Cetuximab plus FOLFOXIRI versus cetuximab plus FOLFOX as conversion regimen in RAS/BRAF wild-type patients with initially unresectable colorectal liver metastases (TRICE trial): A randomized controlled trial.

BACKGROUND: It remains unclear whether intensification of the chemotherapy backbone in tandem with an anti-EGFR can confer superior clinical outcomes in a cohort of RAS/BRAF wild-type colorectal cancer (CRC) patients with initially unresectable colorectal liver metastases (CRLM). To that end, we sought to comparatively evaluate the efficacy and safety of cetuximab plus FOLFOXIRI (triplet arm) versus cetuximab plus FOLFOX (doublet arm) as a conversion regimen (i.e., unresectable to resectable) in CRC patients with unresectable CRLM. METHODS AND FINDINGS: This open-label, randomized clinical trial was conducted from April 2018 to December 2022 in 7 medical centers across China, enrolling 146 RAS/BRAF wild-type CRC patients with initially unresectable CRLM. A stratified blocked randomization method was utilized to assign patients (1:1) to either the cetuximab plus FOLFOXIRI (n = 72) or cetuximab plus FOLFOX (n = 74) treatment arms. Stratification factors were tumor location (left versus right) and resectability (technically unresectable versus ≥5 metastases). The primary outcome was the objective response rate (ORR). Secondary outcomes included the median depth of tumor response (DpR), early tumor shrinkage (ETS), R0 resection rate, progression-free survival (PFS), overall survival (not mature at the time of analysis), and safety profile. Radiological tumor evaluations were conducted by radiologists blinded to the group allocation. Primary efficacy analyses were conducted based on the intention-to-treat population, while safety analyses were performed on patients who received at least 1 line of chemotherapy. A total of 14 patients (9.6%) were lost to follow-up (9 in the doublet arm and 5 in the triplet arm). The ORR was comparable following adjustment for stratification factors, with 84.7% versus 79.7% in the triplet and doublet arms, respectively (odds ratio [OR] 0.70; 95% confidence intervals [CI] [0.30, 1.67], Chi-square p = 0.42). Moreover, the ETS rate showed no significant difference between the triplet and doublet arms (80.6% (58/72) versus 77.0% (57/74), OR 0.82, 95% CI [0.37, 1.83], Chi-square p = 0.63). Although median DpR was higher in the triplet therapy group (59.6%, interquartile range [IQR], [50.0, 69.7] versus 55.0%, IQR [42.8, 63.8], Mann-Whitney p = 0.039), the R0/R1 resection rate with or without radiofrequency ablation/stereotactic body radiation therapy was comparable with 54.2% (39/72) of patients in the triplet arm versus 52.7% (39/74) in the doublet arm. At a median follow-up of 26.2 months (IQR [12.8, 40.5]), the median PFS was 11.8 months in the triplet arm versus 13.4 months in the doublet arm (hazard ratio [HR] 0.74, 95% CI [0.50, 1.11], Log-rank p = 0.14). Grade ≥ 3 events were reported in 47.2% (35/74) of patients in the doublet arm and 55.9% (38/68) of patients in the triplet arm. The triplet arm was associated with a higher incidence of grade ≥ 3 neutropenia (44.1% versus 27.0%, p = 0.03) and diarrhea (5.9% versus 0%, p = 0.03). The primary limitations of the study encompass the inherent bias in subjective surgical decisions regarding resection feasibility, as well as the lack of a centralized assessment for ORR and resection. CONCLUSIONS: The combination of cetuximab with FOLFOXIRI did not significantly improve ORR compared to cetuximab plus FOLFOX. Despite achieving an enhanced DpR, this improvement did not translate into improved R0 resection rates or PFS. Moreover, the triplet arm was associated with an increase in treatment-related toxicity. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT03493048.

Skp2-dependent ubiquitination and activation of LKB1 is essential for cancer cell survival under energy stress.

LKB1 is activated by forming a heterotrimeric complex with STRAD and MO25. Recent studies suggest that LKB1 has pro-oncogenic functions, besides acting as a tumor suppressor. How the LKB1 activity is maintained and how LKB1 regulates cancer development are largely unclear. Here we show that K63-linked LKB1 polyubiquitination by Skp2-SCF ubiquitin ligase is critical for LKB1 activation by maintaining LKB1-STRAD-MO25 complex integrity. We further demonstrate that oncogenic Ras acts upstream of Skp2 to promote LKB1 polyubiquitination by activating Skp2-SCF ubiquitin ligase. Moreover, Skp2-mediated LKB1 polyubiquitination is required for energy-stress-induced cell survival. We also detected overexpression of Skp2 and LKB1 in late-stage hepatocellular carcinoma (HCC), and their overexpression predicts poor survival outcomes. Finally, we show that Skp2-mediated LKB1 polyubiquitination is important for HCC tumor growth in vivo. Our study provides new insights into the upstream regulation of LKB1 activation and suggests a potential target, the Ras/Skp2/LKB1 axis, for cancer therapy.

Histopathological growth patterns correlate with the immunoscore in colorectal cancer liver metastasis patients after hepatectomy.

Various scoring systems have been proposed to predict the postoperative prognosis of colorectal liver metastasis (CRLM), including the clinical risk score (CRS), the immunoscore and so on. Recently, histopathological growth patterns (HGPs) have been recognized. However, the correlation between HGPs and the immunoscore, and their prognostic values in patients with CRLM after liver resection remain undetermined. In this study, HGPs were retrospectively evaluated in H&E-stained slides from 166 CRLM patients. The immunoscore was calculated according to the densities of immunostained CD3 + and CD8 + cells. A risk score combining HGPs, the immunoscore and the CRS was defined and divided patients into the low-, medium- and high-risk group. Our results showed that the densities of CD3 + and CD8 + cells were higher in the desmoplastic HGP (dHGP) group than in the non-dHGP group, and the proportion of high immunoscores was also higher in the dHGP group (51.9% vs. 33.0%, respectively, P = 0.020). Patients with the dHGP had significantly longer relapse-free survival (RFS) and overall survival (OS) than those with the non-HGP. The low-risk group showed significantly higher 2-year RFS and 5-year OS rates than the other two groups (RFS: 76.2%, 43.7% and 33.1%, respectively; P < 0.001; OS: 89.7%, 54.4% and 33.3%, respectively; P < 0.001). In conclusion, the dHGP correlates with relatively high immunoscores, predicting a favorable prognosis independent of the immunoscore and CRS. A novel risk score combining HGPs, the immunoscore and the CRS may be used for the stratification of CRLM patients' survival.

The Immunoscore system predicts prognosis after liver metastasectomy in colorectal cancer liver metastases.

BACKGROUND: The Immunoscore was initially established to evaluate the prognosis of stage I/II/III colorectal cancer patients. However, the feasibility of the Immunoscore for the prognosis of colorectal cancer liver metastases (CRCLM) has not been reported. METHODS: Liver metastases in 249 CRCLM patients were retrospectively analyzed. The Immunoscore was assessed according to the counts and densities of CD3+ and CD8+ T cells in the central- and peritumoral areas by immunohistochemistry. The prognostic role of the Immunoscore for relapse-free survival (RFS) and overall survival (OS) was analyzed with Kaplan-Meier curves and Cox multivariate models, and confirmed via an internal validation. Receiver operating characteristic (ROC) curves were plotted to compare the prognostic values of the Immunoscore and the clinical risk score (CRS) system. RESULTS: CRCLM patients with high Immunoscores (> 2) had significantly longer RFS [median RFS (95% confidence interval; 95% CI) 21.4 (7.8-35.1) vs. 8.7 (6.8-10.5) months, P < 0.001] and OS [median OS (95% CI): not reached vs. 28.7 (23.2-34.2) months, P < 0.001] than those with low Immunoscores (≤ 2). After stratification by CRS, the Immunoscore retained a statistically significant prognostic value for OS. The areas under the ROC curves (AUROCs) of the Immunoscore and the CRS system for RFS were 0.711 [95% CI 0.642-0.781] and 0.675[95% CI 0.601-0.749] (P = 0.492), whereas the AUROC of the Immunoscore system for OS was larger than that of the CRS system [0.759 (95% CI 0.699-0.818) vs. 0.660 (95% CI 0.592-0.727); P = 0.029]. CONCLUSIONS: The Immunoscore of liver metastases can be applied to predict the prognosis of CRCLM patients following liver resection.

Liver hypertrophy and accelerated growth of implanted tumors in nonembolized liver of rabbit after left portal vein embolization.

BACKGROUND: Portal vein embolization (PVE) has become a standard preoperative procedure to promote hypertrophy of the future remnant liver to reduce postoperative liver failure. Whether PVE accelerates tumor growth is still controversial. We developed a left PVE procedure and investigated its effect on liver hypertrophy and tumor growth in a rabbit liver tumor model. MATERIALS AND METHODS: VX2 tumors were implanted in both the external left and right middle lobe (the bilateral group) or in the external left lobe only (the unilateral group) of rabbit liver. Both groups were further divided into a PVE or a sham/control group. Tumor volume and tumor growth rate as volume relative increase were determined by ultrasound. Liver volume-to-body weight index, an index for liver volume, was compared. Serum HGF was measured by ELISA. RESULTS: In the bilateral PVE group, tumor volume and relative increase value in the nonembolized lobe were significantly (71% and 65%, respectively) greater than those in the control group at 5 d post-PVE. In the unilateral PVE group, liver volume-to-body weight index of the nonembolized lobes was significantly increased by 17%. Increase of serum HGF level after PVE was correlated well with both tumor growth and liver hypertrophy. CONCLUSIONS: Left PVE promoted both the growth of implanted tumors and liver hypertrophy in the nonembolized liver, in which serum HGF might play an important role.

LIN28 expression and prognostic value in hepatocellular carcinoma patients who meet the Milan criteria and undergo hepatectomy.

Stem cell marker LIN28, related closely with SOX2 and OCT4, has been studied as a biomarker for the maintainance of pluripotent cells in several malignancies. Our previous study showed that SOX2 and OCT4 were negative predictors for hepatocellular carcinoma (HCC). However, the predictive value of LIN28 in HCC outcome is still undetermined. We hypothesized that LIN28 may also play a role as a biomarker for HCC. To test this hypothesis, we examined the expression of LIN28 in 129 radically resected HCC tissues using reverse transcription-polymerase chain reaction and analyzed the association of LIN28 expression with clinicopathologic features and prognosis. Our study showed that LIN28 was expressed at a higher frequency in tumor tissues than in non-HCC tissues (45.0% vs. 21.7%, P = 0.020). Moreover, LIN28 expression was significantly increased in cases with large tumor size (P = 0.010). Univariate analysis did not reveal a significant correlation between LIN28 expression and overall survival or recurrence-free survival. For HCC patients who met the Milan criteria, stratified analysis revealed shorter overall survival (P = 0.007) and recurrence-free survival (P < 0.001) in those with detectable LIN28 expression compared to those with no detectable LIN28 expression. Furthermore, multivariate analysis revealed that LIN28 was a negative independent predictor for both overall survival (hazard ratio= 7.093, P = 0.017) and recurrence-free survival (hazard ratio=5.518, P = 0.004) in patients who met the Milan criteria. Taken together, our results suggest that LIN28 identifies low-risk and high-risk subsets of HCC patients meeting the Milan criteria who undergo hepatectomy.

More Liver Metastases Detected Intraoperatively Indicates Worse Prognosis for Colorectal Liver Metastases Patients after Resection Combined with Microwave Ablation.

Background: Whether more tumor numbers detected in surgery compared to preoperative image affecting survival of colorectal liver metastases (CRLM) patients after hepatectomy combined with microwave ablation (MWA) remains unclear. Methods: From 2013 to 2018, 85 CRLM patients who underwent hepatectomy combined with MWA were retrospectively assessed. Compared to the tumor numbers in preoperative image, patients with equal intraoperative tumor numbers were defined as the equal number group (n = 45); patients detected more tumor numbers in surgery were defined as the more number group (n = 40). Clinicopathological factors and prognosis were compared between two groups. Results: Compared to the equal number group, the more number group was characterized by more lymphatic metastasis, synchronous metastasis of liver lesion, and tumor numbers over 5 (all P < 0.05). Median survival time was 46.7 months and 26.8 months in the equal and more number group. Significantly worse overall survival (OS) was found in more number group to the equal number group (P = 0.027). In Cox analysis, more tumor number than image and high level of carbohydrate antigen 19-9 (CA19-9) were poor prognostic factors for OS. Conclusion: In patients receiving hepatectomy combined with MWA, detecting more liver metastases in surgery than preoperative image indicates poor long-term survival. These patients were characterized by more lymphatic metastasis, synchronous metastasis of liver lesion, and tumor numbers over 5. Intensive follow-up to detect early recurrence and potent postoperative therapy to improve survival may be justified in patients detected more tumor numbers in surgery with a high CA19-9 level.

A detailed assessment of liver function in patients with hepatocellular carcinoma via the modified albumin-bilirubin (mALBI) grade.

Recently, the albumin-bilirubin (ALBI) score, a continuous index consisting of only albumin and bilirubin, has been developed for objectively assessing liver function in patients with hepatocellular carcinoma (HCC). However, the ALBI score was arbitrarily categorized into three ALBI grades based on two artificially predetermined cutoff points with no explanation and statistical grounds, causing a considerable loss of discriminatory ability. This study aims to propose a modified ALBI (mALBI) grade for offering a detailed evaluation of hepatic reserve and specify its role during clinical practice in the HCC setting. The study population comprised 3540 HCC patients treated with mainstream therapies including hepatectomy (n=2056), thermal ablation (n=550), and transcatheter arterial chemoembolization (n=934) from 2002 to 2017. The ALBI score was stratified into four mALBI grades through a recently proposed statistical method aiming to select the optimal cutoff points of a continuous predictive variable by maximizing the discriminative ability in a multivariable Cox regression model. The mALBI grade had an overall better discriminatory ability than the ALBI grade in predicting overall survival through Harrell's C-index (0.614 vs. 0.598, P<0.001). Both visual inspections of Kaplan-Meier curves and calculation of hazard ratios displayed a more subtle evaluation of liver function using the mALBI grade. Moreover, the newly identified cut-point (ALBI score = -2.29) between the mALBI grade 2a and 2b was much closer to a 30% retention rate of indocyanine green at 15 minutes, an indicator for the performance of a subsegmentectomy. The newly proposed mALBI grade provides a more subtle assessment of liver function to guide clinical decision-making and predicts the prognosis of HCC patients more accurately than the original ALBI grade.

Overexpression of eIF5A-2 is an adverse prognostic marker of survival in stage I non-small cell lung cancer patients.

We have previously isolated an oncogene EIF5A2 (eukaryotic initiation factor 5A2) from a frequently amplified region at 3q of a primary ovarian cancer cell line, and demonstrated its impact on prognosis in human ovarian cancer. Amplification of chromosome 3q has also been detected frequently in non-small cell lung cancer (NSCLC), however, abnormalities of EIF5A2 and its clinicopathologic significance in NSCLC haven't been studied. In our study, the methods of immunohistochemistry and fluorescence in situ hybridization were utilized to examine protein expression and amplification of EIF5A2 in 248 surgically resected NSCLCs (learning cohort) and another validation cohort of 120 stage I NSCLC patients. Overexpression and amplification of EIF5A2 was detected informatively in 48.7% and 13.7% of NSCLCs in learning cohort, 33.3% and 6.0% of NSCLCs in validation cohort. Overexpression of eIF5A-2 was found to correlate with gene amplification, increased cell proliferation and advanced T stage. In learning cohort, eIF5A-2 expression was evaluated as a strong prognostic factor on disease-specific survival, but in subgroup analyses, it only retained its stratified significance in stage I set (Hazards ratio = 2.799, p = 0.001). In validation cohort, the impact of eIF5A-2 expression on survival in stage I NSCLC patients was also observed (Hazard ratio = 2.097, p = 0.014). Our findings suggested that overexpression of eIF5A-2 correlates with local invasion of NSCLC, and might serve as an adverse prognostic marker of survival for stage I NSCLC patients.

Prognostic value of Wnt inhibitory factor-1 expression in hepatocellular carcinoma that is independent of gene methylation.

Recently, Wnt inhibitory factor-1 (WIF-1) was found to be epigenetically inactivated in several solid tumors, but the biological and clinical relevance of WIF-1 methylation and expression status in hepatocellular carcinoma (HCC) are still unclear. In the present study, reverse transcription polymerase chain reaction (PCR) and methylation-specific PCR were used to examine the WIF-1 expression and methylation in HCC cell lines. In addition, methylation and expression status of WIF-1 in 105 HCC cases were correlated with clinicopathological parameters and prognosis after tumor resection. WIF-1 was expressed in one HCC cell line and L02, both of which were not methylated in promoter region. DNA hypermethylation of WIF-1 promoter was identified in the other four HCC cell lines without WIF-1 expression. In neoplastic and non-neoplastic tissue samples, the rates of WIF-1 methylation were 61.9% and 37.1% (P = 0.001), respectively. WIF-1 was significantly downregulated in neoplastic tissues at messenger ribonucleic acid (mRNA) level, as compared to adjacent non-neoplastic tissues (P = 0.006). A significant inverse association was observed between WIF-1 methylation of and WIF-1 expression (P 0.017, R = -0.232). Methylation of WIF-1 was not associated with patient survival. In contrast, patients whose tumors exhibited negative WIF-1 mRNA expression had lower rates of overall survival. These findings suggested that aberrant methylation of WIF-1 is a common event in hepatocarcinogenesis. In addition, expression, but not methylation, of WIF-1 is a predictor of good outcome in patients undergoing resection of HCC.

NADPH oxidase DUOX1 and DUOX2 but not NOX4 are independent predictors in hepatocellular carcinoma after hepatectomy.

NADPH oxidase DUOX1, DUOX2, and NOX4 have recently been gained considerable concerns, owing to the fact that they involve in reactive oxygen species-induced genetic and epigenetic alternations of human carcinogenesis and serve as biomarkers in several cancers. Whether they predict survival in hepatocellular carcinoma (HCC) is still uncertain. Here, we detected the expressions of DUOX1, DUOX2, and NOX4 in one normal liver cell line, seven HCC cell lines, 30 non-cirrhotic normal liver tissues, and 107 paired HCC tissues using reverse transcription-polymerase chain reaction. The correlations of genes expression with prognoses were analyzed. DUOX1 was expressed at high levels in MHCC-97H and MHCC-97L, but at low levels in Bel-7402. In contrast to low expression level at SMMC-7721, DUOX2 was expressed at considerably high levels in MHCC-97H and MHCC-97L. The transcript of NOX4 was only detected in SMMC-7721. All the 30 normal liver tissues failed to express the three candidate markers. Compared with adjacent non-neoplastic tissues, DUOX1, DUOX2, and NOX4 were expressed at higher frequencies in tumor specimens. Both univariate and multivariate analyses revealed that elevated expression of DUOX1 or DUOX2 predicted poorer recurrence-free survival and overall survival. No such significance trend regarding NOX4 predictive value in survival, however, was seen in univariate analysis. These results suggested DUOX1 and DUOX2, but not NOX4, could predict HCC prognoses after hepatectomy.

Identification of a TGF-ß/SMAD/lnc-UTGF positive feedback loop and its role in hepatoma metastasis.

Aberrant activation of the TGF-ß/SMAD signaling pathway is often observed in hepatocellular carcinoma (HCC). Whether lncRNA regulates the TGF-ß/SMAD signaling remains largely unknown. Here, we identified an oncogenic lncRNA that was upregulated in HCC and was transcriptionally induced by TGF-ß (named lnc-UTGF, lncRNA upregulated by TGF-ß). Upon TGF-ß stimulation, SMAD2/3 bound to the lnc-UTGF promoter and activated lnc-UTGF expression. In turn, the TGF-ß/SMAD signaling was augmented by overexpressing lnc-UTGF, but was inhibited by silencing lnc-UTGF. Mechanism investigations revealed that lnc-UTGF interacted with the mRNAs of SMAD2 and SMAD4 via complementary base-pairing, resulting in enhanced stability of SMAD2/4 mRNAs. These data suggest a novel TGF-ß/SMAD/lnc-UTGF positive feedback circuitry. Subsequent gain- and loss-of-function analyses disclosed that lnc-UTGF promoted the migration and invasion of hepatoma cells, and this effect of lnc-UTGF was attenuated by repressing SMAD2/4 expression or by mutating the SMAD2/4-binding sites in lnc-UTGF. Studies using mouse models further confirmed that in vivo metastasis of hepatoma xenografts was inhibited by silencing lnc-UTGF, but was enhanced by ectopic expression of lnc-UTGF. The lnc-UTGF level was positively correlated with the SMAD2/4 levels in xenografts. Consistently, we detected an association of lnc-UTGF upregulation with increase of SMAD2, SMAD4, and their metastasis effector SNAIL1 in human HCC. And high lnc-UTGF level was also significantly associated with enhanced metastasis potential, advanced TNM stages, and worse recurrence-free survival. Conclusion: there exists a lnc-UTGF-mediated positive feedback loop of the TGF-ß signaling and its deregulation promotes hepatoma metastasis. These findings may provide a new therapeutic target for HCC metastasis.

A novel prognostic nomogram for colorectal cancer liver metastasis patients with recurrence after hepatectomy.

PURPOSE: We aimed to construct a nomogram to predict personalized post-recurrence survival (PRS) among colorectal cancer liver metastasis (CRLM) patients with post-hepatectomy recurrence. METHODS: Colorectal cancer liver metastasis patients who received initial hepatectomy and had subsequent recurrence between 2001 and 2019 in Sun Yat-sen University Cancer Center from China were included in the study. Patients were randomly assigned to a training cohort and a validation cohort on a ratio of 2:1. Univariable analysis was first employed to select potential predictive factors for PRS. Then, the multivariable Cox regression model was applied to recognize independent prognostic factors. According to the model, a nomogram to predict PRS was established. The nomogram's predictive capacity was further assessed utilizing concordance index (C-index) values, calibration plots, and Kaplan-Meier curves. RESULTS: About 376 patients were finally enrolled, with a 3-year PRS rate of 37.3% and a 5-year PRS rate of 24.6%. The following five independent predictors for PRS were determined to construct the nomogram: the largest size of liver metastases at initial hepatectomy, relapse-free survival, CEA level at recurrence, recurrent sites, and treatment for recurrence. The nomogram displayed fairly good discrimination and calibration. The C-index value was 0.742 for the training cohort and 0.773 for the validation cohort. Patients were grouped into three risk groups very well by the nomogram, with 5-year PRS rates of 45.2%, 23.3%, and 9.0%, respectively (p < 0.001) in the training cohort and 36.0%, 9.2%, and 4.6%, respectively (p < 0.001) in the validation cohort. CONCLUSION: A novel nomogram was built and validated to enable the prediction of personal PRS in CRLM patients with post-hepatectomy recurrence. The nomogram may help physicians in decision making.

Dynamic monitoring of circulating tumor DNA to predict prognosis and efficacy of adjuvant chemotherapy after resection of colorectal liver metastases.

Rationale: Hepatectomy and adjuvant chemotherapy after resection of colorectal liver metastases (CRLM) may improve survival, however, patients which may benefit cannot currently be identified. Postoperative circulating tumor DNA (ctDNA) analysis can detect minimal residual disease (MRD) and predict the prognosis and efficacy of adjuvant chemotherapy. Our study aims to determine the impact of serial ctDNA analysis to predict the outcome among patients undergoing resection of CRLM. Methods: Between May 2018 and October 2019, 91 CRLM patients were prospectively enrolled. Whole exome sequencing was performed in 50 primary and 48 metastatic liver tissues. Targeted sequencing of 451 cancer relevant genes was performed in 50 baseline plasma to determine plasma-tissue concordance. We prospectively investigated changes in the amount and constitution of ctDNA in 271 serial plasma samples taken at different time points (baseline, pre-operation, post-operation, post-operative adjuvant chemotherapy (post-ACT) and recurrence) during the treatment of CRLM. Results: Detected molecular alterations were highly consistent among baseline ctDNA, primary and liver metastases tissue. Patients with a higher variant allele frequency (VAF) level at baseline ctDNA represent a higher tumor burden, and decreased ctDNA during pre-operative chemotherapy predicted better tumor response. Patients with detectable post-operative and post-ACT ctDNA were associated with significantly shorter recurrence-free survival (RFS). ROC analysis showed that post-ACT ctDNA status was superior to post-operative ctDNA status in predicting RFS with an AUROC of 0.79. A significant difference in overall recurrence rate was observed in patients with detectable vs undetectable levels of ctDNA after resection of CRLM (79.4% vs 41.7%) and after completion of adjuvant chemotherapy (77.3% vs 40.7%). During adjuvant chemotherapy, patients with decreased ctDNA VAF after adjuvant chemotherapy had a recurrence rate of 63.6%, compared to 92.3% in patients with increased ctDNA VAF. Conclusions: We envision that dynamic ctDNA analysis, especially in a post-ACT setting, might be used to not only reflect MRD but also to determine rational personalized adjuvant therapy after the resection of CRLM.

Primary tumor immune score fails to predict the prognosis of colorectal cancer liver metastases after hepatectomy in Chinese populations.

BACKGROUND: Increasing evidence suggests that the immune score is significantly associated with cancer prognosis. However, the prognostic role of primary tumor immune score in colorectal cancer liver metastases (CRLM) after hepatectomy in Chinese patients has not been reported. The present study is designed to investigate whether the immune score of primary tumor can predict the postoperative survival of liver metastases in Chinese patients. METHODS: A total of 131 patients diagnosed with CRLM were included, and the corresponding primary tumor and liver metastasis specimens were acquired. An immune score ranging from 0 to 4 was established based on the counts and densities of CD3+ and CD8+ T cells in the core tumor (CT) and the invasive margin (IM). Relapse-free survival (RFS) and overall survival (OS) were analyzed by Kaplan-Meier curves to assess the prognostic role of primary tumor immune score. Furthermore, we conducted a comprehensive search of the Gene Expression Omnibus (GEO) and selected stage IV colorectal cancer (CRC) patients with liver metastasis to compare the tumor-infiltrating T cell profiles of the primary tumor and liver metastases by CIBERSORT. RESULTS: Patients with high immune scores in the primary tumor has no significantly better RFS and OS after hepatectomy than those with low immune scores [median RFS (95% CI): 19.13 (10.07-28.20) vs. 27.13 (15.97-38.29) months, P=0.604; median OS (95% CI): 64.37 (35.96-92.78) vs. 40.07 (32.54-47.59) months, P=0.652]. Data collected from the GEO indicates that the proportion of CD8+ T cells and total T cells in the primary tumor and liver metastatic lesion are also not significantly correlated (CD8+ T cells: r2 =0.030, P=0.468; total T cells: r2 =0.165, P=0.076). CONCLUSIONS: The immune score of the primary tumor fails to predict the prognosis of CRLM after hepatectomy in Chinese patients.

High expression of EZH2 is associated with tumor aggressiveness and poor prognosis in patients with esophageal squamous cell carcinoma treated with definitive chemoradiotherapy.

The enhancer of zeste homolog 2 (EZH2), a known repressor of gene transcription, has been reported to be associated with biological malignancy in several cancers. The potential oncogenic role of EZH2 and its clinical/prognostic significance, however, in esophageal squamous cell carcinoma (ESCC) are unclear. In this study, the methods of immunohistochemistry and fluorescence in-situ hybridization were used to examine protein expression and amplification of EZH2 in 98 pretreatment biopsy specimens of ESCC who received definitive chemoradiotherapy (CRT). High expression of EZH2 and amplification of EZH2 was found in 54.1% and 12.0% of ESCCs, respectively. High EZH2 expression was significantly correlated with increased cell proliferation (p = 0.009), high histopathological grade (p = 0.002), regional (p = 0.025) and distant lymph node metastasis (p < 0.001) and lack of clinical complete response to CRT (p = 0.028). Univariate analysis revealed that high expression of EZH2 was associated with poor metastasis-free survival (MFS) (p = 0.003), poor progression-free survival (PFS) (p = 0.001) and poor disease-specific survival (DSS) (p < 0.001). In multivariate analysis, high expression of EZH2, together with lack of clinical complete response, were evaluated as significant independent prognostic factors of MFS, PFS and DSS for patients with ESCC. These findings suggest that high expression of EZH2 correlates with tumor aggressiveness and adverse patient outcome in ESCC treated with definitive CRT. Evaluation of EZH2 expressions might be useful for predicting tumor response to CRT and prognosis for patients with ESCC.

[OCT4 expression in hepatocellular carcinoma and its clinical significance].

BACKGROUND AND OBJECTIVE: Recently, many studies have focused on stem cells in hepatocellular carcinoma (HCC) and found some stem cell markers in HCC, which are associated with the prognosis. OCT4, as a member of the POU transcription factor family, is a key factor to maintain self-renewal and pluripotency of embryonic stem cells (ESCs). This study was to explore the expression of the ESCs marker OCT4A in HCC, and its correlations with clinicopathologic features and prognosis of HCC. METHODS: OCT4A mRNA expression was detected in five liver cancer cell lines (SMMC-7721, BEL-7402, Hep-G2, MHCC97-L, and MHCC97-H), one immortalized liver cell line L-O2, tumor tissues with matched non-neoplastic liver tissues in 107 HCC patients, and normal liver tissues of 20 cases using semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR). The correlations between OCT4A mRNA and clinicopathologic features and prognosis of HCC were analyzed. RESULTS: OCT4A mRNA was detected in SMMC-7721, BEL-7402, Hep-G2, MHCC-97L, and MHCC-97H cells, but not in L-O2 cells. The positive rate of OCT4A mRNA expression was significantly higher in the HCC tissues than in the non-neoplastic liver tissues (72.0% vs. 30.8%, P<0.001). No OCT4A mRNA expression was found in the normal liver tissues. OCT4A mRNA expression was correlated with the tumor size, vascular invasion, and TNM stage (P<0.05). Kaplan-Meier survival curves showed that patients with positive expression of OCT4A mRNA had lower overall survival and disease-free survival rates. CONCLUSIONS: OCT4A mRNA, which is highly expressed in a subset of liver cancer cell lines and HCC tissues, may be involved in the carcinogenesis of HCC. OCT4A mRNA may be a valuable biomarker for assessing the prognosis of HCC.

Overexpression of AIB1 predicts resistance to chemoradiotherapy and poor prognosis in patients with primary esophageal squamous cell carcinoma.

AIB1 (amplified in breast cancer 1) is frequently overexpressed in esophageal squamous cell carcinoma (ESCC), but the significance of AIB1 expression in chemoradiotherapy (CRT) sensitivity and its effect on prognosis are still unclear. In this study, the expression of AIB1 was examined by immunohistochemistry in 98 biopsy specimens of primary ESCC patients treated with definitive CRT. AIB1 overexpression was found in 63/98 (64.3%) of the ESCCs. There was a significant association between AIB1 overexpression and distant lymph node metastases (P = 0.011), but not regional lymph node metastases. In the M0 subgroup, overexpression of AIB1 was observed more frequently in stage T4 than in stage T2-3 (66.7%vs 38.5%, P = 0.031). In addition, AIB1 expression was the only factor that showed a significant correlation with CRT response, in which overexpression of AIB1 was observed more frequently in the CRT resistant group than in the CRT effective group (86.5%vs 50.8%, P < 0.001). Univariate analysis revealed that AIB1 overexpression was associated with poor progression-free survival (PFS) (P < 0.001) and poor disease-specific survival (DSS) (P <0.001). Furthermore, AIB1 expression could stratify patient survival in stages T2-3, T4, N1, and M0 (P < 0.05), as well as in the CRT effective group (P < 0.05), and AIB1 overexpression and CRT resistance were evaluated as significant independent prognostic factors for both PFS and DSS in multivariate analysis. These findings suggest that overexpression of AIB1 is a useful predictor of CRT resistance and an independent molecular marker of poor prognosis for ESCC patients.

Clusterin as a predictor for chemoradiotherapy sensitivity and patient survival in esophageal squamous cell carcinoma.

Clusterin (CLU) is frequently overexpressed and correlates closely with chemotherapy and radiotherapy resistance and poor prognosis in many human cancers. However, the significance of CLU expression in chemoradiotherapy (CRT) sensitivity and its effect on the prognosis of esophageal squamous cell carcinoma (ESCC) are still unknown. In the present study, we used the methods of immunohistochemistry and terminal deoxyuridine triphosphate nick-end labeling assay to examine the expression status of CLU and apoptotic index in 110 pretreated biopsy specimens of ESCC patients treated with definitive CRT. High expression of CLU was observed in 42.7% of epithelium and 50.0% of stroma in ESCC. A significant association of high CLU stromal expression with large tumor size (P = 0.012) and locoregional progression (P = 0.001) was observed, and high epithelial expression of CLU showed a significant correlation with the lack of complete response (P = 0.028) and low apoptotic index (P = 0.001). Univariate analysis revealed that high CLU stromal expression was associated with poor locoregional progression-free survival, distant progression-free survival, and overall survival. Furthermore, ESCC patients with high CLU expression in both epithelium and stroma have the shortest survival time among the subgroups of different CLU expression status. In multivariate analysis, CLU stromal expression was evaluated as an independent prognostic factor for locoregional progression-free survival, distant progression-free survival, and overall survival. These findings suggest an important role for CLU, especially in stroma, in ESCC progression, and that high CLU epithelial expression might be a promising predictor of ESCC resistance to CRT.

Prognostic impact of H3K27me3 expression on locoregional progression after chemoradiotherapy in esophageal squamous cell carcinoma.

BACKGROUND: Trimethylation of lysine 27 on histone H3 (H3K27me3) by enhancer of zeste homolog 2 (EZH2) is an epigenetic mark that mediates gene silencing. EZH2 is overexpressed and correlates with poor prognosis in many cancers. However, the clinical implication of H3K27me3 in human malignancies has not been well established. We wished to ascertain whether a correlation exists between the expression of H3K27me3 and clinical outcome in a group of patients with esophageal squamous cell carcinoma (ESCC) treated with definitive chemoradiotherapy (CRT). METHODS: The method of immunohistochemistry (IHC) was utilized to examine the protein expression of H3K27me3 in 98 pretreatment biopsy specimens of ESCC and in 30 samples of normal esophageal mucosa. The clinical/prognostic significance of H3K27me3 expression was statistically analyzed. RESULTS: The expression frequency and expression levels of H3K27me3 were significantly higher in ESCCs than in normal tissues. There was a positive correlation between H3K27me3 expression and WHO grade (P = 0.016), tumor size (P = 0.019), T status (P = 0.024), locoregional progression (P = 0.009) and EZH2 expression (P = 0.036). High H3K27me3 expression was associated with poor locoregional progression-free survival (LPFS) (P = 0.010) in ESCC. Further analysis demonstrated that H3K27me3 could stratify patient outcome in T2-3 (P = 0.048), N0 (P = 0.005) and M0 (P = 0.018) stages as well as in CRT effective group (P = 0.022). CONCLUSIONS: Our data suggests that H3K27me3 expression examined by IHC might be useful for stratifying LPFS for different subsets of ESCC patients treated with definitive CRT.