Anti-GRP (full length) polyclonal antibody

Purpose: The clinical utility of circulating tumor DNA (ctDNA) monitoring has been shown in tumors that harbor highly recurrent mutations. Leiomyosarcoma represents a type of tumor with a wide spectrum of heterogeneous genomic abnormalities; thus, targeting hotspot mutations or a narrow genomic region for ctDNA detection may not be practical. Here, we demonstrate a combinatorial approach that integrates different sequencing protocols for the orthogonal detection of single-nucleotide variants (SNV), small indels, and copy-number alterations (CNA) in ctDNA. Experimental Design: We employed Cancer Personalized Profiling by deep Sequencing (CAPP-Seq) for the analysis of SNVs and indels, together with a genome-wide interrogation of CNAs by Genome Representation Profiling (GRP). We profiled 28 longitudinal plasma samples and 25 tumor specimens from 7 patients with leiomyosarcoma. Results: We detected ctDNA in 6 of 7 of these patients with >98% specificity for mutant allele fractions down to a level of 0.01%. We show that results from CAPP-Seq and GRP are highly concordant, and the combination of these methods allows for more comprehensive monitoring of ctDNA by profiling a wide spectrum of tumor-specific markers. By analyzing multiple tumor specimens in individual patients obtained from different sites and at different times during treatment, we observed clonal evolution of these tumors that was reflected by ctDNA profiles. Conclusions: Our strategy allows for the comprehensive monitoring of a broad spectrum of tumor-specific markers in plasma. Our approach may be clinically useful not only in leiomyosarcoma but also in other tumor types that lack recurrent genomic alterations. (C) 2018 AACR.

The presence in cypress pollen of an important allergen, belonging to the gibberellin-regulated protein (GRP) family, has been suggested for many years. However, it has never been isolated and sometimes the homologous peach allergen, Pru p 7, has been used as a surrogate to perform immunological investigations. The aim of this study has been the isolation and molecular characterization of the GRP contained in the Cupressus sempervirens pollen. This protein, named Cypmaclein, has been purified from the natural source using conventional biochemical methods consisting in different chromatographic separations. Cypmaclein has been identified by direct protein sequencing of the N-terminal region and of internal fragments of the molecule. In SDS-PAGE, its apparent molecular mass is slightly higher than that of Pru p 7. Nevertheless, the mass spectrometry experiments reveal that the exact molecular mass of Cypmaclein (6821.88 Da) is very close to that of Pru p 7 (6909.90 Da). Two regions of Cypmaclein have been sequenced providing 50% of its primary structure. A high overall sequence identity of Cypmaclein with all the analyzed GRP has been observed, although in the N-terminal region the high identity is limited to the homolog of Cryptomeria japonica. In circular dichroism experiments Cypmaclein produced a spectrum overlapping that of Pru p 7. However, the comparative analysis of Cypmaclein, Pm p 7 and Pun g 7 IgE reactivity revealed a behavior that was not completely overlapping, thus suggesting that the IgE epitopes are only partially shared. In single point highest inhibition achievable assays performed with the FABER test, Cypmaclein efficiently competed with the allergenic peach and pomegranate GRP in the binding of specific IgE of patients sensitized to Pru p 7. In conclusion, the natural cypress pollen GRP has been isolated for the first time, its structural features have been investigated and its cross-reactivity with Pm p 7 and Pun g 7 has been demonstrated. This protein is now available for further investigations aimed at understanding its clinical relevance in the allergy to cypress pollen. In addition, the prevalence of sensitization directly to Cypmaclein, and not limited to the homologs, can be defined.