CD55 and CD59 Can Limit the Anti-Tumor Efficacy of Daratumumab in Natural Killer/T-Cell Lymphoma

Complement-dependent cytotoxicity (CDC) is one of the major mechanisms mediating the anti-tumor efficacy of Daratumumab. We have previously demonstrated that a majority of Natural Killer/T- Cell Lymphoma (NKTL) patient samples express CD38 and Daratumumab is highly effective against NKTL cell lines expressing mid-high levels of CD38. In this report we show that subsequent testing in an NKTL mouse xenograft model confirms the potency of Daratumumab in vivo as evidenced by the inhibition in tumour progression and prolongation of mouse survival. When treatment was continued over a month, some tumors began to rapidly enlarge ('Resistant') while the rest remained similar or smaller ('Sensitive') than the tumour volume at the initiation of Daratumumab treatment. An mRNA analysis comparing 'Resistant' and ‘Sensitive’ tumors showed that while both tumours bore similar levels of CD38 expression, resistant tumours displayed an upregulation of complement inhibitory proteins (CIP), CD55 and CD59 but not CD46. This led us to hypothesize that CD59 and CD55 may play a critical role in Daratumumab-mediated CDC in NKTL. FACS analyses demonstrated that the number of membrane molecules of CD55 and CD59 appeared inversely correlated to Daratumumab-mediated CDC. A single CIP knockdown was first performed to delineate the role of each CIP. Silencing CD46 confirmed that it does not have any effect on CDC in NKTL. However, single knockdown of CD55 or CD59 was able to induce cytotoxicity in CDC-resistant cell line CD38midCD55hiCD59lo NKYS, and promote NKS1 CD38hiCD55hiCD59mid to further lysis. Both single and double knockdown of CD55 and CD59 could not enhance Daratumumab-induced CDC in CD38loCD55hiCD59hi HuT78 which recapitulates the importance of CD38 levels. Unexpectedly, the double knockdown did not sensitize CD38hiCD55hiCD59hi KMS12BM either. This led us to conjecture that it may be the ratio of CD38:CIPs which is predictive of response to Daratumumab than CD38 or CIPs alone. All-Trans Retinoic Acid (ATRA) binds the RARE element in CD38 gene leading to upregulation of mRNA and protein expression of CD38. We thus downregulated the expression of CIPs with siRNA followed by amplification of CD38 expression with ATRA in NKTL. This strategy resulted in a significant increase in the CD38:CIP ratio and induced almost a total lysis of NKS1 cells, as well as sensitised HuT78 to a massive amount of Daratumumab-mediated CDC. These experiments suggest that by increasing the CD38: CIP, ra