Assumptions and Validations of Statistical Tests for Functional Neuroimaging

We contrast two statistical methods: three-dimensional cluster analysis and statistical parametric mapping. We show that three-dimensional cluster analysis is based on a neurobiological theory of the regulation of blood flow and, unlike statistical parametric mapping, carries a minimum of assumptions that are tested. Statistical parametric mapping is a formal approach, which is based on a multitude of assumptions of which the majority have not been validated. We also demonstrate that in practice three-dimensional cluster analysis has a reasonable balance between sensitivity and the probability of false positives, giving high reproducibility with data on e.g. colour discrimination.     

Modulation by a moving texture of cat area 18 neuron responses to moving bars

1. The influence of a moving texture on neuronal responses to a moving bar was tested in 103 area 18 neurons of anesthetized and paralyzed cats. The texture was a two-dimensional noise pattern, the bar moved at optimal speed, and its contrast was adjusted to yield 50% of the maximum response. 2. The moving texture exerted two different but related effects: it suppressed the response of area 18 neurons to the moving bar, and it modulated the direction selectivity of parastriate neurons. These effects were strongest when the texture moved at the same speed or faster than the bar. 3. Genuine suppressive effects of the moving texture were distinguished from lack of summation between bar and texture responses. Suppressive effects of either type were observed in 75% of the area 18 cells and occurred more frequently among C family cells, velocity tuned cells, and in layer 5 than in other groups of cells. 4. The modulation of direction selectivity was distinguished from pseudomodulation because of lack of summation of bar and texture responses. The direction selectivity of 35% of the area 18 cells was modulated by the moving texture. Six different relative direction selectivity (RDS) types were observed in area 18. 5. The neurons of which direction selectivity was modulated by the moving texture occurred predominantly in layers 2-3 and 6, suggesting that they represent a further stage of processing within area 18. 6. Many (75%) area 18 cells responded to the texture moving on its own. Most of these cells respond to isolated features ("grains") in the patterns rather than to the movement of the whole pattern. Cells responding to the movement of the whole pattern were generally C family cells, and their direction selectivity was not modulated by the moving texture. 7. These results are compared with those obtained under identical experimental conditions in area 17. Although suppressive effects are similar in both areas, RDS types are differently distributed in the two areas. 8. The possible origins of the interactions and their functional significance are discussed.     

Real-life experience with switching TNF-α inhibitors in ankylosing spondylitis

Background

Published economic evaluations of trastuzumab for the treatment of HER2-positive metastatic breast cancer have arrived at different conclusions regarding the cost-effectiveness of trastuzumab, despite comparative efficacy being demonstrated by a small set of randomised controlled trials (RCTs).

Objectives

This article aims to provide insight into the quality of the evaluations and explore the possible drivers of the conflicting conclusions.

Methods

A systematic literature review was conducted to identify all published economic evaluations that compared the incremental costs and outcomes of trastuzumab versus a comparator.

Results

Fifteen economic evaluations were identified. In the evaluations that estimated efficacy using an RCT, the key drivers of the conclusions regarding cost-effectiveness were: the approach used to estimate overall survival in the control group given crossover to trastuzumab following progression in the trials; the inclusion of treatment beyond progression; inclusion of wastage due to unused vial portions, adverse events, and the cost of HER2 testing. Four evaluations used non-randomised approaches to estimate efficacy, thus introducing the potential for confounding. As a result these evaluations reported relatively optimistic estimates of comparative effectiveness. Finally the evaluations used different thresholds to determine whether treatment with trastuzumab was cost-effective.

Conclusion

There were numerous drivers of the different conclusions regarding the cost-effectiveness of trastuzumab, many of which are due to judgements made by the authors when translating data from RCTs. Many of the potential drivers were not identified by the published systematic reviews of economic evaluations and perhaps more remain unidentified because of inconsistent and limited reporting.     

Potent and long-acting corticotropin releasing factor (CRF) receptor 2 selective peptide competitive antagonists

We present evidence that members of the corticotropin releasing factor (CRF) family assume distinct structures when interacting with the CRF(1) and CRF(2) receptors. Predictive methods, physicochemical measurements, and structure-activity relationship studies have suggested that CRF, its family members, and competitive antagonists such as astressin [cyclo(30-33)[DPhe(12),Nle(21),Glu(30),Lys(33),Nle(38)]hCRF((12-41))] assume an alpha-helical conformation when interacting with their receptors. We had shown that alpha-helical CRF((9-41)) and sauvagine showed some selectivity for CRF receptors other than that responsible for ACTH secretion(1) and later for CRF2.(2) More recently, we suggested the possibility of a helix-turn-helix motif around a turn encompassing residues 30-33(3) that would confer high affinity for both CRF(1) and CRF(2)(2,4) in agonists and antagonists of all members of the CRF family.(3) On the other hand, the substitutions that conferred ca. 100-fold CRF(2) selectivity to the antagonist antisauvagine-30 [[DPhe(11),His(12)]sauvagine((11-40))] did not confer such property to the corresponding N-terminally extended agonists. We find here that a Glu(32)-Lys(35) side chain to side chain covalent lactam constraint in hCRF and the corresponding Glu(31)-Lys(34) side chain to side chain covalent lactam constraint in sauvagine yield potent ligands that are selective for CRF(2). Additionally, we introduced deletions and substitutions known to increase duration of action to yield antagonists such as cyclo(31-34)[DPhe(11),His(12),C(alpha)MeLeu(13,39),Nle(17),Glu(31),Lys(34)]Ac-sauvagine((8-40)) (astressin(2)-B) with CRF(2) selectivities greater than 100-fold. CRF receptor autoradiography was performed in rat tissue known to express CRF(2) and CRF(1) in order to confirm that astressin(2)-B could indeed bind to established CRF(2) but not CRF(1) receptor-expressing tissues. Extended duration of action of astressin(2)-B vs that of antisauvagine-30 is demonstrated in the CRF(2)-mediated animal model whereby the inhibition of gastric emptying of a solid meal in mice by urocortin administered intraperitoneally at time zero is antagonized by the administration of astressin(2)-B but not by antisauvagine-30 at times -3 and -6 h while both peptides are effective when given 10 min before urocortin.     

Segregation of two endocannabinoid-hydrolyzing enzymes into pre- and postsynaptic compartments in the rat hippocampus, cerebellum and amygdala

Fatty acid amide hydrolase (FAAH) and monoglyceride lipase (MGL) catalyse the hydrolysis of the endocannabinoids anandamide and 2-arachidonoyl glycerol. We investigated their ultrastructural distribution in brain areas where the localization and effects of cannabinoid receptor activation are known. In the hippocampus, FAAH was present in somata and dendrites of principal cells, but not in interneurons. It was located mostly on the membrane surface of intracellular organelles known to store Ca(2+) (e.g. mitochondria, smooth endoplasmic reticulum), less frequently on the somatic or dendritic plasma membrane. MGL immunoreactivity was found in axon terminals of granule cells, CA3 pyramidal cells and some interneurons. In the cerebellum, Purkinje cells and their dendrites are intensively immunoreactive for FAAH, together with a sparse axon plexus at the border of the Purkinje cell/granule cell layers. Immunostaining for MGL was complementary, the axons in the molecular layer were intensively labelled leaving the Purkinje cell dendrites blank. FAAH distribution in the amygdala was similar to that of the CB(1) cannabinoid receptor: evident signal in neuronal somata and proximal dendrites in the basolateral nucleus, and hardly any labelling in the central nucleus. MGL staining was restricted to axons in the neuropil, with similar relative signal intensities seen for FAAH in different nuclei. Thus, FAAH is primarily a postsynaptic enzyme, whereas MGL is presynaptic. FAAH is associated with membranes of cytoplasmic organelles. The differential compartmentalization of the two enzymes suggests that anandamide and 2-AG signalling may subserve functional roles that are spatially segregated at least at the stage of metabolism.     

Potent somatostatin undecapeptide agonists selective for somatostatin receptor 1 (sst1)

A family of analogues of des-AA(1,2,5)-[DTrp(8)/D2Nal(8)]-SRIF that contain a 4-(N-isopropyl)-aminomethylphenylalanine (IAmp) at position 9 was identified that has high affinity and selectivity for human somatostatin receptor subtype 1 (sst1). The binding affinities of des-AA(1,2,5)-[DTrp(8),IAmp(9)]-SRIF (c[H-Cys-Lys-Phe-Phe-DTrp-IAmp-Thr-Phe-Thr-Ser-Cys-OH], CH-275) (7), des-AA(1,5)-[Tyr(2),DTrp(8),IAmp(9)]-SRIF (CH-288) (16), des-AA(1,2,5)-[Tyr(7),DTrp(8),IAmp(9)]-SRIF (23), and des-AA(1,2,5)-[DTrp(8),IAmp(9),Tyr(11)]-SRIF (25) are about (1)/(7), (1)/(4), (1)/(125), and (1)/(4) that of SRIF-28 (1) to sst1, respectively, about (1)/(65), (1)/(130), <(1)/(1000), and <(1)/(150) that of 1 to sst3, respectively, and about or less than (1)/(1000) that of 1 to the other three human SRIF receptor subtypes. A substitution of DTrp(8) by D2Nal(8) in 7 to yield des-AA(1,2,5)-[D2Nal(8),IAmp(9)]-SRIF (13) and in 16 to yield des-AA(1,5)-[Tyr(2),D2Nal(8),IAmp(9)]-SRIF (17) was intended to increase chemical stability, selectivity, and affinity and resulted in two analogues that were less potent or equipotent with similar selectivity, respectively. Carbamoylation of the N-terminus as in des-AA(1,2,5)-[DTrp(8),IAmp(9),Tyr(11)]-Cbm-SRIF (27) increased affinity slightly as well as improved selectivity. Monoiodination of 25 to yield 26 and of 27 to yield 28 resulted in an additional 4-fold increase in affinity at sst1. Desamination of the N-terminus of 17 to yield 18, on the other hand, resulted in significant loss of affinity. Attempts at reducing the size of the ring with maintenance of selectivity failed in that des-AA(1,4,5,13)-[Tyr(2),DTrp(8),IAmp(9)]-SRIF (33) and des-AA(1,4,5,6,12,13)-[Tyr(2),DTrp(8),IAmp(9)]-SRIF (34) progressively lost affinity for all receptors. Both des-AA(1,2,5)-[DTrp(8),IAmp(9),Tyr(11)]-Cbm-SRIF (27) and des-AA(1,2,5)-[DCys(3),DTrp(8),IAmp(9),Tyr(11)]-Cbm-SRIF (29) show agonistic activity in a cAMP assay; therefore, the structural basis for the agonist property of this family of analogues is not contingent upon the chirality of the Cys residue at position 3 as shown to be the case in 18-membered ring SRIF octapeptides. None of the high affinity structures described here showed receptor antagonism. We have prepared the radiolabeled des-AA(1,2,5)-[DTrp(8),IAmp(9),(125)ITyr(11)]-SRIF ((125)I-25) and des-AA(1,2,5)-[DTrp(8),IAmp(9), (125)ITyr(11)]-Cbm-SRIF ((125)I-27), used them as in vitro tracers, and found them to be superior to des-AA(1,5)-[(125)ITyr(2),DTrp(8),IAmp(9)]-SRIF ((125)I-16) for the detection of sst1 tumors in receptor autoradiography studies.     

Identification of mammalian sperm surface antigens: II. Characterization of an acrosomal cap protein and a tail protein using monoclonal anti-mouse sperm antibodies

Monoclonal anti-mouse sperm antibodies have been produced by fusing mouse myeloma cells with spleen cells from rats immunized with epididymal sperm of C3H mice. Immunoprecipitation and immunoperoxidase techniques showed that one such monoclonal antibody, AMS IV-33, recognized a 200 000 dalton protein localized on the acrosomal cap of the sperm cell. Two other monoclonal antibodies AMS IV-54 and -76, reacted with a 68 000 dalton component on the surface of the sperm tail. Both antigenic targets were species specific and were present in about equal amounts on sperm from several different strains of mice. The tail protein was sperm specific, whereas the antibody reacting with the acrosomal cap protein also appeared to react somewhat with antigens present in other mouse tissues.     

Influence of a moving textured background on direction selectivity of cat striate neurons

The influence of a moving textured background on direction selectivity for a moving bar was tested in 118 striate neurons and in 19 dorsal lateral geniculate neurons of anesthetized and paralyzed cats. In the standard conditions the background was a two-dimensional noise pattern, the bar moved at optimal speed, and its contrast was adjusted to the level producing 50% of the maximum response. These experiments revealed a new typology of cortical cells based on relative direction selectivity. Six different relative-direction-selectivity types are described. Two types of cells were found to have opposite kinds of relative direction selectivity: antiphase direction-selective cells (5% of the cortical sample) preferred the direction of the bar opposite to the direction of background motion, and absolutely direction-selective cells (20% of the cortical sample) kept their direction selectivity for bar motion independently of the background motion. Three types of cortical cells were direction selective for bar motion only in restricted background motion conditions: conditionally direction-selective cells (20% of cortical sample) only expressed their direction selectivity when the bar and the background moved in antiphase, differencing direction-selective cells (5% of the cortical sample) only expressed their direction selectivity when the bar and the background differed in speed, and limited direction-selective cells (20% of the cortical sample) only expressed their direction selectivity for near zero background speeds. The sixth type, relative nondirection-selective cells (30% of the cortical sample and all of the geniculate cells) were direction selective for none of the background motion conditions. These different relative-direction-selectivity types differed in RF organization, in ocular dominance, velocity sensitivity, in laminar distribution, and in distribution in the visual field. The relative-direction-selectivity types were invariant for changes in the contrast and bar speed. The construction of these relative-direction-selectivity types from the geniculate input requires some inhibitory, but mainly facilitatory, intracortical interactions. These experimental findings suggest that area 17 in the cat has the neuronal machinery to extract depth from motion (limited direction-selective cells) and to segregate visual scenes by motion cues (antiphase, conditionally and differencing direction-selective cells).     

Two <Emphasis Type="Italic">C</Emphasis>-Methyl Derivatives of [<Superscript>11</Superscript>C]WAY-100635 – Effects of an Amido <Emphasis Type="Bold">α</Emphasis>-Methyl Group on Metabolism and Brain 5-HT<Subscript>1A</Subscript> Receptor Radioligand Behavior in Monkey*

PURPOSE: [carbonyl-11C]N-(2-(1-(4-(2-methoxyphenyl)-piperazinyl)ethyl)-N-pyridinyl)cyclohexanecarboxamide ([carbonyl-11C]WAY-100635) is an effective radioligand for imaging brain 5-HT1A receptors with positron emission tomography (PET). However, this radioligand has some drawbacks for deriving relative regional receptor densities, including rapid metabolism, which acts against accurate definition of an arterial input function for compartmental modeling, and very low nonspecific binding in brain, which detracts from the accuracy of modeling by a simplified reference tissue (cerebellum) approach. Here, in a search for a radioligand that overcomes these limitations, we investigated the effects of introducing a single methyl group at either of the carbon atoms alpha to the amide bond in [11C]WAY-100635. PROCEDURES: Ligands with a methyl group on the alpha carbon of the cyclohexyl group (SWAY) or the alpha carbon of the C2H4 linker ((R,S)-JWAY) were synthesized and tested for binding affinity and intrinsic activity at 5-HT1A receptors. SWAY was labeled with carbon-11 (t1/2 = 20.4 minutes; beta+ = 99.8%) in its O-methyl group and (R,S)-JWAY in its carbonyl group. Each radioligand was evaluated by PET experiments in cynomolgus monkey. RESULTS: SWAY and (R,S)-JWAY were found to be high-affinity antagonists at 5-HT1A receptors. After injection of [11C]SWAY into monkey, radioactivity uptake in brain reached a maximum of 3% at 4.5 minutes and decreased to 0.7% at 72 minutes. However, over the time span of the experiment, radioactivity concentrations in 5-HT1A receptor-rich brain regions were only fractionally higher than in cerebellum. Radioactivity represented by parent radioligand in plasma was 39% at 45 minutes. After injection of [11C](R,S)-JWAY alone, radioactivity uptake in brain reached a maximum of 4.8% at 2.5 minutes and decreased to 1.2% at 90 minutes. At this time, radioactivity concentration in 5-HT1A receptor-rich brain regions was markedly greater than in cerebellum. In another PET experiment, the monkey was predosed with WAY-100635 before [11C](R,S)-JWAY injection. At 90 minutes after injection, the ratio of radioactivity in 5-HT1A receptor-rich regions to that in cerebellum was reduced to near unity. Radioactivity represented by parent radioligand in plasma was 12% at 45 minutes. CONCLUSIONS: [11C](R,S)-JWAY, but not [11C]SWAY, gives a sizeable 5-HT1A receptor-selective PET signal in monkey. The presence of a C-methyl group adjacent to the amide bond in SWAY or (R,S)-JWAY fails to counter metabolism.     

Preparation and biological evaluation of 111In-, 177Lu- and 90Y-labeled DOTA analogues conjugated to B72.3

Three 1,4,7,10-tetraazacyclododecane-N,N',N",N"'-tetraacetic acid (DOTA) analogues were evaluated for relative in vivo stability when radiolabeled with (111)In, (90)Y and (177)Lu and conjugated to the monoclonal antibody B72.3. The DOTA analogues evaluated were "NHS-DOTA" [N-hydroxysuccinimdyl (NHS) group activating one carboxylate], "Arm-DOTA" (also known as MeO-DOTA; with a p-NCS, o-MeO-benzyl moiety on the methylene group of one acetic acid arm) and "Back-DOTA" (with a p-NCS-benzyl moiety on a backbone methylene group of the macrocycle). The B72.3 was conjugated to the DOTA analogues to increase the retention time of the radioloabeled conjugates in vivo in mice. The serum stability of the various radiometalated DOTA conjugates showed them to have good stability out to 168 h (all >95% except (111)In-NHS-DOTA-B72.3, which was 91% stable). Hydroxyapatite stability for the (111)In and (177)Lu DOTA-conjugates was >95% at 168 h, while the (90)Y DOTA-conjugates were somewhat less stable (between 90% and 95% at 168 h). The biodistribution studies of the radiometalated DOTA-conjugates showed that no significant differences were observed for the (111)In and (177)Lu analogues; however, the (90)Y analogues showed lower stabilities, as evidenced by their increased bone uptake relative to the other two [2-20% injected dose per gram (% ID/g) for (90)Y and 2-8% ID/g for (111)In and (177)Lu]. The lower stability of the (90)Y analogues could be due to the higher beta energy of (90)Y and/or to the larger ionic radius of Y(3+). Based on the bone uptake observed, the (177)Lu-NHS-DOTA-B72.3 had slightly lower stability than the (177)Lu-Arm-DOTA-B72.3 and (177)Lu-Back-DOTA-B72.3, but not significantly at all time points. For (90)Y, the analogue showing the lowest stability based on bone uptake was (90)Y-Arm-DOTA-B72.3, perhaps because of the metal's larger ionic radius and potential steric interactions minimizing effective complexation. The (111)In analogues all showed similar biological distributions at the various time points. This study suggests that care must be taken when evaluating (90)Y-labeled antibodies and in using NHS-DOTA-antibody conjugates with (177)Lu. All evaluations should be extended to time points relevant to the half-life of the radiometal and the therapy applications.