Clinical proof of concept for a safe and effective NF‐κB‐targeting strategy in multiple myeloma (2018)

Despite the recent introduction of new treatments and improvements in clinical outcomes, multiple myeloma (MM) remains a medical problem. The majority of patients relapse and die from their disease, underscoring the need for new treatments (Kumar et al, 2017). The nuclear factor (NF)-κB pathway is aberrantly activated in virtually all cases of MM, where it enables tumour cell survival, and is therefore considered an effective therapeutic route in MM. Yet, following an aggressive 30-year effort by the pharmaceutical industry, no specific NF-κB inhibitor has been clinically approved, due to the dose-limiting toxicities associated with the general NF-κB suppression. Agents indicated in MM, e.g., proteasome inhibitors and immunomodulatory drugs, inhibit NF-κB, but also many other pathways, and neither specifically target cancer cells nor afford their clinical benefit through NF-κB inhibition (Greten et al, 2007; Di Donato et al, 2012; Begalli et al, 2017; Bennett et al, 2018). Therefore, there is a need for a fresh approach to safely block NF-κB signalling in MM. To overcome the barrier to safe NF-κB inhibition, we developed a novel class of NF-κB-targeting therapeutics. We identified the interaction between the JNK-activating kinase, MKK7, and the NF-κB-regulated antiapoptotic factor, GADD45β (De Smaele et al, 2001; Papa et al, 2004; Tornatore et al, 2014; Capece et al, 2018), as an essential, cancer-restricted survival module downstream of NF-κB in MM. Accordingly, GADD45β is selectively highly expressed in MM cells, where it denotes shorter patient overall survival (Tornatore et al, 2014). We developed the first-in-class GADD45β/MKK7 inhibitor, DTP3, which selectively kills MM cells by inducing MKK7/JNK-dependent apoptosis, ex vivo and in vivo, without toxicity to normal tissues (Tornatore et al, 2014). Further preclinical investigations demonstrated that DTP3 combines on-target-selective pharmacology and favourable drug-like properties with tolerability and none of the preclusive toxicities of conventional IKKβ/NF-κB-targeting agents (Tornatore et al unpublished observations). Due to this unique mode of action, DTP3 represents a significant opportunity for managing MM patients. The success rate for novel drug candidates in oncology from first-in-human trial to market registration is only ~5%, due to the heterogeneity of tumours and absence of drug-specific biomarkers, clearly demonstrating therapeutic target engagement (Kola & Landis, 2004). Considering that any GADD45β/MKK7-targeting drug would potentially benefit only a discrete subset of patients, we conducted a preclinical proof-of-concept study (REC 11/LO/1628) to develop a companion biomarker platform capable of assessing pharmacodynamic response in the earliest stages of the DTP3 clinical development. The current Letter reports the clinical proof-of-concept for an NF-κB-targeting strategy as a safe and mechanistically effective novel therapy in MM patients. We sought to first verify the cancer-selective mechanistic specificity of DTP3 in primary MM cells (Supplementary Methods). DTP3 was effective in inducing JNK phosphorylation, denoting therapeutic target engagement and caspase-3 cleavage, an apoptosis hallmark, in malignant CD138+ cells, ex vivo (Fig 1A). By contrast, DTP3 produced no such effects in healthy CD20+ cells nor in peripheral blood mononuclear cells (PBMCs) from the same patients, even at 30-fold higher concentrations (Fig 1B, C). Given the wide distribution and overall high levels of GADD45B expression in MM cells (Fig 1D, Supplementary Figure S1), we evaluated whether the cancer-selective pharmacodynamic response to DTP3 depended on GADD45B expression. As shown in Fig 1E, in 13 samples of malignant CD138+ cells which responded to DTP3, GADD45B expression was significantly higher than in any of the unresponsive CD138+-cell samples or any sample of normal CD20+ cells or PBMCs. Hence, the capacity of DTP3 to trigger JNK-driven apoptosis in primary MM cells correlates with a high significance with the GADD45B expression levels (Fig 1E). These results demonstrate that the on-target-specific therapeutic response to DTP3 can be monitored and potentially predicted by an objective measurement of GADD45B expression and JNK-dependent apoptosis in tumour cells. Interestingly, upon clinical validation, this approach could inform a patient stratification platform predictive of objective clinical response to support the clinical development of DTP3. Accordingly, we initiated the first-in-human phase-I/IIa trial of DTP3 to evaluate the safety, tolerability, pharmacokinetics and pharmacodynamics of this novel NF-κB-targeting therapeutic in patients with relapsed or refractory MM (EudraCT: 2015-003459-23; Supplementary Methods). Three single-patient, dose-escalation cohorts have been investigated to date, evaluating DTP3 at the dose levels of 0·5, 1 and 2 mg/kg, administered by rapid intravenous infusion three times a week (Fig 2A, Supplementary Figure S2). All patients had progressive disease at the point of entry into the study, despite having received multiple prior lines of therapy. All three patients completed their first 28-day treatment cycle with no complications. No reportable adverse effects, nor any grade 2 or greater toxicity were recorded at any point, in any patient, indicating that DTP3 was tolerated at all dose levels investigated (Fig 2A). Notably, Patient 002 demonstrated a reduction in serum free light-chain and paraprotein levels in response to DTP3, consistent with stable disease, according to the International Myeloma Working Group criteria. This patient was maintained on DTP3 therapy for a total of three 4-week treatment cycles before developing progressive disease, in the absence of any adverse effect (Fig 2A). To evaluate the pharmacodynamic response to DTP3, we analysed blood and bone marrow samples collected 18–36 h following administration of dose 4, on day 8 of treatment. Strikingly, Patient 001, who had been treated at the starting dose of 0·5 mg/kg, demonstrated a distinct shift in the signals corresponding to JNK phosphorylation and caspase-3 cleavage in MM (CD138+) cells, but, crucially, neither in normal CD20+ cells, nor in PBMCs concurrently isolated from blood (Fig 2B–D). As expected, no such signals denoting JNK activation or apoptosis were detected in MM cells, B lymphocytes or PBMCs isolated at screening, prior to treatment with DTP3 (Fig 2B–D). Hence, consistent with its ex-vivo mode of action and on-target-selective pharmacology (Fig 1A–C; Tornatore et al unpublished observations), DTP3 demonstrated the clinical capacity to trigger a cancer-specific pharmacodynamic response, inducing JNK-driven apoptosis in MM, but not normal cells (Fig 2B), thereby establishing clinical proof-of-mechanism. Notably, DTP3 produced these clinical effects in refractory MM patients, as a single agent, with a potential 40-fold margin for dose escalation in the trial. Collectively, the preclinical data and encouraging initial clinical results introduce DTP3 as a first-in-class NF-κB-targeting therapeutic with a novel mode of action in oncology. This work was supported in part by Cancer Research UK programme grant A15115, Medical Research Council (MRC) Biomedical Catalyst grant MR/L005069/1 and Bloodwise project grant 15003 to G.F., and Cancer Research UK Clinician Scientist Fellowship C41494/A15448 to H.W.A. JFA is a UK NIHR Senior Investigator. The infrastructure support for this study was generously provided by the NIHR CRF at Imperial College Healthcare NHS Trust. [Correction added on 25 October 2018, after first online publication: The preceding acknowledgement has been inserted in this current version.] J.F.A. and H.W.A. acknowledge the support of the Imperial College London NIHR Biomedical Research Centre (BRC). H.W.A. also acknowledges the Cancer Research UK Imperial Centre, and the Imperial Experimental Cancer Medicine Centre. G.F., L.T. and M.R. are named inventors on patents relating to this research. L.T. G.A., R.B., H.E.O., M.F.K., A.W. J.F.A. H.W.A. and G.F. designed experiments. L.T., D.C., F.B., D.V. and J.B. performed experiments. L.T., D.D., G.A., L.E.C., J.K., M.T., N.A., J.F.A.; H.W.A. and G.F. analysed data. L.T. G.A., L.E.C., J.K., M.T., N.A., S.B., J.F.A. H.W.A. and G.F. contributed to the preparation of the regulatory documentation for MHRI approval. A.L., A.S., D.R., M.R., A.C., M.O., M.J.A., R.S. K., I.G., R.B. and H.W.A. contributed clinical samples or key reagents. G.F. and L.T. wrote the paper. D.C., D.D. and H.W.A. contributed to writing the paper. Fig S1. Selective therapeutic target engagement by DTP3 in primary multiple myeloma cells expressing GADD45B. Fig S2. List of inclusion and exclusion criteria of the phase I trial (MHRA reference number: 19174/0369/001-0001). Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.