Anti-Drug Antibodies Against Immune

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Author Manuscript Published OnlineFirst on November 22, 2019; DOI: 10.1158/1078-0432.CCR-19-2337 Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited.

1 Title page 2 3 Anti-drug Antibodies Against Immune Checkpoint Blockers: Impairment of drug 4 efficacy or indication of immune activation? 5 6 Authors: Diego Enrico1,2, Angelo Paci1,4,5, Nathalie Chaput1,3,4, Eleni Karamouza1,6, Benjamin Besse1,2,7 7 8 1 Gustave Roussy Cancer Campus, F-94805, Villejuif, France 9 2 Medical Oncology Department, Gustave Roussy Cancer Center, F-94805, Villejuif, France 10 3 Laboratory for Immunomonitoring in Oncology, UMS 3655 CNRS/US 23 INSERM, Gustave Roussy 11 Cancer Campus, F-94805, Villejuif, France 12 4 University Paris-Saclay, Faculty of Pharmacy, Chatenay-Malabry, F-92296, France 13 5 Laboratory of Pharmacology and Drug Analysis, Gustave Roussy Cancer Center, F-94805, Villejuif, France 14 6 Ligue Nationale Contre le Cancer Meta-Analysis Platform, Biostatistics and Epidemiology Unit, INSERM 15 U1018, CESP 16 7 University Paris-Saclay, Faculty of Medicine, F-94270, Le Kremlin-Bicêtre, France 17 18 19 Running tittle: Immunogenicity of cancer immunotherapy 20 21 22 Corresponding Author: Benjamin Besse, MD, PhD, Cancer Medicine Department, Gustave Roussy, 114 23 Rue Edouard Vaillant, 94805 Villejuif, France ([email protected]). 24 25 26 Keywords: Anti-drug Antibodies, Immunogenicity, Cancer, Oncology, Immunotherapy, Immune 27 checkpoint inhibitors, Atezolizumab 28 29 30 Conflict of interest statement 31 The authors declare no potential conflicts of interest 32 33 34

Downloaded from clincancerres.aacrjournals.org on September 30, 2021. © 2019 American Association for Cancer Research. Author Manuscript Published OnlineFirst on November 22, 2019; DOI: 10.1158/1078-0432.CCR-19-2337 Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited.

1 Translational Relevance 2 After therapeutic drugs exposure, the immune system induces a humoral response resulting in the 3 generation of anti-drug antibodies (ADAs). This particular immune response can cause a decreased in the 4 amount of drug available, resulting in some cases in reduced clinical efficacy. For the oncology drugs, there 5 is limited data about this phenomenon and its clinical relevance. Atezolizumab is the only immune 6 checkpoint inhibitor with an expanded analysis of ADAs but the clinical implication data is conflicting. 7 Furthermore, the combination of immune checkpoint inhibitors with chemotherapy, an established 8 standard of care in untreated advanced NSCLC patients, adds more complexity to this field. Given that 9 immune checkpoint inhibitors can modify the immune response, additional basic, translational and clinical 10 research may help to a better understanding of the biological process and the clinical implications of ADAs. 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34

1 Abstract

2 The generation of antibodies following exposure to therapeutic drugs has been widely studied, however in 3 oncology, data in relation to their clinical relevance are limited. Anti-drug antibodies (ADAs) can cause a 4 decrease in the amount of drug available, resulting in some cases in decreased anti-tumour activity and a 5 consequent impact on clinical outcomes. Several immunological factors can influence the development of 6 ADAs, and in addition, the sensitivity of the different testing methods used in different studies can vary, 7 representing an additional potential confounding factor. The reported frequency of ADA-positive patients 8 following treatment with immune checkpoint inhibitors varies from as low as 1.5% for pembrolizumab to 9 54% for atezolizumab. This latter drug is the only immune checkpoint inhibitor to have undergone an 10 expanded analysis of the clinical implications of ADAs, but with discordant results. Given that immune 11 checkpoint inhibitors can modify the immune response and potentially impact ADA formation, data from 12 published as well as prospective trials need to be evaluated for a better understanding of the clinical 13 implications of ADAs in this setting.

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1 Introduction

2 Any biologic agent administered to humans is likely to be recognised by the immune system which induces a 3 humoral response resulting in the generation of antidrug antibodies (ADAs) (Fig. 1)(1). These antibodies 4 may bind to the drug and cause loss of activity via different mechanisms, such as blockade of the drug target 5 binding (neutralising antibodies) or by increasing the clearance of the ADA-drug complex, resulting in 6 reduced drug concentrations in the plasma(2). Furthermore, ADAs may induce toxicities by the generation 7 of an immune response to the ADA-drug complex, with infusion-related reactions being the most frequent 8 events(3).

9 A range of factors can influence the development of this immunogenicity, such as the origin and structure 10 of the drug, impurities with adjuvant activity, route, dose and frequency of administration, 11 immunomodulatory properties of the therapeutic product, formulation of recombinant therapeutic proteins, 12 aggregates formed by shaking during preparation shipment, the patient’s immunologic and genetic status 13 and concomitant treatment(4). By their nature, ADAs are a heterogeneous group of antibodies with 14 different isotypes (IgM, IgG, IgE, or IgA). ADA development seems to be an early event, occurring during 15 the drug infusion, and in some cases as early as the first dose (5). Initially there is a low-affinity and non- 16 neutralising IgM response, followed by higher-affinity and neutralising antibodies mostly IgG1 and IgG4. In 17 patients treated for autoimmune or inflammatory diseases, the titres of ADAs, as well as the persistence of 18 ADAs throughout the treatment, were associated with clinical relevance(6). Adding to the difficulty of 19 interpretation is the fact that there are many testing methods, each with different levels of sensitivity and 20 cut-off points(6). In this context of the difficulty of clinical interpretation of ADA induction, caution must 21 be exercised when comparing ADA incidence between studies and the potential relevance for clinical 22 outcome.

24 ADAs induced by immune checkpoint inhibitors

25 In many tumour types, immune checkpoint inhibitors are now part of the standard of care. Humanized and 26 fully human antibodies against PD-L1 or PD-1 have had the most success to date, with five drugs approved 27 (nivolumab, pembrolizumab, atezolizumab, durvalumab, avelumab). Over a very short period, many trials 28 with very similar designs have been conducted with these drugs and, although their targets are similar, there 29 are some discrepancies in the results that are difficult to explain. As an example, in the subgroup of non- 30 small cell lung cancers (NSCLC) expressing PD-L1 on at least 50% of the tumour cells, pembrolizumab 31 significantly improved both progression-free survival and overall survival (OS) over platinum-based 32 chemotherapy, whereas nivolumab and durvalumab failed to demonstrate superiority (7–9). Similarly, in 33 second-line therapy in platinum-refractory advanced urothelial carcinoma, pembrolizumab was associated

1 with significantly longer OS than chemotherapy, but atezolizumab failed to reach this endpoint in a similar 2 scenario(10,11). Among the hypotheses put forward to explain these discrepancies, those highlighted 3 include, patient selection, tumour phenotype, dose and schedule selection, co-medication such as steroids, 4 or host characteristics (microbiota in particular)(12). It is also feasible that drug immunogenicity also 5 contribute to these conflicting results given that one of the main concerns in ADA development is their 6 clinical relevance and impact on drug efficacy.

7 ADAs have been analysed in various medical fields, with oncology being a particular point of focus(13). 8 Unfortunately, fewer than 50% of published oncology trials have analysed the possible effects of ADAs on 9 pharmacokinetics, efficacy, or safety parameters(14). The incidence of ADAs induced by immune 10 checkpoint inhibitors varies largely between drugs, as summarised in Fig. 2(15–26). Anti-nivolumab 11 antibodies were present in 11.2% of the 2085 patients analysed using the electrochemiluminescence (ECL) 12 immunoassay(17). Interestingly, in patients who were treated with nivolumab and ipilimumab, the presence 13 of anti-nivolumab antibodies was between 23.8% to 37.8% whereas anti-ipilimumab antibodies ranged from 14 4.1% to 8.4%. Agrawal et al (27) demonstrated the development of anti-nivolumab antibodies in 12.7% of 15 1086 patients with advanced melanoma or lung cancer from six phase 2/3 clinical studies using the ECL 16 immunoassay. For atezolizumab, pembrolizumab, durvalumab and avelumab, the maximum ADA-positive 17 rates reported were 54.1%, 2.1%, 2.9% and 5.9%, respectively (15,22,23,25). 18 It is well known that the intrinsic immunogenicity of monoclonal antibodies has been progressively reduced 19 from murine, chimeric to humanized and fully human antibodies, but even these latter two types of 20 antibodies cannot prevent the ADA development(28). Theoretically, fully human antibodies should have 21 significantly lower immunogenicity compared to humanized, but today this principle is still a matter of 22 debate since it could not be widely demonstrated in non-oncologic settings(29). This postulate is also not 23 valid for the immune checkpoints inhibitors considering the frequency of ADAs reported. Humanized Fc- 24 engineered atezolizumab presented higher ADAs frequency (54.1%) compared to the fully human 25 ipilimumab (5.4%), avelumab (5.9%) and durvalumab (2.9%). However, the humanized pembrolizumab 26 reported an ADA incidence of 2.1% compared to 11.2% with the fully human nivolumab. These results 27 demonstrate, with the limitations that all these studies used tests with different sensitivity, that the use of 28 humanised and/or fully human monoclonal antibodies in cancer patients is not systematically associated 29 with reduced induction of ADAs, as previously described in patients with inflammatory diseases(6).

31 Clinical relevance

32 In a study of single-agent ipilimumab (anti-CTLA-4 antibody) in 31 patients treated for melanoma, the 26% 33 of patients who developed ADAs by a bead-based assay, reflecting in this case anti-ipilimumab antibodies,

1 had significantly shorter OS, compared to the ADA-negative patients(30). The authors defined ADA 2 positivity based on both absolute levels and increase from baseline. Ipilimumab effectiveness is dependent 3 on the dose and the corresponding serum concentration, however this study did not demonstrate an 4 association between ADA positivity and low serum levels of ipilimumab. Only free forms of ADAs, as well 5 as ipilimumab, were detected since dissociation of ADA-drug complexes was not performed. Contrary to 6 ipilimumab, PD-1 and PD-L1 inhibitors do not have a clear certified dose-response association(31).

7 While in patients treated with nivolumab, pembrolizumab, durvalumab, or avelumab, there was no evidence 8 that ADAs had clinical consequences (21,23,25,27), atezolizumab is the only checkpoint inhibitor with an 9 expanded analysis of ADAs. The phase 3 OAK trial compared second-line atezolizumab to docetaxel in 850 10 NSCLC patients (32). In a subset analysis of 565 patients, 30% tested positive for ADAs at any post-dose 11 time point, and 21% after 4 weeks. These patients had a 25% higher drug clearance compared to ADA- 12 negative patients. The subset of ADA-positive patients did not have increased toxicity, however an 13 exploratory analysis suggested an impact on OS, with OS in ADA-positive patients similar to that in 14 docetaxel-treated patients (ADA-positive subgroup HR 0.89 [95% CI: 0.61-1.30]; ADA-negative subgroup 15 HR 0.68 [95% CI: 0.55-0.83]) (Fig. 3)(16). These data are not in line with the sister phase 2 study POPLAR 16 that also compared second-line atezolizumab to docetaxel in 287 NSCLC patients(33). When comparing the 17 response rate in a specific subgroup analysis of atezolizumab arm according to ADA status, it was found an 18 ORR of 20.5% (95% CI 12.0-31.6) in the ADA-positive subgroup (n=73), compared to 9.7% (95% CI 3.6- 19 19.9) in the ADA-negative population (n=62). This better response in the ADA-positive subset seems to 20 translate in a progression-free survival benefit, which was 4.1 months (95% CI 2.7-5.7) in ADA-positive 21 versus 2.7 months (1.5-4.2) in ADA-negative patients (15). A higher ORR in ADA-positive patients was 22 also found in the phase 2 FIR trial which evaluated the efficacy and safety of atezolizumab in a PD-L1– 23 selected population with NSCLC. The analysis of 31 patients in cohort 1 revealed an ORR of 31.3% in 24 ADA-positive patients compared to 20% in the ADA-negatives (15,34). In the phase 2 single-arm 25 IMvigor210 trial of atezolizumab in patients with locally advanced and metastatic urothelial carcinoma who 26 had progressed (cohort 2) or who were cisplatin-ineligible (cohort 1), 42% of the 275 patients, and 48% of 27 111 patients respectively, were positive for ADAs (16). Even though these patients also had lower systemic 28 exposure of atezolizumab, it had no impact on efficacy, since a subgroup analysis of cohort 2 found an 29 ORR of 19.3% (95% CI 12.5-27.8) in 114 ADA-positive patients compared to an ORR of 15.5% (95% CI 30 10.3-22.1) in 161 ADA-negative patients(15).

31 While these conflicting results call for prospective evaluations, it could be hypothesized that the difference 32 in clinical efficacy between ADA-positive and negative patients observed in the mentioned trials may 33 explain, at least partly, the lower atezolizumab response rate in advanced lung and urothelial carcinoma trials 34 compared to the other immune checkpoint inhibitors. In the phase 3 OAK trial, the objective response rate

1 (ORR) for atezolizumab was 14%, which is slightly lower than in other studies in NSCLC, including 2 nivolumab in the phase 3 CheckMate 017 (20%) and CheckMate 057 (19%), or with pembrolizumab in the 3 phase 2/3 Keynote-010 (18%) [21–24]. Similarly, in platinum-refractory advanced urothelial carcinoma 4 trials, the ORR with atezolizumab was 13.4% (phase 3 IMvigor211 trial), which was lower compared to 5 pembrolizumab (21.1% in the phase 3 KEYNOTE-045), nivolumab (19.6% in the phase 2 CheckMate 275), 6 durvalumab (17.8% in the phase 1/2 NCT01693562), or avelumab (18.2% in the phase 1b 7 JAVELIN)(10,11,38–40). With respect to cisplatin-ineligible patients, a similar tendency was observed for 8 atezolizumab which had a lower ORR (23% in IMvigor210 cohort 1) compared to pembrolizumab (27% in 9 the phase 2 KEYNOTE-052)(41,42). It should be highlighted that each patient can develop ADAs with 10 different effects (blocking the drug or not). Additionally, one could see the development of ADAs as a 11 surrogate marker of the immune system activation. Even though these provocative comparisons are not 12 supported statistically, all the above clinical observations can provide the background for possible 13 hypotheses of future prospective studies evaluating the clinical implication of ADA status when using 14 immune checkpoint inhibitors. Otherwise, one of the most worrying clinical relevance of ADAs is the 15 relationship with toxicity, mostly with infusion-related reactions. High titers of IgE ADAs, after the first 16 drug exposure, can develop a hypersensitivity reaction mediated by degranulation of histamine(43). 17 However, an increasing frequency of infusion-related reactions was not reported for the ADA-positive 18 patients treated with immune checkpoint inhibitors, revealing that this event may also be induced by a non- 19 antibody-mediated mechanism (non-ADA-dependent), such as the cytokine release syndrome(3).

20 21 22 ADAs in the context of chemo-immunotherapy

23 It could be hypothesized that the combination of immune checkpoint inhibitors with chemotherapy may 24 theoretically decrease the generation of ADAs due to the immunosuppressive effect of chemotherapy. This 25 was demonstrated in rheumatoid arthritis patients concomitantly treated with methotrexate and adalimumab 26 who had a reduced rate of ADA development(44). It is important to consider that low-dose injection of 27 methotrexate may have a different immunomodulatory role compared to doses used for cancer treatment. 28 The phase 3 Impassion130 trial, which combined atezolizumab and nab-paclitaxel in advanced triple- 29 negative breast cancer, supports this hypothesis. It was demonstrated a low incidence of ADA development 30 (13%) among 434 tested patients(15). Discordant results come from the phase 3 IMpower150 study that 31 evaluated chemotherapy (carboplatin and paclitaxel) plus bevacizumab with or without atezoliazumab in 800 32 nonsquamous NSCLC patients (arms B and C)(45). ADA positivity against atezalizumab, among 365 ADA- 33 evaluable patients, was 36% after the first or subsequent doses. An exploratory analysis on efficacy 34 according to ADA status was performed. The hazard ratio for OS in the ADA-positive subgroup was 0.69

1 (95% CI: 0.44, 1.07) comparing to 0.64 (95% CI: 0.46, 0.90) in the ADA-negative subgroup (Fig. 4)(16). The 2 subset of ADA-positive patients appeared to have similar efficacy as compared to the ADA-negative group, 3 but the confidence intervals in ADA-positive subgroup were wide. Some strategies to reduce ADA 4 formation by immune tolerance have been proposed outside cancer treatment, such as increasing the dose 5 and frequency of the therapy or adding corticosteroids(46,47). This last strategy would not seem to be a 6 possible alternative in cancer treatment since the immunosuppressive activity of corticosteroids may reduce 7 the antitumor activity and clinical efficacy of immune checkpoint inhibitors(48,49). The combination of 8 chemotherapy plus immunotherapy has been established as a standard of care in untreated advanced 9 NSCLC patients. Nonetheless, studies supporting this do not report data on ADA incidence, and their 10 clinical impact is unknown in this setting – highlighting a gap in our knowledge.

12 Conclusion

13 Despite the widespread use of immune checkpoint inhibitors in oncology, there is a lack of information 14 concerning their immunogenicity and its implications. Immunogenicity is very heterogeneous across 15 immune checkpoint inhibitors, and this can be explained by the absence of standardized methodology. 16 Although expanded reports of ADAs were made for atezolizumab, the data are conflicting and additional 17 analyses from published as well as prospective trials are required. A better understanding of the clinical 18 impact of ADAs may help to define therapeutic strategies, for example, by modifying the drug dose in the 19 event of ADA development, or adding chemotherapy to prevent ADA formation as was demonstrated in 20 rheumatology. For now, clinical implications of ADAs against immune checkpoint inhibitors remain to be 21 elucidated.

23 24 25 26 27 28 29 30 31 32 33

1 Acknowledgements 2 The authors thank to Sarah MacKenzie for English language edition. The authors received no specific 3 funding for this work. DE was the recipient of a DUERTECC/EURONCO grant for 2018-2019. 4 Sponsored Research at Gustave Roussy Cancer Center: Abbvie, Amgen, AstraZeneca, Biogen, Blueprint 5 Medicines, BMS, Celgene, Eli Lilly, GSK, Ignyta, IPSEN, Merck KGaA, MSD, Nektar, Onxeo, Pfizer, 6 Pharma Mar, Sanofi, Spectrum Pharmaceuticals, Takeda, Tiziana Pharma. 7 8 9 10 11 12 13 14

1 References 2 1. van Meer PJK, Kooijman M, Brinks V, Gispen-de Wied CC, Silva-Lima B, Moors EHM, et al. 3 Immunogenicity of mAbs in non-human primates during nonclinical safety assessment. MAbs. 4 2013;5:810–6.

5 2. Chirmule N, Jawa V, Meibohm B. Immunogenicity to Therapeutic Proteins: Impact on PK/PD and 6 Efficacy. AAPS J. 2012;14:296–302.

7 3. Vultaggio A, Nencini F, Pratesi S, Petroni G, Maggi E, Matucci A. Manifestations of Antidrug 8 Antibodies Response: Hypersensitivity and Infusion Reactions. J Interferon Cytokine Res. 9 2014;34:946–52.

10 4. Faraji F, Karjoo Z, Moghaddam MV, Heidari S, Emameh RZ, Falak R. Challenges related to the 11 immunogenicity of parenteral recombinant proteins: Underlying mechanisms and new approaches to 12 overcome it. Int Rev Immunol. 2018;1–15.

13 5. Kverneland AH, Enevold C, Donia M, Bastholt L, Svane IM, Nielsen CH. Development of anti-drug 14 antibodies is associated with shortened survival in patients with metastatic melanoma treated with 15 ipilimumab. Oncoimmunology. 2018;7:e1424674.

16 6. van Schouwenburg PA, Rispens T, Wolbink GJ. Immunogenicity of anti-TNF biologic therapies for 17 rheumatoid arthritis. Nat Rev Rheumatol. 2013;9:164–72.

18 7. Reck M, Rodríguez-Abreu D, Robinson AG, Hui R, Csőszi T, Fülöp A, et al. Pembrolizumab versus 19 Chemotherapy for PD-L1–Positive Non–Small-Cell Lung Cancer. New England Journal of Medicine. 20 2016;375:1823–33.

21 8. Carbone DP, Reck M, Paz-Ares L, Creelan B, Horn L, Steins M, et al. First-Line Nivolumab in Stage 22 IV or Recurrent Non–Small-Cell Lung Cancer. New England Journal of Medicine. 2017;376:2415– 23 26.

24 9. AstraZeneca provides update on the Phase III MYSTIC trial of Imfinzi and tremelimumab in Stage IV 25 non-small cell lung cancer [Internet]. [cited 2018 Nov 21]. Available from: 26 https://www.astrazeneca.com/media-centre/press-releases/2018/astrazeneca-provides-update-on-the- 27 phase-iii-mystic-trial-of-imfinzi-and-tremelimumab-in-stage-iv-non-small-cell-lung- 28 cancer16112018.html

29 10. Bellmunt J, de Wit R, Vaughn DJ, Fradet Y, Lee J-L, Fong L, et al. Pembrolizumab as Second-Line 30 Therapy for Advanced Urothelial Carcinoma. New England Journal of Medicine. 2017;376:1015–26.

31 11. Powles T, Durán I, van der Heijden MS, Loriot Y, Vogelzang NJ, De Giorgi U, et al. Atezolizumab 32 versus chemotherapy in patients with platinum-treated locally advanced or metastatic urothelial 33 carcinoma (IMvigor211): a multicentre, open-label, phase 3 randomised controlled trial. Lancet. 34 2018;391:748–57.

35 12. Agrawal S, Feng Y, Roy A, Kollia G, Lestini B. Nivolumab dose selection: challenges, opportunities, 36 and lessons learned for cancer immunotherapy. J Immunother Cancer. 2016;4:72.

37 13. Vennegoor A, Rispens T, Strijbis EM, Seewann A, Uitdehaag BM, Balk LJ, et al. Clinical relevance 38 of serum natalizumab concentration and anti-natalizumab antibodies in multiple sclerosis. Mult Scler. 39 2013;19:593–600.

1 14. van Brummelen EMJ, Ros W, Wolbink G, Beijnen JH, Schellens JHM. Antidrug Antibody Formation 2 in Oncology: Clinical Relevance and Challenges. Oncologist. 2016;21:1260–8.

3 15. European Medicines Agency, Committee for Medicinal Products for Human Use (CHMP). 4 Assessment report for Tecentriq (atezolizumab). 5 https://www.ema.europa.eu/en/documents/assessment-report/tecentriq-epar-public-assessment- 6 report_en.pdf. [Accessed 28 June 2019].

7 16. TECENTRIQ (atezolizumab) – FDA, https://www.gene.com/download/pdf/tecentriq_prescribing.pdf. 8 [Accessed 28 June 2019].

9 17. OPDIVO (nivolumab) – FDA, 10 https://www.accessdata.fda.gov/drugsatfda_docs/label/2019/125554s070lbl.pdf. [accessed 28 June 11 2019].

12 18. European Medicines Agency, Committee for Medicinal Products for Human Use (CHMP). 13 Assessment report for Opdivo (nivolumab). https://www.ema.europa.eu/en/documents/assessment- 14 report/opdivo-epar-public-assessment-report_en.pdf. [Accessed 28 June 2019].

15 19. Yervoy (ipilimumab) – FDA, https://packageinserts.bms.com/pi/pi_yervoy.pdf; [Accessed 28 June 16 2019].

17 20. European Medicines Agency, Committee for Medicinal Products for Human Use (CHMP). 18 Assessment report for Yervoy (ipilimumab). https://www.ema.europa.eu/en/documents/assessment- 19 report/yervoy-epar-public-assessment-report_en.pdf. [Accessed 28 June 2019].

20 21. BAVENCIO (avelumab) – FDA, 21 https://www.accessdata.fda.gov/drugsatfda_docs/label/2019/761049s006lbl.pdf. [Accessed 28 June 22 2019].

23 22. European Medicines Agency, Committee for Medicinal Products for Human Use (CHMP). 24 Assessment report for Bavencio (avelumab). https://www.ema.europa.eu/en/documents/assessment- 25 report/bavencio-epar-public-assessment-report_en.pdf. [Accessed 28 June 2019].

26 23. IMFINZI (durvalumab) – FDA, 27 https://www.accessdata.fda.gov/drugsatfda_docs/label/2018/761069s002lbl.pdf. [Accessed 28 June 28 2019].

29 24. European Medicines Agency, Committee for Medicinal Products for Human Use (CHMP). 30 Assessment report for Imfinzi (durvalumab). https://www.ema.europa.eu/en/documents/assessment- 31 report/imfizi-epar-public-assessment-report_en.pdf. [Accessed 28 June 2019].

32 25. KEYTRUDA (pembrolizumab) – FDA, 33 https://www.accessdata.fda.gov/drugsatfda_docs/label/2019/125514s040lbl.pdf. [Accessed 28 June 34 2019].

35 26. European Medicines Agency, Committee for Medicinal Products for Human Use (CHMP). 36 Assessment report for Keytruda (pembrolizumab). 37 https://www.ema.europa.eu/en/documents/assessment-report/keytruda-epar-public-assessment- 38 report_en.pdf. [Accessed 28 June 2019].

1 27. Agrawal S, Statkevich P, Bajaj G, Feng Y, Saeger S, Desai DD, et al. Evaluation of Immunogenicity 2 of Nivolumab Monotherapy and Its Clinical Relevance in Patients With Metastatic Solid Tumors. J 3 Clin Pharmacol. 2017;57:394–400.

4 28. Hwang WYK, Foote J. Immunogenicity of engineered antibodies. Methods. 2005;36:3–10.

5 29. Garcês S, Demengeot J. The Immunogenicity of Biologic Therapies. Curr Probl Dermatol. 6 2018;53:37–48.

7 30. Kverneland AH, Enevold C, Donia M, Bastholt L, Svane IM, Nielsen CH. Development of anti-drug 8 antibodies is associated with shortened survival in patients with metastatic melanoma treated with 9 ipilimumab. Oncoimmunology. 2018;7.

10 31. Centanni M, Moes DJAR, Trocóniz IF, Ciccolini J, van Hasselt JGC. Clinical Pharmacokinetics and 11 Pharmacodynamics of Immune Checkpoint Inhibitors. Clin Pharmacokinet. 2019;58:835–57.

12 32. Rittmeyer A, Barlesi F, Waterkamp D, Park K, Ciardiello F, von Pawel J, et al. Atezolizumab versus 13 docetaxel in patients with previously treated non-small-cell lung cancer (OAK): a phase 3, open-label, 14 multicentre randomised controlled trial. Lancet. 2017;389:255–65.

15 33. Fehrenbacher L, Spira A, Ballinger M, Kowanetz M, Vansteenkiste J, Mazieres J, et al. Atezolizumab 16 versus docetaxel for patients with previously treated non-small-cell lung cancer (POPLAR): a 17 multicentre, open-label, phase 2 randomised controlled trial. The Lancet. 2016;387:1837–46.

18 34. Spigel DR, Chaft JE, Gettinger S, Chao BH, Dirix L, Schmid P, et al. FIR: Efficacy, Safety, and 19 Biomarker Analysis of a Phase II Open-Label Study of Atezolizumab in PD-L1-Selected Patients 20 With NSCLC. J Thorac Oncol. 2018;13:1733–42.

21 35. Brahmer J, Reckamp KL, Baas P, Crinò L, Eberhardt WEE, Poddubskaya E, et al. Nivolumab versus 22 Docetaxel in Advanced Squamous-Cell Non–Small-Cell Lung Cancer. New England Journal of 23 Medicine. 2015;373:123–35.

24 36. Borghaei H, Paz-Ares L, Horn L, Spigel DR, Steins M, Ready NE, et al. Nivolumab versus Docetaxel 25 in Advanced Nonsquamous Non–Small-Cell Lung Cancer. New England Journal of Medicine. 26 2015;373:1627–39.

27 37. Herbst RS, Baas P, Kim D-W, Felip E, Pérez-Gracia JL, Han J-Y, et al. Pembrolizumab versus 28 docetaxel for previously treated, PD-L1-positive, advanced non-small-cell lung cancer (KEYNOTE- 29 010): a randomised controlled trial. Lancet. 2016;387:1540–50.

30 38. Sharma P, Retz M, Siefker-Radtke A, Baron A, Necchi A, Bedke J, et al. Nivolumab in metastatic 31 urothelial carcinoma after platinum therapy (CheckMate 275): a multicentre, single-arm, phase 2 trial. 32 Lancet Oncol. 2017;18:312–22.

33 39. Powles T, O’Donnell PH, Massard C, Arkenau H-T, Friedlander TW, Hoimes CJ, et al. Efficacy and 34 Safety of Durvalumab in Locally Advanced or Metastatic Urothelial Carcinoma: Updated Results 35 From a Phase 1/2 Open-label Study. JAMA Oncol. 2017;3:e172411.

36 40. Apolo AB, Infante JR, Balmanoukian A, Patel MR, Wang D, Kelly K, et al. Avelumab, an Anti- 37 Programmed Death-Ligand 1 Antibody, In Patients With Refractory Metastatic Urothelial Carcinoma: 38 Results From a Multicenter, Phase Ib Study. J Clin Oncol. 2017;35:2117–24.

1 41. Balar AV, Galsky MD, Rosenberg JE, Powles T, Petrylak DP, Bellmunt J, et al. Atezolizumab as 2 first-line treatment in cisplatin-ineligible patients with locally advanced and metastatic urothelial 3 carcinoma: a single-arm, multicentre, phase 2 trial. Lancet. 2017;389:67–76.

4 42. Balar AV, Castellano D, O’Donnell PH, Grivas P, Vuky J, Powles T, et al. First-line pembrolizumab 5 in cisplatin-ineligible patients with locally advanced and unresectable or metastatic urothelial cancer 6 (KEYNOTE-052): a multicentre, single-arm, phase 2 study. Lancet Oncol. 2017;18:1483–92.

7 43. Cheifetz A, Mayer L. Monoclonal antibodies, immunogenicity, and associated infusion reactions. Mt 8 Sinai J Med. 2005;72:250–6.

9 44. Krieckaert CL, Nurmohamed MT, Wolbink GJ. Methotrexate reduces immunogenicity in adalimumab 10 treated rheumatoid arthritis patients in a dose dependent manner. Ann Rheum Dis. 2012;71:1914–5.

11 45. Socinski MA, Jotte RM, Cappuzzo F, Orlandi F, Stroyakovskiy D, Nogami N, et al. Atezolizumab for 12 First-Line Treatment of Metastatic Nonsquamous NSCLC. N Engl J Med. 2018;378:2288–301.

13 46. Brackmann HH, Oldenburg J, Schwaab R. Immune tolerance for the treatment of factor VIII 14 inhibitors--twenty years’ “bonn protocol.” Vox Sang. 1996;70 Suppl 1:30–5.

15 47. Farrell RJ, Alsahli M, Jeen Y-T, Falchuk KR, Peppercorn MA, Michetti P. Intravenous 16 hydrocortisone premedication reduces antibodies to infliximab in Crohn’s disease: a randomized 17 controlled trial. Gastroenterology. 2003;124:917–24.

18 48. Libert C, Dejager L. How steroids steer T cells. Cell Rep. 2014;7:938–9.

19 49. Arbour KC, Mezquita L, Long N, Rizvi H, Auclin E, Ni A, et al. Impact of Baseline Steroids on 20 Efficacy of Programmed Cell Death-1 and Programmed Death-Ligand 1 Blockade in Patients With 21 Non-Small-Cell Lung Cancer. J Clin Oncol. 2018;36:2872–8.

22 50. Socinski MA, Jotte RM, Cappuzzo F, Orlandi FJ, Stroyakovskiy D, Nogami N, et al. Overall survival 23 (OS) analysis of IMpower150, a randomized Ph 3 study of atezolizumab (atezo) + chemotherapy 24 (chemo) ± bevacizumab (bev) vs chemo + bev in 1L nonsquamous (NSQ) NSCLC. JCO. 25 2018;36:9002–9002.

26 27 28 29 30 31 32 33 34 35 36

1 Figure 1. Generation of antidrug antibodies. Schematic process of the mechanism involved in the ADAs 2 formation after therapeutic antibody (or other biologic agent) administration. Therapeutic drug induces the 3 activation of the T-cell pathway following the participation of B-cells and finally plasma cells to produce 4 antibodies through the acquired immunity. MHC II, histocompatibility complex class II; TCR, T-cell receptor; 5 PD-L1, programmed death-ligand 1. 6 7 8 Figure 2. Highest published incidences of ADAs developing with different immune checkpoint 9 inhibitors. Data from U.S. Food and Drug Administration (FDA) and European Medicines Agency (EMA). 10 ADAs, anti-drug antibodies; n, total number of patients tested for ADA; NR, not reported. 11 12 13 Figure 3. Plot of hazard ratios for death. Analysis of overall survival and 95% confidence interval, according 14 to biomarker subgroups and ADA status, generated with data from the OAK clinical study in lung cancer 15 patients (NCT02008227)(16,31). ITT, intention to treat; ADAs, anti-drug antibodies; IC, tumour-infiltrating 16 immune cell; TC, tumour cell; CI, confidence interval. 17 18 19 Figure 4. Plot of hazard ratios for death. Analysis of overall survival and 95% confidence interval, according 20 to biomarker subgroups and ADA status, generated with data from the IMpower150 clinical study in lung 21 cancer patients (NCT02366143) (16,49). ITT-WT, intention to treat wild-type; ADAs, anti-drug antibodies; IC, 22 tumour-infiltrating immune cell; TC, tumour cell; BCP, bevacizumab, carboplatin, paclitaxel; CI, confidence 23 interval. 24 25

Anti-drug Antibodies Against Immune Checkpoint Blockers: Impairment of drug efficacy or indication of immune activation?

Diego Enrico, Angelo Paci, Nathalie Chaput, et al.

Clin Cancer Res Published OnlineFirst November 22, 2019.

Updated version Access the most recent version of this article at: doi:10.1158/1078-0432.CCR-19-2337

Author Author manuscripts have been peer reviewed and accepted for publication but have not yet Manuscript been edited.

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