Killing of CLL and NHL cells by rituximab and ofatumumab under limited availability of complement

Okrój, Marcin; Eriksson, Ingrid; Österborg, Anders; Blom, Anna M.

doi:10.1007/s12032-013-0759-5

Cited by 25 publications

(33 citation statements)

References 19 publications

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“…Ofatumumab (originally designed as 2F2) was first reported in 2004 as a fully human anti-CD20 antibody, which outperformed rituximab in CDC activation [ 14 ]. These results were later confirmed by Beurskens et al, who tested both antibodies within a wide range of tumour cell load and serum concentration [ 10 ] and also by our studies, in which in vitro susceptibility to CDC under limited availability of complement of eighteen CD20 + cell lines and fresh CLL cultures was assessed [ 15 ]. We found that ratio of the target (CD20) to complement inhibitor CD55 distinguished cells highly sensitive to both anti-CD20 mAbs from those of moderate sensitivity, which were killed more efficiently by ofatumumab.…”

Section: Introductionsupporting

confidence: 85%

“…To confirm that alternative complement pathway is relevant to the amplification of anti-CD20 mAb-mediated complement activation, we performed CDC assay in serum with or without FB. Previously, we found Daudi cells to be highly sensitive to ofatumumab at low serum concentrations [ 15 ] and now we tested the same cells at two different, suboptimal concentrations of ofatumumab to better visualize the importance of alternative pathway. In parallel with increasing serum concentration, there was a trend towards higher CDC in FB-reconstituted serum.…”

Section: Resultsmentioning

confidence: 99%

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Mutations resulting in the formation of hyperactive complement convertases support cytocidal effect of anti-CD20 immunotherapeutics

Felberg

Urbán

Borowska

et al. 2019

Cancer Immunol Immunother

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Anti-CD20 monoclonal antibodies (mAbs) rituximab and ofatumumab are potent activators of the classical complement pathway, and have been approved for the treatment of B-cell malignancies. However, complement exhaustion and overexpression of complement inhibitors by cancer cells diminish their therapeutic potential. The strategies of targeting membrane complement inhibitors by function-blocking antibodies and the supplementation with fresh frozen plasma have been proposed to overcome tumour cell resistance. We present a novel approach, which utilizes gain-of-function variants of complement factor B (FB), a component of alternative C3/C5 convertases, which augment mAb-activated reactions through a positive feedback mechanism called an amplification loop. If complement concentration is limited, an addition of quadruple gain-of-function FB mutant p.D279G p.F286L p.K323E p.Y363A (or selected single mutants) results in significantly increased complement-mediated lysis of ofatumumab-resistant tumour cells, as well as the complete lysis of moderately sensitive cells. Importantly, this effect cannot be achieved by further increasing ofatumumab concentration. Potentiation of cytotoxic effect towards moderately sensitive cells was less apparent at physiological serum concentration. However, an addition of hyperactive FB could compensate the loss of cytotoxic potential of serum collected from the NHL and CLL patients after infusion of rituximab. Residual levels of rituximab in such sera, in combination with added FB, were able to efficiently lyse tumour cells. We suggest that the administration of gain-of-function variants of FB can restore cytotoxic potential of complement-exhausted serum and maximize the therapeutic effect of circulating anti-CD20 mAbs. Electronic supplementary material The online version of this article (10.1007/s00262-019-02304-0) contains supplementary material, which is available to authorized users.

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Section: Introductionsupporting

confidence: 85%

Section: Resultsmentioning

confidence: 99%

Mutations resulting in the formation of hyperactive complement convertases support cytocidal effect of anti-CD20 immunotherapeutics

Felberg

Urbán

Borowska

et al. 2019

Cancer Immunol Immunother

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“…Thus according to our ELISAs, there was not a single case of decrease in C4d, sC5b-6 or sC5b-9 after anti-CD20 treatment. As an explanation for increase of sC5b-9 in only 56% of the patients presenting increase of C4d, we propose the effect of membrane bound complement inhibitors expressed on the surface of tumor cells, which mainly target the level of complement convertases engaged in transition from opsonic to lytic stage (Manches et al, 2003;Okroj et al, 2013a).…”

Section: Measurement Of Soluble Markers Of Complement Activation In Pmentioning

confidence: 98%

“…Upon binding to target cell, anti-CD20 mAbs are expected to activate the classical complement pathway but it is not clear, which effector arm of the complement system contribute to the therapeutic effect (Okroj et al, 2013b). Also, it is not always the case that complement attack on tumor cells will proceed to the lytic stage, as malignant B cells may shed CD20-mAb complexes (Taylor and Lindorfer, 2014) and widely express the complement inhibitors CD46 and CD55 preventing C5 cleavage (Okroj et al, 2013a). Two samples were taken from each patient: one just before and the second immediately after CD20 mAb infusion.…”

Section: Measurement Of Soluble Markers Of Complement Activation In Pmentioning

confidence: 99%

Antibodies reactive to cleaved sites in complement proteins enable highly specific measurement of soluble markers of complement activation

Blom

Österborg

Mollnes

et al. 2015

Molecular Immunology

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Antibodies reactive to cleaved sites in complement proteins enable highly specific measurement of soluble markers of complement activation.Blom, Anna; Österborg, Anders; Mollnes, Tom E; Okroj, Marcin Link to publication Citation for published version (APA): Blom, A., Österborg, A., Mollnes, T. E., & Okroj, M. (2015). Antibodies reactive to cleaved sites in complement proteins enable highly specific measurement of soluble markers of complement activation. Molecular Immunology, 66(2), 164-170. DOI: 10.1016164-170. DOI: 10. /j.molimm.2015 General rights Copyright and moral rights for the publications made accessible in the public portal are retained by the authors and/or other copyright owners and it is a condition of accessing publications that users recognise and abide by the legal requirements associated with these rights.• Users may download and print one copy of any publication from the public portal for the purpose of private study or research.• You may not further distribute the material or use it for any profit-making activity or commercial gain • You may freely distribute the URL identifying the publication in the public portal AbstractAn emerging number of diseases and therapeutic approaches with defined involvement of the complement system justify a need for specific markers reflecting activation of particular effector arms of the complement cascade. Measurement of such soluble markers in circulation is a challenge since the specificity of antibodies must be limited to activated complement fragments but not predominant and ubiquitous parental molecules. Existing assays for the measurement of soluble, activated complement proteins are based on the detection of conformational neoepitopes. We tested an alternative approach based on detection of short linear neoepitopes exposed at the cleavage sites after activation of the actual complement component. Obtained antibodies reactive to C4d and C5b fragments enabled us to set up highly specific sandwich ELISAs, which ensured trustful measurements without false positive readouts characteristic for some of the widely used assays.

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“…Ofatumumab induces greater cell lysis than rituximab through increased CDC activity, primarily through greater activation of the classical complement pathway [31,32]. When compared with rituximab, the CDC activity of ofatumumab is augmented by the drug's ability to more efficiently utilize available complement; a nontrivial finding when taking into account the often low complement availability within the blood of CLL patients [33]. Ofatumumab is also able to induce similar ADCC as that seen with rituximab but more potent antibody-dependent cellular phagocytosis (ADCP) and greater direct cell killing than rituximab in CLL cells [26].…”

Section: Pharmacodynamicsmentioning

confidence: 99%

Ofatumumab as Front-Line Therapy in Untreated Chronic Lymphocytic Leukemia

Reagan

Castillo

2014

Future Oncol.

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Ofatumumab is a fully human IgG1 type I anti-CD20 monoclonal antibody that binds to both the small and large loop of the membrane antigen CD20. Much of its therapeutic efficacy is derived through complement-dependent cytotoxicity, although it also appears to operate via induction of caspase-dependent apoptosis and shows potent antibody-dependent cellular phagocytosis. CD20 is an important but sometimes difficult antigen to effectively target in chronic lymphocytic leukemia (CLL) secondary to its overall dim expression in CLL cells. Currently, ofatumumab is approved in the USA and EU for fludarabine- and alemtuzumab-refractory CLL patients. However, the experience with ofatumumab in untreated CLL patients is mounting and shows competitive response and survival rates with an acceptable adverse event profile. Herein, we outline the efficacy and toxicities of ofatumumab alone and in combination for the front-line treatment of CLL.

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Killing of CLL and NHL cells by rituximab and ofatumumab under limited availability of complement

Cited by 25 publications

References 19 publications

Mutations resulting in the formation of hyperactive complement convertases support cytocidal effect of anti-CD20 immunotherapeutics

Mutations resulting in the formation of hyperactive complement convertases support cytocidal effect of anti-CD20 immunotherapeutics

Antibodies reactive to cleaved sites in complement proteins enable highly specific measurement of soluble markers of complement activation

Ofatumumab as Front-Line Therapy in Untreated Chronic Lymphocytic Leukemia

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