Oncolytic virotherapy has been proposed as an ablative and immunostimulatory treatment strategy for solid tumors that are resistant to immunotherapy alone; however, there is a need to optimize host immune activation using preclinical immunocompetent models in previously untested common adult tumors. We studied a modified oncolytic myxoma virus (MYXV) that shows high efficiency for tumor-specific cytotoxicity in small-cell lung cancer (SCLC), a neuroendocrine carcinoma with high mortality and modest response rates to immune checkpoint inhibitors. Using an immunocompetent SCLC mouse model, we demonstrated the safety of intrapulmonary MYXV delivery with efficient tumor-specific viral replication and cytotoxicity associated with induction of immune cell infiltration. We observed increased SCLC survival following intrapulmonary MYXV that was enhanced by combined low-dose cisplatin. We also tested intratumoral MYXV delivery and observed immune cell infiltration associated with tumor necrosis and growth inhibition in syngeneic murine allograft tumors. Freshly collected primary human SCLC tumor cells were permissive to MYXV and intratumoral delivery into patient-derived xenografts resulted in extensive tumor necrosis. We confirmed MYXV cytotoxicity in classic and variant SCLC subtypes as well as cisplatin-resistant cells. Data from 26 SCLC human patients showed negligible immune cell infiltration, supporting testing MYXV as an ablative and immune-enhancing therapy.
Patrick Kellish, Daniil Shabashvili, Masmudur M. Rahman, Akbar Nawab, Maria V. Guijarro, Min Zhang, Chunxia Cao, Nissin Moussatche, Theresa Boyle, Scott Antonia, Mary Reinhard, Connor Hartzell, Michael Jantz, Hiren J. Mehta, Grant McFadden, Frederic J. Kaye, Maria Zajac-Kaye
The mesenchymal (MES) subtype of glioblastoma (GBM) stem cells (GSCs) represents a subpopulation of cancer cells that are notorious for their highly aggressive nature and resistance to conventional therapy. Aldehyde dehydrogenase 1A3 (ALDH1A3) has been recently suggested as a key determinant for the maintenance of MES features of GSCs. However, the mechanisms underpinning aberrant ALDH1A3 expression remain elusive. Here, we identified ubiquitin-specific protease 9X (USP9X) as a bona fide deubiquitinase of ALDH1A3 in MES GSCs. USP9X interacted with, depolyubiquitylated, and stabilized ALDH1A3. Moreover, we showed that FACS-sorted USP9Xhi cells were enriched for MES GSCs with high ALDH1A3 activity and potent tumorigenic capacity. Depletion of USP9X markedly downregulated ALDH1A3, resulting in a loss of self-renewal and tumorigenic capacity of MES GSCs, which could be largely rescued by ectopic expression of ALDH1A3. Furthermore, we demonstrated that the USP9X inhibitor WP1130 induced ALDH1A3 degradation and showed marked therapeutic efficacy in MES GSC–derived orthotopic xenograft models. Additionally, USP9X strongly correlated with ALDH1A3 expression in primary human GBM samples and had a prognostic value for patients with the MES subgroup. Collectively, our findings unveil USP9X as a key deubiquitinase for ALDH1A3 protein stabilization and a potential target for GSC-directed therapy.
Zhengxin Chen, Hong-Wei Wang, Shuai Wang, Ligang Fan, Shuang Feng, Xiaomin Cai, Chenghao Peng, Xiaoting Wu, Jiacheng Lu, Dan Chen, Yuanyuan Chen, Wenting Wu, Daru Lu, Ning Liu, Yongping You, Huibo Wang
Intrinsically disordered proteins (IDPs) are emerging as attractive drug targets by virtue of their prevalence in various diseases including cancer. Drug development targeting IDPs is challenging because they have dynamical structure features and conventional drug design is not applicable. NUPR1 is an IDP playing an important role in pancreatic cancer. We previously reported that Trifluoperazine (TFP), an antipsychotic agent, was capable of binding to NUPR1 and inhibiting tumors growth. Unfortunately, TFP showed strong central nervous system side-effects. In this work, we undertook a multidisciplinary approach to optimize TFP, based on the synergy of computer modeling, chemical synthesis, and a variety of biophysical, biochemical and biological evaluations. A family of TFP-derived compounds was produced and the most active one, named ZZW-115, showed a dose-dependent tumor regression with no neurological effects and induced cell death mainly by necroptosis. This study opens a new perspective for drug development against IDPs, demonstrating the possibility of successful ligand-based drug design for such challenging targets.
Patricia Santofimia-Castaño, Yi Xia, Wenjun Lan, Zhengwei Zhou, Can Huang, Ling Peng, Philippe Soubeyran, Adrian Velazquez-Campoy, Olga Abian, Bruno Rizzuti, Jose L. Neira, Juan Iovanna
BACKGROUND. African American (AA) patients have higher cancer mortality rates and shorter survival times compared to European American (EA) patients. Despite a significant focus on socioeconomic factors, recent findings strongly argue the existence of biological factors driving this disparity. Most of these factors have been described in a cancer-type specific context rather than a pan-cancer setting. METHODS. A novel in silico approach based on Gene Set Enrichment Analysis (GSEA) coupled to Transcription Factor enrichment was carried out to identify common biological drivers of pan-cancer racial disparity using The Cancer Genome Atlas (TCGA) dataset. Mitochondrial content in patient tissues was examined using a multi-cancer tissue microarray approach (TMA). RESULTS. Mitochondrial oxidative phosphorylation was uniquely enriched in AA tumors compared to EA tumors across various cancer types. AA tumors also showed strong enrichment for the ERR1-PGC1α-mediated transcriptional program, which has been implicated in mitochondrial biogenesis. TMA analysis revealed that AA cancers harbor significantly more mitochondria compared to their EA counterparts. CONCLUSIONS. These findings highlight changes in mitochondria as a common distinguishing feature between AA and EA tumors in a pan-cancer setting, and provide the rationale for the repurposing of mitochondrial inhibitors to treat AA cancers.
Danthasinghe Waduge Badrajee Piyarathna, Akhila Balasubramanian, James M. Arnold, Stacy M. Lloyd, Balasubramanyam Karanam, Patricia Castro, Michael M. Ittmann, Nagireddy Putluri, Nora Navone, Jeffrey A. Jones, Wendong Yu, Vlad C. Sandulache, Andrew G. Sikora, George Michailidis, Arun Sreekumar
Rationale Tumor infiltrating lymphocytes are widely associated with positive outcomes, yet carry key indicators of a systemic failed immune response against unresolved cancer. Cancer immunotherapies can reverse their tolerance phenotypes, while preserving tumor-reactivity and neoantigen-specificity shared with circulating immune cells. Objectives We performed comprehensive transcriptomic analyses to identify gene signatures common to circulating and tumor infiltrating lymphocytes in the context of clear cell renal cell carcinoma. Modulated genes also associated with disease outcome were validated in other cancer types. Findings Using bioinformatics, we identified practical diagnostic markers and actionable targets of the failed immune response. On circulating lymphocytes, three genes, LEF1, FASLG, and MMP9, could efficiently stratify patients from healthy control donors. From their associations with resistance to cancer immunotherapies and microbial infections, we uncovered not only pan-cancer, but pan-pathology failed immune response profiles. A prominent lymphocytic matrix metallopeptidase cell migration pathway, is central to a panoply of diseases and tumor immunogenicity, correlates with multi-cancer recurrence, and identifies a feasible, non-invasive approach to pan-pathology diagnoses. Conclusions The non-invasive differently expressed genes we have identified warrant future investigation towards the development of their potential in precision diagnostics and precision pan-disease immunotherapeutics.
Anne Monette, Antigoni Morou, Nadia A. Al-Banna, Louise Rousseau, Jean-Baptiste Lattouf, Sara Rahmati, Tomas Tokar, Jean-Pierre Routy, Jean-Francois Cailhier, Daniel E. Kaufmann, Igor Jurisica, Rejean Lapointe
Because metastasis is associated with the majority of cancer-related deaths, its prevention is a clinical aspiration. Prostanoids are a large family of bioactive lipids derived from the activity of cyclooxygenase-1 (COX-1) and COX-2. Aspirin impairs the biosynthesis of all prostanoids through the irreversible inhibition of both COX isoforms. Long-term administration of aspirin leads to reduced distant metastases in murine models and clinical trials, but the COX isoform, downstream prostanoid, and cell compartment responsible for this effect are yet to be determined. Here, we have shown that aspirin dramatically reduced lung metastasis through inhibition of COX-1 while the cancer cells remained intravascular and that inhibition of platelet COX-1 alone was sufficient to impair metastasis. Thromboxane A2 (TXA2) was the prostanoid product of COX-1 responsible for this antimetastatic effect. Inhibition of the COX-1/TXA2 pathway in platelets decreased aggregation of platelets on tumor cells, endothelial activation, tumor cell adhesion to the endothelium, and recruitment of metastasis-promoting monocytes/macrophages, and diminished the formation of a premetastatic niche. Thus, platelet-derived TXA2 orchestrates the generation of a favorable intravascular metastatic niche that promotes tumor cell seeding and identifies COX-1/TXA2 signaling as a target for the prevention of metastasis.
Serena Lucotti, Camilla Cerutti, Magali Soyer, Ana M. Gil-Bernabé, Ana L. Gomes, Philip D. Allen, Sean Smart, Bostjan Markelc, Karla Watson, Paul C. Armstrong, Jane A. Mitchell, Timothy D. Warner, Anne J. Ridley, Ruth J. Muschel
Background: Chimeric antigen receptor (CAR) T cells are a promising therapy for hematologic malignancies. B-cell maturation antigen (BCMA) is a rational target in multiple myeloma (MM). Methods: We conducted a phase I study of autologous T cells lentivirally-transduced with a fully-human, BCMA-specific CAR containing CD3ζ and 4-1BB signaling domains (CART-BCMA), in subjects with relapsed/refractory MM. Twenty-five subjects were treated in 3 cohorts: 1) 1-5 x 108 CART-BCMA cells alone; 2) Cyclophosphamide (Cy) 1.5 g/m2 + 1-5 x 107 CART-BCMA cells; and 3) Cy 1.5 g/m2 + 1-5 x 108 CART-BCMA cells. No pre-specified BCMA expression level was required. Results: CART-BCMA cells were manufactured and expanded in all subjects. Toxicities included cytokine release syndrome and neurotoxicity, which were grade 3-4 in 8 (32%) and 3 (12%) subjects, respectively, and reversible. One subject died at day 24 from candidemia and progressive myeloma, following treatment for severe CRS and encephalopathy. Responses (based on treated subjects) were seen in 4/9 (44%) in cohort 1, 1/5 (20%) in cohort 2, and 7/11 (64%) in cohort 3, including 5 partial, 5 very good partial, and 2 complete responses, 3 of which were ongoing at 11, 14, and 32 months. Decreased BCMA expression on residual MM cells was noted in responders; expression increased at progression in most. Responses and CART-BCMA expansion were associated with CD4:CD8 T cell ratio and frequency of CD45RO-CD27+CD8+ T cells in the pre-manufacturing leukapheresis product. Conclusion: CART-BCMA infusions with or without lymphodepleting chemotherapy are clinically active in heavily-pretreated MM patients. Trial Registration: NCT02546167. Funding: University of Pennsylvania-Novartis Alliance and NIH.
Adam D. Cohen, Alfred L. Garfall, Edward A. Stadtmauer, J. Joseph Melenhorst, Simon F. Lacey, Eric Lancaster, Dan T. Vogl, Brendan M. Weiss, Karen Dengel, Annemarie Nelson, Gabriela Plesa, Fang Chen, Megan M. Davis, Wei-Ting Hwang, Regina M. Young, Jennifer L. Brogdon, Randi Isaacs, Iulian Pruteanu-Malinici, Don L. Siegel, Bruce L. Levine, Carl H. June, Michael C. Milone
BACKGROUND. Chimeric antigen receptor (CAR) T cells can induce remission in highly refractory leukemia and lymphoma subjects, yet the parameters for achieving sustained relapse-free survival are not fully delineated. METHODS. We analyzed 43 pediatric and young adult subjects participating in a Phase I trial of defined composition CD19CAR T cells (NCT02028455). CAR T cell phenotype, function and expansion, as well as starting material T cell repertoire, were analyzed in relation to therapeutic outcome (defined as achieving complete remission within 63 days) and duration of leukemia free survival and B cell aplasia. RESULTS. These analyses reveal that initial therapeutic failures (n = 5) were associated with attenuated CAR T cell expansion and/or rapid attrition of functional CAR effector cells following adoptive transfer. The CAR T products were similar in phenotype and function when compared to products resulting in sustained remissions. However, the initial apheresed peripheral blood T cells could be distinguished by an increased frequency of LAG-3+/TNF-αlow CD8 T cells and, following adoptive transfer, the rapid expression of exhaustion markers. For the 38 subjects who achieved an initial sustained MRD-neg remission, remission durability correlated with therapeutic products having increased frequencies of TNF-α-secreting CAR CD8+ T cells, and was dependent on a sufficiently high CD19+ antigen load at time of infusion to trigger CAR T cell proliferation. CONCLUSION. These parameters have the potential to prospectively identify patients at risk for therapeutic failure and support the development of approaches to boost CAR T cell activation and proliferation in patients with low levels of CD19 antigen. TRIAL REGISTRATION. ClinicalTrials.gov NCT02028455. FUNDING. Partial funding for this study was provided by Stand Up to Cancer & St. Baldrick’s Pediatric Dream Team Translational Research Grant (SU2C-AACR-DT1113), RO1 CA136551-05, Alex Lemonade Stand Phase I/II Infrastructure Grant, Conquer Cancer Foundation Career Development Award, Washington State Life Sciences Discovery Fund, Ben Towne Foundation, William Lawrence & Blanche Hughes Foundation, and Juno Therapeutics, Inc., a Celgene Company.
Olivia C. Finney, Hannah M. Brakke, Stephanie Rawlings-Rhea, Roxana Hicks, Danielle Doolittle, Marisa Lopez, Robert B. Futrell, Rimas J. Orentas, Daniel Li, Rebecca A. Gardner, Michael C. Jensen
A key mechanism of tumor resistance to immune cells is mediated by expression of peptide-loaded HLA-E in tumor cells, which suppresses natural killer (NK) cell activity via ligation of the NK inhibitory receptor CD94/NKG2A. Gene expression data from approximately 10,000 tumor samples showed widespread HLAE expression, with levels correlating with those of KLRC1 (NKG2A) and KLRD1 (CD94). To bypass HLA-E inhibition, we developed a way to generate highly functional NK cells lacking NKG2A. Constructs containing a single-chain variable fragment derived from an anti-NKG2A antibody were linked to endoplasmic reticulum-retention domains. After retroviral transduction in human peripheral blood NK cells, these NKG2A Protein Expression Blockers (PEBLs) abrogated NKG2A expression. The resulting NKG2Anull NK cells had higher cytotoxicity against HLA-E-expressing tumor cells. Transduction of anti-NKG2A PEBL produced more potent cytotoxicity than interference with an anti-NKG2A antibody and prevented de novo NKG2A expression, without affecting NK cell proliferation. In immunodeficient mice, NKG2Anull NK cells were significantly more powerful than NKG2A+ NK cells against HLA-E-expressing tumors. Thus, NKG2A downregulation evades the HLA-E cancer immune-checkpoint, and increases the anti-tumor activity of NK cell infusions. Because this strategy is easily adaptable to current protocols for clinical-grade immune cell processing, its clinical testing is feasible and warranted.
Takahiro Kamiya, See Voon Seow, Desmond Wong, Murray Robinson, Dario Campana
Mucus-invasive bacterial biofilms are identified on the colon mucosa of approximately 50% of colorectal cancer (CRC) patients and approximately 13% of healthy subjects. Here, we test the hypothesis that human colon biofilms comprise microbial communities that are carcinogenic in CRC mouse models. Homogenates of human biofilm-positive colon mucosa were prepared from tumor patients (tumor and paired normal tissues from surgical resections) or biofilm-positive biopsies from healthy individuals undergoing screening colonoscopy; homogenates of biofilm-negative colon biopsies from healthy individuals undergoing screening colonoscopy served as controls. After 12 weeks, biofilm-positive, but not biofilm-negative, human colon mucosal homogenates induced colon tumor formation in 3 mouse colon tumor models (germ-free ApcMinΔ850/+;Il10–/– or ApcMinΔ850/+ and specific pathogen–free ApcMinΔ716/+ mice). Remarkably, biofilm-positive communities from healthy colonoscopy biopsies induced colon inflammation and tumors similarly to biofilm-positive tumor tissues. By 1 week, biofilm-positive human tumor homogenates, but not healthy biopsies, displayed consistent bacterial mucus invasion and biofilm formation in mouse colons. 16S rRNA gene sequencing and RNA-Seq analyses identified compositional and functional microbiota differences between mice colonized with biofilm-positive and biofilm-negative communities. These results suggest human colon mucosal biofilms, whether from tumor hosts or healthy individuals undergoing screening colonoscopy, are carcinogenic in murine models of CRC.
Sarah Tomkovich, Christine M. Dejea, Kathryn Winglee, Julia L. Drewes, Liam Chung, Franck Housseau, Jillian L. Pope, Josee Gauthier, Xiaolun Sun, Marcus Mühlbauer, Xiuli Liu, Payam Fathi, Robert A. Anders, Sepideh Besharati, Ernesto Perez-Chanona, Ye Yang, Hua Ding, Xinqun Wu, Shaoguang Wu, James R. White, Raad Z. Gharaibeh, Anthony A. Fodor, Hao Wang, Drew M. Pardoll, Christian Jobin, Cynthia L. Sears