21 research outputs found
Localization of diphtheria toxin nuclease activity to fragment A.
We describe a series of experiments that aimed to establish whether nuclease activity is actually associated with diphtheria toxin (DTx) and its A subunit (DTA), as we originally reported (M. P. Chang, R. L. Baldwin, C. Bruce, and B. J. Wisnieski, Science 246:1165-1168, 1989). Here we show that (i) trypsinization of DTx does indeed produce nucleolytically active DTA, (ii) reduction of electroeluted, unreduced, cleaved DTx (58 kDa) yields nuclease-active DTA (24 kDa), and (iii) fractionation of DTx and DTA by anion-exchange chromatography leads to coelution of nuclease activity with both forms of the toxin, even though each form elutes at a distinct salt concentration. In addition, we show that Escherichia coli-derived DTA also expresses nuclease activity. These studies confirm our initial assertion that the nuclease activity observed in DTx preparations is intrinsic to the DTA portion of DTx
Evaluation of integrin αvβ<sub>6</sub> cystine knot PET tracers to detect cancer and idiopathic pulmonary fibrosis.
Advances in precision molecular imaging promise to transform our ability
to detect, diagnose and treat disease. Here, we describe the
engineering and validation of a new cystine knot peptide (knottin) that
selectively recognizes human integrin αvβ6 with single-digit
nanomolar affinity. We solve its 3D structure by NMR and x-ray
crystallography and validate leads with 3 different radiolabels in
pre-clinical models of cancer. We evaluate the lead tracer’s safety,
biodistribution and pharmacokinetics in healthy human volunteers, and
show its ability to detect multiple cancers (pancreatic, cervical and
lung) in patients at two study locations. Additionally, we demonstrate
that the knottin PET tracers can also detect fibrotic lung disease in
idiopathic pulmonary fibrosis patients. Our results indicate that these
cystine knot PET tracers may have potential utility in multiple disease
states that are associated with upregulation of integrin αvβ6