A CB-TE2A bifunctional chelate applicable to immunoPET with ⁶⁴Cu

Objectives CB-TE2A is a highly stable chelate for ⁶⁴Cu which shows improved background clearance. In this work we synthesized and evaluated 1, a side-arm functionalized version of CB-TE2A which yields a six coordinate uncharged Cu complex. The unanticipated room-temperature labeling of 1 allowed evaluation of this bifunctional chelate (BFC) for imaging with antibodies.

Methods 1 and Cu(1) complexes were synthesized and characterized using standard techniques. Radiolabeling was optimized for pH, temperature, time and BFC concentration. 1 was conjugated to trastuzumab and used for ⁶⁴Cu PET imaging RAG2M mice bearing HER2⁺ or HER2^- tumor xenografts.

Results The synthesis of 1 yielded two conformational isomers separable by chromatography, 1a and 1b. Cu(1a) and Cu(1b) were fully characterized, including structural determination by X-ray crystallography. 1a and 1b had significantly different ⁶⁴Cu radiolabeling properties. 1b required radiolabeling conditions similar to the conditions reported for CB-TE2A (pH >6, 90°C, 1 h). Unexpectedly, 1a could be radiolabeled with ⁶⁴Cu in >95% radiochemical yield under mild conditions (pH 6, <30 min, room temperature). The harsh radiolabeling conditions required for CB-TE2A have limited CB-TE2A’s application in ⁶⁴Cu labeling of antibodies. To illustrate that the mild conditions optimized for ⁶⁴Cu labeling of 1a are applicable to ⁶⁴Cu immunoPET, 1a was conjugated to trastuzumab and subsequently used to image HER2⁺ tumors in mice. Within 48 h, the HER2⁺ tumors were clearly visualized, with 7±1 %ID/g measured in the HER2⁺ tumors compared to 3.2±0.4 %ID/g in the HER2^- tumors (p<0.0003).

Conclusions The conformational isomer 1a containing the superior Cu chelating CB-TE2A core was discovered and shown to be applicable to ⁶⁴Cu immunoPET. 1a should also be useful for immunotherapy with ⁶⁷Cu