LAG-3: another brake to release in breast cancer?

Abstract

Upregulation of inhibitory immune checkpoints is critical for the control of T-cell activation in order to prevent autoimmunity and tissue damage. It is now clear that tumors can hijack immune checkpoint mechanisms as protection against the anticancer T-cell response. Blockade of the PD-1/PD-L1 immune checkpoint pathway has created a revolution in the treatment of melanoma, lung cancer and several other cancer types. In metastatic breast cancer patients, the response rates to PD-1 blocking antibodies are modest (4%–25%), but durable responses are seen [1.S Adams, Schmid P, Rugo HS Phase 2 study of pembrolizumab (pembro) monotherapy for previously treated metastatic triple-negative breast cancer (mTNBC): KEYNOTE-086 cohort A. 2017; J Clin Oncol 35(15_suppl): 1008.Google Scholar, 2.Adams S. Phase 2 study of pembrolizumab as first-line therapy for PD-L1–positive metastatic triple-negative breast cancer (mTNBC): Preliminary data from KEYNOTE-086 cohort B.J Clin Oncol 2017. 2017; 35: 1088Google Scholar, 3.L Dirix , Takacs I, Nikolinakos P. Abstract [S1-04] Avelumab (MSB0010718C), an anti-PD-L1 antibody, in patients with locally advanced or metastatic breast cancer: a phase Ib JAVELIN solid tumor trial. In Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium, San Antonio, December 8-12, 2015. AACR 2016; 76(4_suppl): S1-04–S1-04.Google Scholar, 4.Kok M. LBA14. Adaptive phase II randomized non-comparative trial of nivolumab after induction treatment in triple negative breast cancer: TONIC-trial.In ESMO, Madrid, Spain, 2017; Annals of Oncology. 2017; 28: mdx440Google Scholar, 5.Nanda R. et al.Pembrolizumab in patients with advanced triple-negative breast cancer: phase Ib KEYNOTE-012 study.J Clin Oncol. 2016; 34: 2460-2467Crossref PubMed Scopus (967) Google Scholar, 6.HS Rugo , J-PDelord, S-AIm Preliminary efficacy and safety of pembrolizumab (MK-3475) in patients with PD-L1-positive, ER-positive (ER+)/HER-2 negative breast cancer enrolled in Keynote 028. In San Antonio Breast Cancer Symposium, , San Antonio, USA, Dec 2015, Abstract S5–07.Google Scholar, 7.Schmid P. Atezolizumab in metastatic TNBC (mTNBC).Long-term clinical outcomes and biomarker analyses in AACR. 2017; Google Scholar]. Given that the majority of patients do not have clinical benefit, there is an urgent need to improve immunotherapy for breast cancer patients. This optimization is not a paved road and requires the integration of different parallel approaches, such as the search for predictive biomarkers [8.S Loi , Adams S, Schmid P LBA13 - Relationship between tumor infiltrating lymphocyte (TIL) levels and response to pembrolizumab (pembro) in metastatic triple-negative breast cancer (mTNBC): results from KEYNOTE-086. In ESMO 2017 Congress, Madrid, Spain, September 2017. Annals Oncol 2017; 28 (suppl_5): v605-v649.Google Scholar], combination treatment with conventional therapies [9.Adams S. et al.Phase Ib trial of atezolizumab in combination with nab-paclitaxel in patients with metastatic triple-negative breast cancer (mTNBC).J Clin Oncol 2016. 2016; 34: 1009Google Scholar, 10.S Tolaney , Savulsky C, Aktan G Phase 1b/2 Study to Evaluate Eribulin Mesylate in Combination With Pembrolizumab in Patients With Metastatic Triple-negative Breast Cancer. In 2016 San Antonio Breast Cancer Symposium, San Antonio, Texas, December 6-10, 2016; AACR 2017;77(4_suppl): P5-15-02.Google Scholar], strategies to convert cold tumors into hot tumors [4.Kok M. LBA14. Adaptive phase II randomized non-comparative trial of nivolumab after induction treatment in triple negative breast cancer: TONIC-trial.In ESMO, Madrid, Spain, 2017; Annals of Oncology. 2017; 28: mdx440Google Scholar, 11.A Ribas , Dummer R, Puzanov I Oncolytic virotherapy promotes intratumoral T cell infiltration and improves anti-PD-1 immunotherapy. Cell170(6): 1109–1119.e10.Google Scholar] and development of novel immunomodulatory compounds. The next wave of antibodies against co-inhibitory molecules that are currently being tested in clinical trials includes anti-LAG-3 (lymphocyte activation gene-3) antibodies [12.Andrews L.P. Marciscano A.E. Drake C.G. et al.LAG3 (CD223) as a cancer immunotherapy target.Immunol Rev. 2017; 276: 80-96http://dx.doi.org/10.1111/imr.12519Crossref PubMed Scopus (460) Google Scholar]. LAG-3 is expressed on effector T cells, regulatory T cells (Tregs), NK cells, and is considered a marker for T-cell exhaustion [13.AC Anderson , NJoller, K.Vijay Kuchroo, Lag-3, Tim-3, and TIGIT: co-inhibitory receptors with specialized functions in immune regulation . Immunity44(5): 989–1004.Google Scholar, 14.Wherry E.J. Kurachi M. Molecular and cellular insights into T cell exhaustion.Nat Rev Immunol. 2015; 15: 486-499http://dx.doi.org/10.1038/nri3862Crossref PubMed Scopus (2243) Google Scholar]. Exhausted T cells have reduced effector functions and express multiple inhibitory receptors. MHC class II molecules are ligands for LAG-3. LAG-3 is a negative regulator of T-cell expansion and preclinical research suggests that LAG3 is required for optimal Treg function [15.Workman C.J. Cauley L.S. Kim I.-J. et al.Lymphocyte activation gene-3 (CD223) regulates the size of the expanding T cell population following antigen activation in vivo.J Immunol. 2004; 172: 5450-5455http://dx.doi.org/10.4049/jimmunol.172.9.5450Crossref PubMed Scopus (222) Google Scholar, 16.Workman C.J. Vignali DAA. Negative regulation of T cell homeostasis by lymphocyte activation gene-3 (CD223).J Immunol. 2005; 174: 688-695http://dx.doi.org/10.4049/jimmunol.174.2.688Crossref PubMed Scopus (233) Google Scholar]. Preclinical data have also shown a synergy between PD-1 and LAG-3 in tolerance to both self- and tumor antigens [17.Woo S.-R. Turnis M.E. Goldberg M.V. et al.Immune inhibitory molecules LAG-3 and PD-1 synergistically regulate T-cell function to promote tumoral immune escape.Cancer Res. 2012; 72: 917-927http://dx.doi.org/10.1158/0008-5472.CAN-11-1620Crossref PubMed Scopus (1032) Google Scholar]. Currently, there are at least four LAG-3 targeted agents being tested in clinical trials [12.Andrews L.P. Marciscano A.E. Drake C.G. et al.LAG3 (CD223) as a cancer immunotherapy target.Immunol Rev. 2017; 276: 80-96http://dx.doi.org/10.1111/imr.12519Crossref PubMed Scopus (460) Google Scholar]. The first-in-class agent, IMP321, is a LAG-3 fusion protein that can activate DCs, and when combined with paclitaxel in the treatment of metastatic breast cancer, has shown higher response rates compared with historical controls [18.Brignone C. Gutierrez M. Mefti F. et al.First-line chemoimmunotherapy in metastatic breast carcinoma: combination of paclitaxel and IMP321 (LAG-3Ig) enhances immune responses and antitumor activity.J Transl Med. 2010; 8: 71.http://dx.doi.org/10.1186/1479-5876-8-71Crossref PubMed Scopus (186) Google Scholar]. Currently, LAG-3 specific antibodies are under development as monotherapy or in combination with PD-1/PDL-1 blocking agents. In several malignancies, such as ovarian cancer, melanoma and hepatocellular carcinoma, LAG-3 is concurrently expressed with PD-1 in a subset of tumors. In this issue of Annals of Oncology, Buruga et al. [19.Burugu S. et al.LAG-3+ tumor infiltrating lymphocytes in breast cancer: clinical correlates and association with PD-1/PD-L1+ tumors.Ann Oncol. 2017; 28: 2977-2984Abstract Full Text Full Text PDF PubMed Scopus (103) Google Scholar] present data on the co-occurrence of LAG-3 and PD-1 in breast tumors and the association of LAG-3 with prognosis in historical patient cohorts. They found that in 11% (327/2921) of cases LAG-3 was present on intraepithelial tumor infiltrating lymphocytes (iTILs) and PD-1 in 8.7% (244/2796). LAG-3-positive iTILs were more often seen in ER-negative tumors (24%). In 61% (147/244) of the tumors harboring PD-1-positive iTILs, co-occurrence with LAG-3 positivity was seen. This results in 5.3% (147/2796) of the total amount of tumors having iTILs with both PD-1 as well as LAG-3. Even though LAG-3-positive iTILs were associated with favorable prognosis, independent of known prognostic factors, the association between LAG-3 and outcome was no longer significant when PD-1-positive iTILs were added to the model. The current study by Buruga et al. was conducted using a relatively large and well-characterized dataset and though the proportion of LAG-3/PD-1 positive cases was low (5.3%), it provides additional rationale for the development of novel immunomodulatory treatment approaches for breast cancer, in particular for triple negative breast cancer (TNBC) for which targeted treatments are still lacking. Previously, using somewhat smaller datasets (total n = 363), Bottai et al. [20.Bottai G. Raschioni C. Losurdo A. et al.An immune stratification reveals a subset of PD-1/LAG-3 double-positive triple-negative breast cancers.Breast Cancer Res. 2016; 18: 121.http://dx.doi.org/10.1186/s13058-016-0783-4Crossref PubMed Scopus (75) Google Scholar] reported the concurrent expression of PD-1 and LAG-3 in ∼15% of TNBCs. However, in contrast with the current study by Buruga et al., they did not observe a significant improvement in survival for LAG-3 positive cases. Obviously, both studies should be considered exploratory and non-significant results might be due to the small numbers of LAG-3/PD-1 positive cases. In addition, different methods were used. While Buruga et al. used tissue microarrays (TMAs) and focused on iTILs, the paper by Bottai et al. described data using whole slides and stromal TIL (sTILs). The differences between these studies illustrate the limitations and inconsistencies in data analyses generally used in this kind of work. For example, since TMAs are constructed using only small tumor cores that have often been taken in regions with high tumor content and not selected based on the presence of abundant immune infiltration, TMAs may not give the complete picture of the presence of LAG-3 positive cells. Moreover, both studies did not examine actual co-expression of LAG-3 and PD-1 using multiplex staining and the question concerning which immune cell subtype within the TILs have upregulated these immune checkpoints remains unanswered. The association of PD-1 with LAG-3 can be interpreted in at least two ways. First, LAG-3 could be a marker of exhaustion of T cells that are part of the immune suppressive tumor microenvironment. On the other hand, LAG-3 may reflect an ongoing endogenous anticancer T-cell response and PD-1 upregulation that often accompanies LAG3 expression is a downstream effect of this immune response. To further distinguish between these models it may, for instance, be of interest to assess whether LAG3 expression is increased upon therapy in the small fraction of patients with a favorable response to PD(L)1 blockade. While the authors of the current manuscript suggest that patients with a tumor positive for PD-1/PD-L1 and LAG-3 should perhaps be excluded from trials evaluating combinations of immune checkpoint inhibitors, as the disease course of this subgroup is relatively favorable with the current treatment, their data can also be used to argue in the opposite direction. At the ESMO 2017 congress, Ascierto and colleagues presented data on simultaneous blockade of PD-1 and LAG-3 (using relatlimab) resulting in objective responses in 11% of melanoma patients who progressed after anti-PD(L)1 monotherapy [21.Ascierto P.A. et al.LBA18. Efficacy of BMS-986016, a monoclonal antibody that targets lymphocyte activation gene-3 (LAG-3), in combination with nivolumab in pts with melanoma who progressed during prior anti-PD-1/PD-L1 therapy (mel prior IO) in all-comer and biomarker enriched populations.In ESMO 2017 Congress. 2017; Google Scholar]. Their first translational results indicate that these responses were associated with expression of LAG-3 in the tumor microenvironment. This suggests that LAG-3 could be a biomarker to select patients for (combination) treatment with anti-LAG3. Given these recent preliminary data, the current manuscript of Buruga et al. can be placed in a somewhat different spotlight than simply being a descriptive prognostic study and may help to prioritize strategies on how to optimize immune checkpoint blockade for breast cancer patients.