Immune evasion is a pivotal event in tumor progression. To eliminate human cancer cells, current immune checkpoint therapy is set to boost the CD8+ T cell-mediated cytotoxicity. However, this action is eventually dependent on the efficient recognition of tumor-specific antigens via T cell receptors. One primary mechanism by which tumor cells evade immune surveillance is to downregulate their antigen presentation. Little progress has been made towards harnessing potential therapeutic targets for enhancing antigen presentation on the tumor cell. Here, we identified MAL2 as a key player that determines the turnover of the antigen-loaded MHC-I complex and reduces the antigen presentation on tumor cells. MAL2 promotes the endocytosis of tumor antigens via direct interaction with the MHC-I complex and endosome-associated RAB proteins. In preclinical models, depletion of MAL2 in breast tumor cells profoundly enhanced the cytotoxicity of tumor-infiltrating CD8+ T cells and suppressed breast tumor growth, suggesting that MAL2 is a potential therapeutic target for breast cancer immunotherapy.
Yuanzhang Fang, Lifei Wang, Changlin Wan, Yifan Sun, Kevin Van der Jeught, Zhuolong Zhou, Tianhan Dong, Ka Man So, Tao Yu, Yujing Li, Haniyeh Eyvani, Austyn Colter, Edward Dong, Sha Cao, Jin Wang, Bryan P. Schneider, George Sandusky, Yunlong Liu, Chi Zhang, Xiongbin Lu, Xinna Zhang
Diffuse intrinsic pontine glioma (DIPG) kills more children than any other type of brain tumor. Despite clinical trials testing many chemotherapeutic agents, palliative radiotherapy remains the standard treatment. Here, we utilized Cre/loxP technology to show that deleting Ataxia telangiectasia mutated (Atm) in primary mouse models of DIPG can enhance tumor radiosensitivity. Genetic deletion of Atm improved survival of mice with p53 deficient but not p53 wild-type gliomas following radiotherapy. Similar to patients with DIPG, mice with p53 wild-type tumors had improved survival after radiotherapy independent of Atm deletion. Primary p53 wild-type tumor cell lines induced proapoptotic genes after radiation and repressed the NRF2 target, NAD(P)H quinone dehydrogenase 1 (Nqo1). Tumors lacking p53 and Ink4a/Arf expressed the highest level of Nqo1 and were most resistant to radiation, but deletion of Atm enhanced the radiation response. These results suggest that tumor genotype may determine whether inhibition of ATM during radiotherapy will be an effective clinical approach to treat DIPGs.
Katherine Deland, Bryce F. Starr, Joshua S. Mercer, Jovita Byemerwa, Donna M. Crabtree, Nerissa T. Williams, Lixia Luo, Yan Ma, Mark Chen, Oren J. Becher, David G Kirsch
Background. Clear cell renal cell carcinoma (ccRCC) is the most common histologically defined renal cancer. However, it is not a uniform disease and includes several genetic subtypes with different prognosis. ccRCC is also characterized by distinguished metabolic reprogramming. Tobacco smoking (TS) is an established risk factor for ccRCC with unknown effects on tumor pathobiology. Methods. We investigated the landscape of ccRCCs and paired normal kidney tissues (NKTs) using integrated transcriptomic, metabolomic and metallomic approaches in a cohort of never smokers (NS) and long-term current smokers (LTS) Caucasian males. Results. All three Omics domains consistentl identified a distinct metabolic subtype of ccRCCs in LTS, characterized by activation of oxidative phosphorylation (OxPhos) coupled with reprogramming of the malate-aspartate shuttle and metabolism of aspartate, glutamate, glutamine and histidine. Cadmium, copper and inorganic arsenic accumulated in LTS tumors showing redistribution among intracellular pools, including relocation of copper into the cytochrome c oxidase complex. Gene expression signature based on the LTS metabolic subtype provided prognostic stratification of The Cancer Genome Atlas (TCGA) ccRCC tumors that was independent from genomic alterations. Conclusions. The work identifies the TS related metabolic subtype of ccRCC with vulnerabilities that can be exploited for precision medicine approaches targeting metabolic pathways. The results provide rationale for the development of metabolic biomarkers with diagnostic and prognostic applications using evaluation of OxPhos status. The metallomic analysis reveals the role of disrupted metal homeostasis in ccRCC highlighting the importance of studying effects of metals from e-cigarettes and environmental exposures.
James Reigle, Dina Secic, Jacek Biesiada, Collin Wetzel, Behrouz Shamsaei, Johnson Chu, Yuanwei Zang, Xiang Zhang, Nicholas J. Talbot, Megan E. Bischoff, Yongzhen Zhang, Charuhas V. Thakar, Krishnanath Gaitonde, Abhinav Sidana, Hai Bui, John T. Cunningham, Qing Zhang, Laura S. Schmidt, W. Marston Linehan, Mario Medvedovic, David R. Plas, Julio A. Landero Figueroa, Jarek Meller, Maria F. Czyzyk-Krzeska
Estrogen receptor (ER)-negative breast cancer is thought to be more malignant and devastating than ER-positive breast cancer and exhibit elevated NF-κB activity. How abnormally high NF-κB activity is maintained in ER-negative breast cancer is poorly understood. The importance of linear ubiquitination, which is generated by the linear ubiquitin chain assembly complex (LUBAC), is increasingly appreciated in NF-κB signaling, which regulates cell activation and death. Here, we showed that epsin proteins, a family of ubiquitin-binding endocytic adaptors, interacted with LUBAC via its Ubiquitin-Interacting Motif (UIM) and bound LUBAC’s bona fide substrate NEMO via its N-terminal homolog (ENTH) domain. Furthermore, epsins promoted NF-κB essential modulator (NEMO) linear ubiquitination and served as scaffolds for recruiting other components of the IκB kinase (IKK) complex; thereby, resulting in the heightened IKK activation and sustained NF-κB signaling essential for the development of ER-negative breast cancer. Heightened epsin levels in ER-negative human breast cancer are associated with poor, relapse-free survival. We showed that transgenic and pharmacological approaches eliminating epsins potently impeded breast cancer development in both spontaneous and patient-derived xenograft breast cancer mouse models. Our findings established the pivotal role epsins played in promoting breast cancer. Thus, targeting epsins may represent a strategy to restrain NF-κB signaling, and provide an important perspective into ER-negative breast cancer treatment.
Kai Song, Xiaofeng Cai, Yunzhou Dong, Hao Wu, Yong Wei, Uma Shankavaram, Kui Cui, Yang Lee, Bo Zhu, Sudarshan Bhattacharjee, Beibei Wang, Kun Zhang, Aiyun Wen, Scott Wong, Lili Yu, Lijun Xia, Alana L Welm, Diane R. Bielenberg, Kevin Camphausen, Yibin Kang, Hong Chen
Therapeutic strategies designed to target TP53-deficient cancer cells remain elusive. Here, we showed that TP53 loss initiated a pharmacologically actionable secretory process that drove lung adenocarcinoma (LUAD) progression. Molecular, biochemical, and cell biological studies showed that TP53 loss increased the expression of Golgi reassembly and stacking protein 55 kD (G55), a Golgi stacking protein that maintains Golgi organelle integrity and is part of a GOLGIN45/myosin IIA-containing protein complex that activates secretory vesicle biogenesis in the Golgi. TP53 loss activated G55-dependent secretion by relieving G55 and myosin IIA from miR-34a-dependent silencing. G55-dependent secreted proteins enhanced the proliferative and invasive activities of TP53-deficient LUAD cells and promoted angiogenesis and CD8+ T cell exhaustion in the tumor microenvironment. A small molecule that blocks G55/G45 interactions impaired secretion and reduced TP53-deficient LUAD growth and metastasis. These results identified a targetable secretory vulnerability in TP53-deficient LUAD cells.
Xiaochao Tan, Lei Shi, Priyam Banerjee, Xin Liu, Hou-Fu Guo, Jiang Yu, Neus Bota-Rabassedas, B. Leticia Rodriguez, Don L. Gibbons, William K. Russell, Chad J. Creighton, Jonathan M. Kurie
Proteins created from recurrent fusion genes like CBFB-MYH11 are prevalent in acute myeloid leukemia (AML), often necessary for leukemogenesis, persistent throughout the disease course, and highly leukemia specific, making them attractive neoantigen targets for immunotherapy. A nonameric peptide derived from a prevalent CBFB-MYH11 fusion protein was found to be immunogenic in HLA-B*40:01+ donors. High-avidity CD8+ T cell clones isolated from healthy donors killed CBFB-MYH11+ HLA-B*40:01+ AML cell lines and primary human AML samples in vitro. CBFB-MYH11–specific T cells also controlled CBFB-MYH11+ HLA-B*40:01+ AML in vivo in a patient-derived murine xenograft model. High-avidity CBFB-MYH11 epitope–specific T cell receptors (TCRs) transduced into CD8+ T cells conferred antileukemic activity in vitro. Our data indicate that the CBFB-MYH11 fusion neoantigen is naturally presented on AML blasts and enables T cell recognition and killing of AML. We provide proof of principle for immunologically targeting AML-initiating fusions and demonstrate that targeting neoantigens has clinical relevance even in low–mutational frequency cancers like fusion-driven AML. This work also represents a first critical step toward the development of TCR T cell immunotherapy targeting fusion gene–driven AML.
Melinda A. Biernacki, Kimberly A. Foster, Kyle B. Woodward, Michael E. Coon, Carrie Cummings, Tanya M. Cunningham, Robson G. Dossa, Michelle Brault, Jamie Stokke, Tayla M. Olsen, Kelda Gardner, Elihu Estey, Soheil Meshinchi, Anthony Rongvaux, Marie Bleakley
Background: Patients with diffuse midline gliomas (DMG), including diffuse intrinsic pontine glioma (DIPG), have dismal outcomes. We previously described the H3.3K27M mutation as a shared neoantigen in HLA-A*02.01+ H3.3K27M+ DMGs. Within the Pacific Pediatric Neuro-Oncology Consortium, we assessed safety and efficacy of an H3.3K27M-targeted peptide vaccine. Patients and Methods: Newly diagnosed patients aged 3-21 years with HLA-A*02.01+ and H3.3K27M+ status were enrolled into Stratum A (DIPG) and Stratum B (non-pontine DMG). Vaccine was administered in combination with poly-ICLC every three weeks for eight cycles, followed by once every six weeks. Immuno-monitoring and imaging occurred every three months. Imaging was centrally reviewed. Immunological responses were assessed in peripheral blood mononuclear cells using mass cytometry. Results: 19 patients enrolled in Stratum A (median age=11 years) and 10 in Stratum B (median age=13 years). There were no grade 4 treatment-related adverse events (TRAE). Injection site reaction was the most commonly reported TRAE. Overall survival (OS) at 12 months was 40% (95% CI, 22% to 73%) for Stratum A and 39% (95% CI, 16% to 93%) for Stratum B. The median OS was 16.1 months in patients exhibiting an expansion of H3.3K27M-reactive CD8+ T-cells compared to 9.8 months for their counterparts (p=0.05). DIPG patients with below-median baseline levels of myeloid-derived suppressor cells had prolonged OS compared to their counterparts (p<0.01). Immediate pre-treatment dexamethasone administration inversely associated with H3.3K27M-reactive CD8+ T-cell responses. Conclusion: Administration of the H3.3K27M-specific vaccine is well tolerated. Patients with H3.3K27M-specific CD8+ immunological responses demonstrated prolonged OS compared to non-responders.
Sabine Mueller, Jared M. Taitt, Javier E. Villanueva-Meyer, Erin R. Bonner, Takahide Nejo, Rishi R Lulla, Stewart Goldman, Anu Banerjee, Susan N. Chi, Nicholas S. Whipple, John R. Crawford, Karen Gauvain, Kellie J. Nazemi, Payal B. Watchmaker, Neil D. Almeida, Kaori Okada, Andres M. Salazar, Ryan D. Gilbert, Javad Nazarian, Annette M. Molinaro, Lisa H. Butterfield, Michael D. Prados, Hideho Okada
BACKGROUND. Current methods for the detection and surveillance of bladder cancer (BCa) are often invasive and/or possess suboptimal sensitivity and specificity, especially in early stage, minimal, residual tumors. METHODS. We developed a novel method for the detection of urine tumor DNA Methylation at multiple genomic regions by Mass Array, termed utMeMA. We identified the BCa-specific methylation markers by combined analyses of Sun Yat-sen Memorial Hospital (SYSMH), TCGA and GEO cohorts. The BCa diagnostic model was built in a retrospective cohort (n=313) and validated in a multicenter, prospective cohort (n=175). The performance of this diagnostic assay was analyzed and compared with urine cytology and FISH. RESULTS. We first discovered 26 significant methylation markers of BCa in combined analyses. We build and validate a two-marker-based diagnostic model that discriminated patients with BCa with high accuracy (86.7%), sensitivity (90.0%) and specificity (83.1%). Furthermore, utMeMA based assay achieved a great improvement in sensitivity over urine cytology and FISH, especially in the detection of early stage (Ta and low grade tumor, 64.5% vs. 11.8%, 15.8%), minimal (81.0% vs. 14.8%, 37.9%), residual (93.3% vs. 27.3%, 64.3%) and recurrent (89.5% vs. 31.4%, 52.8%) tumors. The urine diagnostic score (UD-score) from this assay was better associated with tumor malignancy and burden. CONCLUSIONS. Urine tumor DNA methylation assessment for early diagnosis, minimal, residual tumor detection and surveillance in bladder cancer is a rapid, high-throughput, non-invasive and promising approach, which may reduce the burden of cystoscopy and blind second surgery.
Xu Chen, Jingtong Zhang, Weimei Ruan, Ming Huang, Chanjuan Wang, Hong Wang, Zeyu Jiang, Shaogang Wang, Zheng Liu, Chunxiao Liu, Wanlong Tan, Jin Yang, Jiaxin Chen, Zhiwei Chen, Xia Li, Xiaoyu Zhang, Peng Xu, Lin Chen, Ruihui Xie, Qianghua Zhou, Shizhong Xu, Darryl Irwin, JIAN-BING FAN, Jian Huang, Tianxin Lin
The transcription factor, Mef2d, is important in the regulation of differentiation and adaptive responses in many cell types. Among T cells, Mef2d gains new functions in Foxp3+ T-regulatory (Treg) cells as a result of its interactions with the transcription factor, Foxp3, and its release from canonical partners, like histone/protein deacetylases. Though not necessary for the generation and maintenance of Tregs, Mef2d is required for the expression of IL-10, Ctla-4 and Icos, and for the acquisition of an effector Treg phenotype. At these loci, Mef2d acts both synergistically and additively to Foxp3, and down-stream of Blimp1. Mice with the conditional deletion in Tregs of the gene encoding Mef2d are unable to maintain long-term allograft survival despite costimulation blockade and have enhanced antitumor immunity in syngeneic models, but they display only minor evidence of autoimmunity when maintained under normal conditions. The role played by Mef2d in sustaining effector Foxp3+ Treg functions without abrogating their basal actions suggests its suitability for drug discovery efforts in cancer therapy.
Eros Di Giorgio, Liqing Wang, Yan Xiong, Tatiana Akimova, Lanette M. Christensen, Rongxiang Han, Arabinda Samanta, Matteo Trevisanut, Tricia R. Bhatti, Ulf H. Beier, Wayne W. Hancock
Background: The recent failure of checkpoint-blockade therapies for glioblastoma multiforme (GBM) in late-phase clinical trials has directed interest towards adoptive cellular immunotherapies (ACT). In this open-label, first-in-human trial, we have assessed the safety and therapeutic potential of cytomegalovirus (CMV)-specific ACT in an adjuvant setting for patients with primary GBM, with an ultimate goal to prevent or delay recurrence and prolong overall survival. Methods: Twenty-eight patients with primary GBM were recruited to this prospective study, 25 of whom were treated with in vitro-expanded autologous CMV-specific T cells. Participants were monitored for safety, progression-free survival (PFS), overall survival (OS) and immune reconstitution. Results: No participants showed evidence of ACT-related toxicities. Of 25 evaluable participants, ten were alive at the completion of follow-up, while five were disease free. Reconstitution of CMV-specific T-cell immunity was evident and CMV-specific ACT may trigger bystander effect leading to additional T-cell responses to non-viral tumour-associated antigens through epitope spreading. Long-term follow-up of participants treated before recurrence showed significantly improved OS when compared to those who progressed before ACT (median 23 months, range 7–65 vs. median 14 months, range 5–19; p=0.018). Gene expression analysis of the ACT products indicated that a favourable T-cell gene signature was associated with improved long-term survival. Conclusion: Data presented in this study demonstrate that CMV-specific ACT can be safely used as an adjuvant therapy for primary GBM and, if offered before recurrence, this therapy may improve overall survival of GBM patients.Trial registration: anzctr.org.au: ACTRN12615000656538Funding Source:National Health & Medical Research Council (Australia) Trial registration: anzctr.org.au: ACTRN12615000656538 Funding Source: Philanthropic funding &National Health & Medical Research Council (Australia)
Corey Smith, Katie E. Lineburg, J. Paulo Martins, George Ambalathingal, Michelle A. Neller, Beth Morrison, Katherine K. Matthews, Sweera Rehan, Pauline Crooks, Archana Panikkar, Leone Beagley, Laetitia Le Texier, Sriganesh Srihari, David Walker, Rajiv Khanna