Abstract: Cancer immunotherapy attempts to exploit the capability of the immune system to attack malignant cells. Recent results suggest that clinical responses in patients point to this new mechanism as potentially beneficial in harnessing the immune system for combating established malignancies. These checkpoint-related immunotherapies rely on engaging a subset of T cells in anti-tumor immune responses. BiTE® (Bi-specific T cell engager) represents a distinct modality that directly engages any T cell and a specific antigen expressing tumor cell. The approach offers the advantage of engaging T cells and patient tumor cells that differentially express a specific cell surface antigen. The specificity confers redirected tumor cell killing and recent clinical data with the BiTE blinatumomab show evidence of clinical remissions. The characteristics of a suitable BiTE with the benefit of CD3 mediated T cell recognition and articulation of tumor specific antigens combined in this therapeutic modality is described here.
Cancer therapy has emerged as a clinical challenge which reflects the complexity of the disease and its etiology with morbidity resulting due to resistance and relapse. While the field has enjoyed some significant improvement in a subset of cancers with patient responses, there is still a large unmet need and a necessity to evaluate new therapeutic paradigms. In the quest to identify therapies that may provide durable responses, investment in cancer immunotherapeutics is yielding an emerging understanding in this relatively new area of investigation. The attempt to harness a patient’s immune system in the treatment of malignancy is forging new paths. Modulation of adaptive and innate immunity provides a therapeutic opportunity for patients with the advent of therapeutics targeting CTLA4 and PD-1/PDL-1 (Hamid et al., 2013; Sharma et al., 2011; Topalian et al., 2011; Wolchok et al., 2013). An alternative mechanism to harnesses the patient’s immune cells in a directed manner such as BiTE® (Bi-specific T cell Engager) and CAR (Chimeric Antigen Receptors) have also emerged. The focus of this review is to discuss the BiTE approach and the properties that engage T cells directly and the targets amenable to such approaches.
The BiTE technology represents the combination of exquisite antibody based selective target recognition and direct engagement of the highly cytotoxic capability of T cells. BiTE therapeutics are currently being evaluated in hematologic and solid tumor malignancies (Amann et al., 2009; Baeuerle and Reinhardt, 2009; Bargou et al., 2008). Several BiTE programs have entered the clinic and published results from these programs show evidence of clinical remissions (Frankel and Baeuerle, 2013; Nagorsen et al., 2012).
BiTE technology embodies a novel molecule with three major components consisting of T cell receptor (TCR) recognition and tumor specific antigen recognition moieties tethered by an optimal linker configuration (Figure 1A-1C). BiTE molecules form a cytolytic synapse between tumor cells and T cells bringing them into close proximity. The resultant activation of T cells elicits a cytolytic response by driving a highly effective target cell killing machinery (Brischwein et al., 2006; Kufer et al., 2004; Mack et al., 1995). This direct engagement of tumor and T cells in the presence of a BiTE molecule can be modulated with delivery of a therapeutic dose and drug withdrawal. To utilize this new mechanism of tumor and T cell engagement, some specific T cell and target selection criteria are important in addition to the optimized linker region.
T Cell Recognition by BiTE®
Direct T cell recognition of BiTE molecules is conferred by single chain Fv (scFv) recognizing CD3ε, the invariant signaling component of the T cell receptor complex. This takes advantage of the ability to harness polyclonal populations of cytotoxic T cells [CD8+, CD4+, including regulatory T cells (Tregs)] and is not dependent on MHC class I for antigen presentation (Offner et al., 2006). The monovalent binding of BiTE molecules to CD3 does not activate the TCR unless target cell binding is engaged. T cell costimulation seems not to be required (Dreier et al., 2002). BiTE engagement of target cells drives activation and resultant proliferation of the T cells which contributes to enhanced efficacy. Moreover BiTE activated T cells contribute to serial lysis which further enhances efficacy (Hoffmann et al., 2005). This mechanism of action is consistent with BiTE molecules being highly potent drivers of redirected target cell lysis. As BiTE molecules are highly potent with demonstrated improved efficacy, the data suggest that low dose treatment should elicit an efficacious response. The cell based EC50 of blinatumomab (AMG 103) is 0.01-0.1 ng/ml for target cell lysis. Clinical observations of blinatumomab treatment in acute lymphoblastic leukemia (ALL) patients who receive 15 µg/m2/kg as the daily dose support this thinking (Topp et al., 2011). In addition the ability to control drug delivery of BiTE therapeutics provides a distinct advantage of obtaining optimal efficacy and managing any potential toxicity.
The ability of BiTE molecules to broadly engage CD3 positive T cells and elicit tumor directed cytotoxicity is in contrast to recruiting T cell subsets dependent on the tumor immune environment for immune modulation and tumor cell targeting. This mode of polyclonal T cell engagement may potentially overcome escape mechanisms of tumor cells evading T cell recognition commonly observed in cancer (Holzel et al., 2013). Immune editing with immune selection of tumor cell variants that escape T cell mediated cytotoxicity may be overcome by this redirected mechanism of T cell engagement.
Target Selection Criteria for BiTE Strategy
Differential target expression in tumor compared to normal tissue offers an advantage in selecting targets for BiTE molecules. This enables an increased therapeutic window as highly overexpressed antigens on tumors will likely display better recognition by BiTE molecules in contrast to normal tissue which has lower or no expression. This is not unique to BiTE target selection alone and is used in guiding target selection for other modalities such as Antibody Drug Conjugates (ADC) as well although internalization of the target bound ADC distinguishes the mechanisms of action. EGFRviii (a mutant epidermal growth factor receptor) and PSMA (prostate-specific membrane antigen) exemplify such differentially expressed targets and representative programs are currently in clinical development.
Expression of targets in tumor and normal tissue with non-essential or regenerative capability is another opportunity for exploration and application of BiTE technology. For example CD19 is expressed on tumor and normal B cells. The BiTE therapeutic blinatumomab in clinical trials for ALL demonstrates efficacy in B cell malignancy while elimination of the normal CD19 expressing B cells is tolerated. The ability to modulate drug exposure, as in the case of BiTE therapeutics, is important in addressing these types of target expression patterns where the normal cell compartment has the capability to regenerate. Potentially, targets expressed in normal tissues such as prostate, breast, ovarian, and endometrium could be suitable for BiTE generation despite similar target expression in tumor and normal tissue alike. Expression in essential tissues such as pancreatic, lung, liver, etc., should be more thoughtfully evaluated as the effect on these tissues may bring toxicities that could be difficult to manage in the clinic.
As BiTE molecules are highly potent the opportunity arises to prosecute targets with lower levels of differential expression not amenable to ADC approaches. ADC usually requires thousands of targets per cell to drive on target cytotoxicity. BiTE molecules are thought to be effective in killing tumor cells with very low target numbers. This potentially influences the target repertoire that could be addressed with BiTE therapeutics.
It is worth noting that the BiTE mechanism is independent of target internalization as cytotoxic activity is elicited by engagement of T cells at the cell surface and not via a mechanism of internalization and release of the warhead as in ADC approaches. Moreover, target internalization does not impact BiTE activity likely due to the prevalence of sufficient target numbers at the cell surface. In this regard, while EGFR is internalized EGFR BiTE molecules are capable of eliciting cytolytic activity in tumor cells (Lutterbuese et al., 2010). The ability to prosecute both internalized and non-internalized targets expands the types of cell surface proteins that can be pursued with BiTE technology. Additionally, BiTE therapeutic efficacy is not dependent on the tumor cell cycle to deliver warhead mediated cytotoxicity. For example microtubule inhibitors in ADC, such as mertansine derivatives or auristatin drugs in approved ADC therapeutics (or currently in clinical trials) depend on tumor cells entering mitosis for efficacy (Polakis, 2005).
Overall, the BiTE technology confers some distinct advantages in addressing expressed tumor cell surface targets as described here. It is worth underscoring the highly effective cytolytic activity of T cells that drive efficacy. The balance of enhanced efficacy with toxicity is currently being evaluated in ongoing clinical studies. While toxicities in the advanced clinical programs are manageable, ongoing and future studies will expand our understanding of therapeutic window using this novel T cell engaging approach. The exploration of the BiTE platform offers future prospects for single agent and combination use with signaling inhibitors and other immunotherapeutics which lack durable responses. The potential opportunity to combat the polyclonal, polygenic, and heterogeneous nature of cancer with BiTE molecules to overcome relapse and resistance is indeed an exciting prospect.
D.W. is a full time employee of Amgen Inc.
Dineli Wickramasinghe, Ph.D., Executive Director, Oncology Research, Amgen Inc., South San Francisco, California 94080, USA.
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