Non-Invasive In Vivo Imaging of Tumor-Associated CD133/Prominin

detection of cancer stem cells is of great importance. detection of CD133/prominin, a cancer stem cell surface marker for a variety of tumor entities. The CD133-specific monoclonal antibody AC133.1 was used for quantitative fluorescence-based optical imaging of mouse xenograft models based on isogenic pairs of CD133 positive and negative cell lines. A first set consisted of wild-type U251 glioblastoma cells, which do not express CD133, and lentivirally transduced CD133-overexpressing U251 cells. A second set made use of HCT116 colon carcinoma cells, which uniformly express CD133 at levels comparable to primary glioblastoma stem cells, and a CD133-negative HCT116 derivative. Not surprisingly, visualization and quantification of CD133 in overexpressing U251 xenografts was successful; more importantly, however, significant differences were also found in matched HCT116 xenograft pairs, despite the lower CD133 expression levels. The binding of i.v.-injected AC133.1 antibodies to CD133 positive, but not negative, tumor cells isolated from xenografts was confirmed by flow cytometry. imaging of tumor-associated CD133 is feasible and that CD133 antibody-based tumor targeting is efficient. This should facilitate developing clinically applicable cancer stem cell imaging methods and CD133 antibody-based therapeutics

Predicting Crystallization of Amorphous Drugs with Terahertz Spectroscopy.

There is a controversy about the extent to which the primary and secondary dielectric relaxations influence the crystallization of amorphous organic compounds below the glass transition temperature. Recent studies also point to the importance of fast molecular dynamics on picosecond-to-nanosecond time scales with respect to the glass stability. In the present study we provide terahertz spectroscopy evidence on the crystallization of amorphous naproxen well below its glass transition temperature and confirm the direct role of Johari-Goldstein (JG) secondary relaxation as a facilitator of the crystallization. We determine the onset temperature Tβ above which the JG relaxation contributes to the fast molecular dynamics and analytically quantify the level of this contribution. We then show there is a strong correlation between the increase in the fast molecular dynamics and onset of crystallization in several chosen amorphous drugs. We believe that this technique has immediate applications to quantify the stability of amorphous drug materials.JS and JAZ would like to acknowledge the UK Engineering and Physical Sciences Research Council for funding (EP/J007803/1).This is the final version of the article. It first appeared from ACS at http://dx.doi.org/10.1021/acs.molpharmaceut.5b0033

Immunophenotypic predictive profiling of BRCA1-associated breast cancer

The immunophenotypic predictive profile of BRCA1-associated cancers including major predictive markers, i.e., PARP-1, EGFR, c-kit, HER-2, and steroid hormones (ER/PR) that may have therapeutic relevance has not yet been reported in a comprehensive study. Using immunohistochemistry, we examined the expression of these proteins in a large cohort of BRCA1-associated breast cancers. PARP-1 immunoreactivity was found in 81.9%, EGFR in 43.6%, ER/PR in 17.9%, c-kit in 14.7%, and overexpression of HER-2 in 3.6% of cancers. For all markers studied, 8.2% of tumors were negative. Expression of only one predictive marker was found in 29.7% of cancers, and most frequently, it was PARP-1 (20.8%). In 62.1% of tumors, more than one predictive marker was expressed: PARP-1 and EGFR in 30.4%, PARP-1, and hormone receptors in 13.3% and PARP-1 with c-kit in 7.5% of all tumors. Coexpression of two or more other predictive markers was rare. There were significant differences in the median age at diagnosis of BRCA1-associated cancer between patients with ER+ vs. ER− and grades 1–2 vs. grade 3 tumors. These results demonstrate that BRCA1-associated cancers differ with respect to expression of proteins that are regarded as targets for specific therapies and that 92% of patients with BRCA1-associated cancers may benefit from one or several options for specific therapy (in addition to DNA damaging agents, e.g., cisplatin). About 8% of cancers which do not express therapeutic target proteins may not respond to such therapies. Knowledge of the immunophenotypic predictive profile may help with the recruitment of patients for trials of targeted therapies

HRDetect is a predictor of BRCA1 and BRCA2 deficiency based on mutational signatures.

Approximately 1-5% of breast cancers are attributed to inherited mutations in BRCA1 or BRCA2 and are selectively sensitive to poly(ADP-ribose) polymerase (PARP) inhibitors. In other cancer types, germline and/or somatic mutations in BRCA1 and/or BRCA2 (BRCA1/BRCA2) also confer selective sensitivity to PARP inhibitors. Thus, assays to detect BRCA1/BRCA2-deficient tumors have been sought. Recently, somatic substitution, insertion/deletion and rearrangement patterns, or 'mutational signatures', were associated with BRCA1/BRCA2 dysfunction. Herein we used a lasso logistic regression model to identify six distinguishing mutational signatures predictive of BRCA1/BRCA2 deficiency. A weighted model called HRDetect was developed to accurately detect BRCA1/BRCA2-deficient samples. HRDetect identifies BRCA1/BRCA2-deficient tumors with 98.7% sensitivity (area under the curve (AUC) = 0.98). Application of this model in a cohort of 560 individuals with breast cancer, of whom 22 were known to carry a germline BRCA1 or BRCA2 mutation, allowed us to identify an additional 22 tumors with somatic loss of BRCA1 or BRCA2 and 47 tumors with functional BRCA1/BRCA2 deficiency where no mutation was detected. We validated HRDetect on independent cohorts of breast, ovarian and pancreatic cancers and demonstrated its efficacy in alternative sequencing strategies. Integrating all of the classes of mutational signatures thus reveals a larger proportion of individuals with breast cancer harboring BRCA1/BRCA2 deficiency (up to 22%) than hitherto appreciated (∼1-5%) who could have selective therapeutic sensitivity to PARP inhibition

X-exome sequencing of 405 unresolved families identifies seven novel intellectual disability genes

X-linked intellectual disability (XLID) is a clinically and genetically heterogeneous disorder. During the past two decades in excess of 100 X-chromosome ID genes have been identified. Yet, a large number of families mapping to the X-chromosome remained unresolved suggesting that more XLID genes or loci are yet to be identified. Here, we have investigated 405 unresolved families with XLID. We employed massively parallel sequencing of all X-chromosome exons in the index males. The majority of these males were previously tested negative for copy number variations and for mutations in a subset of known XLID genes by Sanger sequencing. In total, 745 X-chromosomal genes were screened. After stringent filtering, a total of 1297 non-recurrent exonic variants remained for prioritization. Co-segregation analysis of potential clinically relevant changes revealed that 80 families (20%) carried pathogenic variants in established XLID genes. In 19 families, we detected likely causative protein truncating and missense variants in 7 novel and validated XLID genes (CLCN4, CNKSR2, FRMPD4, KLHL15, LAS1L, RLIM and USP27X) and potentially deleterious variants in 2 novel candidate XLID genes (CDK16 and TAF1). We show that the CLCN4 and CNKSR2 variants impair protein functions as indicated by electrophysiological studies and altered differentiation of cultured primary neurons from Clcn4−/− mice or after mRNA knock-down. The newly identified and candidate XLID proteins belong to pathways and networks with established roles in cognitive function and intellectual disability in particular. We suggest that systematic sequencing of all X-chromosomal genes in a cohort of patients with genetic evidence for X-chromosome locus involvement may resolve up to 58% of Fragile X-negative cases

Assembly, organization, and function of the COPII coat

A full mechanistic understanding of how secretory cargo proteins are exported from the endoplasmic reticulum for passage through the early secretory pathway is essential for us to comprehend how cells are organized, maintain compartment identity, as well as how they selectively secrete proteins and other macromolecules to the extracellular space. This process depends on the function of a multi-subunit complex, the COPII coat. Here we describe progress towards a full mechanistic understanding of COPII coat function, including the latest findings in this area. Much of our understanding of how COPII functions and is regulated comes from studies of yeast genetics, biochemical reconstitution and single cell microscopy. New developments arising from clinical cases and model organism biology and genetics enable us to gain far greater insight in to the role of membrane traffic in the context of a whole organism as well as during embryogenesis and development. A significant outcome of such a full understanding is to reveal how the machinery and processes of membrane trafficking through the early secretory pathway fail in disease states

Overview of the MOSAiC expedition—Atmosphere

With the Arctic rapidly changing, the needs to observe, understand, and model the changes are essential. To support these needs, an annual cycle of observations of atmospheric properties, processes, and interactions were made while drifting with the sea ice across the central Arctic during the Multidisciplinary drifting Observatory for the Study of Arctic Climate (MOSAiC) expedition from October 2019 to September 2020. An international team designed and implemented the comprehensive program to document and characterize all aspects of the Arctic atmospheric system in unprecedented detail, using a variety of approaches, and across multiple scales. These measurements were coordinated with other observational teams to explore cross-cutting and coupled interactions with the Arctic Ocean, sea ice, and ecosystem through a variety of physical and biogeochemical processes. This overview outlines the breadth and complexity of the atmospheric research program, which was organized into 4 subgroups: atmospheric state, clouds and precipitation, gases and aerosols, and energy budgets. Atmospheric variability over the annual cycle revealed important influences from a persistent large-scale winter circulation pattern, leading to some storms with pressure and winds that were outside the interquartile range of past conditions suggested by long-term reanalysis. Similarly, the MOSAiC location was warmer and wetter in summer than the reanalysis climatology, in part due to its close proximity to the sea ice edge. The comprehensiveness of the observational program for characterizing and analyzing atmospheric phenomena is demonstrated via a winter case study examining air mass transitions and a summer case study examining vertical atmospheric evolution. Overall, the MOSAiC atmospheric program successfully met its objectives and was the most comprehensive atmospheric measurement program to date conducted over the Arctic sea ice. The obtained data will support a broad range of coupled-system scientific research and provide an important foundation for advancing multiscale modeling capabilities in the Arctic