High Cytotoxic Effect by Combining Copper-64 with a NOTA–Terpyridine Platinum Conjugate

Abstract

Terpyridine platinum (TP)-based chemotherapeutic agents target three-dimensional structures on DNA known as G-quadruplexes. We report the rational design and synthesis of a TP conjugate combined with copper-64 ( 64 Cu), the decay characteristics of which include emission of β − and Auger electrons for radiotherapy and β + particles for positron emission tomography (PET) imaging. The present experimental studies show that the novel [ 64 Cu]Cu−1,4,7-triazacyclononane-1,4,7triacetic acid (NOTA)−TP is stable, permitting selective killing of cancer cells. The antitumor activity of [ 64 Cu]Cu−NOTA−TP at high apparent molar activity is in the low nanomolar range and 27,800-fold greater than that of nat Cu−NOTA−TP at 24 h post treatment. These results suggest that this combination of a cytotoxic TP agent with 64 Cu has considerable potential for cancer treatment and PET imaging.

Design, Synthesis, and Cytotoxicity Assessment of [64Cu]Cu-NOTA-Terpyridine Platinum Conjugate: A Novel Chemoradiotherapeutic Agent with Flexible Linker

“…internalized fraction increased strikingly from 0.04 ± 0.02% after 15 min incubation to its maximum value (18.7 ± 2.8%) at 24 h, which is consistent with our previous results for a chemoradiotherapeutic agent labeled with 64 Cu [28]. This kinetic is also consistent with those observed with TP compounds as reported by Stafford et al [13].…”

“…As seen in Figure 4A, the total cell-associated and internalization fraction of [ 64 Cu]Cu-NOTA-C3-TP increased over time and reached a maximum value after 24 h of incubation. The internalized fraction increased strikingly from 0.04 ± 0.02% after 15 min incubation to its maximum value (18.7 ± 2.8%) at 24 h, which is consistent with our previous results for a chemoradiotherapeutic agent labeled with 64 Cu [28]. This kinetic is also consistent with those observed with TP compounds as reported by Stafford et al [13].…”

“…In the case of cytoplasmic distribution, a range of percentages between 76 ± 7.3%, 61 ± 11%, and 61 ± 8.3% in HCT116 cancer cells (Figure 6A), and 46 ± 6%, 53 ± 6.5%, and 58.5 ± 8.3% in fibroblasts were calculated at 24 h, 48 h, and 72 h, respectively (Figure 6B). The subcellular localization of [ 64 Cu]Cu(OAc) 2 in HCT116 cells was investigated as a control group, and the results were reported previously [28].…”

“…Interestingly, the cytotoxic activity of [ 64 Cu]Cu-NOTA-C3-TP on GM05757 human fibroblast cells (EC 50 > 200µM) was at least 3.4-fold lower than that of HCT116 cells. Parallel experiments were carried out with [ 64 Cu]Cu(OAc) 2 as control, and the results were reported previously [28]. Accordingly, the measured EC 50 value to induce a 50% reduction in cell viability was 5.6 ± 0.5 in the CRC cells HCT116 at 24 h. This value is about 28 times higher than that for [ 64 Cu]Cu-NOTA-TP at an EC 50 concentration (0.2 MBq), suggesting that 64 Cu alone is much less active than [ 64 Cu]Cu-NOTA-TP toward HCT116 cancer cells.…”

“…For extraction of HCT116 and GM05757 cell nucleus and cytoplasm, we followed a previously reported procedure [28,31]. After treatment of HCT116 cells with 1MBq of [ 64 Cu]Cu-NOTA-C3-TP for 24, 48 and 72 h, CSK buffer (0.5% Triton X-100, 300 mM sucrose, 100 mM NaCl, 1 mM ethylenediaminetetraacetic acid (EDTA), 2 mM MgCl 2 , and 10 mM HEPES, pH 6.8) were employed for nuclear extraction.…”