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Editorials
and also induces apoptosis of CLL cells. In a phase I dose- escalation study, BI 836826 was well-tolerated up to doses of 400 mg and had a similar adverse event profile as other Fc-modified antibodies.10 The objective response rate was 61.5% in patients treated at doses ≥200 mg. Two antibody-drug conjugates and a radioimmunoconjugate targeting CD37 are also undergoing clinical investigation.5
Currently, 36 antibody-based cancer therapies approved by the US Food and Drug Administration (FDA), including 4 biosimilars, are on the market. The majority of these treatments are mAb in IgG1 format that mediate tumor cell killing on their own or in combination with chemotherapy. A key challenge has been the identi- fication of suitable targets for therapeutic mAb as tumor- specific antigens are rare, and tumor-associated antigens are often expressed on essential healthy cells, lowering the therapeutic index. By contrast, lineage-specific anti- gens that are expressed on non-essential healthy cells have emerged as preferred targets of therapeutic mAb. A prime example is CD20, which is expressed on healthy and malignant B cells, and targeted by rituximab (FDA approved in 1997), its biosimilar rituximab-abbs (in 2018), ofatumumab (in 2009), and obinutuzumab (in 2013) for treatment of B-cell malignancies. The same applies to other cell surface antigens, such as CD19, that are restrict- ed to the dispensable B-cell lineage of the hematopoietic system. The mechanism of action (MOA) by which mAb eradicate tumor cells include the induction of apoptosis by interfering with receptor/ligand interactions at the cell surface or by recruiting components of the innate immune system, such as plasma proteins in CDC, NK cells in ADCC, and macrophages in antibody-dependent cellular phagocytosis (ADCP).11 All three principle mech- anisms of innate immune system recruitment, collectively known as effector functions, involve the Fc fragment of IgG1, a homodimer comprising the hinge and the second (CH2) and third (CH3) constant domains of the heavy chain. To mediate CDC, ADCC, and ADCP, the Fc frag- ment interacts with complement protein C1q and FcγRIIIa and FcγRIIa receptors, respectively. It also medi- ates prolonged circulatory half-life through neonatal Fc receptor (FcRn) recycling. All of these mechanisms can be fine tuned by subjecting the Fc fragment to protein or car- bohydrate engineering.12 In fact, several of the FDA- approved mAb for cancer therapy have engineered Fc fragments.
Hexabodies constitute a new class of Fc fragment-engi- neered therapeutic antibodies.13,14 A single amino acid substitution in CH3, E430G, enhances the formation of IgG1 hexamers upon cell surface antigen binding (Figure 1A). As such, hexamerization, which was first discovered for membrane-bound wild-type IgG1,2 facilitates the docking of the hexavalent complement protein C1q initi- ating CDC. Indeed, previous studies revealed that CD20- targeting IgG1 with the E430G mutation mediate signifi- cantly enhanced CDC compared to the parental mAb.13,14 The current study by Oostindie et al.1 makes the same case for a CD37-targeting IgG1. In addition, combining hexameric (E430G) CD37-targeting IgG1 with one of the FDA-approved CD20-targeting IgG1 (rituximab, ofatu- mumab, or obinutuzumab) had a synergistic effect in terms of malignant B-cell lysis by CDC in vitro.
Intriguingly, the authors provide evidence that mixing CD20- and CD37-targeting IgG1 with E430G mutation leads to the formation of hetero-hexamers that are more potent in mediating CDC than the corresponding homo- hexamers on their own or in combination (Figure 1B). This finding is exciting as it suggests that two mAb that target two different cell surface antigens may form bispe- cific hetero-hexamers in the membrane, effectively lead- ing to target clustering and an increase in the density of C1q docking sites. It also sheds a light on a possible con- certed MOA of polyclonal antibodies which might form hetero-hexamers if they target different cell surface anti- gens or different epitopes of the same cell surface antigen. Collectively, the study makes a strong case for investigat- ing multispecific and multiparatopic biclonal, oligoclonal, and polyclonal antibodies for enhancing CDC compared to their parental mAb. Finding co-operative target combi- nations, such as CD20 and CD37 in the current study, that enable hetero-hexamer formation in the presence or absence of hexamerization-enhancing mutations is a key challenge in applying this concept to other hematologic malignancies and solid tumors. In this context, hexameric monoclonal and hetero-hexameric biclonal antibodies should also be tested for enhancing other effector func- tions in addition to CDC. While research into bispecific antibodies has accelerated, with a huge increase in the number of related clinical trials that are now ongoing,15 polyclonal antibodies16 may well be the next wave of antibody-based cancer therapy. Hetero-hexamerization in the membrane is a possible MOA of polyclonal anti- bodies in IgG1 format, providing an incentive to investi- gate their therapeutic utility with and without hexamer- ization-inducing mutations.
In summary, Oostindie et al.1 make a compelling case for further exploration of hexamer-forming antibodies and the combination of two, or possibly even more, tar- geting mAb. The advantages of antibody combinations might include not only increased cytotoxic activity, as described here, but possibly also better tumor-specific tar- geting and mitigation of tumor escape through antigen loss or target internalization.17 However, several aspects of this promising technology need further exploration. How effective is hexamer formation in vivo and what kind of het- ero-hexamers might be formed, especially in tissue sites? The current study is limited to in vitro studies with CLL cells in suspension. It is not immediately clear how these observations will translate to in vivo settings. Furthermore, hetero-hexamers may form in different ratios, some con- taining equal ratios of antibodies, while in others one anti- body may dominate. Will there be an optimal ratio and if so, could a desired composition be engineered into the antibody backbone? Translation of this promising approach into clinical trials may well constitute the next breakthrough in antibody therapy of B-cell malignancies. A first clinical trial with mAb engineered to facilitate hexam- erization is ongoing in solid tumors (clinicaltrials.gov identifi- er: NCT03576131). GEN1029 (also called HexaBody- DR5/DR5) consists of a mixture of two mAb that bind to different epitopes on DR5 and activate this death receptor to induce apoptosis. Results from this and other studies of hexamerization-enhanced mAb and mAb combinations are eagerly awaited.
haematologica | 2019; 104(9)