Cytogenetic and histologic correlations in malignant lymphoma

Although a number of studies have indicated correlations between histologic subtypes of tumors and certain nonrandom chromosome changes, cytogenetic studies of lymphoma are in an early stage compared to those of leukemia. No comprehensive analysis of available data has so far been attempted in the literature either. Here we present an analysis of chromosome changes and their correlation with subtypes of lymphoma studied by conventional histology and cell surface markers, as observed in two sets of data: a group of 65 karyotypically abnormal tumors sequentially ascertained and studied by us during the period January 1, 1984 to April 30, 1985, and a larger data set derived by combining our data with those from two published series from the University of Minnesota that are comparable to our data. These combined data, which comprise the largest data set on the cytogenetics of lymphomas assembled so far, enabled a comprehensive analysis of correlation between chromosome change and tumor histology and the patterns of chromosome instability in these tumors. We found several significant associations, some previously described and others now recognized, between nonrandom chromosome gains, breaks, translocations, and deletions and histologic subtypes of tumors that characterize lymphomas. The data indicate that finding of chromosome breaks at certain sites (eg, 8q24, 14q32, 18q21) is of diagnostic value in dealing with cases of unusual lymphoma. Furthermore, nonrandom chromosome breakage exhibited three distinct patterns that reflected three levels of etiologically relevant genetic change.     

High‐Fiber Orange Juice as a Nutrition Supplement in Women

Background: The daily consumption of dietary fiber is frequently below suggested recommendations. Using a double‐blind, controlled, randomized study, we assessed the efficiency and tolerance of a fiber‐enriched orange juice to supplement fiber intake in women. Materials and Methods: After 1 week of noninterventional observation, 192 healthy adult women ingested 400 mL of orange juice for 21 days, which either was not (placebo group) or was enriched with fiber (fiber group). Orange juice ingestion was registered daily and controlled for each week during the study period. Macronutrient, fiber, and energy intake were determined using a 3‐day food record, validated food chemical composition databases, and the “Pro Diet” software. Gastrointestinal symptoms were self‐evaluated daily by scoring 4 grades of symptom intensity and using a visual analog scale to grade pain severity. Results: No changes were observed for macronutrient and energy ingestion. For the placebo group (n = 97), the total fiber intake record was under the daily recommended value. In contrast, the fiber group (n = 95) displayed higher comparative values of total and soluble fiber consumption (P ≤ .001), achieving the daily recommended values of fiber intake. Both groups reported an increased frequency of slight bloating and rumbles over time (P ≤ .05). The fiber group also experienced a higher frequency of slight flatulence over time (P = .002). Conclusion: Consumption of fiber‐enriched orange juice was efficient to achieve the daily fiber intake recommendation for women, was not accompanied by intense adverse events, and may represent a suitable method to supplement fiber intake in woman.     

From the Cover: Structural basis for the mutual antagonism of cAMP and TRIP8b in regulating HCN channel function

cAMP signaling in the brain mediates several higher order neural processes. Hyperpolarization-activated cyclic nucleotide-gated (HCN) channels directly bind cAMP through their cytoplasmic cyclic nucleotide binding domain (CNBD), thus playing a unique role in brain function. Neuronal HCN channels are also regulated by tetratricopeptide repeat-containing Rab8b interacting protein (TRIP8b), an auxiliary subunit that antagonizes the effects of cAMP by interacting with the channel CNBD. To unravel the molecular mechanisms underlying the dual regulation of HCN channel activity by cAMP/TRIP8b, we determined the NMR solution structure of the HCN2 channel CNBD in the cAMP-free form and mapped on it the TRIP8b interaction site. We reconstruct here the full conformational changes induced by cAMP binding to the HCN channel CNBD. Our results show that TRIP8b does not compete with cAMP for the same binding region; rather, it exerts its inhibitory action through an allosteric mechanism, preventing the cAMP-induced conformational changes in the HCN channel CNBD.Hyperpolarization-activated cyclic nucleotide-gated (HCN1–4) channels are the molecular determinants of the h-current (I_h), which regulates critical neuronal properties, including membrane resting potential, dendritic excitability, and intrinsic rhythmicity (1). HCN channels are dually regulated by voltage and by binding of cAMP to the cyclic nucleotide binding domain (CNBD), which is found on the cytoplasmic C-terminal tail of the channel. The CNBD exerts a tonic inhibition on the channel pore, with the opening transition of the channel being allosterically coupled to the conformational changes in the CNBD induced by cAMP binding (2). Thus, the closed-to-open transition of the channel is thought to reflect the transition from the cAMP-free conformation to the cAMP-bound conformation of the CNBD, which stabilize, respectively, the closed and open states of the channel (2, 3). The C-linker, an α-helical folded domain that connects the CNBD to the pore region, conveys the regulation of channel gating from the CNBD to the pore (4–6). As a result of this allosteric mechanism, the binding of cAMP shifts the voltage dependence of the HCN channel opening to more positive potentials and increases maximal I_h at extreme negative voltages, where voltage gating is complete.In addition to cAMP, HCN channels in the brain are regulated by auxiliary proteins, such as TRIP8b, a cytosolic β-subunit of neuronal HCN channels, which inhibits channel activation by antagonizing the effects of cAMP (7–9). We have previously shown that TRIP8b_core, an 80-aa sequence located in the TRIP8b protein core that directly interacts with the C-linker/CNBD region of HCN channels, is necessary and sufficient to prevent all of the effects of cAMP on the channel (10, 11). TRIP8b_core decreases both the sensitivity of the channel to cAMP [half maximal concentration (k_1/2)] and the efficacy of cAMP in inducing channel opening [half activation voltage (V_1/2)]; conversely, cAMP binding inhibits these actions of TRIP8b. These mutually antagonistic effects are well described by a cyclic allosteric model in which TRIP8b binding reduces the affinity of the channel for cAMP, with the affinity of the open state for cAMP being reduced to a greater extent than the cAMP affinity of the closed state (11).A second important action of TRIP8b is to reduce maximal current through HCN channels in the absence of cAMP (11). As a consequence, application of cAMP produces a larger increase in maximal I_h in the presence of TRIP8b than in its absence. The observation that TRIP8b exerts opposing influences on the two major actions of cAMP on HCN channel function, namely, reduces the effect of cAMP to shift the voltage dependence of channel gating but enhances the effect of cAMP to increase maximal current, has important implications for the ability of cAMP to modulate neuronal excitability in vivo. Thus, the relative extent by which neuromodulatory transmitters alter maximal I_h or shift the voltage dependence of HCN channel gating can vary widely among distinct classes of neurons (12–14). The differential expression of TRIP8b may provide a mechanistic explanation for this finding, because in neurons with high levels of TRIP8b expression, cAMP will exert a larger action to enhance maximal current, and a smaller action to alter the voltage dependence of channel gating, compared with neurons in which TRIP8b expression is low. Such fine-tuning broadens the range of physiological actions that cAMP can exert to modulate neuronal firing.In the present study, we address the structural basis for the mutually antagonistic effects of cAMP and TRIP8b on HCN channel function. Although our previous biochemical and electrophysiological data strongly support the hypothesis that TRIP8b and cAMP binding sites do not overlap, direct structural information on the TRIP8b–CNBD complex is required to validate the allosteric antagonism model of interaction between the two ligands. A plausible hypothesis for the antagonistic effect of TRIP8b and cAMP is that each of the two ligands stabilizes the CNBD in a conformation that decreases the affinity for the other. To test this hypothesis, we first generated the 3D structure of the cAMP-free HCN2 channel CNBD using solution NMR spectroscopy and then characterized its interaction with the TRIP8b_core fragment. By comparing the cAMP-free with the available cAMP-bound HCN2 channel CNBD structure (15, 16), we reconstruct the full conformational changes induced by cAMP binding, revealing critical transitions occurring in the P- and C-helices of the CNBD, and further highlighting the role of the N-terminal helical bundle in transducing the movements of the CNBD to the channel pore. We next identify, through NMR titration, site-directed mutagenesis, and biochemical interaction assays, the binding site of TRIP8b_core on the cAMP-free form of the HCN2 channel CNBD. Our results demonstrate that cAMP and TRIP8b do not directly compete for the same binding region and support a model of mutual allosteric inhibition between cAMP and TRIP8b. Finally, our results clarify the mechanism by which TRIP8b antagonizes the effect of cAMP on channel gating: TRIP8b directly interacts with two mobile elements that drive the ligand-induced conformational changes in the CNBD. TRIP8b binding to the CNBD therefore prevents the cAMP-induced transition and stabilizes the channel in the cAMP-free conformation.     

Prediction of clinically relevant drug‐induced liver injury from structure using machine learning

Drug‐induced liver injury (DILI) is the most common cause of acute liver failure and often responsible for drug withdrawals from the market. Clinical manifestations vary, and toxicity may or may not appear dose‐dependent. We present several machine‐learning models (decision tree induction, k‐nearest neighbor, support vector machines, artificial neural networks) for the prediction of clinically relevant DILI based solely on drug structure, with data taken from published DILI cases. Our models achieved corrected classification rates of up to 89%. We also studied the association of a drug's interaction with carriers, enzymes and transporters, and the relationship of defined daily doses with hepatotoxicity. The results presented here are useful as a screening tool both in a clinical setting in the assessment of DILI as well as in the early stages of drug development to rule out potentially hepatotoxic candidates.