RAISEing VEGF-D’s importance as predictive biomarker for ramucirumab in metastatic colorectal cancer patients (2018)

A key unresolved question in the field of therapeutic antiangiogenesis is the lack of clinically applicable biomarkers to pre-select responding patients and spare resistant patients unnecessary side-effects and costs. The study by Tabernero et al. discovered VEGF-D as a novel biomarker candidate to predict efficacy of the anti-VEGFR-2 antibody ramucirumab as second-line treatment in metastatic colorectal cancer (mCRC) in the phase III RAISE trial [1.Tabernero J. Hozak R.R. Yoshino T. et al.Analysis of angiogenesis biomarkers for ramucirumab efficacy in patients with metastatic colorectal cancer from RAISE, a global, randomized, double-blind, phase III study.Ann Oncol. 2018; 29: 602-609Abstract Full Text Full Text PDF PubMed Scopus (60) Google Scholar]. This trial, which included only patients, who were refractory to bevacizumab, showed a survival benefit upon treatment with ramucirumab + FOLFIRI compared with the placebo + FOLFIRI group [2.Tabernero J. Yoshino T. Cohn A.L. et al.Ramucirumab versus placebo in combination with second-line FOLFIRI in patients with metastatic colorectal carcinoma that progressed during or after first-line therapy with bevacizumab, oxaliplatin, and a fluoropyrimidine (RAISE): a randomised, double-blind, multicentre, phase 3 study.Lancet Oncol. 2015; 16: 499-508Abstract Full Text Full Text PDF PubMed Scopus (623) Google Scholar, 3.Cohn A.L. Yoshino T. Heinemann V. et al.Exposure-response relationship of ramucirumab in patients with advanced second-line colorectal cancer: exploratory analysis of the RAISE trial.Cancer Chemother Pharmacol. 2017; 80: 599-608Crossref PubMed Scopus (22) Google Scholar, 4.Verdaguer H. Tabernero J. Macarulla T. Ramucirumab in metastatic colorectal cancer: evidence to date and place in therapy.Ther Adv Med Oncol. 2016; 8: 230-242Crossref PubMed Scopus (45) Google Scholar, 5.Arnold D. Fuchs C.S. Tabernero J. et al.Meta-analysis of individual patient safety data from six randomized, placebo-controlled trials with the antiangiogenic VEGFR2-binding monoclonal antibody ramucirumab.Ann Oncol. 2017; 28: 2932-2942Abstract Full Text Full Text PDF PubMed Scopus (72) Google Scholar, 6.Obermannova R. Van Cutsem E. Yoshino T. et al.Subgroup analysis in RAISE: a randomized, double-blind phase III study of irinotecan, folinic acid, and 5-fluorouracil (FOLFIRI) plus ramucirumab or placebo in patients with metastatic colorectal carcinoma progression.Ann Oncol. 2016; 27: 2082-2090Abstract Full Text Full Text PDF PubMed Scopus (45) Google Scholar]. These results led to the approval of ramucirumab for the treatment of mCRC besides gastric, gastroesophageal junction and non-small-cell-lung cancer. Consistent with the results from most antiangiogenic therapy (AAT) trials, the survival benefit for the mCRC patients treated with ramucirumab was in the order of months. Unfortunately, until today, very few promising biomarker candidates have been identified, mostly from phase III trials with bevacizumab in different cancer types. These include short VEGF isoforms, modified expression of neuropilin-1, genetic variants and modified expression of VEGF receptor 1 (VEGFR1) [7.Miles D.W. de Haas S.L. Dirix L.Y. et al.Biomarker results from the AVADO phase 3 trial of first-line bevacizumab plus docetaxel for HER2-negative metastatic breast cancer.Br J Cancer. 2013; 108: 1052-1060Crossref PubMed Scopus (129) Google Scholar, 8.Hein A. Lambrechts D. von Minckwitz G. et al.Genetic variants in VEGF pathway genes in neoadjuvant breast cancer patients receiving bevacizumab: results from the randomized phase III GeparQuinto study.Int J Cancer. 2015; 137: 2981-2988Crossref PubMed Scopus (24) Google Scholar, 9.Lambrechts D. Claes B. Delmar P. et al.VEGF pathway genetic variants as biomarkers of treatment outcome with bevacizumab: an analysis of data from the AViTA and AVOREN randomised trials.Lancet Oncol. 2012; 13: 724-733Abstract Full Text Full Text PDF PubMed Scopus (162) Google Scholar, 10.Lambrechts D. Lenz H.J. de Haas S. et al.Markers of response for the antiangiogenic agent bevacizumab.JCO. 2013; 31: 1219-1230Crossref PubMed Scopus (273) Google Scholar, 11.Hatch A.J. Clarke J.M. Nixon A.B. Hurwitz H.I. Identifying blood-based protein biomarkers for antiangiogenic agents in the clinic: a decade of progress.Cancer J. 2015; 21: 322-326Crossref PubMed Scopus (10) Google Scholar]. However, there are no positive trials prospectively validating these candidates yet [12.Miles D. Cameron D. Bondarenko I. et al.Bevacizumab plus paclitaxel versus placebo plus paclitaxel as first-line therapy for HER2-negative metastatic breast cancer (MERiDiAN): a double-blind placebo-controlled randomised phase III trial with prospective biomarker evaluation.Eur J Cancer. 2017; 70: 146-155Abstract Full Text Full Text PDF PubMed Scopus (98) Google Scholar]. The investigators need to be congratulated for conducting a statistically sound prospective biomarker study attached to the trial. Notably, this included randomization of patients in an exploratory and confirmatory cohort, leading to well-balanced clinical characteristics between these subgroups. However, an outstanding question is why in the context of mandatory collection of plasma samples, ∼20% of the patients were not included into the biomarker cohort [1.Tabernero J. Hozak R.R. Yoshino T. et al.Analysis of angiogenesis biomarkers for ramucirumab efficacy in patients with metastatic colorectal cancer from RAISE, a global, randomized, double-blind, phase III study.Ann Oncol. 2018; 29: 602-609Abstract Full Text Full Text PDF PubMed Scopus (60) Google Scholar, 2.Tabernero J. Yoshino T. Cohn A.L. et al.Ramucirumab versus placebo in combination with second-line FOLFIRI in patients with metastatic colorectal carcinoma that progressed during or after first-line therapy with bevacizumab, oxaliplatin, and a fluoropyrimidine (RAISE): a randomised, double-blind, multicentre, phase 3 study.Lancet Oncol. 2015; 16: 499-508Abstract Full Text Full Text PDF PubMed Scopus (623) Google Scholar]. The translational biomarker studies within the RAISE trial included VEGF-C, VEGF-D, soluble (s) VEGFR-1, sVEGFR-2, and sVEGFR-3 in plasma samples and immunohistochemical evaluation of VEGFR-2 and vessel density [1.Tabernero J. Hozak R.R. Yoshino T. et al.Analysis of angiogenesis biomarkers for ramucirumab efficacy in patients with metastatic colorectal cancer from RAISE, a global, randomized, double-blind, phase III study.Ann Oncol. 2018; 29: 602-609Abstract Full Text Full Text PDF PubMed Scopus (60) Google Scholar, 2.Tabernero J. Yoshino T. Cohn A.L. et al.Ramucirumab versus placebo in combination with second-line FOLFIRI in patients with metastatic colorectal carcinoma that progressed during or after first-line therapy with bevacizumab, oxaliplatin, and a fluoropyrimidine (RAISE): a randomised, double-blind, multicentre, phase 3 study.Lancet Oncol. 2015; 16: 499-508Abstract Full Text Full Text PDF PubMed Scopus (623) Google Scholar]. Of all biomarkers analyzed, only VEGF-D displayed predictive and prognostic significance. A cut-off value of 115 pg/mL VEGF-D was identified to discriminate between patients with low and high VEGF-D expression. High expression of VEGF-D was associated with beneficial progression-free survival (PFS) and overall survival (OS), but not objective response rate (ORR) upon addition of ramucirumab to chemotherapy [1.Tabernero J. Hozak R.R. Yoshino T. et al.Analysis of angiogenesis biomarkers for ramucirumab efficacy in patients with metastatic colorectal cancer from RAISE, a global, randomized, double-blind, phase III study.Ann Oncol. 2018; 29: 602-609Abstract Full Text Full Text PDF PubMed Scopus (60) Google Scholar] (Figure 1). In contrast, ramucirumab did not improve PFS nor OS in patients with low VEGF-D plasma levels when compared with the placebo + FOLFIRI arm. VEGF-D binds to VEGFR-2 and VEGFR-3 and has multifaceted tumor-promoting effects including stimulation of tumor growth, angiogenesis, lymphangiogenesis and metastasis in addition to immunosuppression [13.Lohela M. Bry M. Tammela T. Alitalo K. VEGFs and receptors involved in angiogenesis versus lymphangiogenesis.Curr Opin Cell Biol. 2009; 21: 154-165Crossref PubMed Scopus (573) Google Scholar, 14.Carmeliet P. Jain R.K. Molecular mechanisms and clinical applications of angiogenesis.Nature. 2011; 473: 298-307Crossref PubMed Scopus (3721) Google Scholar]. It is thus not surprising that this growth factor of the VEGF family has a prognostic value besides its relevance as a predictive biomarker [1.Tabernero J. Hozak R.R. Yoshino T. et al.Analysis of angiogenesis biomarkers for ramucirumab efficacy in patients with metastatic colorectal cancer from RAISE, a global, randomized, double-blind, phase III study.Ann Oncol. 2018; 29: 602-609Abstract Full Text Full Text PDF PubMed Scopus (60) Google Scholar]. The potential of VEGF-D as predictive biomarker has already been retrospectively studied in two first-line CRC trials (CALGB 80405; AGITG MAX), both combining the anti-VEGF antibody bevacizumab with chemotherapy [15.Weickhardt A.J. Williams D.S. Lee C.K. et al.Vascular endothelial growth factor D expression is a potential biomarker of bevacizumab benefit in colorectal cancer.Br J Cancer. 2015; 113: 37-45Crossref PubMed Scopus (43) Google Scholar, 16.Nixon A. Sibley A. Hatch A. et al.Blood-based biomarkers in patients (pts) with metastatic colorectal cancer (mCRC) treated with FOLFOX or FOLFIRI plus bevacizumab (Bev), cetuximab (Cetux), or bev plus Cetux: results from CALGB 80405 (Alliance).J Clin Oncol. 2016; 34Crossref Google Scholar]. In both trials, low VEGF-D levels were associated with better outcome upon treatment with bevacizumab. At first sight, these findings are apparently in contrast with the outcomes of the RAISE trial. However, although traditionally considered to mediate lymphangiogenesis, VEGF-D can substitute for the bevacizumab target VEGF-A and induce angiogenesis due to its binding capacity to VEGFR-2. Based on this biologic rationale, one can speculate that patients with high expression of VEGF-D would not benefit from bevacizumab but instead from ramucirumab (Figure 1). Along these lines, in the patient cohort enrolled in the RAISE trial, who progressed under therapy with bevacizumab, VEGF-D might be compensatorily upregulated and these patients seem to benefit most from the addition of ramucirumab to second-line treatment. Obviously, this assumption has several caveats, including different methodologies used to detect VEGF-D in the RAISE, CALGB and AGITG MAX trials [15.Weickhardt A.J. Williams D.S. Lee C.K. et al.Vascular endothelial growth factor D expression is a potential biomarker of bevacizumab benefit in colorectal cancer.Br J Cancer. 2015; 113: 37-45Crossref PubMed Scopus (43) Google Scholar, 16.Nixon A. Sibley A. Hatch A. et al.Blood-based biomarkers in patients (pts) with metastatic colorectal cancer (mCRC) treated with FOLFOX or FOLFIRI plus bevacizumab (Bev), cetuximab (Cetux), or bev plus Cetux: results from CALGB 80405 (Alliance).J Clin Oncol. 2016; 34Crossref Google Scholar]. Surprisingly, the VEGF-D levels detected in RAISE were about 10-fold lower than those of VEGF-D in CALGB [1.Tabernero J. Hozak R.R. Yoshino T. et al.Analysis of angiogenesis biomarkers for ramucirumab efficacy in patients with metastatic colorectal cancer from RAISE, a global, randomized, double-blind, phase III study.Ann Oncol. 2018; 29: 602-609Abstract Full Text Full Text PDF PubMed Scopus (60) Google Scholar]. If VEGF-D would mediate resistance to bevacizumab, one could expect higher levels of VEGF-D in the RAISE trial population, because in this second-line setting, all patients included were pre-treated with bevacizumab. These discrepancies in VEGF-D levels illustrate the necessity to develop a clinically applicable diagnostic enzyme-linked immunosorbent assay (ELISA). An interesting future trial design could be to treat mCRC patients, in whom tumors are still progressing after EGFR inhibition, with bevacizumab or ramucirumab according to VEGF-D levels. It is surprising that VEGF-C is neither predictive nor prognostic in RAISE, given the comparable affinity of VEGF-D and VEGF-C for VEGFR-2 [17.Papadopoulos N. Martin J. Ruan Q. et al.Binding and neutralization of vascular endothelial growth factor (VEGF) and related ligands by VEGF Trap, ranibizumab and bevacizumab.Angiogenesis. 2012; 15: 171-185Crossref PubMed Scopus (752) Google Scholar, 18.Leppanen V.M. Jeltsch M. Anisimov A. et al.Structural determinants of vascular endothelial growth factor-D receptor binding and specificity.Blood. 2011; 117: 1507-1515Crossref PubMed Scopus (67) Google Scholar]. Also, both cytokines can induce angiogenesis via VEGFR-2 and VEGFR-3, and mouse data indicate that the combined genetic deletion of VEGF-C and VEGF-D does not have additional inhibitory effects on vascular growth compared with the deletion of VEGF-C alone [19.Haiko P. Makinen T. Keskitalo S. et al.Deletion of vascular endothelial growth factor C (VEGF-C) and VEGF-D is not equivalent to VEGF receptor 3 deletion in mouse embryos.Mol Cell Biol. 2008; 28: 4843-4850Crossref PubMed Scopus (161) Google Scholar]. Perhaps, VEGF-D has another (more important?) biologic role in pathologic angiogenesis mediating resistance to ramucirumab, which might distinguish it from VEGF-C—future work is needed to resolve this issue. One point to consider is that the binding affinities of VEGF-C and VEGF-D for VEGFR-2 are increased by proteolytic cleavage, with only mature forms binding to VEGFR-2 [13.Lohela M. Bry M. Tammela T. Alitalo K. VEGFs and receptors involved in angiogenesis versus lymphangiogenesis.Curr Opin Cell Biol. 2009; 21: 154-165Crossref PubMed Scopus (573) Google Scholar, 20.McColl B.K. Paavonen K. Karnezis T. et al.Proprotein convertases promote processing of VEGF-D, a critical step for binding the angiogenic receptor VEGFR-2.FASEB J. 2007; 21: 1088-1098Crossref PubMed Scopus (90) Google Scholar]. However, recent data indicate that distinct parts of the N-terminus (Phe93–Arg108) of cleaved VEGF-D are critical for binding to VEGFR-2, which is not the case for VEGF-C [21.Davydova N. Harris N.C. Roufail S. et al.Differential Receptor Binding and Regulatory Mechanisms for the Lymphangiogenic Growth Factors Vascular Endothelial Growth Factor (VEGF)-C and -D.J Biol Chem. 2016; 291: 27265-27278Abstract Full Text Full Text PDF PubMed Scopus (25) Google Scholar], indicating functional differences between these growth factors. At this point, it is not clear which of the VEGF-D variants is detected by the ELISA employed. Also, which cells produce VEGF-D in these mCRC patients? While VEGF-D mRNA is detectable in heart, lung and intestines [13.Lohela M. Bry M. Tammela T. Alitalo K. VEGFs and receptors involved in angiogenesis versus lymphangiogenesis.Curr Opin Cell Biol. 2009; 21: 154-165Crossref PubMed Scopus (573) Google Scholar, 22.Achen M.G. Jeltsch M. Kukk E. et al.Vascular endothelial growth factor D (VEGF-D) is a ligand for the tyrosine kinases VEGF receptor 2 (Flk1) and VEGF receptor 3 (Flt4).Proc Natl Acad Sci USA. 1998; 95: 548-553Crossref PubMed Scopus (1017) Google Scholar], possible candidates of VEGF-D-producing cells in mCRC patients can be cancer cells and tumor-infiltrating immune cells [23.Schoppmann S.F. Birner P. Stockl J. et al.Tumor-associated macrophages express lymphatic endothelial growth factors and are related to peritumoral lymphangiogenesis.Am J Pathol. 2002; 161: 947-956Abstract Full Text Full Text PDF PubMed Scopus (671) Google Scholar], though this remains to be confirmed in the future. The prognostic and predictive relevance of VEGF-D raises the question of whether therapeutic targeting of VEGF-D could be an alternative option to blocking VEGFR-2. Monoclonal antibodies targeting VEGF-D were developed and might be useful as a single agent or in combination with bevacizumab to overcome resistance [21.Davydova N. Harris N.C. Roufail S. et al.Differential Receptor Binding and Regulatory Mechanisms for the Lymphangiogenic Growth Factors Vascular Endothelial Growth Factor (VEGF)-C and -D.J Biol Chem. 2016; 291: 27265-27278Abstract Full Text Full Text PDF PubMed Scopus (25) Google Scholar, 24.Tammela T. Zarkada G. Wallgard E. et al.Blocking VEGFR-3 suppresses angiogenic sprouting and vascular network formation.Nature. 2008; 454: 656-660Crossref PubMed Scopus (659) Google Scholar]. Furthermore, combination strategies targeting both VEGFR-2 and VEGFR-3 might be considered, because they proved to be superior in comparison to VEGFR-3 or VEGFR-2 inhibition alone in mice, and inhibition of VEGFR-3 alone has shown to be well-tolerated but displays low efficacy in mCRC patients when given as monotherapy [24.Tammela T. Zarkada G. Wallgard E. et al.Blocking VEGFR-3 suppresses angiogenic sprouting and vascular network formation.Nature. 2008; 454: 656-660Crossref PubMed Scopus (659) Google Scholar, 25.Saif M.W. Knost J.A. Chiorean E.G. et al.Phase 1 study of the anti-vascular endothelial growth factor receptor 3 monoclonal antibody LY3022856/IMC-3C5 in patients with advanced and refractory solid tumors and advanced colorectal cancer.Cancer Chemother Pharmacol. 2016; 78: 815-824Crossref PubMed Scopus (51) Google Scholar]. In conclusion, the RAISE trial identified VEGF-D as a potential predictive biomarker, warranting independent validation. Determination of VEGF-D levels could help to stratify patients for second-line mCRC therapy with ramucirumab and possibly for other indications. In addition to the identification of robust biomarkers for the efficacy of different AATs, alternative strategies, such as targeting endothelial cell metabolism to overcome resistance to current AATs, are warranted [14.Carmeliet P. Jain R.K. Molecular mechanisms and clinical applications of angiogenesis.Nature. 2011; 473: 298-307Crossref PubMed Scopus (3721) Google Scholar, 26.Rohlenova K. Veys K. Miranda-Santos I. et al.Endothelial Cell Metabolism in Health and Disease.Trends Cell Biol. 2017; : 30201-30205Google Scholar]. The work of PC is supported by the Federal Government Belgium (IUAP P7/03), long-term structural Methusalem funding by the Flemish Government, the Research Foundation Flanders (FWO-Vlaanderen), the Foundation against Cancer (2012-175 and 2016-078), Kom op tegen Kanker (Stand up to Cancer) by the Flemish cancer society and an Advanced European Research Council (ERC) Grant (EU-ERC743074). XL is supported by the State Key Laboratory of Ophthalmology at the Zhongshan Ophthalmic Center, Sun Yat-Sen University, and the National Natural Science Foundation of China (81330021, 81670855). LCC is funded by a fellowship of the Else Kröner-Fresenius-Stiftung and the Fritz Thyssen Stiftung (10.16.2.017MN). SL is supported by a Heisenberg Professorship (LO1683/4-1) from the German Research Council (DFG) and by a Starting Grant from the European Research Council (ELIMINATE).