Identification of Optimal Parameters for Assessing Lymph Node Status of Patients with Esophageal Squamous Cell Carcinoma After Neoadjuvant Chemoradiotherapy.

BACKGROUND: This study aimed to compare the prognostic discrimination power of pretreatment pathologic N stage (prepN), lymph node tumor regression grade (LNTRG), and posttreatment pathologic N (ypN) category for esophageal squamous cell carcinoma (ESCC) patients who received neoadjuvant chemoradiotherapy (nCRT) plus surgery. METHODS: The study reviewed 187 ESCC patients from two medical centers who underwent nCRT plus surgery. Pathologic LNTRG was defined by the proportion of viable tumor area within the tumor bed in lymph nodes (LNs). An average LNTRG then was calculated by averaging the tumor regression grade (TRG) score of all resected LNs. Lymph nodes containing regression changes or vital tumor cells were used for interpretation of the prepN stage, which reflects the estimated number of originally involved LNs. RESULTS: The ypN, prepN, and LNTRG categories had significant prognostic stratification power (p < 0.001, log-rank test). Multivariable cox regression showed that all three categories were independent prognostic factors of disease-free survival (DFS) (p < 0.05). The LNTRG category showed a better prognostic value for DFS prediction than the ypN and prepN categories (Akaike information criterion [AIC]: LNTRG [933.69], ypN [937.56], prepN [937.45]). Additionally, the superior predictive capacity of the LNTRG category was demonstrated by decision curve analysis. Similar results were discovered for patients with remaining diseased LNs. CONCLUSIONS: The three staging categories had prognostic relevance for DFS, with the LNTRG category seeming to have better prognostic indication power. Comprehensive consideration of the ypN status, prepN status, and LN regression may allow for better prognostic stratification of patients.

Integration of glucose and cardiolipin anabolism confers radiation resistance of HCC.

BACKGROUND AND AIMS: Poor response to ionizing radiation (IR) due to resistance remains a clinical challenge. Altered metabolism represents a defining characteristic of nearly all types of cancers. However, how radioresistance is linked to metabolic reprogramming remains elusive in hepatocellular carcinoma (HCC). APPROACH AND RESULTS: Baseline radiation responsiveness of different HCC cells were identified and cells with acquired radio-resistance were generated. By performing proteomics, metabolomics, metabolic flux, and other functional studies, we depicted a metabolic phenotype that mediates radiation resistance in HCC, whereby increased glucose flux leads to glucose addiction in radioresistant HCC cells and a corresponding increase in glycerophospholipids biosynthesis to enhance the levels of cardiolipin. Accumulation of cardiolipin dampens the effectiveness of IR by inhibiting cytochrome c release to initiate apoptosis. Mechanistically, mammalian target of rapamycin complex 1 (mTORC1) signaling-mediated translational control of hypoxia inducible factor-1α (HIF-1α) and sterol regulatory element-binding protein-1 (SREBP1) remodels such metabolic cascade. Targeting mTORC1 or glucose to cardiolipin synthesis, in combination with IR, strongly diminishes tumor burden. Finally, activation of glucose metabolism predicts poor response to radiotherapy in cancer patients. CONCLUSIONS: We demonstrate a link between radiation resistance and metabolic integration and suggest that metabolically dismantling the radioresistant features of tumors may provide potential combination approaches for radiotherapy in HCC.

Serum lnc34a is a potential prediction biomarker for bone metastasis in hepatocellular carcinoma patients.

BACKGROUND: Early screening and intervention therapies are crucial to improve the prognosis of hepatocellular carcinoma (HCC) patients with bone metastasis. We aimed to identify serum lncRNA as a prediction biomarker in HCC bone metastasis. METHODS: The expression levels of lnc34a in serum samples from 157 HCC patients were detected by quantitative real-time polymerase chain reaction (PCR). Univariate analysis and multivariate analysis were performed to determine statistically significant variables. RESULTS: Expression levels of lnc34a in serum from HCC patients with bone metastasis were significantly higher than those without bone metastasis. The high expressions of lnc34a, vascular invasion and Barcelona Clinic Liver Cancer (BCLC) stage were associated with bone metastasis by analysis. Moreover, lnc34a expression was specifically associated with bone metastasis rather than lung or lymph node metastasis in HCC. CONCLUSIONS: High serum lnc34a expression was a independent risk factor for developing bone metastasis in HCC.

Circular RNA RSF1 promotes inflammatory and fibrotic phenotypes of irradiated hepatic stellate cell by modulating miR-146a-5p.

The role of circular RNA (circRNA) in radiation-induced liver disease (RILD) remains largely unknown. In this study, Ras-related C3 botulinum toxin substrate 1 (RAC1) was elevated in irradiated human hepatic stellate cell (HSC) line LX2, the important effector cell mediating RILD. Overexpression of RAC1 promotes cell proliferation, proinflammatory cytokines production, and α-smooth muscle actin expression, which were blocked by microRNA (miR)-146a-5p mimics. CircRNA RSF1 (circRSF1) was upregulated in irradiated LX2 cells and predicted to harbor binding site for miR-146a-5p. Biotinylated-RNA pull down and dual-luciferase reporter detection confirmed the direct interaction of circRSF1 and miR-146a-5p. Enforced expression of circRSF1 increased RAC1 expression by acting as miR-146a-5p sponge to inhibit miR-146a-5p activity, and thus enhanced the cell viability, and promoted inflammatory and fibrotic phenotype of irradiated LX2 cells. These findings indicate a functional regulatory axis composing of circRSF1, miR-146a-5p, and RAC1 in irradiated HSC, which may provide attractive therapeutic targets for RILD.

Proteomic Profiling of Human Hepatic Stellate Cell Line LX2 Responses to Irradiation and TGF-ß1.

Hepatic stellate cells (HSCs) are the main target of radiation damage and primarily contribute to the development of radiation-induced liver fibrosis. However, the molecular events underlying the radiation-induced activation of HSCs are not fully elucidated. In the present study, human HSC line LX2 was treated with X-ray irradiation and/or TGF-ß1, and profibrogenic molecules were evaluated. The iTRAQ LC-MS/MS technology was performed to identify global protein expression profiles in LX2 following exposure to different stimuli. Irradiation or TGF-ß1 alone increased expression of α-SMA, collagen 1, CTGF, PAI-1, and fibronectin. Irradiation and TGF-ß1 cooperatively induced expression of these profibrotic markers. In total, 102, 137, 155 dysregulated proteins were identified in LX2 cell samples affected by irradiation, TGF-ß1, or cotreatment, respectively. Bioinformatic analyses showed that the three differentially expressed protein sets were commonly associated with cell cycle and protein processing in endoplasmic reticulum. The expression of a set of proteins was properly validated: CDC20, PRC1, KIF20A, CCNB1, SHCBP, TACC3 were upregulated upon irradiation or irradiation and TGF-ß1 costimulation, whereas SPARC and THBS1 were elevated by TGF-ß1 or TGF-ß1 plus irradiation treatment. Furthermore, CDC20 inhibition suppressed expression of profibrotic markers in irradiated and TGF-ß1-stimulated LX2 cells. Detailed data on potential molecular mechanisms causing the radiation-induced HSC activation presented here would be instrumental in developing radiotherapy strategies that minimize radiation-induced liver fibrosis.

The molecular mechanism of LncRNA34a-mediated regulation of bone metastasis in hepatocellular carcinoma.

BACKGROUND: Bone metastasis (BM) has long been recognized as a major threat to the quality of life of hepatocellular cancer (HCC) patients. While LncRNA34a (Lnc34a) has been shown to regulate colon cancer stem cell asymmetric division, its effect on HCC BM remains unknown. METHODS: In situ hybridization and quantitative real-time polymerase chain reaction (qRT-PCR) were used to detect the expression of Lnc34a in HCC tissues and cell lines. Ventricle injection model was constructed to explore the effect of Lnc34a on BM in vivo. The methylation of miR-34a promoter and histones deacetylation were examined by using bisulfate-sequencing PCR and chromatin immunoprecipitation assays. RNA pull down and RNA immunoprecipitation were performed to investigated the interaction between Lnc34a and epigenetic regulators. Dual-luciferase reporter assay was conducted to find miR-34a target. The involvement of TGF-ß pathway in the BM from HCC was determined by qRT-PCR, western, and elisa assays. RESULTS: We found that Lnc34a was significantly overexpressed in HCC tissues and associated with BM. Both in vitro and in vivo experiments indicate that the restoration or knockdown of Lnc34a expression in HCC cells had a marked effect on cellular migration, invasion, and metastasis. Mechanistic analyses suggested that Lnc34a epigenetically suppresses miR-34a expression through recruiting DNMT3a via PHB2 to methylate miR-34a promoter and HDAC1 to promote histones deacetylation. On the other hand, miR-34a targets Smad4 via the TGF-ß pathway, followed by altering the transcription of the downstream genes (i.e., CTGF and IL-11) that are associated with BM. CONCLUSIONS: Our study is the first to document the pro-bone metastatic role of Lnc34a in BM of HCC and reveal a novel mechanism for the activation of the TGF-ß signaling pathway in HCC BM, providing evidence of a potential therapeutic strategy in HCC BM.

Prognostic index score predicts outcome of patients with Stage I non-small cell lung cancer after stereotactic body radiation therapy.

OBJECTIVE: Lung cancer is the most common causes of cancer death worldwide and patients with non-small-cell lung cancer (NSCLC) have various prognosis. We conducted this study to identify the prognostic predictors and establish a prognostic index score (PIS) for patients with Stage I NSCLC after stereotactic body radiation therapy (SBRT). METHODS: A total of 131 consecutive patients with Stage I NSCLC who underwent SBRT in our institute were analyzed retrospectively. The Cox proportional hazards regression model was applied to identify the prognostic predictors. Time-dependent receiver operating characteristic analysis was performed to examine cutoff values for survival. The Kaplan-Meier method with log-rank test was used to compare survival curves. RESULTS: Univariate analysis indicated that tumor location, maximum standardized uptake value (SUVmax), monocyte counts and platelet-to-lymphocyte ratio (PLR) were prognostic factors of overall survival (OS). SUVmax and PLR remained significant in multivariate analysis. Survival analysis indicated both high-SUVmax and PLR correlated with inferior OS and PFS. A PIS was constructed based on pretreatment SUVmax and PLR and a high PIS was also significantly associated with poor outcome. CONCLUSION: The pretreatment SUVmax and PLR were independent prognostic factors of OS in patients with Stage I NSCLC after SBRT. PIS provides a convenient and accurate tool for predicting outcome of patients after SBRT.

Nomograms for predicting disease progression in patients of Stage I non-small cell lung cancer treated with stereotactic body radiotherapy.

OBJECTIVE: Non-local progression is a major concern in non-small cell lung cancer (NSCLC) treated with stereotactic body radiotherapy (SBRT). Herein we aimed to create a pre-treatment prognostic nomogram for patients with Stage I NSCLC receiving SBRT. METHODS: We retrospectively studied 182 eligible patients. Patients were randomly divided into a model (70%) group and a validation (30%) group. In the model group, thirteen parameters consisting of patient, treatment, and tumor factors were studied and multivariate Cox proportional hazards regression was performed to identify independent predictors for survival outcome, based on which we developed clinical nomogram. The nomogram was externally validated in the validation group. RESULTS: Multivariate analysis showed that tumor size (P = 0.011) was the only factor correlated with 2-year overall survival, whereas 2-year locoregional control (LRC) was significantly related to tumor size (P = 0.024) and the maximum standardized uptake value (SUVmax) (P = 0.044), so does 2-year progression-free survival (PFS) (tumor size: P = 0.026; SUVmax: P = 0.038). Nomogram for 2-year LRC and 2-year PFS were created based on aforementioned results. The C-indexes for the nomograms to predict 2-year LRC and PFS were 0.816 and 0.804, respectively, in model group, and were 0.729 and 0.731, respectively, in the validation group. Calibration plots also showed that the model performed well. CONCLUSIONS: Tumor of larger size and higher SUVmax predisposed patients to early onset of locoregional and distant progression. The nomogram developed in our study would be helpful in clinical decision-making and selection of patients who may benefit from more rigorous follow-up and aggressive systemic treatment plan.

A knowledge-based approach to automated planning for hepatocellular carcinoma.

PURPOSE: To build a knowledge-based model of liver cancer for Auto-Planning, a function in Pinnacle, which is used as an automated inverse intensity modulated radiation therapy (IMRT) planning system. METHODS AND MATERIALS: Fifty Tomotherapy patients were enrolled to extract the dose-volume histograms (DVHs) information and construct the protocol for Auto-Planning model. Twenty more patients were chosen additionally to test the model. Manual planning and automatic planning were performed blindly for all twenty test patients with the same machine and treatment planning system. The dose distributions of target and organs at risks (OARs), along with the working time for planning, were evaluated. RESULTS: Statistically significant results showed that automated plans performed better in target conformity index (CI) while mean target dose was 0.5 Gy higher than manual plans. The differences between target homogeneity indexes (HI) of the two methods were not statistically significant. Additionally, the doses of normal liver, left kidney, and small bowel were significantly reduced with automated plan. Particularly, mean dose and V15 of normal liver were 1.4 Gy and 40.5 cc lower with automated plans respectively. Mean doses of left kidney and small bowel were reduced with automated plans by 1.2 Gy and 2.1 Gy respectively. In contrast, working time was also significantly reduced with automated planning. CONCLUSIONS: Auto-Planning shows availability and effectiveness in our knowledge-based model for liver cancer.

CMISG1701: a multicenter prospective randomized phase III clinical trial comparing neoadjuvant chemoradiotherapy to neoadjuvant chemotherapy followed by minimally invasive esophagectomy in patients with locally advanced resectable esophageal squamous cell carcinoma (cT3-4aN0-1M0) (NCT03001596).

BACKGROUND: Neoadjuvant chemoradiation is not recommended as an approach for treatment of esophageal squamous cell carcinoma due to its significant postoperative mortality. However, it is assumed the combination of neoadjuvant chemoradiation with minimally invasive esophagectomy (MIE) may reduce postoperative mortality, which can revive preoperative chemoradiation. No randomized controlled studies comparing neoadjuvant chemoradiation plus MIE with neoadjuvant chemotherapy plus MIE have been performed so far. The present trial is initiated to obtain valid information whether neoadjuvant chemoradiation plus MIE yields better survival without worse postoperative morbidity and mortality in the treatment of locally advanced resectable esophageal squamous cell carcinoma(cT3-4aN0-1M0). METHODS/DESIGN: CMISG1701 is a multicenter, prospective, randomized, phase III clinical trial, investigating the safety and efficacy of neoadjuvant chemoradiation plus MIE compared with neoadjuvant chemotherapy plus MIE. Patients with locally advanced resectable esophageal squamous cell carcinoma (cT3-4aN0-1M0) are eligible for the study. A total of 264 patients are randomly assigned to neoadjuvant chemoradiation (arm A) or neoadjuvant chemotherapy (arm B) with a 1:1 allocation ratio. The primary outcome is overall survival assessed with a minimum follow-up of 36 months. Secondary outcomes are progression-free survival, recurrence-free survival, postoperative pathologic stage, treatment-related complications, postoperative mortality as well as quality of life. DISCUSSION: The objective of this trial is to identify the superior protocol with regard to patient survival, treatment morbidity/mortality and quality of life between neoadjuvant chemoradiation plus MIE and neoadjuvant chemotherapy plus MIE. TRIAL REGISTRATION: NCT03001596 (December 17, 2016).

Does liver resection/transplantation affect respiratory induced liver motion in patients with hepatocellular carcinoma?

The purpose of this study was to evaluate the changes in magnitude of three-dimensional (3D) liver motion after liver resection/transplantation in patients with hepatocellular carcinoma (HCC) using four-dimensional (4D)-computed tomography (CT) images. From January 2012 to April 2016, 74 HCC patients underwent 4D-CT scans under a free-breathing state to assess respiratory liver motion. Of the 74 patients, 40 did not have a liver resection/transplantation (Group A), 34 with liver resection/transplantation. 15 underwent major or minor resection in the right liver lobe (Group B), 14 underwent major or minor resection in the left liver lobe (Group C), and five underwent liver transplantation (Group D). The 4D-CT images were sorted into 10 image series according to the respiratory phase from the end inspiration to the end expiration, and then transferred to treatment planning software. All liver contours were drawn by a single physician and confirmed by a second. Liver relative coordinates were automatically generated to calculate liver respiratory motion in different axial directions and compiled into a single composite image. Differences in respiratory liver motion were assessed using one-way ANOVA. The average liver respiratory motion in the cranial-caudal direction and 3D magnitude were 10.46 ± 2.78 mm (range, 5.60-18.80 mm) and 11.74 ± 2.65 mm (range, 7.45-20.79 mm) for patients without liver resection/transplantation, and 7.74 ± 2.79 mm (range, 2.20-12.90 mm) and 9.07 ± 2.38 mm (range, 4.79-14.08 mm) for posthepatectomy/post-transplant patients respectively. There were significant differences between Group A and B, Group A and C, Group A and D. However, there were no significant differences among Group B, C, and D. Liver resection/transplantation greatly affected respiratory-induced liver motion in patients with HCC. We, therefore, recommend discriminatory internal target volume (ITV) determination for patients with or without liver resection/transplantation undergoing external radiotherapy for hepatic tumors while respiratory motion management is unavailable.

High dose radiotherapy with image-guided hypo-IMRT for hepatocellular carcinoma with portal vein and/or inferior vena cava tumor thrombi is more feasible and efficacious than conventional 3D-CRT.

OBJECTIVE: To compare the efficacies of conventional three-dimensional conformal radiotherapy and image-guided hypofractionated intensity-modulated radiotherapy treatments in advanced hepatocellular carcinoma patients with portal vein and/or inferior vena cava tumor thrombi. METHODS: A total of 118 hepatocellular carcinoma patients with portal vein and/or inferior vena cava tumor thrombi who received external beam radiation therapy focused on tumor thrombi and intrahepatic tumors were retrospectively reviewed. During the three-dimensional conformal radiotherapy treatments, a median total dose of 54 Gy with a conventional fraction (1.8-2.0 Gy/fx) was delivered. During the image-guided hypofractionated intensity-modulated radiotherapy treatments, a median total dose of 60 Gy with fractions of 2.5-4.0 Gy/fx was delivered. RESULTS: The median follow-up time was 11.8 months (range, 1.7-43.7 months). Higher radiation doses were delivered by image-guided hypofractionated intensity-modulated radiotherapy than by three-dimensional conformal radiotherapy (average dose 57.86 ± 7.03 versus 50.88 ± 6.60 Gy, P ≤ 0.001; average biological effective dose 72.35 ± 9.62 versus 61.45 ± 6.64 Gy, P < 0.001). A longer median survival was found with image-guided hypofractionated intensity-modulated radiotherapy than with three-dimensional conformal radiotherapy (15.47 versus 10.46 months, P = 0.005). Multivariate analysis showed that image-guided hypofractionated intensity-modulated radiotherapy is a significant prognostic factor for overall survival. Toxicity was mild for both image-guided hypofractionated intensity-modulated radiotherapy and three-dimensional conformal radiotherapy. CONCLUSIONS: High dose radiotherapy delivered by image-guided hypofractionated intensity-modulated radiotherapy appears to be an effective treatment that provides a survival benefit without increasing severe toxicity in hepatocellular carcinoma patients with portal vein and/or inferior vena cava tumor thrombi.

MicroRNA-146a-5p Negatively Regulates Pro-Inflammatory Cytokine Secretion and Cell Activation in Lipopolysaccharide Stimulated Human Hepatic Stellate Cells through Inhibition of Toll-Like Receptor 4 Signaling Pathways.

Lipopolysaccharide (LPS)/toll-like receptor 4 (TLR4) signaling pathway is demonstrated to be involved in the hepatic fibrosis. MicroRNA (miR)-146a-5p is a key regulator of the innate immune response. The functional significance of miR-146a-5p during the LPS/TLR4 mediated hepatic fibrosis process remains unclear. In this study, we found that TLR4 and α-smooth muscle actin (α-SMA) were up-regulated and miR-146a-5p was down-regulated in human hepatic stellate cell (HSC) line LX2 after LPS stimulation. Overexpression of miR-146a-5p inhibited LPS induced pro-inflammatory cytokines secretion through down-regulating the expression levels of TLR-4, IL-1 receptor-associated kinase 1 (IRAK1), TNF receptor associated factor-6 (TRAF6) and phosphorylation of nuclear factor-kappa B (NF-κB). Knockdown of IRAK1 and TRAF6 also suppressed pro-inflammatory cytokine production by inhibiting NF-κB phosphorylation. In addition, miR-146a-5p mimic blocked LPS induced TRAF6 dependent c-Jun N-terminal kinase (JNK) and Smad2 activation as well as α-SMA production. Taken together, these results suggest that miR-146a-5p suppresses pro-inflammatory cytokine secretion and cell activation of HSC through inhibition of TLR4/NF-κB and TLR4/TRAF6/JNK pathway.

TLR4-dependant immune response, but not hepatitis B virus reactivation, is important in radiation-induced liver disease of liver cancer radiotherapy.

Toll-like receptor 4 (TLR4) is an important trigger of the immune response against hepatitis B virus (HBV) infection and liver injuries. The roles of HBV reactivation versus TLR4-dependant immune response may be critical factors in preventing radiation-induced liver diseases (RILDs) after liver cancer radiotherapy. This study consists of three phases. In the primary phase, livers of mutant TLR4 (TLR4(-)) mice were irradiated with 30 Gy in either the absence or presence of HBV infection. The latter was done by introduction of plasmid pAAV/HBV 1.2. In the advanced phase, RILDs were compared in normal TLR4 (TLR4(+)) versus TLR4(-) mice. In the validation phase, 28 liver cancer patients who had undergone radiotherapy before hepatectomy were enrolled. Liver biopsies near tumors, irradiated with 35-48 Gy, were used to construct tissue microarrays. HBV reactivation, TLR4 expression, and severity of RILDs were studied in both mouse and human. More HBV reactivation, without significant RILD, was observed in irradiated versus unirradiated TLR4(-) mice. RILD scores of TLR4(+) mice were higher than TLR4(-) mice. In humans, serious RILDs tended to develop in patients with high TLR4 expression, but not in patients with low TLR4 or high HBV surface antigen expression. High TLR4 expression was seen in only 2 of 12 HBV-reactive patients, but in HBV-nonreactive patients, it was seen in 6 of 9 (P < 0.03). In summary, RILDs correlated with high TLR4 expression, but not with HBV reactivation, which is inhibited in liver with high TLR4 expression after liver cancer radiotherapy.

Radiotherapy treatment of adrenal gland metastases from hepatocellular carcinoma: clinical features and prognostic factors.

BACKGROUND: The optimal treatment for adrenal metastases from hepatocellular carcinoma (HCC) has not been established. This study analyzed the effects of radiation therapy (RT) for such metastases and identified clinical features and predictors of survival in these patients. METHODS: We retrospectively investigated 55 patients with adrenal metastasis from HCC who had been treated with RT. Radiation doses to the adrenal lesions ranged from 26 to 60 Gy, while the intrahepatic lesions were treated by surgical resection, transarterial chemoembolization (TACE), liver transplantation, and/or RT. RT was conducted to adrenal lesions after their intrahepatic lesions were controlled more than 2 months. The parameters studied included survival rates and tumor responses to RT. The Kaplan-Meier method was used to evaluate survival rate and the Cox regression model was used to identify potential predictors of outcome. RESULTS: The patients treated by RT had adrenal metastasis on the right side (41), the left (6), or on both sides (8). In all 55 patients, the median survival duration was 13.6 months and there was 100% pain relief after completion of RT. Adverse effects were mild to moderate. Unfavorable pretreatment predictors determined by univariate analysis were associated with multiple intrahepatic foci, metastases to additional organs, high γ-glutamyltransferase and alpha-fetoprotein levels, liver function of Child-Pugh classification B and uncontrolled primary HCC. By multivariate analysis, unfavorable predictors were multiple intrahepatic foci, metastases to additional organs and uncontrolled primary HCC. CONCLUSIONS: Radiotherapy as treatment for adrenal metastases in HCC is a good palliative therapy that is associated with reasonable safety. It appears reasonable that such patients should be considered to be treated with radiotherapy. Multiple intrahepatic foci, metastases to additional organs and uncontrolled primary HCC were unfavorable predictors.

Olaparib enhances radiation-induced systemic anti-tumor effects via activating STING-chemokine signaling in hepatocellular carcinoma.

Although Poly (ADP-ribose) polymerase (PARP) inhibitors have been clinically approved for cancers with BRCA mutations and are known to augment radiotherapy responses, their roles in promoting the abscopal effect and mediating immunotherapy in BRCA-proficient hepatocellular carcinoma (HCC) remain underexplored. Our study elucidates that olaparib enhances the radio-sensitivity of HCC cells. Coadministration of olaparib and irradiation induces significant DNA damage by generating double-strand breaks (DSBs), as revealed both in vitro and in immune-deficient mice. These DSBs activate the cGAS-STING pathway, initiating immunogenic cell death in abscopal tumors. STING activation reprograms the immune microenvironment in the abscopal tumors, triggering the release of type I interferon and chemokines, including CXCL9, CXCL10, CXCL11, and CCL5. This in turn amplifies T cell priming against tumor neoantigens, leading to an influx of activated, neoantigen-specific CD8+ T-cells within the abscopal tumors. Furthermore, olaparib attenuated the immune exhaustion induced by radiation and enhances the responsiveness of HCC to immune checkpoint inhibitors. Collectively, our data advocate that a synergistic regimen of PARP inhibitors and radiotherapy can strategically reinforce both local (primary) and systemic (abscopal) tumor control, bolstering HCC susceptibility to immunotherapy.

METTL3-Mediated STING Upregulation and Activation in Kupffer Cells Contribute to Radiation-Induced Liver Disease via Pyroptosis.

PURPOSE: Radiation therapy is a vital adjuvant treatment for liver cancer, although the challenge of radiation-induced liver diseases (RILDs) limits its implementation. Kupffer cells (KCs) are a crucial cell population of the hepatic immune system, and their biologic function can be modulated by multiple epigenetic RNA modifications, including N6-methyladenosine (m6A) methylation. However, the mechanism for m6A methylation in KC-induced inflammatory responses in RILD remains unclear. The present study investigated the function of m6A modification in KCs contributing to RILD. METHODS AND MATERIALS: Methylated RNA-immunoprecipitation sequencing and RNA transcriptome sequencing were used to explore the m6A methylation profile of primary KCs isolated from mice after irradiation with 3 × 8 Gy. Western blotting and quantitative real-time PCR were used to evaluate gene expression. DNA pulldown and chromatin immunoprecipitation assays were performed to verify target gene binding and identify binding sites. RESULTS: Methylated RNA-immunoprecipitation sequencing revealed significantly increased m6A modification levels in human KCs after irradiation, suggesting the potential role of upregulated m6A in RILD. In addition, the study results corroborated that methyltransferase-like 3 (METTL3) acts as a main modulator to promote the methylation and gene expression of TEAD1, leading to STING-NLRP3 signaling activation. Importantly, it was shown that IGF2BP2 functions as an m6A "reader" to recognize methylated TEAD1 mRNA and promote its stability. METTL3/TEAD1 knockdown abolished the activation of STING-NLRP3 signaling, protected against RILD, and suppressed inflammatory cytokines and hepatocyte apoptosis. Moreover, clinical human normal liver tissue samples collected after irradiation showed increased expression of STING and interleukin-1ß in KCs compared with nonirradiated samples. Notably, STING pharmacologic inhibition alleviated irradiation-induced liver injury in mice, indicating its potential therapeutic role in RILD. CONCLUSIONS: The results of our study reveal that TEAD1-STING-NLRP3 signaling activation contributes to RILD via METTL3-dependent m6A modification.

Radiotherapy enhances efficacy of PD-1 inhibitors in advanced hepatocellular carcinoma: A propensity-matched real-world study.

BACKGROUND: To address the need for immunotherapy in patients with advanced primary hepatocellular carcinoma (HCC), combination with radiotherapy (RT) has emerged as a promising strategy. In preclinical studies, irradiated tumors released tumor antigens to synergistically increase the antitumor effect of immunotherapy. Hence, we investigated whether RT enhances the efficacy of anti-programmed death receptor-1 (PD-1) inhibitors in advanced HCC in real-world practice. METHODS: Between August 2018 and June 2021, 172 patients with advanced primary HCC were enrolled in the tertiary center (Zhongshan Hospital of Fudan University); 95 were treated with a combination of RT and the inhibitor of PD-1 (RT-PD1 cohort), and 77 were administered anti-PD-1 therapy (PD1 cohort). The first cycle of PD-1 inhibitors was administered within 60 days or concurrently with RT. Propensity score matching for bias reduction was used to evaluate the clinical outcomes. RESULTS: Among 71 propensity-matched pairs, median progression-free survival was 5.7 months in the RT-PD1 cohort vs. 2.9 months in the PD1 cohort ( P  <0.001). Median overall survival was 20.9 months in the RT-PD1 cohort vs. 11.2 months in the PD1 cohort ( P  = 0.018). Compared with patients in the PD1 cohort, patients in the RT-PD1 cohort had significantly higher objective response rates (40.8%, 29/71 vs. 19.7%, 14/71, P  = 0.006) and disease control rates (62.0%, 44/71 vs. 31.0%, 22/71, P  <0.001). The incidences of toxic effects were not significantly different between the two cohorts. CONCLUSIONS: RT plus anti-PD-1 therapy is well tolerated. RT enhances the efficacy of anti-PD-1 therapy in patients with advanced primary HCC by improving survival outcomes without increased toxic effects.

Stereotactic body radiation therapy in patients with centrally located hepatocellular carcinoma: A retrospective, single-arm, multi-center study.

Centrally located hepatocellular carcinoma (HCC) is difficult to be radically resected due to its special location close to major hepatic vessels. Thus, we aimed to assess whether stereotactic body radiation therapy (SBRT) can be an effective and safe approach for centrally located HCC. This retrospective study included 172 patients with centrally located HCC who were treated with SBRT. Overall survival (OS) was analyzed as the primary endpoint. Rates of progression-free survival (PFS), local control, intrahepatic relapse, extrahepatic metastasis and toxicities were analyzed as secondary endpoints. The OS rates of 1-, 3-, and 5-year were 97.7%, 86.7%, and 76.3%, respectively. The PFS/local control rates of 1-, 3-, and 5-year were 94.1%/98.2%, 76.8%/94.9%, and 59.3%/92.3%, respectively. The cumulative incidence of intrahepatic relapse/extrahepatic metastases of 1-, 3-, and 5-year were 3.7%/2.9%, 25.0%/7.4%, and 33.3%/9.8%, respectively. Both univariate and multivariate analyses revealed that patients received BED10 at 100 Gy or more had better OS. Radiation-related adverse events were mild to moderate according to Common Terminology Criteria for Adverse Events, and no toxicities over grade 3 were observed. Patients with centrally located HCC in our cohort who received SBRT had similar OS and PFS rates compared to those reported in literatures who received surgery with neoadjuvant or adjuvant intensity-modulated radiation therapy. These results indicate that SBRT is an effective and well-tolerated method for patients with centrally located HCC, suggesting that it may serve as a reasonable alternative treatment for these kind of patients.