Deciphering the pathogenic role of rare RAF1 heterozygous missense mutation in the late-presenting DDH.

Background: Developmental Dysplasia of the Hip (DDH) is a skeletal disorder where late-presenting forms often escape early diagnosis, leading to limb and pain in adults. The genetic basis of DDH is not fully understood despite known genetic predispositions. Methods: We employed Whole Genome Sequencing (WGS) to explore the genetic factors in late-presenting DDH in two unrelated families, supported by phenotypic analyses and in vitro validation. Results: In both cases, a novel de novo heterozygous missense mutation in RAF1 (c.193A>G [p.Lys65Glu]) was identified. This mutation impacted RAF1 protein structure and function, altering downstream signaling in the Ras/ERK pathway, as demonstrated by bioinformatics, molecular dynamics simulations, and in vitro validations. Conclusion: This study contributes to our understanding of the genetic factors involved in DDH by identifying a novel mutation in RAF1. The identification of the RAF1 mutation suggests a possible involvement of the Ras/ERK pathway in the pathogenesis of late-presenting DDH, indicating its potential role in skeletal development.

Valence-Change MnO2-Coated Arsenene Nanosheets as a Pin1 Inhibitor for Hepatocellular Carcinoma Treatment.

The heterogeneity of hepatocellular carcinoma (HCC) can prevent effective treatment, emphasizing the need for more effective therapies. Herein, we employed arsenene nanosheets coated with manganese dioxide and polyethylene glycol (AMPNs) for the degradation of Pin1, which is universally overexpressed in HCC. By employing an "AND gate", AMPNs exhibited responsiveness toward excessive glutathione and hydrogen peroxide within the tumor microenvironment, thereby selectively releasing AsxOy to mitigate potential side effects of As2O3. Notably, AMPNs induced the suppressing Pin1 expression while simultaneously upregulation PD-L1, thereby eliciting a robust antitumor immune response and enhancing the efficacy of anti-PD-1/anti-PD-L1 therapy. The combination of AMPNs and anti-PD-1 synergistically enhanced tumor suppression and effectively induced long-lasting immune memory. This approach did not reveal As2O3-associated toxicity, indicating that arsenene-based nanotherapeutic could be employed to amplify the response rate of anti-PD-1/anti-PD-L1 therapy to improve the clinical outcomes of HCC patients and potentially other solid tumors (e.g., breast cancer) that are refractory to anti-PD-1/anti-PD-L1 therapy.

Buoyant Metal-Organic Framework Corona-Driven Fast Isolation and Ultrasensitive Profiling of Circulating Extracellular Vesicles.

Accurately assaying tumor-derived circulating extracellular vesicles (EVs) is fundamental in noninvasive cancer diagnosis and therapeutic monitoring but limited by challenges in efficient EV isolation and profiling. Here, we report a bioinspired buoyancy-driven metal-organic framework (MOF) corona that leverages on-bubble coordination and dual-encoded surface-enhanced Raman scattering (SERS) nanotags to streamline rapid isolation and ultrasensitive profiling of plasma EVs in a single assay for cancer diagnostics. This integrated bubble-MOF-SERS EV assay (IBMsv) allows barnacle-like high-density adhesion of MOFs on a self-floating bubble surface to enable fast isolation (2 min, near 90% capture efficiency) of tumor EVs via enhanced EV-MOF binding. Also, IBMsv harnesses four-plexed SERS nanotags to profile the captured EV surface protein markers at a single-particle level. Such a sensitive assay allows multiplexed profiling of EVs across five cancer types, revealing heterogeneous EV surface expression patterns. Furthermore, the IBMsv assay enables cancer diagnosis in a pilot clinical cohort (n = 55) with accuracies >95%, improves discrimination between cancer and noncancer patients via an algorithm, and monitors the surgical treatment response from hepatocellular carcinoma patients. This assay provides a fast, sensitive, streamlined, multiplexed, and portable blood test tool to enable cancer diagnosis and response monitoring in clinical settings.

Flexible DNA Nanoclaws Offer Multivalent and Powerful Spatial Pattern-Recognition for Tumor Cells.

Multivalent receptor-ligand interactions (RLIs) exhibit excellent affinity for binding when targeting cell membrane receptors with low expression. However, existing strategies only allow for limited control of the valency and spacing of ligands for a certain receptor, lacking recognition patterns for multiple interested receptors with complex spatial distributions. Here, we developed flexible DNA nanoclaws with multivalent aptamers to achieve powerful cell recognition by controlling the spacing of aptamers to match the spatial patterns of receptors. The DNA nanoclaw with spacing-controllable binding sites was constructed via hybrid chain reaction (HCR), enabling dual targeting of HER2 and EpCAM molecules. The results demonstrate that the binding affinity of multivalent DNA nanoclaws to tumor cells is enhanced. We speculate that the flexible structure may conform better to irregularly shaped membrane surfaces, increasing the probability of intermolecular contact. The capture efficiency of circulating tumor cells successfully verified the high affinity and selectivity of this spatial pattern. This strategy will further promote the potential application of DNA frameworks in future disease diagnosis and treatment.

Low-dose cadmium induces lymphangiogenesis through activation of the STAT3 signaling pathway.

Cadmium (Cd) is a widespread environmental toxicant that poses significant threat to public health. After intake, Cd is distributed throughout the body via blood and lymphatic circulation. However, the effect of Cd on lymphatic vessels has not been revealed. In this study, mice were exposed to 10 µM cadmium chloride through drinking water immediately after corneal alkali burn. In vivo analyses showed that Cd treatment enhances the alkali burn-induced corneal lymphangiogenesis, which is characterized by increased expression of lymphatic vessel endothelial hyaluronan receptor 1 (LYVE-1), prospero-related homeobox 1 (PROX-1) and vascular endothelial growth factor receptor 3 (VEGFR3). In vitro, the proliferation and migration of human dermal lymphatic endothelial cells (HDLECs) are increased by 1 µM Cd treatment, while inhibited by 10 µM Cd treatment. At a concentration of 1 µM, Cd specifically induces phosphorylation of signal transducer and activator of transcription 3 (STAT3), but has no effect on the MAPK, AKT, or NF-κB signaling pathway. In the presence of the STAT3 inhibitor STATTIC, Cd fails to induce HDLECs proliferation and migration. In addition, Cd upregulates VEGFR3 expression and its gene promoter activity in a STAT3-dependent manner. Our study suggests that low-dose Cd promotes lymphangiogenesis through activation of the STAT3 signaling pathway.

Micro-Engineered Organoid-on-a-Chip Based on Mesenchymal Stromal Cells to Predict Immunotherapy Responses of HCC Patients.

Hepatocellular carcinoma (HCC) is one of the most lethal cancers worldwide. Patient-derived organoid (PDO) has great potential in precision oncology, but low success rate, time-consuming culture, and lack of tumor microenvironment (TME) limit its application. Mesenchymal stromal cells (MSC) accumulate in primary site to support tumor growth and recruit immune cells to form TME. Here, MSC and peripheral blood mononuclear cells (PBMC) coculture is used to construct HCC organoid-on-a-chip mimicking original TME and provide a high-throughput drug-screening platform to predict outcomes of anti-HCC immunotherapies. HCC-PDOs and PBMC are co-cultured with MSC and Cancer-associated fibroblasts (CAF). MSC increases success rate of biopsy-derived PDO culture, accelerates PDO growth, and promotes monocyte survival and differentiation into tumor-associated macrophages. A multi-layer microfluidic chip is designed to achieve high-throughput co-culture for drug screening. Compared to conventional PDOs, MSC-PDO-PBMC and CAF-PDO-PBMC models show comparable responses to chemotherapeutic or targeted anti-tumor drugs but more precise prediction potential in assessing patients' responses to anti-PD-L1 drugs. Moreover, this microfluidic platform shortens PDO growth time and improves dimensional uniformity of organoids. In conclusion, the study successfully constructs microengineered organoid-on-a-chip to mimic TME for high-throughput drug screening, providing novel platform to predict immunotherapy response of HCC patients.

pH-sensitive tumor-tropism hybrid membrane-coated nanoparticles for reprogramming the tumor microenvironment and boosting the antitumor immunity.

Metabolic dysregulation contributes not only to cancer development but also to a tumor immune microenvironment (TIME), which poses great challenges to chemo- and immunotherapy. Targeting metabolic reprogramming has recently emerged as a promising strategy for cancer treatment, but the lethality against solid tumors appears to be fairly restricted, partially due to the poor solubility of small molecule drugs. Herein, we construct a versatile biomimetic nanoplatform (referred to as HM-BPT) employing pH-sensitive tumor-tropism hybrid membrane-coated Manganese oxide (MnO2) nanoparticles for the delivery of BPTES, a glutamine metabolism inhibitor. Basically, hybrid membranes consisting of mesenchymal stem cell membranes (MSCm) and pH-sensitive liposomes (pSL) enable the biomimetic nanoplatform to target TME and escape from endo/lysosomes after endocytosis. The results reveal that HM-BPT treatment leads to remarkable tumor inhibition, cytotoxic T lymphocyte (CTL) infiltration, as well as M1 phenotype repolarization and stimulator of IFN genes (STING) pathway activation in macrophages in a 4T1 xenograft model. Furthermore, glutathione (GSH) depletion and oxygen (O2) supply synergistically ameliorate the immunosuppressive status of the TME, boosting potent antitumor immune responses. Overall, our study explores an integrated therapeutic platform for TME reprogramming and immune activation, offering tremendous promise for cancer combination therapy. STATEMENT OF SIGNIFICANCE: Metabolic abnormalities and the tumor immune microenvironment (TIME) lead to hyporesponsiveness to conventional therapies, ultimately resulting in refractory malignancies. In the current work, a biomimetic nanoplatform (HM-BPT) was developed for TME metabolic reprogramming in favor of immunotherapy. Particularly, hybrid membrane camouflage endowed the nanoplatform with TME targeting, endo/lysosomal escape, and sensitive release properties. The impact of hybrid membrane fusion ratio on cellular uptake and cell viability was explored, yielding beneficial references for the future development of bioactive nanomaterials. Intravenous administration of HM-BPT substantially relieved tumor burden and restored innate and acquired immune activation in 4T1 xenograft models. In conclusion, the created HM-BPT system has the potential to be a promising nanoplatform for combining cancer therapies.

Anti-PD-L1 antibody enhances curative effect of cryoablation via antibody-dependent cell-mediated cytotoxicity mediating PD-L1highCD11b+ cells elimination in hepatocellular carcinoma.

Cryoablation (CRA) and microwave ablation (MWA) are two main local treatments for hepatocellular carcinoma (HCC). However, which one is more curative and suitable for combining with immunotherapy is still controversial. Herein, CRA induced higher tumoral PD-L1 expression and more T cells infiltration, but less PD-L1highCD11b+ myeloid cells infiltration than MWA in HCC. Furthermore, CRA had better curative effect than MWA for anti-PD-L1 combination therapy in mouse models. Mechanistically, anti-PD-L1 antibody facilitated infiltration of CD8+ T cells by enhancing the secretion of CXCL9 from cDC1 cells after CRA therapy. On the other hand, anti-PD-L1 antibody promoted the infiltration of NK cells to eliminate PD-L1highCD11b+ myeloid cells by antibody-dependent cell-mediated cytotoxicity (ADCC) effect after CRA therapy. Both aspects relieved the immunosuppressive microenvironment after CRA therapy. Notably, the wild-type PD-L1 Avelumab (Bavencio), compared to the mutant PD-L1 atezolizumab (Tecentriq), was better at inducing the ADCC effect to target PD-L1highCD11b+ myeloid cells. Collectively, our study uncovered the novel insights that CRA showed superior curative effect than MWA in combining with anti-PD-L1 antibody by strengthening CTL/NK cell immune responses, which provided a strong rationale for combining CRA and PD-L1 blockade in the clinical treatment for HCC.

Modular DNA-Origami-Based Nanoarrays Enhance Cell Binding Affinity through the "Lock-and-Key" Interaction.

Surface proteins of cells are generally recognized through receptor-ligand interactions (RLIs) in disease diagnosis, but their nonuniform spatial distribution and higher-order structure lead to low binding affinity. Constructing nanotopologies that match the spatial distribution of membrane proteins to improve the binding affinity remains a challenge. Inspired by the multiantigen recognition of immune synapses, we developed modular DNA-origami-based nanoarrays with multivalent aptamers. By adjusting the valency and interspacing of the aptamers, we constructed specific nanotopology to match the spatial distribution of target protein clusters and avoid potential steric hindrance. We found that the nanoarrays significantly enhanced the binding affinity of target cells and synergistically recognized low-affinity antigen-specific cells. In addition, DNA nanoarrays used for the clinical detection of circulating tumor cells successfully verified their precise recognition ability and high-affinity RLIs. Such nanoarrays will further promote the potential application of DNA materials in clinical detection and even cell membrane engineering.

Synthesis of dihydrofuran-3-one and 9,10-phenanthrenequinone hybrid molecules and biological evaluation against colon cancer cells as selective Akt kinase inhibitors.

A series of dihydrofuran-3-one and 9,10-phenanthrenequinone hybrid compounds were synthetized through a one-pot gold-catalyzed oxidative cyclization and Aldol-type addition cascade reaction of homopropargylic alcohols with 9,10-phenanthrenequinone. The cytotoxicity of newly synthesized compounds was evaluated in CCK8 assay against different human cancer cells, showing significantly antiproliferative activity against tested tumor cell lines with a lowest IC50 value of 0.92 µM over HCT-116. Further investigation revealed that the treatment of HCT-116 cell line with the promising compound 4c induced cell death as a selective Akt inhibitor. In addition, controlled experiments and molecular docking study suggested that the significant antitumor activity might be attributed to the unique hybrid structure, which implied the promising potential of this dual heterocycle hybrid method in the discovery of novel bioactive molecules with structural diversity.

Identifying the Phenotypes of Tumor-Derived Extracellular Vesicles Using Size-Coded Affinity Microbeads.

Tumor-derived extracellular vesicle (tEV) biomarkers can reflect cancer cell phenotypes and have great potential for cancer diagnosis and treatment. However, tEVs display high heterogeneity, and rapid and sensitive identification of EV biomarkers remains challenging due to their low expression. Spectral overlap also significantly limits the multiplex analysis of EV biomarkers by fluorescent probes. Herein, we developed a method for highly sensitive tEV phenotyping that uses size-coded microbeads that carry hairpin probes that can bind to aptamers targeting distinct tEV biomarkers. We also designed a microfluidic chip containing spacer arrays that segregate these microbeads in distinct chip regions according to their size to generate location-specific signals indicating the level of different EV biomarkers. The EV biomarker signal on these microbeads was amplified by in situ rolling cyclic amplification (RCA). This strategy permits the simultaneous detection of multiple tEV phenotypes by fluorescence spectroscopy without the limitations of spectral overlap. This study demonstrates that this tEV phenotyping method can rapidly and simultaneously detect six different tEV phenotypes with high sensitivity. Due to the programmability of the sensing platform, this method can be rapidly adapted to detect different tEV phenotype substitutions of the detected biomarkers. Notably, clinical cohort studies show that this strategy may provide new ideas for the precise diagnosis and personalized treatment of cancer patients.

TREM2/ß-catenin attenuates NLRP3 inflammasome-mediated macrophage pyroptosis to promote bacterial clearance of pyogenic bacteria.

Triggering receptors expressed on myeloid cells 2 (TREM2) is considered a protective factor to protect host from bacterial infection, while how it elicits this role is unclear. In the present study, we demonstrate that deficiency of triggering receptors expressed on myeloid cells 2 (TREM2) significantly enhanced macrophage pyroptosis induced by four common pyogenic bacteria including Staphylococcus aureus, Pseudomonas aeruginosa, Streptococcus pneumoniae, and Escherichia coli. TREM2 deficiency also decreased bacterial killing ratio of macrophage, while Caspase-1 or GSDMD inhibition promoted macrophage-mediated clearance to these bacteria. Further study demonstrated that the effect of TREM2 on macrophage pyroptosis and bacterial eradication mainly dependents on the activated status of NLRP3 inflammasome. Moreover, as the key downstream of TREM2, ß-catenin phosphorylated at Ser675 by TREM2 signal and accumulated in nucleus and cytoplasm. ß-catenin mediated the effect of TREM2 on NLRP3 inflammasome and macrophage pyroptosis by reducing NLRP3 expression, and inhibiting inflammasome complex assembly by interacting with ASC. Collectively, TREM2/ß-catenin inhibits NLRP3 inflammasome to regulate macrophage pyroptosis, and enhances macrophage-mediated pyogenic bacterial clearance.

In situ signal amplification improves the capture efficiency of circulating tumor cells with low expression of EpCAM.

Circulating tumor cells (CTCs) as non-invasive biomarkers have great potential in evaluating tumor progression and prognosis. However, effective enrichment of CTCs and minimizing phenotypic bias remain a serious challenge. Herein, a DNA tetrahedron-aptamer complex-mediated rolling circle amplification (TDN-RCA) strategy is developed for cell surface protein signal amplification and CTC enrichment, employing DNA tetrahedron-EpCAM aptamer complex as a scaffold and initiating rolling circle amplification (RCA) reaction on the surface of CTCs in situ. The DNA tetrahedron-aptamer complex enables the cell-specific recognition and enhances cell membrane anchoring ability, generating a large number of magnetic beads binding sites through the RCA reaction in situ. Thus, the signals of cell surface markers with low expression levels are amplified in situ and then efficient CTC enrichment is achieved. This method improves the capture efficiency of CTCs with low expression of EpCAM, which has great potential in clinical application.

A Dual-Encoded Bead-Based Immunoassay with Tunable Detection Range for COVID-19 Serum Evaluation.

Serological assay for coronavirus 2019 (COVID-19) patients including asymptomatic cases can inform on disease progression and prognosis. A detection method taking into account multiplex, high sensitivity, and a wider detection range will help to identify and treat COVID-19. Here we integrated color-size dual-encoded beads and rolling circle amplification (RCA) into a bead-based fluorescence immunoassay implemented in a size sorting chip to achieve high-throughput and sensitive detection. We used the assay for quantifying COVID-19 antibodies against spike S1, nucleocapsid, the receptor binding domain antigens. It also detected inflammatory biomarkers including interleukin-6, interleukin-1ß, procalcitonin, C-reactive protein whose concentrations range from pg mL-1 to µg mL-1 . Use of different size beads integrating with RCA results in a tunable detection range. The assay can be readily modified to simultaneously measure more COVID-19 serological molecules differing by orders of magnitude.

Tetrahedral DNA nanostructures synergize with MnO2 to enhance antitumor immunity via promoting STING activation and M1 polarization.

Stimulator of interferon genes (STING) is a cytosolic DNA sensor which is regarded as a potential target for antitumor immunotherapy. However, clinical trials of STING agonists display limited anti-tumor effects and dose-dependent side-effects like inflammatory damage and cell toxicity. Here, we showed that tetrahedral DNA nanostructures (TDNs) actively enter macrophages to promote STING activation and M1 polarization in a size-dependent manner, and synergized with Mn2+ to enhance the expressions of IFN-ß and iNOS, as well as the co-stimulatory molecules for antigen presentation. Moreover, to reduce the cytotoxicity of Mn2+, we constructed a TDN-MnO2 complex and found that it displayed a much higher efficacy than TDN plus Mn2+ to initiate macrophage activation and anti-tumor response both in vitro and in vivo. Together, our studies explored a novel immune activation effect of TDN in cancer therapy and its synergistic therapeutic outcomes with MnO2. These findings provide new therapeutic opportunities for cancer therapy.

Doxorubicin-Loaded UiO-66/Bi2S3 Nanocomposite-Enhanced Synergistic Transarterial Chemoembolization and Photothermal Therapy against Hepatocellular Carcinoma.

Transcatheter arterial chemoembolization (TACE) is the first choice for patients with intermediate hepatocellular carcinoma (HCC), but clinical applications still face some problems, such as the difficulties in clearing all cancer cells and lack of targeting, which would damage normal liver cells. Recently, photothermal therapy (PTT) and nanodelivery systems have been used to improve the efficacy of TACE. However, most of these strategies achieve only a single function, and the synthesis process is complicated. Here, a simple one-step solvothermal method was used to develop multifunctional nanoparticles (UiO-66/Bi2S3@DOX), which can simultaneously achieve photothermal effects and low pH-triggered DOX release. UiO-66/Bi2S3 exhibited a pH-responsive release behavior and an excellent photothermal effect in a series of in vitro and in vivo studies. Biocompatibility was confirmed by cytotoxicity and hemocompatibility evaluations. The rat N1S1 liver tumor model was established to investigate the therapeutic effect and biosafety of the nanoplatforms using TACE. The results revealed that the combination of TACE and PTT resulted in remarkable tumor growth inhibition, and the histopathological assay further revealed extensive necrosis, downregulated angiogenesis, increased apoptosis, and proliferation in the tumor response. These results demonstrated that this nanosystem platform was a promising therapeutic agent for enhancing TACE therapy for HCC treatment.

Accurate Isolation of Circulating Tumor Cells via a Heterovalent DNA Framework Recognition Element-Functionalized Microfluidic Chip.

Detection of circulating tumor cells (CTCs) has provided a noninvasive and efficient approach for early diagnosis, treatment, and prognosis of cancer. However, efficient capture of CTCs in the clinical environment is very challenging because of the extremely rare and heterogeneous expression of CTCs. Herein, we fabricated a multimarker microfluidic chip for the enrichment of heterogeneous CTCs from peripheral blood samples of breast cancer patients. The multimarker aptamer cocktail DNA nanostructures (TP-multimarker) were modified on a deterministic lateral displacement (DLD)-patterned microfluidic chip to enhance the capture efficiency through the size selection effect of DLD arrays and the synergistic effect of multivalent aptamers. As compared to a monovalent aptamer-modified chip, the multimarker chip exhibits enhanced capture efficiency toward both high and low epithelial cell adhesion molecule expression cell lines, and the DNA nanostructure-functionalized chip enables the accurate capture of different phenotypes of CTCs. In addition, the DNA nanoscaffold makes nucleases more accessible to the aptamers to release cells with molecular integrity and outstanding cell viability.

Unbiased Enrichment of Circulating Tumor Cells Via DNAzyme-Catalyzed Proximal Protein Biotinylation.

Circulating tumor cells (CTCs) are noninvasive biomarkers with great potential for assessing neoplastic diseases. However, the enrichment bias toward heterogeneous CTCs remains to be minimized. Herein, a DNAzyme-catalyzed proximal protein biotinylation (DPPB) strategy is established for unbiased CTCs enrichment, employing DNA-framework-based, aptamer-coupled DNAzymes that bind to the surface marker of CTCs and subsequently biotinylated membrane proteins in situ. The DNA framework enables the construction of multivalent DNAzyme and serves as steric hindrance to avoid undesired interaction between DNAzymes and aptamer, leading to efficient binding and biotinylation. Compared with a biotinylated-aptamer strategy, fivefold lower bias of cell subpopulations was achieved by DPPB before and after capture, which enabled a 4.6-fold performance for CTCs analysis in clinic blood samples. DPPB is envisioned to offer a new solution for CTC-based cancer diagnostics.

MR imaging features of Lhermitte-Duclos disease: Case reports and literature review.

RATIONALE: Lhermitte-Duclos disease (LDD) is a rare tumor of the nervous system with a typical "tiger striped'" sign, but its features on functional magnetic resonance imaging (fMRI) are still inconclusive. PATIENT CONCERNS: To explore the characteristics of LDDs using fMRI. DIAGNOSES: We report 3 cases of pathologically confirmed LDDs. INTERVENTIONS: Three patients underwent brain tumor surgery. OUTCOMES: All the patients had a good prognosis. LESSONS: Magnetic resonance spectroscopy and susceptibility-weighted imaging combined with conventional MRI can be used to better diagnose LDDs. Perfusion-weighted imaging is not specific for distinguishing cerebellar tumors. The combined application of fMRI and conventional MRI can improve the accuracy of LDD diagnoses.

CXCR4-Directed PET/CT with [68Ga]Pentixafor in Central Nervous System Lymphoma: A Comparison with [18F]FDG PET/CT.

PURPOSE: This study aimed to evaluate the value of [68 Ga]Pentixafor PET/CT for the detection of lesions in central nervous system lymphoma (CNSL) patients before chemotherapy, during treatment and suspected CSNL recurrence, compared with 2-deoxy-2-[18F]fluoro-D-glucose ([18F]FDG) positron emission tomography/computed tomography (PET/CT). PROCEDURES: Twenty-six patients with newly or previously diagnosed CNSL who underwent [68 Ga]Pentixafor PET/CT were included retrospectively. Histopathological results, magnetic resonance imaging (MRI), and follow-up were used as the standard reference. The accuracy of lesion detection, maximum standardized uptake value (SUVmax) of tumors, and ratio of tumor-to-normal brain (T/N) with [68 Ga]Pentixafor PET/CT were calculated and compared to those obtained with [18F]FDG PET/CT. CXCR4 expression was analyzed through immunohistochemistry. RESULTS: Of 26 patients, 18 were newly diagnosed with a total of 23 lesions, 4 had recurrent with 4 lesions, and 4 underwent a mid-term treatment assessment after 4 cycles of chemotherapy (3 achieved complete response (CR), 1 experienced progressive disease (PD) with a total of 8 lesions). Thirty-five lesions were all clearly detected with favorable contrast by [68 Ga]Pentixafor PET/CT (accuracy, 100%), consistent with the results of contrast-enhanced magnetic resonance imaging (CE-MRI). The SUVmax of positive lesions in [68 Ga]Pentixafor PET/CT was correlated with tumor size (r = 0.555, P = 0.001). In 21 patients, compared with [18F]FDG PET/CT, [68 Ga]Pentixafor PET/CT showed a remarkably higher T/N ratio (21.93 ± 10.77 vs 4.29 ± 2.16, P = 0.000) and detected 5 more lesions in the mid-term treatment assessment of patients (P = 0.026). The CXCR4 expression of CNSL lesions was correlated with SUVmax of [68 Ga]Pentixafor PET/CT (r = 0.772, P = 0.000). CONCLUSIONS: CXCR4-directed PET/CT using [68 Ga]Pentixafor, with excellent tumor-to-background contrast, might be a more promising agent for the detection of lesions in CNSL patients than [18F]FDG PET/CT.