Immunotherapy of hepatocellular carcinoma: strategies for combinatorial intervention

Hepatocellular carcinoma(HCC) is the most frequent primary liver cancer, leading to 74.6 thousand deaths annually. The prognosis of HCC over the last few decades has remained unsatisfactory, and over half of patients with early-stage HCC develop recurrence by the time of follow-up. Immunotherapeutic intervention has emerged as a novel, effective treatment to delay the progression of aggressive tumors and suppress tumor recurrence and metastasis. However, few clinical immunotherapy trials have been conducted in HCC patients, and there is an unmet need for novel therapeutic strategies. The combination of conventional treatments with specific immunotherapeutic approaches may dramatically improve the efficacy of HCC treatment and the clinical outcome of HCC patients. In this review, we briefly summarize immunotherapy strategies and discuss new advances in combined immunotherapeutic approaches for the treatment of patients with liver cancer.     

Perillyl alcohol as a radio-/chemosensitizer in malignant glioma

The prognosis for patients with malignant glioma has not significantly changed in two decades, despite advances in surgery, radiation, and chemotherapy, emphasizing the growing need for novel approaches to glioma therapy. Perillyl alcohol (POH) is a naturally occurring monoterpene that has been shown to possess chemotherapeutic as well as chemopreventive activity in animal tumor models and is currently in Phase I and Phase II clinical trials. In the present study, we have demonstrated that POH is an effective radiosensitizer at clinically relevant doses of radiation using established glioma cell lines. POH caused a transient arrest in the G2/M phase of the cell cycle and induced apoptosis in glioma cells. POH treatment sensitized glioma cells to Fas-mediated apoptosis, which was further augmented in the presence of ionizing radiation and abrogated in the presence of antagonistic antibody. POH-induced radiosensitization was partially inhibited in glioma cells expressing dominant negative Fas-associated death domain and completely inhibited in glioma cells overexpressing the cytokine response modifier A. In addition, POH treatment resulted in a dose-dependent sensitization to cisplatin and doxorubicin induced cytotoxicity in glioma cells, highlighting its usefulness as a potent radio/chemosensitizer in the treatment of malignant glioma.     

综述与专论: 脊髓星形细胞瘤研究进展

脊髓星形细胞瘤是一种罕见的中枢神经系统恶性肿瘤,在流行病学、肿瘤临床学表型、分子遗传标记、治疗及研究方面有着独特特征。虽然随着手术技术的进步以及分子病理的发展,脑胶质瘤的研究和治疗取得较大进展,但脊髓星形细胞瘤的研究和治疗却发展缓慢。其原因一方面在于临床样本较少,难以开展研究,另一方面因其分子遗传独特性,对脑胶质瘤一线化疗药替莫唑胺敏感性差。因而亟需理清脊髓星形细胞瘤的研究现状,为改善其临床疗效梳理潜在方向。基于此,本文综述脊髓星形细胞瘤的临床特征、病理分型、分子遗传特征和当前治疗方法等方面的研究进展,在描绘脊髓星形细胞瘤的临床治疗现状和研究进展的基础上,提出了未来研究和治疗潜在方向。     

胶质瘤的免疫治疗策略及进展

胶质瘤是最常见的原发性颅内肿瘤之一,具有较高的发病率及死亡率。目前,传统的手术治疗辅助放化疗效果不佳。肿瘤的免疫治疗作为一种新兴的疗法在胶质瘤的治疗中已经取得了一定的疗效。本文将全面的总结归纳胶质瘤的三类免疫治疗方法:主动免疫疗法、被动免疫疗法和免疫调节疗法的内容以及现阶段的进展,并提出胶质瘤的免疫治疗潜在的问题:如接受免疫治疗的患者的免疫状态评估体系不健全、缺乏个性化治疗手段、经济效益以及伦理等问题。同时本文将提出免疫治疗在胶质瘤应用上的展望:1)采用手术治疗、放化疗与免疫治疗相结合的立体式鸡尾酒疗法,在增强各种治疗方法疗效的同时弥补传统治疗方法的不足;2)寻找合适的免疫治疗靶点;3)重点研究结合了过继性细胞免疫治疗与体液免疫治疗优势的治疗方法:嵌合抗原受体修饰的T细胞治疗胶质瘤。     

A novel vaccine containing EphA2 epitope and LIGHT plasmid induces robust cellular immunity against glioma U251 cells

EphA2 is a receptor tyrosine kinase and can be acted as an attractive antigen for glioma vaccines. In addition, LIGHT plays an important role on enhancing T cell proliferation and cytokine production. To improve the CTL mediated immune response against glioma cells, we prepared the novel vaccine containing EphA2_883–891 peptide (TLADFDPRV) and LIGHT plasmid and utilized it to immunize the HLA-A2 transgenic HHD mice. In addition, trimera mice were immunized with the novel vaccine to elicit the antitumor immune response. The results demonstrated that the novel vaccine could induce robust cellular immunity against glioma U251 cells without lysing autologous lymphocytes. Moreover, the novel vaccine could significantly inhibit the tumor growth and prolong the life span of tumor bearing mice. These findings suggested that the novel vaccine containing EphA2 epitope and LIGHT plasmid could induce anti-tumor immunity against U251 cells expressing EphA2, and provided a promising strategy for glioma immunotherapy.     

Targeted nitric oxide delivery preferentially induces glioma cell chemosensitivity via altered p53 and O6‐Methylguanine‐DNA Methyltransferase activity

Glioblastoma multiforme is the most common malignant central nervous system tumor, and also among the most difficult to treat due to a lack of response to chemotherapeutics. New methods of countering the mechanisms that confer chemoresistance to malignant gliomas could lead to significant advances in the quest to identify novel drug combinations or targeted drug delivery systems for cancer therapy. In this study, we investigate the use of a targeted nitric oxide (NO) donor as a pretreatment to sensitize glioma cells to chemotherapy. The protein chlorotoxin (CTX) has been shown to preferentially target glioma cells, and we have developed CTX–NO, a glioma‐specific, NO‐donating CTX derivative. Pretreatment of cells with CTX–NO followed by 48‐h exposure to either carmustine (BCNU) or temozolomide (TMZ), both common chemotherapeutics used in glioma treatment, resulted in increased efficacy of both therapeutics. After CTX–NO exposure, both T98G and U‐87MG human malignant glioma cells show increased sensitivity to BCNU and TMZ. Further investigation revealed that the consequences of this combination therapy was a reduction in active levels of the cytoprotective enzyme MGMT and altered p53 activity, both of which are essential in DNA repair and tumor cell resistance to chemotherapy. The combination of CTX–NO and chemotherapeutics also led to decreased cell invasion. These studies indicate that this targeted NO donor could be an invaluable tool in the development of novel approaches to treat cancer. Biotechnol. Bioeng. 2013; 110: 1211–1220. © 2012 Wiley Periodicals, Inc.     

Dendritic cells pulsed with tumor extract–cationic liposome complex increase the induction of cytotoxic T lymphocytes in mouse brain tumor

Dendritic cells (DCs) are professional antigen-presenting cells (APCs) that locate in peripheral organs. It has been thought that a systemic immune response does not play a role in regression of central nervous system (CNS) tumors, because the CNS is an immunologically privileged site. However, recent advances in immunology have led to the possibility of immunotherapy using peripheral DCs against CNS tumors. Here, we investigated whether DCs pulsed with tumor extract could induce an antitumor effect against malignant glioma. Furthermore, we also investigated whether the antitumor effect become higher by pulsation with tumor extract-liposome complex, compared to pulsation with tumor extract alone. As a liposome, we used cationic small unilamellar vesicles composed of N-(alpha-trimethylammonioacetyl)-didodecyl-D-glutamate chloride (TMAG), dilauroylphosphatidylcholine (DLPC), and dioleoylphosphatidylethanolamine (DOPE) in a molar ratio of 1:2:2. After intracerebral inoculation of mouse malignant glioma GL261 cells into syngeneic C57BL/6 mice, DCs pulsed with extract from the glioma cells by sonication were administered intraperitoneally thrice weekly on days 7, 14 and 21. Tumor growth inhibition was evaluated by measuring the tumor size 1 month after the tumor inoculation. The group treated with DCs pulsed by tumor extract was inhibited in tumor progression compared with the control non-pulsed DCs group, and the group treated with DCs pulsed by tumor extract and liposomes showed substantial tumor volume reductions in all the mice. Among the mice, there were several with no visible masses in their brains. Immunohistochemical study showed that the CD8-positive cytotoxic T cells (CTLs) were strongly recognized among the almost disappearing tumor cells of pulsed DCs groups. The CTLs showed a specific antitumor activity for GL261 mouse glioma cells. These findings indicated that DCs pulsed with tumor extract and liposomes might play an important role in the activation of an immune response in malignant glioma.     

Identification of immune-related hub genes and construction of an immune-related gene prognostic index for low-grade glioma

Low-grade glioma (LGG) poses significant management challenges and has a dismal prognosis. While immunotherapy has shown significant promise in cancer treatment, its progress in glioma has confronted with challenges. In our study, we aimed to develop an immune-related gene prognostic index (IRGPI) which could be used to evaluate the response and efficacy of LGG patients with immunotherapy. We included a total of 529 LGG samples from TCGA database and 1152 normal brain tissue samples from the GTEx database. Immune-related differentially expressed genes (DEGs) were screened. Then, we used weighted gene co-expression network analysis (WGCNA) to identify immune-related hub genes in LGG patients and performed Cox regression analysis to construct an IRGPI. The median IRGPI was used as the cut-off value to categorize LGG patients into IRGPI-high and low subgroups, and the molecular and immune mechanism in IRGPI-defined subgroups were analysed. Finally, we explored the relationship between IRGPI-defined subgroups and immunotherapy related indicators in patients after immunotherapy. Three genes (RHOA, NFKBIA and CCL3) were selected to construct the IRGPI. In a survival analysis using TCGA cohort as a training set, patients in the IRGPI-low subgroup had a better OS than those in IRGPI-high subgroup, consistent with the results in CGGA cohort. The comprehensive results showed that IRGPI-low subgroup had a more abundant activated immune cell population and lower TIDE score, higher MSI, higher TMB score, lower T cell dysfunction score, more likely benefit from ICIs therapy. IRGPI is a promising biomarker in the field of LGG ICIs therapy to distinguish the prognosis, the molecular and immunological characteristics of patients.     

Adenoviral-mediated interleukin-18 expression in mesenchymal stem cells effectively suppresses the growth of glioma in rats

Glioma is the most common primary intracranial malignant tumor. Despite advances in surgical techniques and adjuvant radio- and chemotherapies, the prognosis for patients with glioma remains poor. We have explored the effects of using genetically modified mesenchymal stem cells (MSCs) to treat malignant glioma in rats. Mesenchymal stem cells isolated from Sprague-Dawley rats can directly suppress the growth of C6 cells in vitro. MSCs transplanted intratumorally can also significantly inhibit the growth of glioma and prolong survival in C6 glioma-bearing models. MSCs producing Interleukin-18 infected by adenoviral vector inhibited glioma growth and prolonged the survival of glioma-bearing rats. Transplantation of IL-18 secreting MSCs was associated with enhanced T cell infiltration and long-term anti-tumor immunity. Thus, IL-18 may be an effective adoptive immunotherapy for malignant glioma. When used in conjunction with MSCs as targeting vehicles in vivo, IL-18 may offer a promising new treatment option for malignant glioma.     

Gene therapy for high-grade glioma: Current approaches and future directions

The treatment of high-grade gliomas remains difficult despite recent advances in surgery, radiotherapy and chemotherapy. True advances may emerge from the increasing understanding in molecular biology and discovery of novel mechanisms for the delivery of tumoricidal agents. In an attempt to overcome this formidable neoplasm, molecular approaches using gene therapy have been investigated clinically since 1992. The clinical trials have mainly been classified into three approaches: suicide gene therapy, immune gene therapy and oncolytic viral therapy. In this article, we review these approaches, which have been studied in previous and ongoing clinical trials.Key words: glioma, gene therapy, suicide gene, cytokine gene, oncolytic viruses     

Results of a phase I clinical trial of vaccination of glioma patients with fusions of dendritic and glioma cells

Several reports of clinical trials of immunotherapy using dendritic cells have been published to date. In this study, we investigated the safety and clinical response of immunotherapy with fusions of dendritic and glioma cells for the treatment of patients with malignant glioma. Eight patients with malignant glioma, ranging in age from 4 to 63 years old, participated in this study. Dendritic cells were generated from peripheral blood. Cultured autologous glioma cells were established from surgical specimens in each case. Fusion cells of dendritic and glioma cells were prepared with polyethylene glycol, and the fusion efficiency ranged from 9.2 to 35.3% (mean, 21.9%). All patients received the fusion cells every three weeks for a minimum of 3, and a maximum of 7, immunizations. Fusion cells were injected intradermally, close to a cervical lymph node. The percentage of CD16- and CD56-positive cells in peripheral blood lymphocytes slightly increased after immunization in 4 out of 5 cases investigated. Peripheral blood mononuclear cells were incubated with irradiated autologous glioma or U87MG cells and supernatants were harvested. In 6 cases analyzed, the concentration of interferon-gamma in the supernatant increased after immunization. Clinical results showed that there were no serious adverse effects and two partial responses. Although the results of the phase I clinical trial of fusion cells indicated that this treatment safely induced immune responses. we were unable to establish a statistically significant treatment-associated response rate, due to the limited sample population. Therefore, further evaluation of the role of adjuvant cytokines is necessary.     

Radiotherapy: Brightness and darkness in the era of immunotherapy

The introduction of immunotherapy into cancer treatment has radically changed clinical management of tumors. However, only a minority of patients (approximately 10 to 30%) exhibit long-term response to monotherapy with immunotherapy. Moreover, there are still many cancer types, including pancreatic cancer and glioma, which are resistant to immunotherapy. Due to the immunomodulatory effects of radiotherapy, the combination of radiotherapy and immunotherapy has achieved better therapeutic effects in a number of clinical trials. However, radiotherapy is a double-edged sword in the sense that it also attenuates the immune system under certain doses and fractionation schedules, not all clinical trials show improved survival in the combination of radiotherapy and immunotherapy. Therefore, elucidation of the interactions between radiotherapy and the immune system is warranted to optimize the synergistic effects of radiotherapy and immunotherapy. In this review, we highlight the dark side as well as bright side of radiotherapy on tumor immune microenvironment and immune system. We also elucidate current status of radioimmunotherapy, both in preclinical and clinical studies, and highlight that combination of radiotherapy and immunotherapy attenuates combinatorial effects in some circumstances. Moreover, we provide insights for better combination of radiotherapy and immunotherapy.     

'Clinical trials in Alzheimer's disease': immunotherapy approaches

Recent advances in the understanding of Alzheimer's disease pathogenesis have led to the development of numerous compounds that might modify the disease process. Amyloid β (Aβ) peptide represents an important molecular target for intervention in Alzheimer's disease. Several types of Aβ peptide immunotherapy for Alzheimer's disease are under investigation, direct immunization with synthetic intact Aβ(42) , active immunization involving the administration of synthetic fragments of Aβ peptide conjugated to a carrier protein and passive administration with monoclonal antibodies directed against Aβ peptide. Pre-clinical studies showed that immunization against Aβ peptide can provide protection and reversal of the pathology of Alzheimer's disease in animal models. Indeed, several adverse events have been described like meningoencephalitis with AN1792, vasogenic edema and microhemorrhages with bapineuzumab. Although immunotherapy approaches resulted in clearance of amyloid plaques in patients with Alzheimer's disease, this clearance did not show significant cognitive effect for the moment. Currently, several Aβ peptide immunotherapy approaches are under investigation but also against tau pathology.     

Extracellular vesicles in ovarian cancer: applications to tumor biology,immunotherapy and biomarker discovery

In recent years there has been tremendous interest in both the basic biology and applications of extracellular vesicles (EVs) in translational cancer research. This includes a better understanding of their biogenesis and mechanisms of selective cargo packaging, their precise roles in horizontal communication, and their application as non-invasive biomarkers. The rapid advances in next-generation omics technologies are the driving forces for these discoveries. In this review, the authors focus on recent results of EV research in ovarian cancer. A deeper understanding of ovarian cancer-derived EVs, the types of cargo molecules and their biological roles in cancer growth, metastases and drug resistance, could have significant impact on the discovery of novel biomarkers and innovative therapeutics. Insights into the role of EVs in immune regulation could lead to novel approaches built on EV-based immunotherapy.     

Molecular and clinical characterization of Galectin-9 in glioma through 1,027 samples

In recent years, research on glioma immunotherapy have grown rapidly. However, the autoimmune-like side effects that are caused by blocking immunological checkpoints hinder their clinical application in gliomas currently. Galectin-9, a ligand for T-cell immunoglobulin mucin 3, has shed a new light on the treatment of malignant glioma. However, the potential mechanism of Galectin-9 is still under discussion. In this study, first, we methodically gathered 1,027 glioma patients with RNA-seq and 986 patients with survival data to explore the role and mechanism of Galectin-9 in gliomas. Second, we analyzed glioma samples from 50 patients in the Department of Neurosurgery, Tianjin Medical University General Hospital. Finally, we found that Galectin-9 was strongly upregulated in glioblastoma multiforme compared with normal brain tissues and lower-grade glioma. Patients with Galectin-9 overexpression had a significantly shorter overall survival. Moreover, the tissue microarray data displayed that the expression of Galectin-9 in the core of tumor is higher than that in the border and was correlated with the shorter survival in glioma patients. Galectin-9 is more highly expressed in the mesenchymal subtype of glioblastoma multiforme than in the other subtypes. Simultaneously, Galectin-9 was closely associated with the immune response and lymphocyte activation, especially T-cell activation. To further determine the underlying role of Galectin-9 in the immune response, we selected seven immune metagenes. Through cluster analysis and correlation analysis, we discovered that Galectin-9 was highly correlated with immune checkpoint molecules and M2 tumor-associated macrophages. In summary, Galectin-9 serves as a potential therapeutic target to treat glioblastoma multiforme.     

The combination of DNA methyltransferase inhibitor therapy and immunotherapy for acute myeloid leukemia

Acute myeloid leukemia (AML), the most common form of acute leukemia in adults, is characterized by abnormal proliferation and blocked maturation and differentiation of myeloid precursor cells. AML is an aggressive cancer that progresses rapidly without treatment. Therefore, effective treatment modalities should be implemented immediately after diagnosis. The mainstay of classical AML therapy has been chemotherapy, which is not suitable for relapsing or refractory patients, especially elderly patients. Among emerging novel therapeutic approaches for AML, epigenetic therapy and immunotherapy represent two exciting therapeutic developments. This review focuses on discussion of the therapeutic considerations for AML from the perspective of combination treatment, which incorporates both DNA methyltransferase inhibitor therapy, as one of the most promising epigenetic therapies, and immune checkpoint inhibitor or dendritic cell-based vaccination treatments, as examples of immunotherapy. Both challenges and rationale in the optimization of therapeutic approaches, as well as recent clinical trial developments, along this line are summarized.     

Comparative analysis of peripheral and localised cytokine secretion in glioblastoma patients

BACKGROUND: Malignant gliomas are the most common primary brain tumours of both children and adults. The unique aspects of their biology and anatomic site render them refractory to conventional therapeutic strategies such as surgery and chemotherapy. Significant attention has been given, recently, to immunotherapy which, although promising in preclinical studies, has not yet enhanced the survival of patients with glioblastomas. METHODS: To further understand the immunobiology of glioblastomas in clinical settings, we examined the secretion of four main cytokines in the peripheral blood and in primary cell cultures of 33 human glioblastoma patients. An ELISPOT methodology was used for the first time to examine Th1, and Th2 cytokine secretion from both peripheral lymphocytes and glioma tumour cells. RESULTS: Th1 cytokines (tumour necrosis factor (TNF-alpha), interferon (IFN-gamma) were markedly reduced compared to control levels (P=0.01 and P<0.001, respectively), whereas in contrast, Th2 (interleukin (IL)-4 and IL-10) were strongly expressed in both peripheral lymphocytes and glioma cell cultures (P=0.05 and P<0.001, respectively). CONCLUSION: This pattern indicates an 'immunosuppressive status' in glioblastomas which is related to their origination and the evasion of glioma cells from immune surveillance and could account for the failure of immunotherapy in such tumours. Furthermore, ELISPOT methodology can be used for monitoring of cytokine secretion from tumour cells, in addition to the well-established peripheral cytokine secretion.     

A novel recombinant protein of IP10-EGFRvIIIscFv and CD8+ cytotoxic T lymphocytes synergistically inhibits the growth of implanted glioma in mice

The epidermal growth factor receptor (EGFR) mutant of EGFRvIII is highly expressed on glioma cells and has been thought to be an excellent target molecule for immunotherapy. IP-10 is a potent chemokine and can recruit CXCR3⁺ T cells, including CD8⁺ T cells that are important for the control of tumor growth. This study is aimed at investigating the therapeutic efficacy of a novel fusion protein of IP10-EGFRvIIIscFv (IP10-scFv) in combination with glioma lysate-pulsed DCs-activated CD8⁺ cytotoxic T lymphocytes (CTLs) in a mouse model of glioma. A plasmid of pET-IP10-scFv was generated by linking mouse IP-10 gene with the DNA fragment for anti-EGFRvIIIscFv, a (Gly₄Ser)₃ flexible linker and a His-tag. The recombinant IP10-scFv in E. coli was purified by affinity chromatography and characterized for its anti-EGFRvIII immunoreactivity and chemotactic activity. C57BL/6 mice were inoculated with mouse glioma GL261 cells in the brain and treated intracranially with IP10-scFv and/or intravenously with CTL for evaluating the therapeutic effect. The glioma-specific immune responses were examined. The IP10-scFv retained anti-EGFRvIII immunoreactivity and IP-10-like chemotactic activity. Treatment with both IP10-scFv and CTL synergistically inhibited the growth of glioma and prolonged the survival of tumor-bearing mice, accompanied by increasing the numbers of brain-infiltrating lymphocytes (BILs) and the frequency of CXCR3⁺CD8⁺ T cells, enhancing glioma-specific IFN-γ responses and cytotoxicity, and promoting glioma cell apoptosis in mice. Our novel data indicate that IP10-scFv and CTL have synergistic therapeutic effects on inhibiting the growth of mouse glioma in vivo.