68Ga-DOTA-DiPSMA PET/CT Imaging: Biodistribution, Dosimetry and First Comparison with 68Ga-PSMA-11 in Prostate Cancer

Purpose This prospective trial aimed to evaluate the safety, dosimetry, biodistribution, and diagnostic ecacy of a novel theranostic probe 68 Ga-DOTA-DiPSMA. Also, we have performed the rst comparison with 68 Ga-PSMA-11 in prostate cancer (PCa) patients. Methods Five healthy volunteers and ten PCa patients with a previous clinical 68 Ga-PSMA-11 PET/CT were injected with an intravenous bolus of 68 Ga-DOTA-DiPSMA with a dose of 1.85MBq/kg. Healthy volunteers received serial whole-body PET scans from the time of injection up to 60 min post-injection, with a second PET/CT scanning at 120 min post-injection. In PCa patients, low-dose CT scan, whole-body PET was performed with 2 min per bed position in 40 min post-injection. In addition, 68 Ga-PSMA-11 scanning was performed on PCa patients within 10 days under the same acquisition procedure. Absorbed organ doses and effective doses were calculated using OLINDA/EXM. Normal organ uptake and tumor lesion uptake (SUV max ) were measured. A lesion-by-lesion analysis was performed. shows comparable pharmacokinetics and detection ability in PCa patients that warrant further head-to-head comparison. Low non-specic uptake in salivary glands and kidneys of 68 Ga-DOTA-DiPSMA indicates potential radioligand therapy (RLT) application when labeled with 177 Lu, 90 Y, or 225 Ac.


Introduction
Prostate cancer (PCa) is the most frequently diagnosed cancer in men and one of the lethal malignant diseases leading to male cancer-related death worldwide [1]. The accurate presence and location of primary or recurrent tumors are critical for planning effective patient management [2].
The diagnostic capability of conventional anatomic imaging such as MRI and CT to determine prostate cancer is limited in metastases and speci city [3]. Only prostate biopsy is the de nitive way to con rm prostate cancer [1]. Multiple needle biopsy will increase the positive rate of lesions determination signi cantly. However, it is di cult to determine distant metastases and increase the risk of complications resulting from biopsy operation [1]. There has been an unmet need for more advanced imaging modalities to determine primary and metastatic lesions that can be helpful to prostate cancer patient management (observation, salvage local therapy, systemic therapy). PET with 18 F-FDG is effective for most malignant tumors, but it has insu cient speci city for prostate cancer and is prone to have false-positive results. Therefore, it is urgent to discover new nuclear medicine imaging agents with more speci city for prostate cancer.
Prostate-Speci c Membrane Antigen (PSMA) is a transmembrane glycoprotein enzyme selectively overexpressed in prostate cancer cells, with its expression increasing in higher-grade malignancy [4]. PET imaging with PSMA probes targeting various prostate cancer-speci c markers will provide additional molecular information to facilitate lesion detection and staging [5].
Recently, a relatively new nuclear imaging modality 68 Ga-PSMA PET/CT imaging with good PCa diagnosis and staging performance, has become increasingly utilized to evaluate PCa aggressiveness, especially in patients with biochemical recurrence after surgery [6-8]. PSMA can be coupled with different chelators and labeled with corresponding radionuclides for different purposes. The most widely used PSMA ligands in clinical examination are PSMA-11 and PSMA-617 containing different linkers and chelators. According to the previously published papers, 68 Ga-PSMA-11 and 177 Lu-PSMA-617 are molecular-pair in metastatic castration-resistant PCa (mCRPC) diagnosis and radioligand therapy (RLT) [9,10]. However, considering the high nonspeci c uptake in the salivary, kidney, and bone marrow of 68 Ga-PSMA-11 and 177 Lu-PSMA-617, novel PSMA tracers with low accumulation in normal organs are urgently needed.
We have discovered a PSMA dimer (DOTA-DiPSMA, Fig. 1), and proved its imaging ability and low unspeci c uptake (salivary, kidney) in preclinical experiments, which will be published later. In this study, we aimed to evaluate the safety, biodistribution and dosimetry of 68 Ga-DOTA-DiPSMA in healthy volunteers. Furthermore, a head-to-head comparison of 68 Ga-DOTA-DiPSMA vs. 68 Ga-PSMA-11, including lesion detection rate, tumor uptake, and biodistribution in PCa patients were performed.  Fig. 2). Five healthy volunteers (5 men, age range 42-76 y mean age ± SD, 59.83 ±11.65 y]; weight range, 55.0-78.0 kg [mean weight ±SD, 70.27 ± 13.05 kg]) were enrolled to validate the safety, biodistribution and radiation dosimetry of 68 Ga-DOTA-DiPSMA in this study. Exclusion criteria consisted of mental illness conditions, severe liver or kidney disease with serum creatinine greater than 3.0 mg/dL (270 mM), or any hepatic enzyme level 5 times or more than the standard upper limit. Participants were also excluded if they were known to have severe allergy or hypersensitivity to intravenous radiographic contrast or claustrophobia during PET/CT scanning.

Methods
A total of 10 patients who were newly diagnosed as having prostate cancer by sextant core-needle biopsy and had not received any prior therapy were enrolled with written informed consent. The inclusion criteria were being between 40 and 80 years old, having a prostate neoplasm identi ed by ultrasound or MRI, being diagnosed by needle biopsy as having prostate cancer. The exclusion criteria included claustrophobia, kidney or liver failure, and inability to ful ll the study. The demographics of healthy volunteers and patients are listed in Table 1

Safety Assessment
Patient safety was assessed and graded according to Common Terminology Criteria for Adverse Events (version 5.0), electrocardiograms, physical examination, and vital signs (blood pressure, respiratory rate, heart rate, body temperature). Within the rst 72 h after 68 Ga-DOTA-DiPSMA injection, the research team kept phone contact with each subject monitoring their adverse event (AE) responses.

Radiopharmaceutical Preparation
Precursors were supplied by Prof. Cui from Key Laboratory of Radiopharmaceuticals, Ministry of Education, Beijing Normal University. 68 Ga 3+ was eluted from a 68 Ge/ 68 Ga generator (JSC Isotope, Russian) using 0.1 M HCl and mixed with 1.25 M NaOAc buffer to adjust pH to around 4.0. Radiolabeling of DiPSMA-DOTA-COOH was performed in a sterile hot cell. The radiochemical purity of the product 68 Ga-DOTA-DiPSMA exceeded 99%. 68 Ga-PSMA-11 was synthesized following a procedure reported previously [12].

Examination Procedures
For healthy volunteers, the blood pressure, pulse, respiratory frequency, and temperature were measured, and routine blood and urine tests, liver function, and renal function were examined immediately before and 24 h after the scan. In addition, any possible side effects during 68  volunteers. The whole-body background of 68 Ga-DOTA-DiPSMA was low. The highest uptake was observed in the kidney with a SUV mean of 43.4 ± 26.8 at 5 min p.i. and further decreased to 11.4 ± 6.5 at 120 min p.i. The spleen, liver, salivary gland, and small intestine showed moderate uptake, with SUV mean of 2.90 ± 1.5, 1.89 ± 0.75, 2.30 ± 0.87, and 2.42 ± 0.64 at 30 min after injection, respectively. Low background uptake was observed in the brain, lungs, muscle, red marrow, heart, thyroid, gall bladder, pancreas, stomach, bone, and large intestine. The rapid presence in the kidneys, followed by a passage toward the urinary bladder, illustrates the tracer's fast and mainly renal excretion.

Dosimetry
The average estimated absorbed organ in healthy volunteers was summarized in Table 2. The highest absorbed dose was received by the kidneys (114.46 ± 29.28 uSv/MBq), followed by the urinary bladder wall (100.82 ± 46.22 uSv/MBq). The mean effective dose was 19.46 ± 1.73 μSv/MBq.

Detection of Primary Prostate Cancer
For the 10 patients with primary prostate cancer, 68 Ga-DOTA-DiPSMA PET/CT showed 27 positive ndings including 16 prostate lesions, 4 bone metastases, 5 lymph node metastases and 2 seminal vesicle metastases. The primary lesions were con rmed by needle biopsy. SUV max for prostate lesions, bone metastases, and lymph node metastases were 4.41 ± 2.72, 2.95 ± 1.11 and 3.26 ± 1.2, respectively.

Discussion
To our knowledge, this is the rst human study to evaluate the novel tracer 68 Ga-DOTA-DiPSMA in healthy volunteers and patients with PCas. This tracer is a new type of Ga-68-labeled dimer PSMA imaging agent with a simple structure, easy synthesis, and low synthesis cost. 68 Ga-DOTA-DiPSMA can be prepared by a one-step labeling reaction in high yield greater than 95% between 68 Ga 3+ ions eluted from a germaniumgallium generator and the precursor DiPSMA-DOTA-COOH.
Here, we presented the results of an independently performed rst clinical evaluation of 68 Ga-DOTA-DiPSMA in ve healthy volunteers, including biodistribution, dosimetry and safety. Also, we have performed the head-to-head lesion comparison of 68 Ga-DOTA-DiPSMA to 68 Ga-PSMA-11 in ten PCa patients.
The results showed that this tracer displayed favorable biodistribution, dosimetry features and well- respectively. 68 Ga-DOTA-DiPSMA seems to have a lower uptake in small lesions and distant metastasis [21][22]. However, the lower uptake of 68 Ga-DOTA-DiPSMA in normal organs than 68 Ga-PSMA-11 maybe its advantage. The small lesion near the urinary bladder would be more apparent with this relatively low background.
The primary limitation of our study is the sample size which did not enable accurate multivariate regression analysis about comparing 68 Ga-DOTA-DiPSMA to 68 Ga-PSMA-11. Besides, neither blood nor urine samples were collected in our study, which will allow for the stability test in vivo. Further detailed and head-to-head comparison studies are required. Con ict of interest The authors declare that they have no con ict of interest.
Availability of data and materials Not applicable.
Code availability Not applicable.
Ethics approval All procedures involving human participants were carried out in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 Helsinki Declaration and its later amendments or comparable ethical standards. This article does not contain any experiments with animals.    Study owchart with excluded healthy volunteers and patients.

Figure 3
Biodistribution of 68Ga-DOTA-DiPSMA in healthy volunteers. A maximum intensity projection PET images at several time points post-injection of a 56-year-old male healthy volunteer.   Normal organs SUVmax (Average in 10 patients) of 68Ga-DOTA-DiPSMA as compared with 68Ga-PSMA-11