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Abstract Small extracellular vesicles (sEVs) provide major promise for advances in cancer diagnostics, prognostics, and therapeutics, ascribed to their distinctive cargo reflective of pathophysiological status, active involvement in intercellular communication, as well as their ubiquity and stability in bodily fluids. As a result, the field of sEV research has expanded exponentially. Nevertheless, there is a lack of standardisation in methods for sEV isolation from cells grown in serum‐containing media. The majority of researchers use serum‐containing media for sEV harvest and employ ultracentrifugation as the primary isolation method. Ultracentrifugation is inefficient as it is devoid of the capacity to isolate high sEV yields without contamination of non‐sEV materials or disruption of sEV integrity. We comprehensively evaluated a protocol using tangential flow filtration and size exclusion chromatography to isolate sEVs from a variety of human and murine cancer cell lines, including HeLa, MDA‐MB‐231, EO771 and B16F10. We directly compared the performance of traditional ultracentrifugation and tangential flow filtration methods, that had undergone further purification by size exclusion chromatography, in their capacity to separate sEVs, and rigorously characterised sEV properties using multiple quantification devices, protein analyses and both image and nano‐flow cytometry. Ultracentrifugation and tangential flow filtration both enrich consistent sEV populations, with similar size distributions of particles ranging up to 200 nm. However, tangential flow filtration exceeds ultracentrifugation in isolating significantly higher yields of sEVs, making it more suitable for large‐scale research applications. Our results demonstrate that tangential flow filtration is a reliable and robust sEV isolation approach that surpasses ultracentrifugation in yield, reproducibility, time, costs and scalability. These advantages allow for implementation in comprehensive research applications and downstream investigations.

Background: Lenvatinib and atezolizumab plus bevacizumab(A + B) have been used for unresectable hepatocellular carcinoma (HCC) as first‐line therapy. Real‐world studies comparison of efficacy and safety in these two regimens are limited, we therefore conduct this study to investigate these issues. Methods: We retrospectively reviewed patients received lenvatinib (n = 46) and A + B (n = 46) as first‐line systemic therapy for unresectable HCC in a tertiary medical center. Objective response rate (ORR), progression free survival (PFS), and overall survival (OS) were evaluated according to modified Response Evaluation Criteria in Solid Tumors (mRECIST). Inverse probability weighting (IPW) was performed for baseline clinical features balance. Results: A total of 92 patients with median age of 63.8 year‐old, 78.3% male, 85.9% viral hepatitis infected, 67.4% BCLC stage C were enrolled. The median treatment and follow‐up duration were 4.7 months and 9.4 months, respectively. There was no significant difference in ORR (26.1% vs. 41.3%, p = 0.1226), PFS (5.9 vs. 5.3 months, p = 0.4066), and OS (not reached vs. not reached, p = 0.7128) between the lenvatinib and A + B groups. After IPW, the results of survival and response rate were also compared. Subgroup analysis suggested that using lenvatinib was not inferior to A + B in regards of PFS, including those with elder, Child‐Pugh class B, beyond up‐to‐seven, or portal vein invasion VP4 patients. Among the lenvatinib treated patients, multivariate analysis showed patients elder than 65‐year‐old was an independent predictor associated with shorter PFS (adjust HR: 2.085[0.914–4.753], p = 0.0213). The incidence rates of adverse events were similar between two groups (76 vs. 63%, p = 0.1740). Both of two regimens had similarly few impact on liver function by comparison of baseline, third month, and sixth month albumin‐bilirubin index and Child‐Pugh score. Conclusions: The efficacy and safety of lenvatinib are similar to A + B as a first‐line systemic therapy for unresectable HCC. [ABSTRACT FROM AUTHOR]

Background Purpose Methods Results Conclusions Long‐axis positron emission tomography (PET) has emerged as one of the recent research directions in PET due to its ability to significantly enhance sensitivity and counting performance for low‐dose imaging, rapid imaging, and whole‐body dynamic imaging.The PET system presented in this study is a long‐axis animal PET based on lutetium–yttrium orthosilicate and silicon photomultiplier, designed for whole‐body imaging in rats. It features a diameter of 143 mm and an axial length of 213.3 mm. This study evaluated the performance of this PET system in accordance with the National Electrical Manufacturers Association (NEMA) NU 4‐2008 standards.The performance evaluation was conducted according to the NEMA NU 4‐2008 standards in terms of spatial resolution, sensitivity, counting rate performance, scatter fraction (SF) and image quality. In addition, a rat imaging study was conducted to assess the imaging capability of this PET system.The average energy resolution of the PET system was 12.87%, the average coincidence timing resolution was 751 ps. The FWHM of spatial resolution reconstructed by filtered back projection and 3D‐OSEM‐PSF algorithm at 5 mm radial offset from the axial center were 1.65 and 0.88 mm. The peak absolute sensitivity measured by a point source at the center of the field of view was evaluated as 6.71% (361–661 keV) and 10.31% (250–750 keV). For the mouse‐like phantom, the SF was 11.0% and the peak noise equivalent counting rate (NECR) was 1193 kcps at 94.2 MBq (2.54 mCi). For the rat‐like phantom, the SF was 26.8% and the NECR was 682.5 kcps at 78.6 MBq (2.12 mCi).The performance measurement results demonstrate that this PET system exhibits high sensitivity and count rate performance, making it potential for high‐quality whole‐body dynamic imaging of rats. [ABSTRACT FROM AUTHOR]

Background: The combination of anti-angiogenic therapy and immune checkpoint inhibitors has revolutionized the management of unresectable hepatocellular carcinoma (uHCC). While an early-phase study demonstrated promising outcomes for lenvatinib plus pembrolizumab (L+P) in treating uHCC, the LEAP-002 trial did not meet its primary endpoint. However, the comparative efficacy between L+P and atezolizumab plus bevacizumab (A+B) as first-line treatment remains a topic of uncertainty. This study aimed to assess the effectiveness and safety of L+P in contrast to A+B among patients diagnosed with uHCC., Methods: We conducted a retrospective analysis of enrolled patients with uHCC who received L+P or A+B as initial systemic treatment at Chang Gung Memorial Hospital from June 2019 to December 2022. The overall survival (OS), progression-free survival (PFS), objective response rate (ORR), and disease control rate (DCR) by modified RECIST were compared., Results: 121 patients were recruited, with 37 receiving L+P and 84 receiving A+B. Among them, 95 (78.5%) patients were BCLC stage C, and 99 (81.8%) patients had viral etiology for HCC, predominantly chronic HBV (68.6%). Both the L+P and the A+B groups demonstrated comparable OS (18.2 months versus 14.6 months, p = 0.35) and PFS (7.3 months versus 8.9 months, p = 0.75). The ORR and DCR were similar. After propensity score matching, the results remained consistent between the matched patients. Treatment-related adverse events of any grade occurred in 30 (81.1%) in the L+P group and 62 (73.8%) in the A+B group., Conclusions: Our findings suggest that L+P and A+B exhibit comparable efficacy and safety profiles in real-world settings.

For advanced hepatocellular carcinoma (HCC), the best second-line treatment after first-line treatment with sorafenib is unclear. This study aimed to compared the efficacy of second-line regorafenib (a tyrosine kinase inhibitor) and immune checkpoint inhibitors (ICIs) in patients with advanced HCC after sorafenib therapy. This retrospective study included 89 patients with HCC treated with sorafenib, and then regorafenib (n = 58) or an ICI (n = 31). Treatment response, overall survival (OS) and progression-free survival (PFS) of the 2 groups were compared, and factors associated with post-treatment mortality or disease progression were evaluated. During follow-up period, compared to regorafenib, treatment with an ICI results in a slight increase in a 20% decrease of AFP (35.7% vs. 31.8%), complete response rate (6.5% vs. 0%), objective response rate (16.1% vs. 6.9%), median overall survival (13.3 vs. 5 months), and median PFS (3.0 vs. 2.6 months). Combined locoregional treatment (LRT) (hazard ratio [HR] = 0.40, 95% confidence interval [CI]: 0.15-0.99) during second-line treatment was associated with a decreased risk of post-treatment mortality. After propensity scoring matching, combined LRT during second-line treatment had longer post-treatment OS than patients without combined LRT. A 20% decrease of AFP (HR = 0.54, 95% CI: 0.31-0.94) was associated with a decreased risk of post-treatment disease progression. In conclusions, second-line treatment with regorafenib or ICI prolongs OS in patients with advanced HCC treated with sorafenib. Combined LRT during second-line treatment is associated with decreased post-treatment mortality. A 20% decrease of AFP level may be predictive of a lower rate of disease progression., Competing Interests: None., (AJCR Copyright © 2024.)

Immune checkpoint inhibitors (ICIs) with atezolizumab plus bevacizumab are promising agents for unresectable hepatocellular carcinoma (HCC). We tried to guide the treatment based on recent developed CRAFITY score combining with on-treatment AFP response. Eighty-nine patients who received atezolizumab plus bevacizumab regardless of as a first-line therapy or not for unresectable HCC were enrolled for analyses. Radiologic evaluation was based on modified Response Evaluation Criteria in Solid Tumors (mRECIST). The objective response rate (ORR) and disease control rate (DCR) were 25.0% and 65.5%, respectively. Multivariate analysis showed that low CRAFITY score (AFP<100 ng/ml or CRP<10 mg/l) and satisfactory AFP response at 6 weeks (≥75% decrease or ≤10% increase from baseline) were independent factors determining good overall survival (OS) (hazard ratio [HR]=0.143, P=0.002 & HR=0.337, P=0.031), progression-free survival (PFS) (HR=0.419, P=0.022 & HR=0.429, P=0.025) and good responder (odds ratio [OR]=1.763, P=0.044 & OR=3.881, P=0.011). Patients were further divided into three classes by combination of CRAFITY score and AFP response at 6 weeks [The CAR (CRAFITY score and AFP-Response) classification)]: low CRAFITY score with satisfactory AFP response at 6 weeks (class I), either high CRAFITY score or unsatisfactory AFP response at 6 weeks (class II) and high CRAFITY score together with unsatisfactory AFP response at 6 weeks (class III). ORR was 35.0%, 18.2%, and 0% in class I, II and III patients, respectively (overall P=0.034). Patients in the class I had the best OS and PFS, followed by class II and class III (median OS: not reached vs. 11.1 vs. 4.3 months, log-rank P<0.001; median PFS: 7.9 vs. 6.6 vs. 2.6 months, log-rank P=0.001). Combination CRAFITY score and AFP response at 6 weeks with AUROC predicts OS and tumor response to be 0.809 and 0.798, respectively, better than either CRAFITY score (0.771 & 0.750) or AFP response at 6 weeks (0.725 & 0.680) alone. In conclusions, the CAR classification which combining CRAFITY score and AFP response at 6 weeks provides a practical guidance for atezolizumab plus bevacizumab therapy in unresectable HCC patients., Competing Interests: None., (AJCR Copyright © 2022.)