Oncology
Abstract
While cytotoxic CD4+ tumor-infiltrating lymphocytes have anti-cancer activity in patients, whether these can be non-invasively monitored and how these are regulated remains obscure. By matching single cells with T cell receptors (TCR) in tumor and blood of bladder cancer patients, we identified distinct pools of tumor-matching cytotoxic CD4+ T cells in the periphery directly reflecting the predominant antigenic specificities of intratumoral CD4+ TILs. On one hand, the granzyme B (GZMB)-expressing cytotoxic CD4+ subset proliferated in blood in response to PD-1 blockade, but was separately regulated by the killer cell lectin-like receptor G1 (KLRG1) which inhibited their killing by interacting with E-cadherin. Conversely, a clonally related, granzyme K (GZMK)-expressing circulating CD4+ population demonstrated basal proliferation and a memory phenotype that may result from activation of GZMB+ cells, but was not directly mobilized by PD-1 blockade. As KLRG1 marked the majority of circulating tumor TCR-matched cytotoxic CD4+ T cells, this work nominates KLRG1 as a means to isolate them from blood and provide a window into intratumoral CD4+ recognition, as well as a putative regulatory receptor to mobilize the cytolytic GZMB+ subset for therapeutic benefit. Our findings also underscore ontogenic relationships of GZMB- and GZMK-expressing populations and the distinct cues that regulate their activity.
Authors
Serena S. Kwek, Hai Yang, Tony Li, Arielle Ilano, Eric D. Chow, Li Zhang, Hewitt Chang, Diamond Luong, Averey Lea, Matthew Clark, Alec Starzinski, Yimin Shi, Elizabeth McCarthy, Sima Porten, Maxwell V. Meng, Chun Jimmie Ye, Lawrence Fong, David Y. Oh
Abstract
Bariatric surgery is associated with improved breast cancer (BC) outcomes, including greater immunotherapy effectiveness in a pre-clinical BC model. A potential mechanism of bariatric surgery-associated protection is the gut microbiota. Here, we demonstrate the dependency of improved immunotherapy response on the post-bariatric surgery gut microbiome via fecal microbial transplant (FMT). Response to αPD-1 immunotherapy was significantly improved following FMT from formerly obese bariatric surgery-treated mice. When stool from post-bariatric surgery patients was transplanted into recipient mice and compared to the patients’ pre-surgery transplants, post-surgery microbes significantly reduced tumor burden and doubled immunotherapy effectiveness. Microbes impact tumor burden through microbially derived metabolites, including branched chain amino acids (BCAA). Circulating BCAAs correlated significantly with natural killer T (NKT) cell content in the tumor microenvironment in donor mice after bariatric surgery and FMT recipients of donor cecal content after bariatric surgery compared to obese controls. BCAA supplementation replicated improved αPD-1 effectiveness in two BC models, supporting the role of BCAAs in increased immunotherapy effectiveness after bariatric surgery. Ex vivo exposure increased primary NKT cell expression of anti-tumor cytokines, demonstrating direct activation of NKT cells by BCAAs. Together, findings suggest that reinvigorating anti-tumor immunity may depend upon bariatric surgery-associated microbially derived metabolites, namely BCAAs.
Authors
Margaret S. Bohm, Sydney C. Joseph, Laura M. Sipe, Minjeong Kim, Cameron T. Leathem, Tahliyah S. Mims, Nathaniel B. Willis, Ubaid A. Tanveer, Joel H. Elasy, Emily W. Grey, Madeline E. Pye, Zeid T. Mustafa, Barbara Anne Harper, Logan G. McGrath, Deidre Daria, Brenda Landvoigt Schmitt, Jelissa A. Myers, Patricia Pantoja Newman, Brandt D. Pence, Marie Van der Merwe, Matthew J. Davis, Joseph F. Pierre, Liza Makowski
Abstract
It is well documented that impaired DNA damage repair (DDR) induces genomic instability that can efficiently increase the sensitivity of prostate cancer (PCa) cells to PARP inhibitors; however, the underlying mechanism remains elusive. Here, we found profound genomic instability in PCa cells with SPOP gene mutations and confirmed the sensitivity of SPOP-mutated PCa cells to olaparib-induced apoptosis. Mechanistically, we identified olaparib-induced CK2-mediated phosphorylation of PIAS1-S468, which in turn mediated SUMOylation of SPOP, thus promoting its E3 ligase activity in the DDR. Moreover, an abnormal CK2/PIAS1/SPOP axis due to SPOP mutations or defects in CK2-mediated phosphorylation of PIAS1, as well as SPOP inhibitor treatment, led to impaired DDR, thus increasing olaparib-induced apoptosis of PCa cells and enhancing olaparib sensitivity in animal models and patient-derived organoids. This suggested that disruption of the CK2/PIAS1/SPOP signaling axis could serve as an indicator for targeted therapy of PCa using a PARP inhibitor.
Authors
Hui Zhang, Lili Kong, Jinhui Li, Zhihan Liu, Yiting Zhao, Xiuyi Lv, Liangpei Wu, Lin Chai, Hongjie You, Jiabei Jin, Xinyi Cao, Zhong Zheng, Yadong Liu, Zejun Yan, Xiaofeng Jin
Abstract
Aneuploidy, a cancer hallmark, drives chromosomal instability, drug resistance, and clinically-aggressive tumors. Cyclin-dependent kinase 2 (CDK2) antagonism with independent inhibitors or CDK2 knock-down triggered anaphase catastrophe. This disrupts supernumerary centrosome clustering, causing multipolar division and apoptosis. Time-lapse fluorescent microscopy of FUCCI cell cycle probes transduced into aneuploid lung cancer cells revealed distinct fates of bipolar and polyploid cells after CDK2 inhibition. Apoptosis occurred in multipolar progeny but was repressed in persistent polyploid cancer cells. RNA-seq analyses after CDK2 inhibition of 4N versus 2N lung cancer cells were enriched for CDK1 pathway and KIF family members. The Cancer Genome Atlas (TCGA) analysis of lung cancers indicated CDK1 and KIF family member overexpression was associated with an unfavorable survival. Intravital microscopy of transplanted lung cancer cells in mice extended findings from the in vitro to in vivo settings. CDK2 inhibition of tumor-bearing mice produced polyploid cancer cells in vivo. These cancer cells were resistant to apoptosis and proliferated despite CDK2 inhibition. In contrast, polyploid populations were rarely detected in CDK2 inhibited human alveolar epithelial cells. These findings are translationally relevant. Combined targeting of CDK2 with CDK1 or kinesin family member antagonists should eliminate polyploid cancer cells, promote apoptosis and augment antineoplastic effects.
Authors
Liliya Tyutyunyk-Massey, Zibo Chen, Xiuxia Liu, Masanori Kawakami, Adam Harned, Yeap Ng, Brian Luke, Samuel C. Okpechi, Blessing Ogunlade, Yair Alfaro-Mora, Roberto Weigert, Kedar Narayan, Xi Liu, Ethan Dmitrovsky
Abstract
Metastatic outgrowth in distant microscopic niches requires sufficient nutrients, including fatty acids (FAs), to support tumor growth and to generate an immunosuppressive tumor microenvironment (TME). However, despite the important role of FAs in metastasis, the regulation of FA supply in metastatic niches has not been defined. In this report, we show that tumor endothelium actively promotes outgrowth and restricts anti-tumor cytolysis by transferring FA into developing metastatic tumors. We describe a process of transendothelial FA delivery via endosomes that requires mTORC1 activity. Thus, endothelial-specific targeted deletion of Raptor (RptorECKO), a unique component of the mTORC1 complex, significantly reduced metastatic tumor burden that was associated with improved markers of T cell cytotoxicity. Low dose everolimus that selectively inhibited endothelial mTORC1 improves immune checkpoint responses in metastatic disease models. This work reveals the importance of transendothelial nutrient delivery to the TME, highlighting a future target for therapeutic development.
Authors
Deanna N Edwards, Shan Wang, Kelby Kane, Wenqiang Song, Laura C. Kim, Verra M. Ngwa, Yoonha Hwang, Kevin C. Ess, Mark R. Boothby, Jin Chen
Abstract
Dipeptidase-1 (DPEP1) is highly upregulated in colorectal cancer (CRC), with its enzymatic function linked to invasion and metastasis. More recently, DPEP1 was found to serve as a receptor for neutrophils when expressed by activated endothelial cells. It is unknown whether neutrophils bind to DPEP1-expressing CRC cells and whether this impacts features of CRC. Neutrophils have been shown to be tumor-promoting in cancers including CRC, where they act to exclude CD8+ T cells. Herein, we show that neutrophils bind DPEP1-expressing CRC cells. In addition, DPEP1 is preferentially expressed in microsatellite stable (MSS) CRC, in which there are a paucity of CD8+ T cells, whereas DPEP1 is negatively correlated with microsatellite unstable (MSI-H) CRC, which are T cell-rich and are more responsive to immunotherapy. Remarkably, carcinogen-treated Dpep1 null mice develop multiple, large, plaque-like, locally invasive adenocarcinomas and squamous cell cancers in the distal colon. These adenocarcinomas exhibit a marked reduction of neutrophils and an influx CD8+ T cells, along with reduced expression of mismatch repair proteins, consistent with features of MSI-H CRC. These results establish DPEP1’s importance in maintaining MSS CRC and its ability to shape the tumor microenvironment.
Authors
Sarah E. Glass, Matthew E. Bechard, Zheng Cao, Radhika Aramandla, Ping Zhao, Samuel T. Ellis, Emily H. Green, Elizabeth G. Fisher, Ryan T. Smith, Chelsie K. Sievers, Maria Johnson Irudayam, Frank Revetta, M. Kay Washington, Gregory D. Ayers, Cody N. Heiser, Alan J. Simmons, Yanwen Xu, Yu Wang, Annika Windon, Martha J. Shrubsole, Nicholas O. Markham, Qi Liu, Ken S. Lau, Robert J. Coffey
Abstract
An estimated 5-10% of cancer results from an underlying genetic predisposition, yet for the majority of these cases the genes in question remain unknown, suggesting a critical need to identify new cancer predisposition genes. The protein phosphatase 2A (PP2A) family exists as a trimeric holoenzyme and is a vital negative regulator of multiple oncogenic pathways. PP2A inhibition by somatic mutation, loss of expression, and upregulation of its exogenous inhibitors in tumors has been well described. However, it remains unknown whether germline loss of any PP2A subunits results in a predisposition to cancer in humans. In this study, we identified nine cancer patients with germline loss-of-function (LOF) variants in PPP2R1B (Aβ), the beta isoform of the PP2A scaffold subunit. All four patients for whom documentation was available also had a family history of cancer, including multiple indicators of hereditary cancer. The most highly represented cancer among the Aβ germline patients was breast cancer. Overexpression of these mutant forms of Aβ resulted in truncated proteins that were rapidly turned over. Characterization of an additional missense germline Aβ variant, R233C, that is also recurrently mutated at the somatic level found that it disrupts PP2A catalytic subunit binding resulting in loss of phosphatase activity. An analysis of Aβ expression among multiple breast cancer cohorts revealed that somatic, heterozygous loss of Aβ was a frequent event in this disease and decreased Aβ expression correlated with shorter disease-free and overall survival. Furthermore, Aβ levels were significantly lower in multiple histological subtypes of both in situ and malignant breast cancer compared to adjacent normal breast tissue, suggesting that Aβ loss is an early event in breast cancer development. Together, this highlights a role for Aβ as a predisposition gene in breast cancer and potentially additional cancers.
Authors
Sahar Mazhar, Caitlin M. O’Connor, Alexis Harold, Amanda C. Dowdican, Peter J. Ulintz, Erika N. Hanson, Yuping Zhang, Michelle F. Jacobs, Sofia D. Merajver, Mark W. Jackson, Anthony Scott, Anieta M. Sieuwerts, Arul M. Chinnaiyan, Goutham Narla
Abstract
BACKGROUND. A priori knowledge of recurrence risk in patients with non-metastatic (FIGO stage I) uterine serous carcinoma (USC) would enable a risk-stratified approach to the use of adjuvant chemotherapy. This would greatly reduce treatment-related morbidity and be predicted to improve survival. METHODS. GATA2 expression was scored by immunohistochemistry (IHC) across a retrospective multi-institutional cohort of 195 primary USCs. Associations between GATA2 levels and clinicopathologic metrics were evaluated using Student’s t-test, Fisher’s exact test, Kaplan-Meier method, and Cox proportional hazards ratio. Invasion in patient-derived USC cells was assessed by Student’s t-test. RNA-seq, anti-GATA2 ChIP-seq, and confirmatory western blotting enabled identification of GATA2 targets. RESULTS. Patients with FIGO stage I GATA2high USCs had 100% recurrence-free and 100% cancer related survival, which was significantly better than patients with GATA2low USCs. In patients for whom adjuvant chemotherapy was omitted, patients with GATA2high USC had 100% recurrence free 5-year survival compared to 60% recurrence free survival in patients with GATA2low USC. Depletion of GATA2 in patient-derived USC cells increased invasion in vitro. CONCLUSIONS. Routine GATA2 IHC identifies 33% of FIGO stage I USC patients who have a greatly reduced risk of post-hysterectomy USC recurrence. Our results suggest that a GATA2 guided personalized medicine approach could be rapidly implemented in most hospital settings, would reduce treatment-related morbidity, and likely improve outcomes in USC patients. FUNDING. NIH grants R01 DK068634, P30 CA014520, S10 OD023526, K08 DK127244, T32 HL007899, the UW-Madison Department of Pathology and Laboratory Medicine, the UW-Madison Centennial Scholars Program, the Diane Lindstrom Foundation, the American Cancer Society, the V Foundation, The Hartwell Foundation, and the UMN Department of Obstetrics, Gynecology, and Women's Health.
Authors
Usha S. Polaki, Trey E. Gilpin, Apoorva T. Patil, Emily Chiu, Ruth Baker, Peng Liu, Tatiana S. Pavletich, Morteza Seifi, Paula M. Mañán-Mejías, Jordan Morrissey, Jenna Port, Rene Welch Schwartz, Irene M. Ong, Dina El-Rayes, Mahmoud A. Khalifa, Pei Hui, Vanessa L. Horner, María Virumbrales-Muñoz, Britt K. Erickson, Lisa Barroilhet, Stephanie M. McGregor, Emery H. Bresnick, Daniel R. Matson
Abstract
Non–small cell lung cancer (NSCLC) is a common cause of cancer-related deaths worldwide, and its incidence has been increasing in recent years. While targeted therapies like osimertinib, an epidermal growth factor receptor tyrosine kinase inhibitor, have brought about notable improvements in patient outcomes for advanced NSCLC, the challenge of acquired drug resistance persists. Here, we found that cellular mesenchymal-epithelial transition factor (c-Met) was highly expressed in osimertinib-resistant cells, and depletion of c-Met markedly inhibited the growth of osimertinib-resistant cells ex vivo and in vivo, suggesting that c-Met is a potential target to address osimertinib resistance. Through a screening process using a natural product compound library, we identified piperlongumine as a potent inhibitor to overcome osimertinib resistance. Furthermore, the combined treatment of piperlongumine and osimertinib exhibited robust antitumor effects in resistant cells, partially restoring their sensitivity to osimertinib. Additionally, we discovered that piperlongumine could enhance the interaction between E3 ligase RNF4 and Sp1, inhibit the phosphorylation of Sp1 at Thr739, facilitate the ubiquitination and degradation of Sp1, lead to c-Met destabilization, and trigger intrinsic apoptosis in resistant cells. In summary, our study sheds light on the potential of piperlongumine in overcoming osimertinib resistance, offering new strategies and perspectives for the clinical management of drug-resistant NSCLC.
Authors
Ruirui Wang, Qiang Wang, Jinzhuang Liao, Xinfang Yu, Wei Li
Abstract
Hypoxia/hypoxia-inducible factor 1α–driven immunosuppressive transcription and cAMP-elevating signaling through A2A adenosine receptors (A2ARs) represent a major tumor-protecting pathway that enables immune evasion. Recent promising clinical outcomes due to the blockade of the adenosine-generating enzyme CD73 and A2AR in patients refractory to all other therapies have confirmed the importance of targeting hypoxia-adenosinergic signaling. We report a feasible approach to target the upstream stage of hypoxia-adenosinergic immunosuppression using an oxygen-carrying nanoemulsion (perfluorocarbon blood substitute). We show that oxygenation agent therapy (a) eliminates tumor hypoxia, (b) improves efficacy of endogenously developed and adoptively transferred T cells, and thereby (c) promotes regression of tumors in different anatomical locations. We show that both T cells and NK cells avoid hypoxic tumor areas and that reversal of hypoxia by oxygenation agent therapy increases intratumoral infiltration of activated T cells and NK cells due to reprogramming of the tumor microenvironment (TME). Thus, repurposing oxygenation agents in combination with supplemental oxygen may improve current cancer immunotherapies by preventing hypoxia-adenosinergic suppression, promoting immune cell infiltration and enhancing effector responses. These data also suggest that pretreating patients with oxygenation agent therapy may reprogram the TME from immunosuppressive to immune-permissive prior to adoptive cell therapy, or other forms of immunotherapy.
Authors
Katarina Halpin-Veszeleiova, Michael P. Mallouh, Lucy M. Williamson, Ashley C. Apro, Nuria R. Botticello-Romero, Camille Bahr, Maureen Shin, Kelly M. Ward, Laura Rosenberg, Vladimir B. Ritov, Michail V. Sitkovsky, Edwin K. Jackson, Bruce D. Spiess, Stephen M. Hatfield
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