In vitro study: binding of 99mTc-DPD to synthetic amyloid fibrils

This paper is an report of the investigation of the in vitro binding of ^99mTc-DPD for synthetic amyloid fibrils used for the diagnosis of cardiac amyloidosis (CA), as compared with the use of ^99mTc-HMDP and 99mTc-PPI. It also includes an inquiry into the role played by Ca²⁺ ions and serum proteins on binding to amyloid like materials, as well as the saturability and specificity of DPD for fibrils versus amorphous precipitates (AP).

In the work, synthetic insulin fibrils (SIF) and AP were characterized by Congo red staining and TEM imaging. An equal amount of three radiopharmaceuticals were then added to fibrils in Ca²⁺ (0-4.2 mmol/L) or human serum (HS) adjoined samples and radiopharmaceutical uptake was assessed. To test the saturability of amyloid binding sites, a displacement assays with cold DPD was performed, while adding 50-1500 nmol of ^99mTc-DPD to SIF or AP, saturation binding tests were subsequently carried out for evaluating its specificity for amyloid.

Herein, synthetic fibrils and AP showed conformational differences at TEM and polarized microscopy analysis. In our study, ^99mTc-DPD fibrils uptake was seen to be the highest and increased with calcium ions concentration. What is more, serum proteins reduced the bound fraction to the amyloid deposits of about 15%, and the Kd values of 90 nM and 114 nM relative to SIF and AP, respectively, did not significantly differ.

We saw that ^99mTc-DPD is the best seeker for amyloid fibrils in cardiac amyloidosis, and that Ca²⁺ concentration positively influenced DPD fibrils binding. Furthermore, the radioactivity bound to the serum protein clear up the idea of nuclide exchanging dynamic balance between amyloid and circulating proteins. Moreover, non-labeled DPD did not exert a competition for ^99mTc-DPD binding sites, and, finally, DPD cannot be defined a radiopharmaceutical specific for amyloid deposits.