Abstract
Background: The precise mechanism by which the immune system is adversely affected in cancer patients remains poorly understood, but the accumulation of immunosuppressive/protumorigenic myeloid-derived suppressor cells (MDSCs) is thought to be a prominent mechanism contributing to immunologic tolerance of malignant cells in epithelial ovarian cancer (EOC). To this end, we hypothesized genetic variation in MDSC pathway genes would be associated with survival after EOC diagnoses.
Methods: We measured the hazard of death due to EOC within 10 years of diagnosis, overall and by invasive subtype, attributable to SNPs in 24 genes relevant in the MDSC pathway in 10,751 women diagnosed with invasive EOC. Versatile Gene-based Association Study and the admixture likelihood method were used to test gene and pathway associations with survival.
Results: We did not identify individual SNPs that were significantly associated with survival after correction for multiple testing (P < 3.5 × 10−5), nor did we identify significant associations between the MDSC pathway overall, or the 24 individual genes and EOC survival.
Conclusions: In this well-powered analysis, we observed no evidence that inherited variations in MDSC-associated SNPs, individual genes, or the collective genetic pathway contributed to EOC survival outcomes.
Impact: Common inherited variation in genes relevant to MDSCs was not associated with survival in women diagnosed with invasive EOC. Cancer Epidemiol Biomarkers Prev; 26(3); 420–4. ©2016 AACR.
Introduction
Survival after a diagnosis of epithelial ovarian cancer (EOC) has seen only modest improvements in recent decades, making the identification of novel mechanisms and pathways associated with EOC prognosis imperative. EOC is associated with immunosuppressive pathways, including regulatory T cells and myeloid-derived suppressor cells (MDSC) that can be barriers to antitumor immunity and adversely affect clinical outcomes. To this end, MDSCs suppress the antigen-specific T-cell response by both CD4+ and CD8+ T cells, and elevated concentrations of MDSCs have been detected in the peripheral blood of cancer patients when compared with normal controls (1, 2). We hypothesized that common inherited genetic variation in genes involved in the MDSC pathway is associated with survival following ovarian cancer diagnosis.
Materials and Methods
We conducted a pooled analysis utilizing individual-level data from 28 studies in the Ovarian Cancer Association Consortium to assess the association of genes in the MDSC-associated pathway with EOC survival. Participants included 11,034 women ages 18 years and older with a histologically confirmed primary diagnosis of invasive EOC, fallopian tube cancer, or primary peritoneal cancer who were genotyped on the Ilumina iSelect array designed for the Collaborative Oncological Gene-environment Study (3). Clinical, epidemiologic, and follow-up data were made available for all analyses.
To assess the association between invasive EOC outcome and inherited variation in the MDSC pathway, we conducted SNP, gene, and pathway-based analyses of 24 candidate genes relevant to the biology of MDSCs, as established from an extensive literature review utilizing the PubMed database (ARG1, CD274, CSF2, CSF3, EIF2AK4, FLT3, IL10RA, IL13RA2, IL4, IL4R, IL5RA, IL6R, IDO, IRF8, KITLG, MMP1, MMP12, MMP3, MMP9, NOS2A, PSME4, STAT1, STAT3, and VEGFA). SNP selection and quality control were performed as described previously, yielding a total of 736 SNPs for analyses (4). We calculated the effective number of independent SNPs tested; this value was used in a Bonferroni correction to determine single-SNP significance (4). We utilized Cox proportional hazards regression models adjusted for age, tumor stage, and grade to estimate HRs and 95% confidence intervals (CI) representing SNP associations with EOC overall and by invasive histotype. Survival time was defined as the time from diagnosis of invasive EOC until death from any cause or time of last follow-up. Analyses accommodated left truncation to account for prevalent cases where appropriate and right censoring was done at >10 years follow-up time. Analyses and graphics were done using R (https://www.r-project.org). Gene- and pathway-based tests of association with hazard of death were performed using Versatile Gene-based Association Study and the admixture likelihood method, respectively (5, 6).
Results
The clinical characteristics of the study population are presented in Table 1. As expected, the majority of patients were diagnosed with serous EOCs, had poorly differentiated tumors, and were diagnosed with distant disease.
Clinical characteristics of invasive EOC cases from the Ovarian Cancer Association Consortium analyzed for association with MSDC genetic variation
We considered P < 3.5 × 10−5 as the threshold for significance, based on a Bonferroni correction for the estimated number of independent SNPs (n = 288) across five histotypes. Single SNP associations for EOC overall and by invasive histotype are shown in circular Manhattan style plots in Fig. 1 with SNPs showing P < 0.01 highlighted in red. The most significant single SNP was the C allele of rs6492925 in EIF2AK4 on chromosome 15, with a reduction in hazard of death in women with mucinous tumors (HR = 0.57; 95% CI, 0.42–0.78; P = 3.7 × 10−4).
These five concentric circles are circular standard Manhattan plots. The chromosome is on the outer circle, −log10 P values are on the y-axis (vertical), with each circle representing the P value from single SNP tests of association with overall survival adjusted for age, stage, and grade. A–E, The Manhattan plots are as follows: all ovarian cancer cases (A); high-grade serous (B); mucinous cell (C); endometrioid (D); and clear cell (E). The red dashed line designates P = 0.01, −log10(P value) = 2, with all red-colored SNPs above that line reflecting SNPs P < 0.01.
The most significant gene-based associations for all invasive ovarian cancer cases (KITLG, P = 0.07), high-grade serous (VEGFA, P = 0.11), mucinous (EIF2AK4, P = 0.015), endometrioid (CSF, P = 0.02), and clear cell (CD274, P = 0.037) did not the pass multiple test correction threshold set for testing the 24 genes. Taken together, the 24 genes showed no significant association with any histotype; mucinous cell tumors showed the most significant MDSC pathway association with survival (P = 0.11).
Discussion
Assuming genotyping captures, on average, 70% of the variation in each gene for tests of association with overall EOC and given the proportion of events at 51%, our study had 80% power at P < 3.5 × 10−5 to detect an HR of 1.11 to 1.24 for minor allele frequencies between 40% and 10%, respectively. We conducted a well-powered, hypothesis-driven study to evaluate a role for common inherited variation in MDSC pathway genes with EOC survival; we observed no evidence of an association at the SNP, gene or pathway level with EOC survival. To date, neither genome wide analyses of single SNP association with progression-free survival nor copy number variation with overall survival showed significant findings and did not report suggestive associations in these genes (7, 8). It is possible that rare variation in MDSC-associated genes not captured by these analyses could be correlated with EOC outcomes or that the magnitude of effect sizes was below detection. In addition, recent work has identified an expanding list of genes associated with MDSCs; thus, future studies should consider the importance of this emerging knowledge of MDSC biology.
Disclosure of Potential Conflicts of Interest
P.A. Fasching reports receiving commercial research grants from Amgen and Novartis and has received speakers bureau honoraria from Novartis and Pfizer. No potential conflicts of interest were disclosed by the other authors.
Authors' Contributions
Conception and design: L.E. Sucheston-Campbell, E. Zsiros, H. Anton-Culver, M.W. Beckmann, S.A. Engelholm, A. Ziogas, E.L. Goode, K.B. Moysich
Development of methodology: L.E. Sucheston-Campbell, P. Mayor, P.K. Wallace, S.A. Engelholm, T. Huzarski, K.B. Moysich
Acquisition of data (provided animals, acquired and managed patients, provided facilities, etc.): P. Mayor, P.K. Wallace, K.T. Soh, E. Zsiros, H. Anton-Culver, E.V. Bandera, M.W. Beckmann, A. Berchuck, L. Bjorge, I.G. Campbell, G. Chenevix-Trench, D.W. Cramer, A. Dansonka-Mieszkowska, F. Dao, B. Diergaarde, T. Doerk, A. du Bois, D. Eccles, S.A. Engelholm, P.A. Fasching, S.A. Gayther, A. Gentry-Maharaj, R.M. Glasspool, M.T. Goodman, J. Gronwald, P. Harter, A. Hein, F. Heitz, P. Hillemmanns, C. Høgdall, T. Huzarski, A. Jensen, B.Y. Karlan, R. Klapdor, T. Kluz, B. Konopka, S.K. Kjær, J. Kupryjanczyk, D. Lambrechts, J. Lester, D.A. Levine, V. McGuire, U. Menon, F. Modugno, R.B. Ness, S. Orsulic, J. Paul, T. Pejovic, P. Pharoah, S.J. Ramus, M.A. Rossing, M. Rübner, J.M. Schildkraut, N. Siddiqui, P. Sobiczewski, H. Song, K.L. Terry, E. Van Nieuwenhuysen, A. Vanderstichele, I. Vergote, C.S. Walsh, P.M. Webb, N. Wentzensen, A.S. Whittemore, A.H. Wu, A. Ziogas, K. Odunsi, J. Chang-Claude, E.L. Goode, K.B. Moysich
Analysis and interpretation of data (e.g., statistical analysis, biostatistics, computational analysis): L.E. Sucheston-Campbell, A.I. Clay, J.L. Etter, K.H. Eng, S. Liu, S. Battaglia, Q. Hu, J.B. Szender, P. Mayor, S.I. Abrams, B.H. Segal, P.K. Wallace, D. Lambrechts, R.B. Ness, P. Pharoah, M.A. Rossing, B. Schmalfeldt, W. Sieh, I. Vergote, A. Ziogas, K. Odunsi, K.B. Moysich
Writing, review, and/or revision of the manuscript: L.E. Sucheston-Campbell, R. Cannioto, A.I. Clay, J.L. Etter, K.H. Eng, S. Battaglia, J.B. Szender, A. Minlikeeva, J.M. Joseph, P. Mayor, S.I. Abrams, B.H. Segal, P.K. Wallace, K.T. Soh, E. Zsiros, H. Anton-Culver, E.V. Bandera, M.W. Beckmann, A. Berchuck, L. Bjorge, A. Bruegl, G. Chenevix-Trench, D.W. Cramer, B. Diergaarde, J.A. Doherty, A. du Bois, D. Eccles, P.A. Fasching, A. Gentry-Maharaj, R.M. Glasspool, M.T. Goodman, J. Gronwald, P. Harter, F. Heitz, P. Hillemmanns, C. Høgdall, E.V.S. Høgdall, A. Jensen, S.E. Johnatty, A. Jong, B.Y. Karlan, R. Klapdor, T. Kluz, B. Konopka, S.K. Kjær, D. Lambrechts, L. Lundvall, V. McGuire, I.A. McNeish, U. Menon, F. Modugno, R.B. Ness, C.L. Pearce, P. Pharoah, S.J. Ramus, M.A. Rossing, B. Schmalfeldt, N. Siddiqui, W. Sieh, H. Song, I. Vergote, P.M. Webb, N. Wentzensen, A.H. Wu, K. Odunsi, J. Chang-Claude, E.L. Goode, K.B. Moysich
Administrative, technical, or material support (i.e., reporting or organizing data, constructing databases): L.E. Sucheston-Campbell, R. Cannioto, J.L. Etter, J.M. Joseph, A. Berchuck, S.P. Campbell, G. Chenevix-Trench, A. Dansonka-Mieszkowska, T. Doerk, S.A. Engelholm, C. Høgdall, E.V.S. Høgdall, S.E. Johnatty, B.Y. Karlan, T. Kluz, D. Lambrechts, J. Lester, F. Modugno, S.J. Ramus, J. Rothstein, I. Schwaab
Study supervision: L.E. Sucheston-Campbell, S. Battaglia, P. Mayor, M.W. Beckmann, S.K. Kjær, F. Modugno, K.B. Moysich
Other (pathologist, clinical geneticist): J. Lubiński
Other (acquisition of clinical information): P. Sobiczewski
Grant Support
This work was supported by the following grants:
The Ovarian Cancer Association Consortium is supported by a grant from the Ovarian Cancer Research Fund.
This study used shared resources supported by RPCI's Cancer Center Support grant from the NCI (P30CA016056) and was also supported by the NCI Ovarian SPORE grant P50CA159981 and Roswell Park Alliance Foundation.
L.E. Sucheston-Campbell is supported by P50CA159981 and Roswell Park Alliance Foundation.
K.B. Moysich is supported by P50CA159981 and Roswell Park Alliance Foundation, NIH/NCIR01CA095023, and NIH/NCIR01CA126841.
K.H. Eng was supported by the Roswell Park Alliance Foundation.
S.I. Abrams was supported by R01CA140622.
B.H. Segal was supported by R01CA188900.
P.K. Wallace and this work was supported by Roswell Park Cancer Institute Ovarian Spore (1P50CA159981-01A1).
J.B. Szender was supported by 5T32CA108456.
Albina Minlikeeva was supported by Interdisciplinary Training Grant in Cancer Epidemiology (R25CA113951).
AUS (G. Chenevix-Trench and P.M. Webb): U.S. Army Medical Research and Materiel Command (DAMD17-01-1-0729), National Health & Medical Research Council of Australia (199600 and 400281), Cancer Councils of New South Wales, Victoria, Queensland, South Australia, and Tasmania and Cancer Foundation of Western Australia (under Multi-State Applications 191, 211, and 182).
BAV (P.A. Fasching): ELAN Funds of the University of Erlangen-Nuremberg.
BEL (D. Lambrechts): Nationaal Kankerplan.
DOV (M.A. Rossing): NIH (R01-CA112523 and R01-CA87538).
GER (J. Chang-Claude): German Federal Ministry of Education and Research, Programme of Clinical Biomedical Research (01 GB 9401), and the German Cancer Research Center (DKFZ).
HAW (M. Goodman): NIH (R01-CA58598, N01-CN-55424, and N01-PC-67001).
HOP (F. Modugno, K. Moysich, and R. Ness): DOD (DAMD17-02-1-0669) and NCI (K07-CA080668, R01-CA95023, and P50-CA159981); NIH/National Center for Research Resources/General Clinical Research Center grant MO1-RR000056; R01-CA126841.
LAX (B.Y. Karlan): American Cancer Society Early Detection Professorship (SIOP-06-258-01-COUN) and the National Center for Advancing Translational Sciences (NCATS), grant UL1TR000124.
MAL (S.K. Kjær): Funding for this study was provided by research grant R01- CA61107 from the NCI, research grant 94 222 52 from the Danish Cancer Society, Copenhagen, Denmark, and the Mermaid I project.
MAY (E.L. Goode): NIH (R01-CA122443, P30-CA15083, and P50-CA136393), Mayo Foundation, Minnesota Ovarian Cancer Alliance, and Fred C. and Katherine B. Andersen Foundation.
NCO (J. Schildkraut and A. Berchuck): NIH (R01-CA76016) and the Department of Defense (DAMD17-02-1-0666).
NEC (D. Cramer and K. Terry): NIH (R01-CA54419 and P50-CA105009) and Department of Defense (W81XWH-10-1-02802).
NJO (E.V. Bandera): NCI (NIH-K07 CA095666, NIH-K22-CA138563, and P30-CA072720) and the Cancer Institute of New Jersey.
NOR (L. Bjorge): Helse Vest, The Norwegian Cancer Society, and The Research Council of Norway.
ORE (T. Pejovic): OHSU Foundation.
POC (J. Gronwald): Pomeranian Medical University.
POL (N. Wentzensen): Intramural Research Program of the NCI.
PVD (E. Høgdall and C. Høgdall): Herlev Hospitals Forskningsråd, Direktør Jacob Madsens og Hustru Olga Madsens fond, Arvid Nilssons fond, Gangsted fonden, Herlev Hospitals Forskningsråd, and Danish Cancer Society.
RMH (P. Pharoah): Cancer Research UK (no grant number is available) and Royal Marsden Hospital.
SEA (P. Pharoah): Cancer Research UK (C490/A10119 and C490/A10124) and UK National Institute for Health Research Biomedical Research Centres at the University of Cambridge.
SRO (S. Banerjee, J. Paul, N. Siddiqui, R. Glasspool, and I. McNeish): Cancer Research UK (C536/A13086 and C536/A6689) and Imperial Experimental Cancer Research Centre (C1312/A15589).
STA (A.S. Whittemore and W. Sieh): NIH (U01-CA71966, R01-CA16056, K07-CA143047, and U01-CA69417) for recruitment of controls by the Cancer Prevention Institute of California.
UCI (H. Anton-Culver): NIH (R01-CA058860) and the Lon V Smith Foundation grant LVS-39420.
UKO (U Menon, A Gentry-Maharaj, and S. Gayther): The UKOPS study was funded by The Eve Appeal (The Oak Foundation) and supported by the National Institute for Health Research University College London Hospitals Biomedical Research Centre.
UKR (P. Pharaoh): Cancer Research UK (C490/A6187) and UK National Institute for Health Research Biomedical Research Centres at the University of Cambridge.
USC (C.L. Pearce): P01CA17054, P30CA14089, R01CA61132, N01PC67010, R03CA113148, R03CA115195, N01CN025403, and California Cancer Research Program (00-01389V-20170, 2II0200).
WOC (J. Kupryjanczyk): National Science Centren (N N301 5645 40) The Maria Sklodowska-Curie Memorial Cancer Center and Institute of Oncology, Warsaw, Poland.
The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
Footnotes
L.E. Sucheston-Campbell and R. Cannioto are first co-authors of this article.
- Received August 10, 2016.
- Accepted August 26, 2016.
- ©2016 American Association for Cancer Research.
Volume 26, Issue 3
