Abstract
BackgroundSensitive and specific detection of nodal status, sites of metastases and low-volume recurrent disease could greatly improve management of patients with advanced prostate cancer. Prostate-specific membrane antigen (PSMA) is a well-established marker for prostate carcinoma with increased levels of expression in high-grade, hormone-refractory and metastatic disease. The monoclonal antibody (mAb) J591 is directed against an extracellular epitope of PSMA and has been shown to efficiently target disseminated disease including metastases in lymph nodes and bone. Its use as a diagnostic imaging agent however is limited due to its slow pharmacokinetics. In this study a diabody derived from mAb J591 was developed as a single photon emission computed tomography (SPECT) tracer with improved pharmacokinetics for the detection of PSMA expression in prostate cancer.MethodsA diabody in VH-VL orientation and with a C-terminal cysteine was expressed in HEK293T cells and purified by a combination of metal ion affinity and size exclusion chromatography. Specificity and affinity were determined in cell binding studies. For SPECT imaging, the diabody was site-specifically labelled with [99mTc(CO)3]+ via the C-terminal His tag and evaluated in a subcutaneous DU145/DU145-PSMA prostate carcinoma xenograft model.ResultsJ591C diabody binds to PSMA-expressing cells with low nanomolar affinity (3.3 ± 0.2 nM). SPECT studies allowed imaging of tumour xenografts with high contrast from 4 h post injection (p.i.). Ex vivo biodistribution studies showed peak tumour uptake of the tracer of 12.1% ± 1.7% injected dose (ID)/g at 8 h p.i. with a tumour to blood ratio of 8.0. Uptake in PSMA-negative tumours was significantly lower with 6.3% ± 0.5% at 8 h p.i. (p < 0.001).ConclusionThe presented diabody has favourable properties required to warrant its further development for antibody-based imaging of PSMA expression in prostate cancer, including PSMA-specific uptake, favourable pharmacokinetics compared to the parental antibody and efficient site-specific radiolabelling with 99mTc.
Highlights
Sensitive and specific detection of nodal status, sites of metastases and low-volume recurrent disease could greatly improve management of patients with advanced prostate cancer
We describe the development and preclinical evaluation of a diabody derived from the anti-Prostate-specific membrane antigen (PSMA) antibody J591, site- labelled with 99mTc, for single photon emission computed tomography (SPECT) imaging of PSMA expression in prostate cancer
The diabody was cloned into pSEC-tag2 (Invitrogen, Carlsbad, CA, USA)-based mammalian expression vector pMS-C with N-terminal Ig-kappa leader and a C-terminal (His)6-tag followed by a cysteine (J591Cdia)
Summary
Sensitive and specific detection of nodal status, sites of metastases and low-volume recurrent disease could greatly improve management of patients with advanced prostate cancer. Prostate-specific membrane antigen (PSMA) is a well-established marker for prostate carcinoma with increased levels of expression in high-grade, hormone-refractory and metastatic disease. Elevated expression of PSMA is found in virtually all prostate cancers with the highest levels found in high-grade, hormone-refractory and metastatic disease [2,3,4,5,6]. Imaging studies with PSMA-specific small molecules in man have further shown accumulation in lacrimal and salivary glands [9,10]. This very restricted expression pattern makes PSMA an excellent target for detection and targeted therapy of prostate cancer. Not validated as a predictive marker, it has been suggested that PSMA expression levels in the primary tumour can predict disease outcome [11,12]
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