility of the CD4 receptor binding site; (iv) the binding site for MTCPP lies within the central portion of the V3 loop (KSIHIGPGRAFY for the HIV-1 subtype B consensus sequence) and does not involve directly the GPG apex of the loop. These results may help in designing antiviral compounds with improved activity.

Allosteric Regulation↗

HIV type 1 diversity in France, 1999-2001: molecular characterization of non-B HIV type 1 subtypes and potential impact on susceptibility to antiretroviral drugs.

Non-B HIV-1 samples collected in France between 1999 and 2001 were sequenced in the env, reverse transcriptase (RT), and protease genes (1) to characterize further the non-B strains circulating in the country, (2) to assess the importance of recombination, and (3) to describe the polymorphism of RT and protease genes and appreciate a possible impact on susceptibility to antiretroviral (ARV) drugs. The results show that, within a background of CRF02_AG predominance, there is a high genetic diversity of non-B isolates, including intersubtype recombinants. There is an extensive polymorphism of protease and RT genes compared with B consensus sequences; we have so far no data indicating that these non-B isolates may have reduced sensitivity to ARV drugs.

Adult↗

Role of alpha adrenergic receptors in erectile function.

Penile erection is a complex physiological process in which the regulation of penile hemodynamics depends on signal input from central and peripheral nervous systems, and on the balance and interplay between several local physiological mediators (neurotransmitters, vasoactive agents and endocrine factors). A role for the sympathetic nervous system in attaining or maintaining penile flaccidity and detumescence is well recognized. However, the exact mechanisms of alpha-adrenergic receptor mediated erectile function remain poorly defined. The objective of this review is to summarize data presented in the literature and from our laboratory on alpha-adrenergic receptors, and to discuss the conceptual framework by which the alpha-adrenergic receptor pathway modulates penile corpus cavernosum smooth muscle contractility. We will integrate the current state of knowledge of penile erection into one framework encompassing the biochemical and physiological pathways responsible for trabecular smooth muscle relaxation (erection) and contraction (detumescence). We will focus our discussion on local control mechanisms of alpha-adrenergic receptors and explore the following topics: (1) functional activity of alpha-1 and alpha-2 adrenergic receptors in the physiology of human penile erection; (2) identification, classification and characterization of alpha-1 and alpha-2 adrenergic receptor subtypes in human penile erectile tissue; (3) molecular mechanism of action of alpha-1 and alpha-2 adrenergic receptors in human penile erectile tissue; (4) blockade of alpha-1 and alpha-2 adrenergic receptor action by selective and non-selective alpha-1 and alpha-2 adrenergic receptor antagonists (blockers); (5) physiologic (functional) antagonism of alpha-1 and alpha-2 adrenergic receptor activity by vasodilators mediating smooth muscle relaxation; and (6) key areas of consensus, as well as critical gaps in the existing scientific knowledge concerning the rationale and the potential use of alpha-blockers in erectile function.

Adrenergic alpha-Antagonists↗

Role of alpha adrenergic receptors in erectile function.

Penile erection is a complex physiological process in which the regulation of penile hemodynamics depends on signal input from central and peripheral nervous systems, and on the balance and interplay between several local physiological mediators (neurotransmitters, vasoactive agents and endocrine factors). A role for the sympathetic nervous system in attaining or maintaining penile flaccidity and detumescence is well recognized. However, the exact mechanisms of alpha-adrenergic receptor mediated erectile function remain poorly defined. The objective of this review is to summarize data presented in the literature and from our laboratory on alpha-adrenergic receptors, and to discuss the conceptual framework by which the alpha-adrenergic receptor pathway modulates penile corpus cavernosum smooth muscle contractility. We will integrate the current state of knowledge of penile erection into one framework encompassing the biochemical and physiological pathways responsible for trabecular smooth muscle relaxation (erection) and contraction (detumescence). We will focus our discussion on local control mechanisms of alpha-adrenergic receptors and explore the following topics: (1) functional activity of alpha-1 and alpha-2 adrenergic receptors in the physiology of human penile erection; (2) identification, classification and characterization of alpha-1 and alpha-2 adrenergic receptor subtypes in human penile erectile tissue; (3) molecular mechanism of action of alpha-1 and alpha-2 adrenergic receptors in human penile erectile tissue; (4) blockade of alpha-1 and alpha-2 adrenergic receptor action by selective and non-selective alpha-1 and alpha-2 adrenergic receptor antagonists (blockers); (5) physiologic (functional) antagonism of alpha-1 and alpha-2 adrenergic receptor activity by vasodilators mediating smooth muscle relaxation; and (6) key areas of consensus, as well as critical gaps in the existing scientific knowledge concerning the rationale and the potential use of alpha-blockers in erectile function. International Journal of Impotence Research (2000) 12, Suppl 1, S48-63

Journal Article↗

Advances in the diagnosis and classification of B-ALL: comparative insights from updated guidelines.

Accurate molecular classification is essential for diagnosis, risk stratification, and treatment selection in B-cell lymphoblastic leukemia (B-ALL). In this study, we performed a comprehensive, real-world reclassification of 1015 consecutively diagnosed B-ALL patients using the fifth edition of the World Health Organization Classification of Haematolymphoid Tumours (WHO-HAEM5) and the International Consensus Classification (ICC). An integrative genomic strategy that combined whole transcriptome sequencing, fusion detection, mutational analysis, and cytogenetics enabled reclassification according to both the WHO-HAEM5 and ICC frameworks, thereby substantially reducing the proportion of unclassifiable B-ALL from 41.9% (2016 WHO revision [WHO-HAEM4R]) to 15.9% (WHO-HAEM5) and 11.9% (ICC). Distinct clinical and prognostic features were identified across newly defined subtypes. Multivariable analysis confirmed that this genomic classification is a robust, independent predictor of survival after adjusting for age, minimal residual disease status, and transplant intervention. Specifically, HLF-rearranged and MEF2D-rearranged B-ALL conferred a persistently poor prognosis across all age groups despite allogeneic hematopoietic stem cell transplantation, highlighting an urgent need for novel therapeutic strategies. Gene expression profiling resolved cryptic subtypes, including ETV6::RUNX1-like, ZNF384-rearranged-like, and BCR::ABL1-like B-ALL, and uncovered diagnostic ambiguity in patients with concurrent lesions. In addition, we report emerging high-risk groups, including IDH1/2- and ZEB2 Q1072-mutated B-ALL, that may warrant recognition as distinct molecular entities. Our findings demonstrate the clinical use of integrative transcriptomic profiling in refining B-ALL taxonomy in guiding risk-adapted therapies and informing future revisions of diagnostic standards. This study supports the incorporation of high-throughput molecular diagnostics into routine leukemia classification and precision treatment planning.

Humans↗

Sodium channel mRNA in the B104 neuroblastoma cell line.

B104 neuroblastoma cells are excitable, but the ion channels underlying electrogenesis in these cells have not been identified. RT-PCR, restriction enzyme analysis and in situ hybridization were used to study sodium channel mRNAs in B104 cells. High levels of sodium channel alpha-subunit mRNAs III, NaG and Na6 and beta 1-subunit mRNA were detected by RT-PCR in B104 cells. Low levels of types I and II alpha-subunit mRNAs were also present. In situ hybridization with subtype-specific riboprobes detected sodium channel alpha-subunit mRNAs III, NaG and Na6 and beta 1-subunit mRNA in B104 cells; analysis of the percentage of B104 cells expressing each alpha-subunit mRNA subtype suggests that some cells express the mRNAs for several alpha-subunits.

Actins↗

Envelope sequence variability and serologic characterization of HIV type 1 group O isolates from equatorial guinea.

Four sera from Equatorial Guinea (EG) suspected to contain antibody against HIV-1 group O-related viruses were identified on the basis of unusual and differential serologic reactivity in selected commercial assays and Western blot. Degenerate primers, designed from HIV-1 group O published sequences, were used to PCR amplify envelope (env) gene sequences from the suspect EG sera. A complete envelope gene sequence from each serum was determined from the overlapping env gene fragments. Analysis (PHYLIP package of programs) of Env amino acid sequences (translated from nucleotide sequences) indicated that the amino acid sequences obtained from EG sera clustered more closely with HIV Env sequences of group O compared to group M. The amino acid sequences at the octameric tip of the V3 loop were either RIGPLAWY (one isolate), RIGPMAWY (two isolates), or GLGPLAVY (one isolate). The V3 tip tetrameric sequence GPLA is represented only once in the 1995 HIV (Los Alamos) database, but was present in two of our group O-related EG samples. The gp41 immunodominant regions (IDR) protein sequences were identical for sequences from three of the sera, RLLALETLIQNQQLLNLWGCKGR(K)L(I)VCYTSVK(T)W, whereas sequence from the fourth serum contained three changes as noted in parentheses. IDR sequences derived from EG sera were unique compared to those reported for other HIV-1 group O isolate ANT70, VAU, or MVP5180. Antibody in each EG serum directed against the IDR could be detected using synthetic peptides comprising sequences from the ANT70 or MVP5180 IDRs, but were most reactive against the sequences derived from the samples themselves. Little or no serologic reactivity was detected when EG sera were reacted against peptides comprising the IDR of HIV-1 group M (subtype B consensus) or HIV-2 (consensus).

Amino Acid Sequence↗