Digital Genomic Quantification of Tumor-Infiltrating Lymphocytes
Science Translational Medicine • 4 Dec 2013 • Vol 5, Issue 214 • p. 214ra169 • DOI: 10.1126/scitranslmed.3007247
TILling the Soil
The immune system is geared to fight off foreign invaders, but its record against cancer is less than stellar. The recent successes of immunotherapy for cancer add urgency to the need to not only understand exactly what is happening with immune cells in the tumor—tumor-infiltrating lymphocytes (TILs)—but also how to evaluate them as metrics for an active response. However, these evaluations are limited by the tools at hand. Immunohistochemistry is one approach that may give us a big picture but not be able to be quantified precisely enough for consistent diagnostic use between laboratories. Now, Robins et al. report the advent of a digital polymerase chain reaction–based assay that can count and assess clonality of TILs within tumors.
The authors used their approach for diagnosis in acute lymphoblastic leukemia, as well as to measure TIL number in both primary and metastatic ovarian cancer. They show that higher TIL number associates improved survival in ovarian cancer patients. If verified in larger cohorts and paired with immunohistochemistry, this approach could help determine if TIL number could be a biomarker for immune response in tumors.
Abstract
Infiltrating T lymphocytes are frequently found in malignant tumors and are suggestive of a host cancer immune response. Multiple independent studies have documented that the presence and quantity of tumor-infiltrating lymphocytes (TILs) are strongly correlated with increased survival. However, because of methodological factors, the exact effect of TILs on prognosis has remained enigmatic, and inclusion of TILs in standard prognostic panels has been limited. For example, some reports enumerate all CD3+ cells, some count only cytotoxic CD8+ T cells, and the criteria used to score tumors as TIL-positive or TIL-negative are inconsistent among studies. To address this limitation, we introduce a robust digital DNA-based assay, termed QuanTILfy, to count TILs and assess T cell clonality in tissue samples, including tumors. We demonstrate the clonal specificity of this approach by the diagnosis of T cell acute lymphoblastic leukemia and the accurate, sensitive, and highly reproducible measurement of TILs in primary and metastatic ovarian cancer. Our experiments demonstrate an association between higher TIL counts and improved survival among women with ovarian cancer, and are consistent with previous observations that the immune response against ovarian cancer is a meaningful and independent prognostic factor. Surprisingly, the TIL repertoire is diverse for all tumors in the study with no notable oligoclonal expansions. Furthermore, because variability in the measurement and characterization of TILs has limited their clinical utility as biomarkers, these results highlight the significant translational potential of a robust, standardizable DNA-based assay to assess TILs in a variety of cancer types.
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Supplementary Material
Summary
Fig. S1. TIL fraction in patient-matched primary tumor and metastasis.
Table S1. Variable gene forward primers.
Table S2. Joining gene reverse primers.
Table S3. Variable gene TaqMan probes (each with FAM fluorophore).
Table S4. Reference assay oligo sequences.
Table S5. Assay subgroup assignments.
Table S6. TCRβ clone identified by sequencing.
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Information & Authors
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Published In
Science Translational Medicine
Volume 5 | Issue 214
December 2013
December 2013
Copyright
Copyright © 2013, American Association for the Advancement of Science.
Submission history
Received: 5 August 2013
Accepted: 11 October 2013
Acknowledgments
We acknowledge support from the Listwin Family Foundation (to J.H.B.), an Ellison Medical Foundation New Scholar Award (AG-NS-0577-09 to J.H.B.), an Outstanding New Environmental Scientist Award (ONES) (R01) from the National Institute of Environmental Health Sciences (R01ES019319 to J.H.B.), a grant from the Congressionally Directed Medical Research Programs/U.S. Department of Defense (W81XWH-10-1-0563 to J.H.B.), the Pacific Ovarian Cancer Research Consortium Ovarian Cancer SPORE Award (P50 CA083636), a Department of Defense Ovarian Cancer Idea Award (OC093221 to M.T.), a Susan G. Komen postdoctoral fellowship (to J.G.), and the Canary Foundation (to M.T.). The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Cancer Institute, the NIH, or any of the other granting agencies. Author contributions: H.S.R. helped design the experiments and co-wrote the manuscript. N.G.E. performed most of the digital genomic quantification of T lymphocytes and data analysis. M.T. designed the collection of, and J.G. processed the biospecimens used to assess T cell heterogeneity in ovarian cancer. C.W.D. surgically removed and, with K.C.O. and M.T., oversaw the collection of biospecimens, their processing, storage, annotation, and distribution. J.H.B. designed the experiments, developed the QuanTILfy assay, performed digital genomic quantification of T lymphocytes during assay development, analyzed the data, and co-wrote the manuscript. All authors reviewed and edited the manuscript. Competing interests: J.H.B. has equity in Adaptive Biotechnologies. H.S.R. has consultancy, equity ownership, patents, and royalties with Adaptive Biotechnologies. The FHCRC and Adaptive Biotechnologies have jointly filed for a patent application on this technology, entitled “Quantification of adaptive immune cell genomes in a complex mixture of cells” (US 13/656,265).
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