Potentiating radiotherapy and chemotherapy by inhibiting DNA damage repair is proposed as a therapeutic strategy to improve outcomes for patients with solid tumors. However, this approach risks enhancing normal tissue toxicity as much as tumor toxicity, thereby limiting its translational impact. Using NU5455, a newly-identified highly-selective oral inhibitor of DNA-dependent protein kinase catalytic subunit (DNA-PKcs) activity, we found that it was indeed possible to preferentially augment the effect of targeted-radiotherapy on human orthotopic lung tumors without influencing acute DNA-damage or a late radiation-induced toxicity (fibrosis) to normal mouse lung. Furthermore, while NU5455 administration increased both the efficacy and toxicity of a parenterally-administered topoisomerase inhibitor, it enhanced the activity of doxorubicin released locally in liver tumor xenografts without inducing any adverse effect. This strategy is particularly relevant to hepatocellular cancer which is treated clinically with localized drug-eluting beads and for which DNA-PKcs activity is reported to confer resistance to treatment. We conclude that transient pharmacological inhibition of DNA-PKcs activity is effective and tolerable when combined with localized DNA-damaging therapies and thus has promising clinical potential.
Catherine E. Willoughby, Yanyan Jiang, Huw D. Thomas, Elaine Willmore, Suzanne Kyle, Anita Wittner, Nicole Phillips, Yan Zhao, Susan J. Tudhope, Lisa Prendergast, Gesa Junge, Luiza Madia Lourenco, M. Raymond V. Finlay, Paul Turner, Joanne M. Munck, Roger J. Griffin, Tommy Rennison, James Pickles, Celine Cano, David R. Newell, Helen L. Reeves, Anderson J. Ryan, Stephen R. Wedge
Catecholamines released by sympathetic nerves can activate adrenergic receptors present on nearly every cell type, including myeloid derived suppressor cells (MDSCs). Using in vitro systems and murine tumor models, in wild-type mice and genetically modified (β2-AR–/–) mice, as well adoptive transfer approaches, we found that the degree of β2-AR signaling significantly influences MDSC frequency and survival in tumors and other tissues, modulates their expression of immunosuppressive molecules such as arginase-I and PDL-1 and alters their ability to suppress the proliferation of T cells. The regulatory functions of β-AR signaling in MDSCs were found to be dependent upon STAT3 phosphorylation. Moreover, we observed that the β2-AR-mediated increase in survival of MDSCs is dependent upon Fas-FasL interactions, and this is consistent with gene expression analyses which reveal a greater expression of apoptosis-related genes in β2-AR–/– MDSCs. Our data reveals the potential of β2-AR signaling to increase the generation of MDSCs from both murine and human peripheral blood cells and that the immunosuppressive function of MDSCs could be mitigated by treatment with β-AR antagonists, or enhanced by β-AR agonists, strongly supporting the possibility that reducing stress-induced activation of β2-ARs could help to overcome immune suppression and enhance the efficacy of immunotherapy and other cancer therapies.
Hemn Mohammadpour, Cameron R. MacDonald, Guanxi Qiao, Minhui Chen, Bowen Dong, Bonnie L. Hylander, Philip L. McCarthy, Scott I. Abrams, Elizabeth A. Repasky
The Microphthalmia family of transcription factors (MiT/TFE) controls lysosomal biogenesis and is negatively regulated by the nutrient sensor mTORC1. However, the mechanisms by which cells with constitutive mTORC1 signaling maintain lysosomal catabolism remain to be elucidated. Using the murine epidermis as a model system, we found that epidermal Tsc1 deletion resulted in a phenotype characterized by wavy hair and curly whiskers, and was associated with increased EGFR and HER2 degradation. Unexpectedly, constitutive mTORC1 activation with Tsc1 loss increased lysosomal content via up-regulated expression and activity of MiT/TFEs, while genetic deletion of Rheb or Rptor or prolonged pharmacologic mTORC1 inactivation had the reverse effect. This paradoxical increase in lysosomal biogenesis by mTORC1 was mediated by feedback inhibition of AKT, and a resulting suppression of AKT-induced MiT/TFE down-regulation. Thus, inhibiting hyperactive AKT signaling in the context of mTORC1 loss-of-function fully restored MiT/TFE expression and activity. These data suggest that signaling feedback loops work to restrain or maintain cellular lysosomal content during chronically inhibited or constitutively active mTORC1 signaling respectively, and reveal a mechanism by which mTORC1 regulates upstream receptor tyrosine kinase signaling.
Kaushal Asrani, Sanjana Murali, Brandon Lam, Chan-Hyun Na, Pornima Phatak, Akshay Sood, Harsimar Kaur, Zoya Khan, Michaël Noë, Ravi K. Anchoori, C. Conover Talbot Jr., Barbara Smith, Michael Skaro, Tamara L. Lotan
The prognostic value of immune cell infiltration within the tumor microenvironment (TME) has been extensively investigated via histological and genomic approaches. Based on the positive prognostic value of T cell infiltration, Immunoscore has been developed and validated for predicting risk of recurrence for colorectal cancer (CRC). Also, association between a consensus T helper 1 (Th-1) immune response and favorable clinical outcomes has been observed across multiple cancer types. Here, we reanalyzed public genomic data sets from The Cancer Genome Atlas (TCGA) and NCBI Gene Expression Omnibus (NCBI-GEO) and performed multispectral immunohistochemistry (IHC) on a cohort of colorectal tumors. We identified and characterized a risk group, representing approximately 10% of CRC patients, with high intratumoral CD8+ T cell infiltration, but poor prognosis. These tumors included both microsatellite instable (MSI) and stable (MSS) phenotypes and had a high density of tumor-associated macrophages (TAMs) that expressed CD274 (programmed death-ligand 1 [PD-L1]), TGF-β activation, and an immune overdrive signature characterized by the overexpression of immune response and checkpoint genes. Our findings illustrate that CRC patients may have poor prognosis despite high CD8+ T cell infiltration and provide CD274 as a simple biomarker for identifying these patients.
Marwan Fakih, Ching Ouyang, Chongkai Wang, Travis Yiwey Tu, Maricel C. Gozo, May Cho, Marvin Sy, Jeffrey A. Longmate, Peter P. Lee
Beclin 1 (Becn1) is a key molecule of the autophagy pathway and has been implicated in cancer development. Due to the embryonic lethality of Becn1 homozygous deficient mice, the precise mechanisms and cell-type-specific role of Becn1 in the regulation of inflammation and tumor immunity remain elusive. Here, we report that myeloid-deficient Becn1 (Becn1ΔM) mice develop neutrophilia and hypersusceptible to LPS-induced septic shock, with a high risk of developing spontaneous precursor (pre)-B cell lymphoma with elevated expressions of immunosuppressive molecules PD-L1 and IL-10. Becn1 deficiency results in stabilization of neutrophil MEKK3, aberrant p38 activation, and neutrophil-B cell interaction through Cxcl9/Cxcr3 chemotaxis. Neutrophil-B cell interplay leads to activations of IL-21/STAT3/IRF1 and CD40L/ERK signaling, together regulates the programmed death ligand 1 (PD-L1) expression, and suppresses CD8+ T cell function. Ablation of p38 in Becn1ΔM mice prevents neutrophil-inflammation and B cell tumorigenesis. Importantly, low Becn1 expression in human neutrophils correlates with PD-L1 levels in pre-B ALL patients. Our findings have identified myeloid Becn1 as a therapeutic target of cancer immunity and immunotherapy for pre-B lymphomas.
Peng Tan, Lian He, Changsheng Xing, Jingrong Mao, Xiao Yu, Motao Zhu, Lixia Diao, Leng Han, Yubin Zhou, James M. You, Helen Y. Wang, Rong-Fu Wang
Despite progress in intensification of therapy, outcomes for patients with metastatic osteosarcoma (OS) have not improved in thirty years. We developed a system that enabled preclinical screening of compounds against metastatic OS cells in the context of the native lung microenvironment. Using this strategy to screen a library of epigenetically targeted compounds, we identified inhibitors of CDK12 to be most effective, reducing OS cell outgrowth in the lung by more than 90% at submicromolar doses. We found that knockout of CDK12 in an in vivo model of lung metastasis significantly decreased the ability of OS to colonize the lung. CDK12 inhibition led to defects in transcription elongation in a gene length– and expression-dependent manner. These effects were accompanied by defects in RNA processing and altered the expression of genes involved in transcription regulation and the DNA damage response. We further identified OS models that differ in their sensitivity to CDK12 inhibition in the lung and provided evidence that upregulated MYC levels may mediate these differences. Our studies provided a framework for rapid preclinical testing of compounds with antimetastatic activity and highlighted CDK12 as a potential therapeutic target in OS.
Ian Bayles, Malgorzata Krajewska, W. Dean Pontius, Alina Saiakhova, James J. Morrow, Cynthia Bartels, Jim Lu, Zachary J. Faber, Yuriy Fedorov, Ellen S. Hong, Jaret M. Karnuta, Brian Rubin, Drew J. Adams, Rani E. George, Peter C. Scacheri
Oral squamous cell carcinoma (OSCC) frequently invades the maxillary or mandibular bone, and this bone invasion is closely associated with poor prognosis and survival. Here, we show that CCL28 functions as a negative regulator of OSCC bone invasion. CCL28 inhibited invasion and epithelial-mesenchymal transition (EMT), and its inhibition of EMT was characterized by induced E-cadherin expression and reduced nuclear localization of beta-catenin in OSCC cells with detectable RUNX3 expression levels. CCL28 signaling via CCR10 increased retinoic acid receptor (RAR)β expression by reducing the interaction between RARα and HDAC1. In addition, CCL28 reduced RANKL production in OSCC and osteoblastic cells and blocked RANKL-induced osteoclastogenesis in osteoclast precursors. Intraperitoneally administered CCL28 inhibited tumor growth and osteolysis in mouse calvaria and tibia inoculated with OSCC cells. RARβ expression was also increased in tumor tissues. In OSCC patients, low CCL28, CCR10, and RARβ expression levels were highly correlated with bone invasion. OSCC patients with higher expression of CCL28, CCR10, or RARβ had significantly better overall survival. These findings suggest that CCL28, CCR10, and RARβ are useful markers for the prediction and treatment of OSCC bone invasion. Furthermore, CCL28 upregulation in OSCC cells or CCL28 treatment can be a therapeutic strategy for OSCC bone invasion.
Junhee Park, Xianglan Zhang, Sun Kyoung Lee, Na-Young Song, Seung Hwa Son, Ki Rim Kim, Jae Hoon Shim, Kwang-Kyun Park, Won-Yoon Chung
Treatment of tumors with ionizing radiation stimulates an antitumor immune response partly dependent on induction of IFNs. These IFNs directly enhance dendritic cell and CD8+ T cell activity. Here we show that resistance to an effective antitumor immune response is also a result of IFN signaling in a different cellular compartment of the tumor, the cancer cells themselves. We abolished type I IFN signaling in cancer cells by genetic elimination of its receptor, IFNAR1. Pronounced immune responses were provoked after ionizing radiation of tumors from 4 mouse cancer cell lines with Ifnar1 knockout. This enhanced response depended on CD8+ T cells and was mediated by enhanced susceptibility to T cell–mediated killing. Induction of Serpinb9 proved to be the mechanism underlying control of susceptibility to T cell killing after radiation. Ifnar1-deficient tumors had an augmented response to anti–PD-L1 immunotherapy with or without radiation. We conclude that type I IFN can protect cancer cells from T cell–mediated cytotoxicity through regulation of Serpinb9. This result helps explain why radiation of tumors can stimulate antitumor immunity yet also result in resistance. It further suggests potential targets for intervention to improve therapy and to predict responses.
Jianzhou Chen, Yunhong Cao, Bostjan Markelc, Jakob Kaeppler, Jenny A.F. Vermeer, Ruth J. Muschel
BACKGROUND Adenoid cystic carcinoma (ACC) is a rare malignancy arising in salivary glands and other sites, characterized by high rates of relapse and distant spread. Recurrent/metastatic (R/M) ACCs are generally incurable, due to a lack of active systemic therapies. To improve outcomes, deeper understanding of genetic alterations and vulnerabilities in R/M tumors is needed.METHODS An integrated genomic analysis of 1,045 ACCs (177 primary, 868 R/M) was performed to identify alterations associated with advanced and metastatic tumors. Intratumoral genetic heterogeneity, germline mutations, and therapeutic actionability were assessed.RESULTS Compared with primary tumors, R/M tumors were enriched for alterations in key Notch (NOTCH1, 26.3% vs. 8.5%; NOTCH2, 4.6% vs. 2.3%; NOTCH3, 5.7% vs. 2.3%; NOTCH4, 3.6% vs. 0.6%) and chromatin-remodeling (KDM6A, 15.2% vs. 3.4%; KMT2C/MLL3, 14.3% vs. 4.0%; ARID1B, 14.1% vs. 4.0%) genes. TERT promoter mutations (13.1% of R/M cases) were mutually exclusive with both NOTCH1 mutations (q = 3.3 × 10–4) and MYB/MYBL1 fusions (q = 5.6 × 10–3), suggesting discrete, alternative mechanisms of tumorigenesis. This network of alterations defined 4 distinct ACC subgroups: MYB+NOTCH1+, MYB+/other, MYBWTNOTCH1+, and MYBWTTERT+. Despite low mutational load, we identified numerous samples with marked intratumoral genetic heterogeneity, including branching evolution across multiregion sequencing.CONCLUSION These observations collectively redefine the molecular underpinnings of ACC progression and identify further targets for precision therapies.FUNDING Adenoid Cystic Carcinoma Research Foundation, Pershing Square Sohn Cancer Research grant, the PaineWebber Chair, Stand Up 2 Cancer, NIH R01 CA205426, the STARR Cancer Consortium, NCI R35 CA232097, the Frederick Adler Chair, Cycle for Survival, the Jayme Flowers Fund, The Sebastian Nativo Fund, NIH K08 DE024774 and R01 DE027738, and MSKCC through NIH/NCI Cancer Center Support Grant (P30 CA008748).
Allen S. Ho, Angelica Ochoa, Gowtham Jayakumaran, Ahmet Zehir, Cristina Valero Mayor, Justin Tepe, Vladimir Makarov, Martin G. Dalin, Jie He, Mark Bailey, Meagan Montesion, Jeffrey S. Ross, Vincent A. Miller, Lindsay Chan, Ian Ganly, Snjezana Dogan, Nora Katabi, Petros Tsipouras, Patrick Ha, Nishant Agrawal, David B. Solit, P. Andrew Futreal, Adel K. El Naggar, Jorge S. Reis-Filho, Britta Weigelt, Alan L. Ho, Nikolaus Schultz, Timothy A. Chan, Luc G.T. Morris
Microtubule-associated serine/threonine kinase 1 (MAST1) is a central driver of cisplatin resistance in human cancers. However, the molecular mechanism regulating MAST1 levels in cisplatin-resistant tumors is unknown. Through a proteomics screen, we identified the heat shock protein 90 B (hsp90B) chaperone as a direct MAST1 binding partner essential for its stabilization. Targeting hsp90B sensitized cancer cells to cisplatin predominantly through MAST1 destabilization. Mechanistically, interaction of hsp90B with MAST1 blocked ubiquitination of MAST1 at lysines 317 and 545 by the E3 ubiquitin ligase CHIP and prevented proteasomal degradation. The hsp90B-MAST1-CHIP signaling axis and its relationship with cisplatin response were clinically validated in cancer patients. Furthermore, combined treatment with a hsp90 inhibitor and the MAST1 inhibitor lestaurtinib further abrogated MAST1 activity and consequently enhanced cisplatin-induced tumor growth arrest in a patient-derived xenograft model. Our study not only uncovers the regulatory mechanism of MAST1 in tumors but also suggests a promising combinatorial therapy to overcome cisplatin resistance in human cancers.
Chaoyun Pan, Jaemoo Chun, Dan Li, Austin C. Boese, Jie Li, JiHoon Kang, Anna Umano, Yunhan Jiang, Lina Song, Kelly R. Magliocca, Zhuo G. Chen, Nabil F. Saba, Dong M. Shin, Taofeek K. Owonikoko, Sagar Lonial, Lingtao Jin, Sumin Kang