A novel engineered IL-21 receptor arms T-cell receptor-engineered T cells (TCR-T cells) against hepatocellular carcinoma.

Strategies to improve T cell therapy efficacy in solid tumors such as hepatocellular carcinoma (HCC) are urgently needed. The common cytokine receptor γ chain (γc) family cytokines such as IL-2, IL-7, IL-15 and IL-21 play fundamental roles in T cell development, differentiation and effector phases. This study aims to determine the combination effects of IL-21 in T cell therapy against HCC and investigate optimized strategies to utilize the effect of IL-21 signal in T cell therapy. The antitumor function of AFP-specific T cell receptor-engineered T cells (TCR-T) was augmented by exogenous IL-21 in vitro and in vivo. IL-21 enhanced proliferation capacity, promoted memory differentiation, downregulated PD-1 expression and alleviated apoptosis in TCR-T after activation. A novel engineered IL-21 receptor was established, and TCR-T armed with the novel engineered IL-21 receptors (IL-21R-TCR-T) showed upregulated phosphorylated STAT3 expression without exogenous IL-21 ligand. IL-21R-TCR-T showed better proliferation upon activation and superior antitumor function in vitro and in vivo. IL-21R-TCR-T exhibited a less differentiated, exhausted and apoptotic phenotype than conventional TCR-T upon repetitive tumor antigen stimulation. The novel IL-21 receptor in our study programs powerful TCR-T and can avoid side effects induced by IL-21 systemic utilization. The novel IL-21 receptor creates new opportunities for next-generation TCR-T against HCC.

TCR T cells overexpressing c-Jun have better functionality with improved tumor infiltration and persistence in hepatocellular carcinoma.

Background: The overall 5-year survival rate of hepatocellular carcinoma (HCC), a major form of liver cancer, is merely 20%, underscoring the need for more effective therapies. We recently identified T cell receptors (TCR) specific for the HLA-A2/alpha fetoprotein amino acids 158-166 (AFP158) and showed that these TCR engineered T cells could control HCC xenografts in NSG mice. However, their efficacy was limited by poor expansion, loss of function, and short persistence of the TCR T cells. Here, we studied whether overexpression of c-Jun, a transcription factor required for T cell activation, in the TCR T cells could enhance their expansion, function, and persistence in HCC tumor models. Methods: Recombinant lentiviral vectors (lv), expressing either the HLA-A2/AFP158-specific TCR or both the TCR and c-Jun (TCR-JUN), were constructed and used to transduce primary human T cells to generate the TCR or TCR-JUN T cells, respectively. We compared the expansion, effector function, and exhaustion status of the TCR and TCR-JUN T cells in vitro after HCC tumor stimulation. Additionally, we studied the persistence and antitumor effects of the TCR and TCR-JUN T cells using the HCC xenografts in NSG mice. Results: We could effectively transduce primary human T cells to express both TCR and c-Jun. Compared to the HLA-A2/AFP158 TCR T cells, the TCR-JUN T cells have better expansion potential in culture, with enhanced functional capacity against HCC tumor cells. In addition, the TCR-JUN T cells were less apoptotic and more resistant to exhaustion after HepG2 tumor stimulation. In the HCC xenograft tumor model, c-Jun overexpression enhanced the TCR T cell expansion and increased the overall survival rate of the treated mice. Importantly, the TCR-JUN T cells were less exhausted in the tumor lesions and demonstrated enhanced tumor infiltration, functionality, and persistence. Conclusion: c-Jun overexpression can enhance the expansion, function, and persistence of the A2/AFP158 TCR engineered T cells. The c-Jun gene co-delivery has the potential to enhance the antitumor efficacy of AFP specific TCR T cells when treating patients with HCC.

Combination of AFP vaccine and immune checkpoint inhibitors slows hepatocellular carcinoma progression in preclinical models.

Many patients with hepatocellular carcinoma (HCC) do not respond to the first-line immune checkpoint inhibitor treatment. Immunization with effective cancer vaccines is an attractive alternative approach to immunotherapy. However, its efficacy remains insufficiently evaluated in preclinical studies. Here, we investigated HCC-associated self/tumor antigen, α-fetoprotein-based (AFP-based) vaccine immunization for treating AFP (+) HCC mouse models. We found that AFP immunization effectively induced AFP-specific CD8+ T cells in vivo. However, these CD8+ T cells expressed exhaustion markers, including PD1, LAG3, and Tim3. Furthermore, the AFP vaccine effectively prevented c-MYC/Mcl1 HCC initiation when administered before tumor formation, while it was ineffective against full-blown c-MYC/Mcl1 tumors. Similarly, anti-PD1 and anti-PD-L1 monotherapy showed no efficacy in this murine HCC model. In striking contrast, AFP immunization combined with anti-PD-L1 treatment triggered significant inhibition of HCC progression in most liver tumor nodules, while in combination with anti-PD1, it induced slower tumor progression. Mechanistically, we demonstrated that HCC-intrinsic PD-L1 expression was the primary target of anti-PD-L1 in this combination therapy. Notably, the combination therapy had a similar therapeutic effect in the cMet/ß-catenin mouse HCC model. These findings suggest that combining the AFP vaccine and immune checkpoint inhibitors may be effective for AFP (+) HCC treatment.

The affinity of antigen-binding domain on the antitumor efficacy of CAR T cells: Moderate is better.

The overall efficacy of chimeric antigen receptor modified T cells (CARTs) remain limited in solid tumors despite intensive studies that aim at targeting multiple antigens, enhancing migration, reducing tonic signaling, and improving tumor microenvironment. On the other hand, how the affinity and engaging kinetics of antigen-binding domain (ABD) affects the CART's efficacy has not been carefully investigated. In this article, we first analyzed 38 published solid tumor CART trials and correlated the response rate to their ABD affinity. Not surprisingly, majority (25 trials) of the CARTs utilized high-affinity ABDs, but generated merely 5.7% response rate. In contrast, 35% of the patients treated with the CARTs built from moderate-affinity ABDs had clinical responses. Thus, CARTs with moderate-affinity ABDs not only have less off-target toxicity, but also are more effective. We then reviewed the effects of ABD affinity on the biology and function of CARTs, providing further evidence that moderate-affinity ABDs may be better in CART development. In the end, we propose that a fast-on/fast-off (high Kon and Koff ) kinetics of CART-target engagement in solid tumor allow CARTs to generate sufficient signaling to kill tumor cells without being driven to exhaustion. We believe that studying the ABD affinity and the kinetics of CART-tumor interaction may hold a key to designing effective CARTs for solid tumors.

Chimeric antigen receptor engineered T cells and their application in the immunotherapy of solid tumours.

In this article, we reviewed the current literature studies and our understanding of the parameters that affect the chimeric antigen receptor T cells (CAR-T's) activation, effector function, in vivo persistence, and antitumour effects. These factors include T cell subsets and their differentiation stages, the components of chimeric antigen receptors (CAR) design, the expression promoters and delivery vectors, and the CAR-T production process. The CAR signalling and CAR-T activation were also studied in comparison to TCR. The last section of the review gave special consideration of CAR design for solid tumours, focusing on strategies to improve CAR-T tumour infiltration and survival in the hostile tumour microenvironment. With several hundred clinical trials undergoing worldwide, the pace of CAR-T immunotherapy moves from bench to bedside is unprecedented. We hope that the article will provide readers a clear and comprehensive view of this rapidly evolving field and will help scientists and physician to design effective CAR-Ts immunotherapy for solid tumours.

Novel low-avidity glypican-3 specific CARTs resist exhaustion and mediate durable antitumor effects against HCC.

BACKGROUND AND AIMS: Chimeric antigen receptor engineered T cells (CARTs) for HCC and other solid tumors are not as effective as they are for blood cancers. CARTs may lose function inside tumors due to persistent antigen engagement. The aims of this study are to develop low-affinity monoclonal antibodies (mAbs) and low-avidity CARTs for HCC and to test the hypothesis that low-avidity CARTs can resist exhaustion and maintain functions in solid tumors, generating durable antitumor effects. METHODS AND RESULTS: New human glypican-3 (hGPC3) mAbs were developed from immunized mice. We obtained three hGPC3-specific mAbs that stained HCC tumors, but not the adjacent normal liver tissues. One of them, 8F8, bound an epitope close to that of GC33, the frequently used high-affinity mAb, but with approximately 17-fold lower affinity. We then compared the 8F8 CARTs to GC33 CARTs for their in vitro function and in vivo antitumor effects. In vitro, low-avidity 8F8 CARTs killed both hGPC3high and hGPC3low HCC tumor cells to the same extent as high-avidity GC33 CARTs. 8F8 CARTs expanded and persisted to a greater extent than GC33 CARTs, resulting in durable responses against HCC xenografts. Importantly, compared with GC33 CARTs, there were 5-fold more of 8F8-BBz CARTs in the tumor mass for a longer period of time. Remarkably, the tumor-infiltrating 8F8 CARTs were less exhausted and apoptotic, and more functional than GC33 CARTs. CONCLUSION: The low-avidity 8F8-BBz CART resists exhaustion and apoptosis inside tumor lesions, demonstrating a greater therapeutic potential than high-avidity CARTs.

Parthenolide plays a protective role in the liver of mice with metabolic dysfunction­associated fatty liver disease through the activation of the HIPPO pathway.

Metabolic dysfunction­associated fatty liver disease (MAFLD) is a serious threat to human health. Parthenolide (PAR) displays several important pharmacological activities, including the promotion of liver function recovery during hepatitis. The aim of the present study was to assess the effect of PAR on MAFLD in a mouse model. Body weight, liver to body weight ratios, histological score, alanine transaminase, aspartate transaminase, total cholesterol and triglyceride levels were determined to evaluate liver injury. Liver hydroxyproline concentrations were also assessed. The expression levels of lipid metabolism­related genes (sterol regulatory element binding protein­1c, fatty acid synthase, acetyl CoA carboxylase 1, stearoyl CoA desaturase 1 and carbohydrate response element­binding protein, peroxisome proliferator­activated receptor α, carnitine palmitoyl transferase 1α and acyl­CoA dehydrogenase short chain), liver fibrosis­associated genes (α­smooth muscle actin, tissue inhibitor of metalloproteinase 1 and TGF­ß1), pro­inflammatory cytokines (TNF­α, IL­1ß and IL­6) and oxidative stress­associated enzymes (malondialdehyde, superoxide dismutase and glutathione peroxidase) were measured in mice with MAFLD. The expression levels of genes associated with the HIPPO pathway were also measured. In vivo experiments using a specific inhibitor of HIPPO signalling were performed to verify the role of this pathway in the effects of PAR. PAR exerted beneficial effects on liver injury, lipid metabolism, fibrosis, inflammation and oxidative stress in mice with MAFLD, which was mediated by activation of the HIPPO pathway.

AKT-induced lncRNA VAL promotes EMT-independent metastasis through diminishing Trim16-dependent Vimentin degradation.

Despite the importance of AKT overactivation in tumor progression, results from clinical trials of various AKT inhibitors remain suboptimal, suggesting that AKT-driven tumor metastasis needs to be further understood. Herein, based on long non-coding RNA (lncRNA) profiling induced by active AKT, we identify that VAL (Vimentin associated lncRNA, LINC01546), which is directly induced by AKT/STAT3 signaling, functions as a potent pro-metastatic molecule and is essential for active AKT-induced tumor invasion, metastasis and anoikis resistance in lung adenocarcinoma (LAD). Impressively, chemosynthetic siRNAs against VAL shows great therapeutic potential in AKT overactivation-driven metastasis. Interestingly, similar to activated AKT in LAD cells, although unable to induce epithelial-mesenchymal transition (EMT), VAL exerts potent pro-invasive and pro-metastatic effects through directly binding to Vimentin and competitively abrogating Trim16-depedent Vimentin polyubiquitination and degradation. Taken together, our study provides an interesting demonstration of a lncRNA-mediated mechanism for active AKT-driven EMT-independent LAD metastasis and indicates the great potential of targeting VAL or Vimentin stability as a therapeutic approach.

Comprehensive Exome Analysis of Immunocompetent Metastatic Head and Neck Cancer Models Reveals Patient Relevant Landscapes.

Immunocompetent metastatic head and neck cancer (HNC) models, although scarce, can help understanding cancer progression and therapy responses in vivo. Their comprehensive genome characterizations are essential for translational research. We first exome-sequenced the two most widely used spontaneous metastatic immunocompetent models, namely AT-84 and SCC VII, followed by comprehensive genomic analyses with three prior-sequenced models (MOC2, MOC2-10, and 4MOSC2), together with patient tumors for utility assessment. AT-84 and SCC VII bear high HNC tumor resemblance regarding mutational signatures-Trp53, Fanconi anemia, and MAPK and PI3K pathway defects. Collectively, the five models harbor genetic aberrations across 10 cancer hallmarks and 14 signaling pathways and machineries (metabolic, epigenetic, immune evasion), to extents similar in patients. Immune defects in HLA-A (H2-Q10, H2-Q4, H2-Q7, and H2-K1), Pdcd1, Tgfb1, Il2ra, Il12a, Cd40, and Tnfrsf14 are identified. Invasion/metastatic genome analyses first highlight potential druggable ERBB4 and KRAS mutations, for advanced/metastatic oral cavity cancer, as well as known metastasis players (Muc5ac, Trem3, Trp53, and Ttn) frequently captured by all models. Notable immunotherapy and precision druggable targets (Pdcd1, Erbb4, Fgfr1, H/Kras, Jak1, and Map2k2) and three druggable hubs (RTK family, MAPK, and DNA repair pathways) are frequently represented by these models. Immunocompetent metastatic HNC models are worth developing to address therapy- and invasion/metastasis-related questions in host immunity contexts.

Selection of a Clinical Lead TCR Targeting Alpha-Fetoprotein-Positive Liver Cancer Based on a Balance of Risk and Benefit.

Hepatocellular carcinoma (HCC) is the most common type of primary liver cancer with a poor prognosis and limited therapeutic options. Alpha-fetoprotein (AFP), an established clinical biomarker of HCC, has been employed as an attractive target for T cell-based immunotherapy against this disease given its high expression in the tumor and restricted expression in normal tissues. We have identified a number of T cell receptors (TCRs) recognizing the HLA-A*02:01 restricted AFP158-166 peptide FMNKFIYEI, providing a TCR candidate pool for identifying TCRs with optimal clinical benefit. To select the ideal AFP TCR for clinical use, we evaluated the efficacy and safety profile of 7 TCRs by testing their potency toward AFP-expressing HCC cells and their specificity based upon reactivity to normal and transformed cells covering a wide variety of primary cell types and HLA serotypes. Furthermore, we assessed their cross-reactivity to potential protein candidates in the human genome by an extensive alanine scan (X-scan). We first selected three TCR candidates based on the in vitro anti-tumor activity. Next we eliminated two potential cross-reactive TCRs based on their reactivity against normal and transformed cells covering a variety of primary cell types and HLA serotypes, respectively. We then excluded the potential cross-reactivity of the selected TCR with a protein candidate identified by X-scan. At present we have selected an AFP TCR with the optimal affinity, function, and safety profile, bearing properties that are expected to allow AFP TCR redirected T cells to specifically differentiate between AFP levels on tumor and normal tissues. An early phase clinical trial using T cells transduced with this TCR to treat HCC patients (NCT03971747) has been initiated.

Preclinical Studies of the Off-Target Reactivity of AFP158-Specific TCR Engineered T Cells.

Autologous T cells engineered with T receptor genes (TCR) are being studied to treat cancers. We have recently identified a panel of mouse TCRs specific for the HLA-A0201/alpha fetoprotein epitope (AFP158) complex and have shown that human T cells engineered with these TCR genes (TCR-Ts) can eradicate hepatocellular carcinoma (HCC) xenografts in NSG mice. However, due to TCR's promiscuity, their off-target cross-reactivity must be studied prior to conducting clinical trials. In this study, we conducted in vitro X-scan assay and in silico analysis to determine the off-target cross-reactivity of 3 AFP158-specific TCR-Ts. We found that the 3 AFP158-specific TCR-Ts could be cross-activated by ENPP1436 peptide and that the TCR3-Ts could also be activated by another off-target peptide, RCL1215. However, compared to AFP158, it requires 250 times more ENPP1436 and 10,000 times more RCL1215 peptides to achieve the same level of activation. The EC50 of ENPP1436 peptide for activating TCR-Ts is approximately 17-33 times higher than AFP158. Importantly, the ENPP1+ tumor cells did not activate TCR1-Ts and TCR2-Ts, and only weakly activated TCR3-Ts. The IFNγ produced by TCR3-Ts after ENPP1+ cell stimulation was >22x lower than that after HepG2 cells. And, all TCR-Ts did not kill ENPP1 + tumor cells. Furthermore, ectopic over-expression of ENPP1 protein in HLA-A2+ tumor cells did not activate TCR-Ts. In silico analysis showed that the ENPP1436 peptide affinity for HLA-A0201 was ranked 40 times lower than AFP158 and the chance of ENPP1436 peptide being processed and presented by HLA-A0201 was 100 times less likely than AFP158. In contrast, the two off-targets (Titin and MAGE-A3) that did cause severe toxicity in previous trials have the same or higher MHC-binding affinity and the same or higher chance of being processed and presented. In conclusion, our data shows that TCR-Ts can be activated by off-target ENPP1436 peptide. But, compared to target AFP158, it requires at least 250 times more ENPP1436 to achieve the same level of activation. Importantly, ENPP1436 peptide in human cells is not processed and presented to a sufficient level to activate the AFP158-specific TCR-Ts. Thus, these TCR-Ts, especially the TCR1-Ts and TCR2-Ts, will unlikely cause significant off-target toxicity.

Engineering T cells for immunotherapy of primary human hepatocellular carcinoma.

Liver cancers, majority of which are primary hepatocellular carcinoma (HCC), continue to be on the rise in the world. Furthermore, due to the lack of effective treatments, liver cancer ranks the 4th most common cause of male cancer deaths. Novel therapies are urgently needed. Over the last few years, immunotherapies, especially the checkpoint blockades and adoptive cell therapies of engineered T cells, have demonstrated a great potential for treating malignant tumors including HCC. In this review, we summarize the current ongoing research of antigen-specific immunotherapies including cancer vaccines and adoptive cell therapies for HCC. We briefly discuss the HCC cancer vaccine and then focus on the antigen-specific T cells genetically engineered with the T cell receptor genes (TCRTs) and the chimeric antigen receptor genes (CARTs). We first review the current options of TCRTs and CARTs immunotherapies for HCC, and then analyze the factors and parameters that may help to improve the design of TCRTs and CARTs to enhance their antitumor efficacy and safety. Our goals are to render readers a panoramic view of the current stand of HCC immunotherapies and provide some strategies to design better TCRTs and CARTs to achieve more effective and durable antitumor effects.

Vaccinia virus-mediated cancer immunotherapy: cancer vaccines and oncolytics.

Cancer vaccines and oncolytic immunotherapy are promising treatment strategies with potential to provide greater clinical benefit to patients with advanced-stage cancer. In particular, recombinant vaccinia viruses (VV) hold great promise as interventional agents. In this article, we first summarize the current understanding of virus biology and viral genes involved in host-virus interactions to further improve the utility of these agents in therapeutic applications. We then discuss recent findings from basic and clinical studies using VV as cancer vaccines and oncolytic immunotherapies. Despite encouraging results gleaned from translational studies in animal models, clinical trials implementing VV vectors alone as cancer vaccines have yielded largely disappointing results. However, the combination of VV vaccines with alternate forms of standard therapies has resulted in superior clinical efficacy. For instance, combination regimens using TG4010 (MVA-MUC1-IL2) with first-line chemotherapy in advanced-stage non-small cell lung cancer or combining PANVAC with docetaxel in the setting of metastatic breast cancer have clearly provided enhanced clinical benefits to patients. Another novel cancer vaccine approach is to stimulate anti-tumor immunity via STING activation in Batf3-dependent dendritic cells (DC) through the use of replication-attenuated VV vectors. Oncolytic VVs have now been engineered for improved safety and superior therapeutic efficacy by arming them with immune-stimulatory genes or pro-apoptotic molecules to facilitate tumor immunogenic cell death, leading to enhanced DC-mediated cross-priming of T cells recognizing tumor antigens, including neoantigens. Encouraging translational and early phase clinical results with Pexa-Vec have matured into an ongoing global phase III trial for patients with hepatocellular carcinoma. Combinatorial approaches, most notably those using immune checkpoint blockade, have produced exciting pre-clinical results and warrant the development of innovative clinical studies. Finally, we discuss major hurdles that remain in the field and offer some perspectives regarding the development of next generation VV vectors for use as cancer therapeutics.

Establishment and evaluation of a model for predicting 3-month mortality in Chinese patients with hepatic encephalopathy.

Hepatic encephalopathy (HE) is a serious complication of liver disease. To establish a model for predicting 3-month mortality in patients with HE in China. This retrospective study included 609 patients with HE admitted to the Peoples' Hospital, Liaocheng City, China (August 2006 to January 2016). Patients were allocated to a modeling (n = 409) or validation (n = 200) group. Demographic/clinical characteristics, laboratory test results, Model for End Stage Liver Disease (MELD) score and Child-Turcotte-Pugh (CTP) score were extracted from medical records. A model for predicting death within 3 months after admission was established using logistic regression analysis (modeling group). Model validity (validation group) was assessed using receiver operating characteristic (ROC) curve analysis. 270/409(66.0%) patients died in the modeling group and 142/203(70.0%) died in the validation group. Compared with survivors, patients who died had more severe HE, and higher MELD score, CTP score, incidence of complications including hepatorenal syndrome (HRS) and upper gastrointestinal bleeding, and values for laboratory parameters including red blood cell count(RBC) and total bilirubin(TBIL)(P < 0.05). Regression analysis revealed RBC, TBIL, HE stage, HRS and upper gastrointestinal bleeding as independent factors associated with death (P < 0.05). The area under the ROC curve (AUC) for the model was 0.931.The model had a higher Youden index than MELD or CTP scores and predicted death in the validation group with a sensitivity of 83.1% and specificity of 93.4%. The established model has superior performance to MELD and CTP scores for predicting mortality in patients with HE.

CK1α suppresses lung tumour growth by stabilizing PTEN and inducing autophagy.

The contribution of autophagy to cancer development remains controversial, largely owing to the fact that autophagy can be tumour suppressive or oncogenic in different biological contexts. Here, we show that in non-small-cell lung cancer (NSCLC), casein kinase 1 alpha 1 (CK1α) suppresses tumour growth by functioning as an autophagy inducer to activate an autophagy-regulating, tumour-suppressive PTEN/AKT/FOXO3a/Atg7 axis. Specifically, CK1α bound the C-terminal tail of PTEN and enhanced both PTEN stability and activity by competitively antagonizing NEDD4-1-induced PTEN polyubiquitination and abrogating PTEN phosphorylation, thereby inhibiting AKT activity and activating FOXO3a-induced transcription of Atg7. Notably, blocking CK1α-induced Atg7-dependent autophagy cooperates with oncogenic HRasV12 to initiate tumorigenesis of lung epithelial cells. An association of a CK1α-modulated autophagic program with the anti-neoplastic activities of the CK1α/PTEN/FOXO3a/Atg7 axis was demonstrated in xenografted tumour models and human NSCLC specimens. This provides insights into the biological and potentially clinical significance of autophagy in NSCLC.

Publisher Correction: Simultaneous overactivation of Wnt/ß-catenin and TGFß signalling by miR-128-3p confers chemoresistance-associated metastasis in NSCLC.

This corrects the article DOI: 10.1038/ncomms15870.

Prognostic analysis of hepatocellular carcinoma on the background of liver cirrhosis via contrast-enhanced ultrasound and pathology.

The aim of this study was to analyze the correlation between the quantitative parameters of contrast-enhancement ultrasound for primary hepatocellular carcinoma (HCC) and biological manifestations of tumor (Ki-67), and to explore the related risk factors of primary hepatocellular carcinoma, so as to provide the theoretical basis for the further study on contrast-enhancement ultrasound manifestations, clinical features and prognosis of HCC. The patients with HCC confirmed by operation or puncture were collected, and those with the background of liver cirrhosis and immunohistochemical staining for tumor sample sections were selected. H&E staining sections of pathological tissues of tumor samples were observed, whether there was any microvessel invasion (MVI) was recorded, the microvessel density (MVD) was counted and the recurrence situations after liver cancer operation was followed up. The change in size of tumor at arterial phase in contrast-enhancement ultrasound, enhancement mode and form at arterial phase, and whether there were tortuous vessels inside or not, and the enhancement intensity, extinction time and extinction intensity at portal phase were observed. The relationship between the parameters of contrast-enhancement ultrasound and Ki-67, AFP, MVD, MVI, tissue differentiation degree of tumor samples and recurrence was analyzed. Under the background of liver cirrhosis, there were significant differences in different enhancement modes and quantification parameters of contrast-enhancement ultrasound for HCC with different expression of Ki-67. Those with obvious tumor enlargement, inhomogeneous enhancement at arterial phase and irregular enhancement form at arterial phase after contrast-enhancement ultrasound had a high incidence of positive Ki-67 and a high early recurrence rate. The inhomogeneous enhancement at arterial phase might predict the proliferative activity and recurrence time of tumor cells; irregular enhancement form at arterial phase might indicate tumor MVI; and the low enhancement of tumor at portal phase may predict a lower degree of tissue differentiation, a higher tumor malignancy and poor prognosis. The incidence of positive Ki-67 under the background of liver cirrhosis is high, indicating poor prognosis. The enhancement mode and parameters of contrast-enhancement ultrasound for HCC may help evaluate the clinical biological manifestations of HCC and predict the postoperative recurrence of HCC.

Identification of α-fetoprotein-specific T-cell receptors for hepatocellular carcinoma immunotherapy.

Hepatocellular carcinoma (HCC) is the major form of liver cancer for which there is no effective therapy. Genetic modification with T-cell receptors (TCRs) specific for HCC-associated antigens, such as α-fetoprotein (AFP), can potentially redirect human T cells to specifically recognize and kill HCC tumor cells to achieve antitumor effects. In this study, using lentivector and peptide immunization, we identified a population of cluster of differentiation 8 (CD8) T cells in human leukocyte antigen (HLA)-A2 transgenic AAD mice that recognized AFP158 epitope on human HCC cells. Adoptive transfer of the AFP158 -specific mouse CD8 T cells eradicated HepG2 tumor xenografts as large as 2 cm in diameter in immunocompromised nonobese diabetic severe combined immunodeficient gamma knockout (NSG) mice. We then established T-cell hybridoma clones from the AFP158 -specific mouse CD8 T cells and identified three sets of paired TCR genes out of five hybridomas. Expression of the murine TCR genes redirected primary human T cells to bind HLA-A2/AFP158 tetramer. TCR gene-engineered human T (TCR-T) cells also specifically recognized HLA-A2+ AFP+ HepG2 HCC tumor cells and produced effector cytokines. Importantly, the TCR-T cells could specifically kill HLA-A2+ AFP+ HepG2 tumor cells without significant toxicity to normal primary hepatocytes in vitro. Adoptive transfer of the AFP-specific TCR-T cells could eradicate HepG2 tumors in NSG mice. CONCLUSION: We have identified AFP-specific murine TCR genes that can redirect human T cells to specifically recognize and kill HCC tumor cells, and those AFP158 -specific TCRs have a great potential to engineer a patient's autologous T cells to treat HCC tumors. (Hepatology 2018).