Pretargeting with Cucurbituril-Adamantane Host-Guest Pair in Xenograft Models.

The goal of reducing the total-body radiation dose of macromolecule-based nuclear medicine with a 2-step pretargeting strategy has been achieved with several pretargeting methodologies in preclinical and clinical settings. However, the lack of modularity, biocompatibility, and in vivo stability in existing pretargeting agents obstructs their respective platforms' wide clinical use. We hypothesized that host-guest chemistry would provide an optimal pretargeting methodology. A cucurbit[7]uril host and an adamantane guest molecule form a high-affinity host-guest complex (association constant, ∼1014 M-1), and in this work, we explored the use of this noncovalent interaction as the basis for antibody-based pretargeted PET. Along with the straightforward modularity of these agents, cucurbit[7]uril and adamantane are recognized to have high in vivo stability and suitability for human use, which is why we proposed this methodology as the ideal approach for pretargeted nuclear medicine. Methods: Three 64Cu-labeled adamantane guest radioligands were developed, and their in vitro stability, lipophilicity, and in vivo blood half-lives were compared. The adamantane radioligands were analyzed for pretargeting using a cucurbit[7]uril-modified carcinoembryonic antigen-targeting full-length antibody, hT84.66-M5A, as the macromolecule pretargeting agent with 2 different dosing schedules. These molecules were evaluated for pretargeting in human pancreatic cancer BxPC3 and MIAPaCa-2 mouse xenografts using PET and in vivo biodistribution studies. The dosimetry of the cucurbit[7]uril-adamantane (CB7-Adma) pretargeting approach in men was calculated and compared with that of the directly 89Zr-labeled hT84.66-M5A. Results: The adamantane radioligands possessed high in vitro stability up to 24 h (>90%). Pretargeted PET with CB7-Adma methodology resulted in specific tumor uptake (P < 0.05) with low background signal. The in vivo formed CB7-Adma complex was demonstrated to be stable, with high tumor uptake up to 24 h after radioligand injection (12.0 ± 0.9 percentage injected dose/g). The total-body radiation dose of the pretargeting strategy was only 3.3% that of the directly 89Zr-labeled hT84.66-M5A. Conclusion: The CB7-Adma strategy is highly suitable for pretargeted PET. The exceptional stability of the pretargeting agents and the specific and high tumor uptake of the pretargeted adamantane radioligands provide great potential for the platform.

Quantitative liver SPECT/CT is a novel tool to assess liver function, prognosis, and response to treatment in cirrhosis.

Background: Functional liver reserve is an important determinant of survival in cirrhosis. The traditional indocyanine green test (ICG) is cumbersome. Hence, we developed and validated a novel liver imaging, a hybrid of SPECT and CT (Q-SPECT/CT), for evaluating disease severity, outcomes, and response to treatment in decompensated cirrhosis (DC). Methods: We recruited a cohort of DC patients at a tertiary institute between 2016-2019. First, we standardized the Q-SPECT/CT across a predefined range of volumes through phantom experiments. Then we performed clinical and laboratory evaluations, ICG test (retention at 15 min), and Q-SPECT/CT at baseline and 12 months of granulocyte colony-stimulating factor (G-CSF) and standard medical treatment (SMT). Results: In 109 DC patients, 87.1% males, aged 51 ± 10 years, MELD: 14 (7-21), the percent quantitative liver uptake (%QLU) on Q-SPECT/CT exhibited a strong correlation with CTP (r = -0.728, p < 0.001), MELD (r = -0.743; p < 0.001) and ICG-R-15 (r = -0.720, p < 0.001) at baseline. %QLU had the maximum discrimination (AUC: 0.890-0.920), sensitivity (88.9-90.3%), specificity (81.2-90.7%), and accuracy (85.8-89.4%) than liver volumes on Q-SPECT/CT or ICG test for classifying patients in CTP/MELD based prognostic categories. A significant increase in %QLU (26.09 ± 10.06 to 31.2 ± 12.19, p = 0.001) and improvement in CTP/MELD correlated with better survival of G-CSF treated DC patients (p < 0.05). SMT did not show any improvement in Q-SPECT/CT or clinical severity scores (p > 0.05). %QLU > 25 (adj.H.R.: 0.234, p = 0.003) and G-CSF treatment (adj.H.R.: 0.414, p = 0.009) were independent predictors of better 12-months survival in DC. Conclusion: Q-SPECT/CT (%QLU) is a novel non-invasive, diagnostic, prognostic, and theragnostic marker of liver reserve and its functions in cirrhosis patients. Clinical trial registration: Clinicaltrials.gov, NCT02451033 and NCT03415698.

Evaluation of Reconstruction Algorithms to Validate the NEMA Phantom Results in Clinical Scenario - A Comparative Study Using Time-of-Flight versus Non-Time-of-Flight Positron Emission Tomography Imaging.

Objectives: The objective is to standardize the reconstruction parameters for the time-of-flight (TOF) versus non-TOF positron-emission tomography/computed tomography (PET/CT) imaging data and validation of the same in a clinical setting. Methods: The four spheres (10.0/13.0/17.0/22.0 mm) of the PET phantom (NEMA IQ Nu 2-2001) were filled with four times higher activity of [18F]-NaF than the background (5.3kBq/mL). Imaging (image matrix - 128 × 128 × 47, 2 min, 3D model) was done using two different (TOF/non-TOF) PET scanners. Phantom data were reconstructed in TOF and non-TOF modes for lutetium-yttrium oxyorthosilicate and non-TOF mode for bismuth germanate-based PET scanners. The reconstructed data (by varying iteration and subsets) that provided the best image contrast and signal-to-noise ratio (SNR) were evaluated. The whole-body [18F]-fludeoxyglucose (FDG) PET/CT scans (7-8 frames; 2.0 min/frame) in 16 lymphoma patients were acquired at 60 min after injecting the radioactivity (370.0-444.0 MBq of [18F]-FDG. The clinical PET/CT data were reconstructed using phantom-derived reconstruction parameters and evaluated for image contrast and SNR of the detected lesions. Results: TOF reconstruction at second iteration provided significantly (P ≤ 0.02) higher SNR (20.7) and contrast (contrast recovery coefficient/background variability = 3.21) for the smallest hot lesions (10.0 mm) in the phantom than the non-TOF system. Similarly, in patient data analysis for the selected FDG avid lesions, the SNR values were significantly (P = 0.02) higher (13.3 ± 6.49) in TOF than (11 ± 6.48) in non-TOF system. Further, the small (≤10.0 mm) lesions were seen more distinctly in TOF system. Conclusion: It is thus observed that TOF reconstruction converged faster than the non-TOF, and the applicability of the same may impact the image quality and interpretation in the clinical PET data. The validation of the phantom-based experimental reconstruction parameters to clinical PET imaging data is highly warranted.

Hand-grip strength as a screening tool for sarcopenia in males with decompensated cirrhosis.

BACKGROUND: Skeletal muscle index (SMI), the gold standard for sarcopenia, cannot measure muscle strength and requires specialized software and training. Hand-grip strength (HGS) measurement is cheap, requires minimal training and directly reflects muscle strength. We assessed the performance of HGS as a screening tool for sarcopenia in male patients with decompensated cirrhosis (DC). METHODS: Consecutive male DC patients (n=155) were enrolled. Baseline liver functions, etiologic work-up and anthropometric measurements were done. SMI was determined from computed tomography (CT) images at L3 level using ImageJ software. Sarcopenia was diagnosed using SMI cut-off <42 cm2/m2 as suggested by the Indian National Association for Study of the Liver. HGS was assessed using a hand-grip dynamometer. Diagnostic performance of HGS for discriminating sarcopenia was described using receiver operating characteristic (ROC) analysis. Diagnostic performance of different HGS cut-offs was assessed. Findings were internally validated using bootstrapping. RESULTS: Mean HGS and SMI were 25.73 ± 5.94 kg and 47.72 ± 8.71 cm2/m2, respectively. HGS showed modest correlation with SMI (tau: 0.31, p <0.001). Sarcopenia was seen in 41 (26.5%) patients. Age and HGS were independent predictors of sarcopenia on multivariate analysis. Area under the ROC curve (AUROC) of HGS for detecting sarcopenia was 0.73 (p<0.001). Optimal cut-off for using HGS as a screening tool was ≤31 kg (sensitivity: 37/41 [90.2%]; specificity: 29/114 [25.4%]; positive predictive value [PPV] : 37/122 [30.3%]; and negative predictive value [NPV]: 29/33 [87.9%]). CONCLUSION: Prevalence of sarcopenia in Indian male patients with DC is 26.5%. HGS is an independent predictor of sarcopenia and can be used as a screening tool to stratify the need for confirmatory CT-based assessment of sarcopenia.

Multiple Cycles of Granulocyte Colony-Stimulating Factor Increase Survival Times of Patients With Decompensated Cirrhosis in a Randomized Trial.

BACKGROUND & AIMS: There is controversy regarding the inclusion of granulocyte colony stimulating factor (G-CSF) in the treatment of decompensated cirrhosis. Previous studies tested only a single cycle of G-CSF administration or were underpowered to detect changes in survival time. We performed an adequately powered study to determine whether multiple cycles of G-CSF increased the survival of patients 1 year after the start of therapy. METHODS: We conducted an open-label trial of 100 patients with decompensated cirrhosis without acute-on-chronic liver failure at a tertiary center from July 2016 through June 2018. The patients were assigned randomly to a group given 5 days of G-CSF every 3 months, with standard medical therapy, in 4 cycles (group A, n = 50), or standard medical therapy alone (group B, n = 50). The primary outcome was survival for 12 months after treatment began. Secondary outcomes were an increase in the number of CD34+ cells at day 6 compared with day 0, along with reductions in Child-Turcotte-Pugh and model for end-stage liver disease scores, increased control of ascites, reduced decompensation and episodes of infection, fewer hospitalizations, lower liver stiffness measurements, increased quality of life and nutrition, fulfilment of liver transplant criteria, and fewer adverse events at 12 months after the start of treatment. RESULTS: Groups A and B were comparable at baseline. Survival at 12 months after initiation of treatment was significantly higher in group A (74%) than in group B (42%) (P < .001). Blood samples from patients in group A had significantly more CD34+ cells on day 6 than on day 0 (P < .001); there was no significant change in group B. Compared with patients in group B, patients in group A had significant reductions in Child-Turcotte-Pugh and model for end-stage liver disease scores, increased ascites control, fewer infections and hospitalizations, lower liver stiffness measurements, an increased quality of life, and a lower number fulfilled the liver transplant criteria (P < .05). There was no improvement in nutrition in either group compared with baseline. G-CSF was safe and well tolerated. CONCLUSIONS: Administration of multiple cycles of G-CSF increases the numbers of hematopoietic stem cells and survival of patients with decompensated cirrhosis receiving standard medical treatment. The addition of G-CSF to medical treatment might provide a bridge to liver transplantation for these patients. ClincialTrials.gov no: NCT03415698.

Radiosynthesis and pre-clinical evaluation of [68Ga] labeled antimicrobial peptide fragment GF-17 as a potential infection imaging PET radiotracer.

The antimicrobial peptide fragment GF-17 was synthesized in-house and conjugated with DOTA and measured molecular mass of DOTA-GF-17 conjugate was 2489 Da. The peptide conjugate was purified and labeled with [68Ga]. The best radiolabeling efficiency (95.0%) of [68Ga]DOTA-GF-17 was achieved at pH 4 with peptide conjugate amount of 20.0 nmol with 30 min of heating at 95 °C. The product remained stable for up to 3 h. The plasma protein binding and lipophilicity for [68Ga]DOTA-GF-17 were 80.98% and -3.12 respectively. The uptake studies with [68Ga]DOTA- GF-17 in S.aureus and P.aeruginosa bacterial strains demonstrated binding of 69.08% and 43.69% respectively. The animal bio-distribution and PET imaging studies were in agreement showing similar pattern for organs' tracer distribution and renal excretion. The tracer had rapid blood clearance and uptake in bone marrow and muscles was very low. The highest uptake of [68Ga]DOTA-GF-17 was observed at 45 min and 120 min in S.aureus and P.aeruginosa infections respectively. [68Ga]DOTA-GF-17 could be a promising PET tracer and holds a great scope for taking the product further to perform extensive PET studies in animal infection (using gram negative/positive strains) models to prove the diagnostic utility of this novel PET tracer for futuristic clinical applications.

Outcomes after multiple courses of granulocyte colony-stimulating factor and growth hormone in decompensated cirrhosis: A randomized trial.

Decompensated cirrhosis (DC) carries a high mortality. Liver transplantation (LT) is the treatment of choice; however, the limited availability of donor organs has resulted in high waitlist mortality. The present study investigated the impact of multiple courses of granulocyte-colony stimulating factor (G-CSF) with or without growth hormone (GH) in these patients. Sixty-five patients with DC were randomized to standard medical therapy (SMT) plus G-CSF 3 monthly plus GH daily (group A; n = 23) or SMT plus G-CSF (group B; n = 21) or SMT alone (group C; n = 21). The primary outcome was transplant-free survival (TFS) at 12 months. Secondary outcomes were mobilization of CD34+ cells at day 6 and improvement in clinical scores, liver stiffness, nutrition, episodes of infection, and quality of life (QOL) at 12 months. There was significantly better 12-month TFS in groups A and B than in group C (P = 0.001). At day 6 of therapy, CD34+ cells increased in groups A and B compared to baseline (P < 0.001). There was a significant decrease in clinical scores, improvement in nutrition, better control of ascites, reduction in liver stiffness, lesser infection episodes, and improvement in QOL scores in groups A and B at 12 months as compared to baseline (P < 0.05). The therapies were well tolerated. CONCLUSION: Multiple courses of G-CSF improved 12-month TFS, mobilized hematopoietic stem cells, improved disease severity scores, nutrition, fibrosis, QOL scores, ascites control, reduced infections, and the need for LT in patients with DC. However, the use of GH was not found to have any additional benefit. (Hepatology 2017).

Radiosynthesis of [18F]-fluorobenzoate-doxorubicin Using Acylation Approach.

BACKGROUND: Previously, we have labeled doxorubicin with [99Tc] and evaluated its potential as a SPECT agent to detect cancer in tumor bearing mice. In this study, we sought to radiolabel doxorubicin with [18F] using acylation method. METHODS: A quaternary salt of the precursor pentamethylbenzyl-4-(trimethylammonium trifluoromethanesulfonate) benzoate was synthesized and characterized by 1H-NMR. As a first step, 4-[18F]- fluorobenzoic acid (FBA) was synthesized from precursor. In second step, [18F]-FBA was further converted to its corresponding acyl form to radiolabel doxorubicin via acylation reaction. RESULTS: The total reaction time for the synthesis of [18F]-fluorobenzoate-doxorubicin was about 60 minutes. The radiolabeling efficiency of the final product was estimated to be about 59.0% The radiochemical yield for the synthesis of [18F]-FBA and [18F]-fluorobenzoate-doxorubicin were 19.0- 29.0% and 12.0-14.0% respectively. CONCLUSION: Radio synthesis of [18F]-fluorobenzoate-doxorubicin by acylation is a convenient method. However, further improvement in the labeling strategy is required to increase the radiolabeling efficiency and radio synthesis yield. Also, the radiolabeled product needs a detailed pre-clinical evaluation.