Assessment Methods
UAS7 and CU-QoL Total Score: At each visit, the subjects
provided their UAS7 and CU-QoL total scores for the previous 4 weeks.
Blood Sampling: Blood samples were collected prior to any
treatment intervention at all visits in EDTA tubes for total and
differential counts and at visits 2 (post-placebo) and 5 (4-weeks after
3rd dose of benralizumab) in PAXgene Blood RNA
(Qiagen) tubes for RNA-sequencing.
Total RNA Extraction and
RNA-Seq For Differential Gene Expression Profiling from Whole Blood:Directional RNA-seq was performed by the Genomics, Epigenomics and
Sequencing Core at the University of Cincinnati.27,28RNA extraction and sequencing methods are described in the supplement.
Skin Biopsy Collection, Processing, and Analysis : Two
full-thickness skin biopsies were taken using Acu-Punch Kit Ultra (4mm
Acu-Punch, Acuderm) from lesional (active hives) and nearby non-lesional
(normal) skin from CSU subjects at visit 2 and at visit 5 for histologic
and transriptomic analysis. Lesional biopsies (if present in
non-responders or partial responders) and non-lesional biopsies at visit
5 were performed at locations close to the baseline samples. For
complete benralizumab responders (i.e., subjects with UAS7=0 at visit
5), only non-lesional skin was biopsied. Normal skin biopsies were
collected once from non-CSU control subjects using identical methods.
Biopsies for histopathological analysis were transported prior to
processing in formalin and for RNA-sequencing in RNAlaterâ„¢ tubes.
Skin biopsy transcriptomic analyses will be presented in a subsequent
manuscript.