Genome-wide association studies and Mendelian randomization analyses provide insights into the causes of early-onset colorectal cancer.

BACKGROUND: The incidence of early-onset colorectal cancer (EOCRC; diagnosed <50 years of age) is rising globally; however, the causes underlying this trend are largely unknown. CRC has strong genetic and environmental determinants, yet common genetic variants and causal modifiable risk factors underlying EOCRC are unknown. We conducted the first EOCRC-specific genome-wide association study (GWAS) and Mendelian randomization (MR) analyses to explore germline genetic and causal modifiable risk factors associated with EOCRC. PATIENTS AND METHODS: We conducted a GWAS meta-analysis of 6176 EOCRC cases and 65 829 controls from the Genetics and Epidemiology of Colorectal Cancer Consortium (GECCO), the Colorectal Transdisciplinary Study (CORECT), the Colon Cancer Family Registry (CCFR), and the UK Biobank. We then used the EOCRC GWAS to investigate 28 modifiable risk factors using two-sample MR. RESULTS: We found two novel risk loci for EOCRC at 1p34.1 and 4p15.33, which were not previously associated with CRC risk. We identified a deleterious coding variant (rs36053993, G396D) at polyposis-associated DNA repair gene MUTYH (odds ratio 1.80, 95% confidence interval 1.47-2.22) but show that most of the common genetic susceptibility was from noncoding signals enriched in epigenetic markers present in gastrointestinal tract cells. We identified new EOCRC-susceptibility genes, and in addition to pathways such as transforming growth factor (TGF) ß, suppressor of Mothers Against Decapentaplegic (SMAD), bone morphogenetic protein (BMP) and phosphatidylinositol kinase (PI3K) signaling, our study highlights a role for insulin signaling and immune/infection-related pathways in EOCRC. In our MR analyses, we found novel evidence of probable causal associations for higher levels of body size and metabolic factors-such as body fat percentage, waist circumference, waist-to-hip ratio, basal metabolic rate, and fasting insulin-higher alcohol drinking, and lower education attainment with increased EOCRC risk. CONCLUSIONS: Our novel findings indicate inherited susceptibility to EOCRC and suggest modifiable lifestyle and metabolic targets that could also be used to risk-stratify individuals for personalized screening strategies or other interventions.

Protocol for the treatment of cystoid macular edema secondary to retinitis pigmentosa and other inherited retinal dystrophies.

Inherited retinal dystrophies (IRD) are the leading cause of legal blindness in the working population. Cystic macular edema (CME) is one of the treatable causes of visual loss, affecting up to 50% of the patients. A bibliographic review has been carried out combining "inherited retinal dystrophy", "retinitis pigmentosa", "macular oedema" and a diagnostic-therapeutic protocol according to the levels of evidence and recommendations of the "US Agency for Healthcare Research and Quality". This protocol has been discussed in the monthly meetings of the XAREA DHR group with the participation of more than 25 ophthalmologists, creating a consensus document. The etiology of CME is multifactorial: dysfunction of the blood-retinal barrier, retinal pigment epithelium, and Müller cells, inflammation, and vitreous traction. OCT is the test of choice for the diagnosis and follow-up of CME associated with IRD. The drugs with the highest degree of scientific evidence are carbonic anhydrase inhibitors (IAC). Intravitreal corticosteroids, anti-VEGF, and vitrectomy with peeling of the internal limiting membrane do not have sufficient evidence. A treatment scheme is proposed for the CME in IRD in adults, another for pediatric patients and another for IRD and cataract surgery. Oral and topical IACs are effective in the treatment of CME secondary to IRD. Treatment with corticosteroids, anti-VEGF, and vitrectomy are second-line options. Randomized clinical trials are required to establish the therapeutic scale in these patients.

Evaluation of non-exudative tomographic signs in cases of exudative age-related macular degeneration.

OBJECTIVE: To analyse the prevalence of non-exudative tomographic signs (onion sign, pseudoswelling, external retinal tubulation, pseudocysts, subretinal clefts and macular atrophy) in patients with neovascular age-related macular degeneration. MATERIAL AND METHODS: A total of 174 eyes of patients with neovascular age-related macular degeneration who had not received previous treatment were included in the study. Visual acuity, neovascularization activity, and the appearance or not of the different signs under study were assessed at times 0 (initial visit), 4 months, one year, year and a half, and at 2 and 3 years of follow-up. The following were also evaluated: age, sex, affected eye and type of neovascularization (1, 2, 3, polypoid or mixed). The analysis were performed using the statistical software R (version 3.3.2) and the glmmADMB package (version 0.8.3.3). RESULTS: The presence of pseudocysts and external retinal tubulation increases throughout the follow-up. The onion sign begins with an ascending frequency up to 12 months, then decreases at 18 months and increases again at 24 months. Regarding pseudowelling, it maintains an increase until 18 months to finally decrease. Subretinal clefts is the rarest sign, presenting in 1.1% on the first visit. Finally, macular atrophy, present in 12.6% of the eyes initially, is found in 25% after 2 years. CONCLUSION: Pseudocysts, external retinal tubulation and macular atrophy were the most prevalent signs, while subretinal clefts were the most infrequent.

Discovery of the Potent and Selective MC4R Antagonist PF-07258669 for the Potential Treatment of Appetite Loss.

The melanocortin-4 receptor (MC4R) is a centrally expressed, class A GPCR that plays a key role in the regulation of appetite and food intake. Deficiencies in MC4R signaling result in hyperphagia and increased body mass in humans. Antagonism of MC4R signaling has the potential to mitigate decreased appetite and body weight loss in the setting of anorexia or cachexia due to underlying disease. Herein, we report on the identification of a series of orally bioavailable, small-molecule MC4R antagonists using a focused hit identification effort and the optimization of these antagonists to provide clinical candidate 23. Introduction of a spirocyclic conformational constraint allowed for simultaneous optimization of MC4R potency and ADME attributes while avoiding the production of hERG active metabolites observed in early series leads. Compound 23 is a potent and selective MC4R antagonist with robust efficacy in an aged rat model of cachexia and has progressed into clinical trials.

Concurrent J-evolving refocusing pulses.

Conventional refocusing pulses are optimised for a single spin without considering any type of coupling. However, despite the fact that most couplings will result in undesired distortions, refocusing in delay-pulse-delay-type sequences with desired heteronuclear coherence transfer might be enhanced considerably by including coupling evolution into pulse design. We provide a proof of principle study for a Hydrogen-Carbon refocusing pulse sandwich with inherent J-evolution following the previously reported ICEBERG-principle with improved performance in terms of refocusing performance and/or overall effective coherence transfer time. Pulses are optimised using optimal control theory with a newly derived quality factor and z-controls as an efficient tool to speed up calculations. Pulses are characterised in theory and experiment and compared to conventional concurrent refocusing pulses, clearly showing an improvement for the J-evolving pulse sandwich. As a side-product, also efficient J-compensated resfocusing pulse sandwiches - termed BUBU pulses following the nomenclature of previous J-compensated BUBI and BEBEtr pulse sandwiches - have been optimised.

Cross-Linked Poly-4-Acrylomorpholine: A Flexible and Reversibly Compressible Aligning Gel for Anisotropic NMR Analysis of Peptides and Small Molecules in Water.

Determination of the solution conformation of both small organic molecules and peptides in water remains a substantial hurdle in using NMR solution conformations to guide drug design due to the lack of easy to use alignment media. Herein we report the design of a flexible compressible chemically cross-linked poly-4-acrylomorpholine gel that can be used for the alignment of both small molecules and cyclic peptides in water. To test the new gel, residual dipolar couplings (RDCs) and J-coupling constants were used in the configurational analysis of strychnine hydrochloride, a molecule that has been studied extensively in organic solvents as well as a small cyclic peptide that is known to form an α-helix in water. The conformational ensembles for each molecule with the best fit to the data are reported. Identification of minor conformers in water that cannot easily be determined by conventional NOE measurements will facilitate the use of RDC experiments in structure-based drug design.

Men's experiences of radiotherapy treatment for localized prostate cancer and its long-term treatment side effects: a longitudinal qualitative study.

PURPOSE: To investigate men's experiences of receiving external-beam radiotherapy (EBRT) with neoadjuvant Androgen Deprivation Therapy (ADT) for localized prostate cancer (LPCa) in the ProtecT trial. METHODS: A longitudinal qualitative interview study was embedded in the ProtecT RCT. Sixteen men with clinically LPCa who underwent EBRT in ProtecT were purposively sampled to include a range of socio-demographic and clinical characteristics. They participated in serial in-depth qualitative interviews for up to 8 years post-treatment, exploring experiences of treatment and its side effects over time. RESULTS: Men experienced bowel, sexual, and urinary side effects, mostly in the short term but some persisted and were bothersome. Most men downplayed the impacts, voicing expectations of age-related decline, and normalizing these changes. There was some reticence to seek help, with men prioritizing their relationships and overall health and well-being over returning to pretreatment levels of function. Some unmet needs with regard to information about treatment schedules and side effects were reported, particularly among men with continuing functional symptoms. CONCLUSIONS: These findings reinforce the importance of providing universal clear, concise, and timely information and supportive resources in the short term, and more targeted and detailed information and care in the longer term to maintain and improve treatment experiences for men undergoing EBRT.

Modelling the lifetime cost-effectiveness of radical prostatectomy, radiotherapy and active monitoring for men with clinically localised prostate cancer from median 10-year outcomes in the ProtecT randomised trial.

BACKGROUND: Optimal management strategies for clinically localised prostate cancer are debated. Using median 10-year data from the largest randomised controlled trial to date (ProtecT), the lifetime cost-effectiveness of three major treatments (radical radiotherapy, radical prostatectomy and active monitoring) was explored according to age and risk subgroups. METHODS: A decision-analytic (Markov) model was developed and informed by clinical input. The economic evaluation adopted a UK NHS perspective and the outcome was cost per Quality-Adjusted Life Year (QALY) gained (reported in UK£), estimated using EQ-5D-3L. RESULTS: Costs and QALYs extrapolated over the lifetime were mostly similar between the three randomised strategies and their subgroups, but with some important differences. Across all analyses, active monitoring was associated with higher costs, probably associated with higher rates of metastatic disease and changes to radical treatments. When comparing the value of the strategies (QALY gains and costs) in monetary terms, for both low-risk prostate cancer subgroups, radiotherapy generated the greatest net monetary benefit (£293,446 [95% CI £282,811 to £299,451] by D'Amico and £292,736 [95% CI £284,074 to £297,719] by Grade group 1). However, the sensitivity analysis highlighted uncertainty in the finding when stratified by Grade group, as radiotherapy had 53% probability of cost-effectiveness and prostatectomy had 43%. In intermediate/high risk groups, using D'Amico and Grade group > = 2, prostatectomy generated the greatest net monetary benefit (£275,977 [95% CI £258,630 to £285,474] by D'Amico and £271,933 [95% CI £237,864 to £287,784] by Grade group). This finding was supported by the sensitivity analysis. Prostatectomy had the greatest net benefit (£290,487 [95% CI £280,781 to £296,281]) for men younger than 65 and radical radiotherapy (£201,311 [95% CI £195,161 to £205,049]) for men older than 65, but sensitivity analysis showed considerable uncertainty in both findings. CONCLUSION: Over the lifetime, extrapolating from the ProtecT trial, radical radiotherapy and prostatectomy appeared to be cost-effective for low risk prostate cancer, and radical prostatectomy for intermediate/high risk prostate cancer, but there was uncertainty in some estimates. Longer ProtecT trial follow-up is required to reduce uncertainty in the model. TRIAL REGISTRATION: Current Controlled Trials number, ISRCTN20141297: http://isrctn.org (14/10/2002); ClinicalTrials.gov number, NCT02044172: http://www.clinicaltrials.gov (23/01/2014).

Testing for causality between systematically identified risk factors and glioma: a Mendelian randomization study.

BACKGROUND: Whilst epidemiological studies have provided evidence of associations between certain risk factors and glioma onset, inferring causality has proven challenging. Using Mendelian randomization (MR), we assessed whether associations of 36 reported glioma risk factors showed evidence of a causal relationship. METHODS: We performed a systematic search of MEDLINE from inception to October 2018 to identify candidate risk factors and conducted a meta-analysis of two glioma genome-wide association studies (5739 cases and 5501 controls) to form our exposure and outcome datasets. MR analyses were performed using genetic variants to proxy for candidate risk factors. We investigated whether risk factors differed by subtype diagnosis (either glioblastoma (n = 3112) or non-glioblastoma (n = 2411)). MR estimates for each risk factor were determined using multiplicative random effects inverse-variance weighting (IVW). Sensitivity analyses investigated potential pleiotropy using MR-Egger regression, the weighted median estimator, and the mode-based estimator. To increase power, trait-specific polygenic risk scores were used to test the association of a genetically predicated increase in each risk factor with glioma onset. RESULTS: Our systematic search identified 36 risk factors that could be proxied using genetic variants. Using MR, we found evidence that four genetically predicted traits increased risk of glioma, glioblastoma or non-glioblastoma: longer leukocyte telomere length, liability to allergic disease, increased alcohol consumption and liability to childhood extreme obesity (> 3 standard deviations from the mean). Two traits decreased risk of non-glioblastoma cancers: increased low-density lipoprotein cholesterol (LDLc) and triglyceride levels. Our findings were similar across sensitivity analyses that made allowance for pleiotropy (genetic confounding). CONCLUSIONS: Our comprehensive investigation provides evidence of a causal link between both genetically predicted leukocyte telomere length, allergic disease, alcohol consumption, childhood extreme obesity, and LDLc and triglyceride levels, and glioma. The findings from our study warrant further research to uncover mechanisms that implicate these traits in glioma onset.

Insulin-like growth factor-1, insulin-like growth factor-binding protein-3, and breast cancer risk: observational and Mendelian randomization analyses with ∼430 000 women.

BACKGROUND: Epidemiological evidence supports a positive association between circulating insulin-like growth factor-1 (IGF-1) concentrations and breast cancer risk, but both the magnitude and causality of this relationship are uncertain. We conducted observational analyses with adjustment for regression dilution bias, and Mendelian randomization (MR) analyses allowed for causal inference. PATIENTS AND METHODS: We investigated the associations between circulating IGF-1 concentrations and incident breast cancer risk in 206 263 women in the UK Biobank. Multivariable hazard ratios (HRs) and 95% confidence intervals (CI) were estimated using Cox proportional hazards models. HRs were corrected for regression dilution using repeat IGF-1 measures available in a subsample of 6711 women. For the MR analyses, genetic variants associated with circulating IGF-1 and IGF-binding protein-3 (IGFBP-3) levels were identified and their association with breast cancer was examined with two-sample MR methods using genome-wide data from 122 977 cases and 105 974 controls. RESULTS: In the UK Biobank, after a median follow-up of 7.1 years, 4360 incident breast cancer cases occurred. In the multivariable-adjusted models corrected for regression dilution, higher IGF-1 concentrations were associated with a greater risk of breast cancer (HR per 5 nmol/l increment of IGF-1 = 1.11, 95% CI = 1.07-1.16). Similar positive associations were found by follow-up time, menopausal status, body mass index, and other risk factors. In the MR analyses, a 5 nmol/l increment in genetically-predicted IGF-1 concentration was associated with a greater breast cancer risk (odds ratio = 1.05, 95% CI = 1.01-1.10; P = 0.02), with a similar effect estimate for estrogen-positive (ER+) tumours, but no effect found for estrogen-negative (ER-) tumours. Genetically-predicted IGFBP-3 concentrations were not associated with breast cancer risk (odds ratio per 1-standard deviation increment = 1.00, 95% CI = 0.97-1.04; P = 0.98). CONCLUSION: Our results support a probable causal relationship between circulating IGF-1 concentrations and breast cancer, suggesting that interventions targeting the IGF pathway may be beneficial in preventing breast tumorigenesis.

The molecular landscape of osteogenesis imperfecta in a Brazilian tertiary service cohort.

We have sought the molecular diagnosis of OI in 38 Brazilian cases through targeted sequencing of 15 candidate genes. While 71% had type 1 collagen-related OI, defects in FKBP10, PLOD2 and SERPINF1, and a potential digenic P3H1/WNT1 interaction were prominent causes of OI in this underrepresented population. INTRODUCTION: Defects in type 1 collagen reportedly account for 85-90% of osteogenesis imperfecta (OI) cases, but most available molecular data has derived from Sanger sequencing-based approaches in developed countries. Massively parallel sequencing (MPS) allows for systematic and comprehensive analysis of OI genes simultaneously. Our objective was to obtain the molecular diagnosis of OI in a single Brazilian tertiary center cohort. METHODS: Forty-nine individuals (84% adults) with a clinical diagnosis of OI, corresponding to 30 sporadic and 8 familial cases, were studied. Sixty-three percent had moderate to severe OI, and consanguinity was common (26%). Coding regions and 25-bp boundaries of 15 OI genes (COL1A1, COL1A2, IFITM5 [plus 5'UTR], SERPINF1, CRTAP, P3H1, PPIB, SERPINH1, FKBP10, PLOD2, BMP1, SP7, TMEM38B, WNT1, CREB3L1) were analyzed by targeted MPS and variants of interest were confirmed by Sanger sequencing or SNP array. RESULTS: A molecular diagnosis was obtained in 97% of cases. COL1A1/COL1A2 variants were identified in 71%, whereas 26% had variants in other genes, predominantly FKBP10, PLOD2, and SERPINF1. A potential digenic interaction involving P3H1 and WNT1 was identified in one case. Phenotypic variability with collagen defects could not be explained by evident modifying variants. Four consanguineous cases were associated to heterozygous COL1A1/COL1A2 variants, and two nonconsanguineous cases had compound PLOD2 heterozygosity. CONCLUSIONS: Novel disease-causing variants were identified in 29%, and a higher proportion of non-collagen defects was seen. Obtaining a precise diagnosis of OI in underrepresented populations allows expanding our understanding of its molecular landscape, potentially leading to improved personalized care in the future.

Real-world impact of glucocorticoid replacement therapy on bone mineral density: retrospective experience of a large single-center CAH cohort spanning 24 years.

The congenital adrenal hyperplasia population seems to have an intrinsic tendency to a high frequency of low bone mass. However in this single-center and long-term evaluated cohort, the simplified corticoid regimen, with exclusive dexamethasone single dose reposition during adulthood, did not represent a risk factor for decrease in bone health. INTRODUCTION: The impact of long-term and supposedly physiological doses of gluco and mineralocorticoid (GC/MC) on bone mineral density (BMD) in congenital adrenal hyperplasia (CAH) remains discordant among studies, which contain different clinical forms and corticoid regimens. Our aim was to evaluate the BMD in CAH adults receiving similar GC regimen since childhood and to correlate it with GC/MC cumulative doses. METHODS: Only patients with good compliance, who used cortisone acetate (CA) during childhood and dexamethasone after the final height achievement. Cumulative GC/MC doses were calculated from diagnosis until last evaluation. BMD was analyzed by the first and last energy X-ray absorptiometry (DXA) scans performed. RESULTS: Twenty simple virilizing (SV) and 14 salt wasting (WS) whose mean age was 26 ± 6 years, mean CA, dexamethasone, and fludrocortisone cumulative doses were 63,813 ± 32,767, 812 ± 558, and 319 ± 325 mg/m2, respectively. Based on the last DXA, low BMD was observed in 11% of patients, total hip Z-score was lower in the SW than SV form (p = 0.04). Cumulative CA dose had an inverse correlation with femoral neck Z-score (p < 0.01). Total cumulative GC and MC doses had an inverse correlation with total hip Z-score (p < 0.01). In the analysis of sequential BMD during dexamethasone therapy, no association was observed among cumulative GC/MC doses, clinical forms, sex, and lumbar Z-score delta. CONCLUSIONS: Even though a low CA regimen during growth periods in addition to MC replacement appears to have an influence on BMD at femoral sites, interestingly a low dexamethasone one does not seem to be deleterious for bone health in adulthood.

Computer-Assisted 3D Structure Elucidation (CASE-3D): The Structural Value of 2 JCH in Addition to 3 JCH Coupling Constants.

We present a method to use long-range CH coupling constants to derive the correct diastereoisomer from the molecular constitution of small molecules. A set of 79 2 JCH and 3 JCH values collected from a single HSQMBC experiment on a sample of strychnine were used in the CASE-3D (computer-assisted 3D structure elucidation) protocol. In addition to the most commonly used 3 JCH coupling constants, the subset of 32 2 JCH values alone showed an excellent degree of configuration selection. The study is mainly based on comparison of DFT-calculated 2,3 JCH values with experimental ones, critical for the case of 2 JCH . But the configuration selection also works well using 3 JCH values predicted from a semi-empirical Karplus-based equation limited to H-C-C-C fragments. The robustness, shown using strychnine as a proof of concept, makes the J-based CASE-3D analysis a viable option for the application in fields such as peptide and carbohydrate research, organic synthesis, natural-product identification and analysis, as well as medicinal chemistry.

Reference-free NOE NMR analysis.

Nuclear Overhauser Effect (NOE) methods in NMR are an important tool for 3D structural analysis of small molecules. Quantitative NOE methods conventionally rely on reference distances, known distances that have to be spectrally separated and are not always available. Here we present a new method for evaluation and 3D structure selection that does not require a reference distance, instead utilizing structures optimized by molecular mechanics, enabling NOE evaluation even on molecules without suitable reference groups.

[Thyroid effects of treatment with biotin]. / Efectos tiroideos del tratamiento con biotina.

TITLE: Efectos tiroideos del tratamiento con biotina.

Polarization recovery during ASAP and SOFAST/ALSOFAST-type experiments.

Experiments with fast repetition schemes significantly enhance the capabilities of modern NMR spectroscopy. Two schemes for heteronuclear correlation experiments that have been presented are the ASAP and the ALSOFAST method. The first method is Acceleration by Sharing Adjacent Polarization (ASAP) for samples at natural abundance isotope level. It was originally derived in the ASAP-HMQC and recently received renewed attention in the ASAP-HSQC. Sharing the polarization of active protons with the surrounding reservoir can result in seemingly instant polarization recovery and therefore enormous gains in sensitivity, but can also lead to a slight reduction of polarization and spectral intensity, depending on sample and setup. A second type of setup has been introduced with the so-called Alternate SOFAST (ALSOFAST-) HMQC and ALSOFAST-HSQC for natural abundance 1H,13C-correlation experiments and in the SOFAST-HMQC for 1H,15N-correlations. In these cases, the reservoir spins are only maintained through the pulse sequence without Hartmann-Hahn-type mixing. A model for the estimation of the available polarization in the fast repetition schemes could be a valuable tool for experimentalists and pulse sequence developers. Starting from the well-known Ernst angle model, we derive in this article several mathematical models that describe the polarization over the course of ALSOFAST and ASAP type experiments. The models can be used to visualize the initial scans of an experiment and even more importantly, show the polarization and achievable signal intensity in the steady state of an experiment. In this way the two extreme applications of ASAP- and ALSOFAST-type acquisition schemes are covered: (i) acquisition using progressive excitation for experiments with few increments and shortest possible overall acquisition times and (ii) steady-state-type experiments with ultrahigh resolution and correspondingly large number of increments. The two resulting excitation strategies are applied to maximize SNR in different situations. To test the models, experimental data was obtained by special pulse sequences and examples are shown for different spin environments. The results show good agreement between theory and experiment.

ASAP-HSQC-TOCSY for fast spin system identification and extraction of long-range couplings.

Based on Ernst-angle-type excitation and Acceleration by Sharing Adjacent Polarization (ASAP), a fast HSQC-TOCSY experiment is introduced. In the approach, the DIPSI-2 isotropic mixing period of the ASAP-HSQC is simply shifted, which provides a TOCSY period without additional application of rf-energy. The ASAP-HSQC-TOCSY allows the acquisition of a conventional 2D in about 30 s. Alternatively, it allows the acquisition of highly carbon-resolved spectra (several Hz digital resolution) on the order of minutes. An ASAP-HSQC-TOCSY-IPAP variant, finally, allows the sign-sensitive extraction of heteronuclear long-range coupling constants from a pair of highly resolved spectra in less than an hour. Pulse sequences, several example spectra, and a discussion of results are given.

1,5-Cyclooctadienyl alcohols and ketones generate a new class of COD Pt complexes.

A variety of new racemic alcohol and ketone cyclooctadiene derivatives was prepared for their complexation with platinum to generate a new class of platinum(ii) complexes.

Boosting the NMR Assignment of Carbohydrates with Clean In-Phase Correlation Experiments.

Novel CLIP-COSY based homo- and heteronuclear correlation experiments are reported for the rapid, semi-automated NMR assignment of small to medium-sized molecules. The homonuclear CLIP-COSY and corresponding relayed experiments employ the perfect-echo based mixing sequence for in-phase coherence transfer between directly and/or indirectly coupled proton spins. The combined analysis of the resulting CLIP-COSY and relayed spectra made it possible to easily track down, layer by layer, the proton-proton connectivity network. In larger molecules the narrow chemical shift range of protons may, however, compromise the efficacy of the homonuclear correlation based assignment strategy. To overcome this limitation, an HSQC variant of the CLIP-COSY experiment has now been devised. Combined treatment of HSQC-CLIP-COSY (relayed) and standard HSQC spectra facilitates simultaneous and semi-automatic assignment of 1 H and 13 C resonances of medium-sized molecules, such as pentasaccharides. The recently introduced PSYCHE broadband homonuclear decoupling scheme has been also implemented into the devised homo- and heteronuclear CLIP-COSY based experiments, resulting in fully decoupled high-resolution pure-shift correlation spectra.