A 177Lu-labeled albumin-binder-conjugated PSMA-617 derivative with greatly enhanced radiation dose delivered to LNCaP tumor xenografts

7 Objectives: Various radiolabeled prostate specific membrane antigen (PSMA)-targeting radiotracers have been designed for radioligand therapy of prostate cancer, notably 177Lu-PSMA-617. Clinical data showed that 177Lu-PSMA-617 can be effective in treating metastatic prostate cancer and reducing patient PSA levels with no severe side effects. However, the complete response rate was low, and many patients still have progressive disease after 177Lu-PSMA-617 treatment. To improve treatment efficacy, we designed and synthesized a novel albumin-binder-conjugated 177Lu-PSMA-617 derivative, 177Lu-HTK01169, with an extended retention time in blood to maximize tumor uptake. Methods: PSMA-617 was produced by solid phase synthesis following reported procedures. For HTK01169, Fmoc-Lys(ivDde)-OH was coupled to the sequence after Fmoc-tranexamic acid. Elongation was continued with the addition of Fmoc-Glu(OtBu)-OH and 4-(p-iodophenyl)butyric acid to the N-terminus. Subsequently, the ivDde-protecting group was removed with 2% hydrazine in DMF, and DOTA-tris(t-Bu)ester was coupled to the amino group of Lys side chain. Cold standards were prepared in solution by incubating PSMA-617 and HTK01169 with LuCl3. Binding affinity to PSMA was determined by in vitro competition assays using LNCaP prostate cancer cells and 18F-DCFPyL as the hot ligand. 177Lu labeling was performed in acetate buffer (pH 4.5) at 90°C followed by HPLC purification. SPECT/CT imaging and biodistribution studies were conducted in mice bearing PSMA-expressing LNCaP prostate cancer xenografts. Radiation dosimetry was calculated using OLINDA (v.2.0) software. Results: HTK01169 and natLu-HTK01169 were obtained in 21 and 31% yield, respectively. 177Lu-PSMA-617 and 177Lu-HTK01169 were obtained in 86.5 ± 2.1% (n = 2) and 67.7 ± 16.2% (n = 3) decay-corrected radiochemical yields with > 99% radiochemical purity. The specific activities of 177Lu-PSMA-617 and 177Lu-HTK01169 were 766 ± 48.1 and 170 ± 88.8 GBq/µmol, respectively. 177Lu-PSMA-617 and 177Lu-HTK01169 bound PSMA with high affinity, and their calculated Ki values were 0.24 ± 0.06 and 0.04 ± 0.01 nM, respectively. SPECT imaging and biodistribution studies showed that 177Lu-PSMA-617 and 177Lu-HTK01169 were excreted mainly via the renal pathway. With fast blood clearance (0.68 %ID/g at 1 h post-injection), the tumor uptake of 177Lu-PSMA-617 peaked at 1 h post-injection (15.1 ± 5.58 %ID/g), and gradually decreased to 7.91 ± 2.82 %ID/g at 120 h post-injection. With an extended retention time in blood (16.6 ± 1.85 and 2.10 ± 0.41 %ID/g in blood at 1 and 24 h, respectively), the tumor uptake of 177Lu-HTK01169 peaked at 24 h post-injection (55.9 ± 12.5 %ID/g), and remained at the same level by the end of the study. Based on dosimetry calculations, 177Lu-HTK01169 delivered 8.35-fold higher radiation dose than 177Lu-PSMA-617 to LNCaP tumor xenografts. CONCLUSION: We successfully synthesized and evaluated a novel PSMA-targeted endoradiotherapeutic 177Lu-HTK01169 for prostate cancer. The novel albumin-binding motif N-[4-(p-iodophenyl)butanoyl]-Glu significantly extended the retention time of 177Lu-HTK01169 in blood, and dramatically improved the uptake into PSMA-expressing LNCaP tumor xenografts. Based on dosimetry calculations, 177Lu-HTK01169 is expected to produce similar or improved efficacy at only a fraction of 177Lu activity required with 177Lu-PSMA-617.