Per Norlén

The human agonistic CD40 antibody ADC-1013 eradicates bladder tumors and generates T cell dependent tumor immunity

Purpose: Local administration of immune-activating antibodies may increase the efficacy and reduce the immune-related adverse events associated with systemic immunotherapy of cancer. Here, we report the development and affinity maturation of a fully human agonistic CD40 antibody (IgG1), ADC-1013. Experimental Design: We have used molecular engineering to generate an agonistic antibody with high affinity for CD40. The functional activity of ADC-1013 was investigated in human and murine in vitro models. The in vivo effect was investigated in two separate bladder cancer models, both using human xenograft tumors in immune deficient NSG mice and using a syngeneic bladder cancer model in a novel human CD40 transgenic mouse. Results: Activation of dendritic cells (DC) by ADC-1013 results in upregulation of the costimulatory molecules CD80 and CD86, and secretion of IL12. ADC-1013 also activates DCs from human CD40 transgenic mice, and peptide-pulsed and ADC-1013–stimulated DCs induce antigen-specific T-cell proliferation in vitro. In vivo, treatment with ADC-1013 in a syngeneic bladder cancer model, negative for hCD40, induces significant antitumor effects and long-term tumor-specific immunity. Furthermore, ADC-1013 demonstrates significant antitumor effects in a human bladder cancer transplanted into immunodeficient NSG mice. Conclusions: Our data demonstrate that ADC-1013 induces long-lasting antitumor responses and immunologic memory mediated by CD40 stimulation. To the best of our knowledge, ADC-1013 represents the first immunomodulatory antibody developed for local immunotherapy of cancer. Clin Cancer Res; 21(5); 1115–26. ©2014 AACR. See related commentary by Dronca and Dong, p. 944

Kick-starting the cancer-immunity cycle by targeting CD40

Stimulation of CD40 on dendritic cells to expand and activate tumor-specific T cells and generate anticancer immunity is an attractive therapeutic approach. Since CD40 agonists exert their effects upstream of checkpoint inhibitors, including PD-1 or PD-L1 antagonists, they are ideal candidates for combination regimens.

Tumor-directed immunotherapy can generate tumor-specific T cell responses through localized co-stimulation

The most important goals for the field of immuno-oncology are to improve the response rate and increase the number of tumor indications that respond to immunotherapy, without increasing adverse side effects. One approach to achieve these goals is to use tumor-directed immunotherapy, i.e., to focus the immune activation to the most relevant part of the immune system. This may improve anti-tumor efficacy as well as reduce immune-related adverse events. Tumor-directed immune activation can be achieved by local injections of immune modulators in the tumor area or by directing the immune modulator to the tumor using bispecific antibodies. In this review, we focus on therapies targeting checkpoint inhibitors and co-stimulatory receptors that can generate tumor-specific T cell responses through localized immune activation.

ADC-1013, an agonistic CD40 antibody optimized for local immunotherapy of cancer

Local administration of immune activating antibodies may increase the efficacy and reduce the immune-related adverse events associated with systemic immunotherapy of cancer. Here we report the development of a fully human agonistic CD40 antibody (IgG1), ADC-1013, which has been optimized for local immunotherapy by increasing potency and tumor retention. ADC-1013 activates CD40 receptors on antigen-presenting cells such as dendritic cells, resulting in up-regulation of the co-stimulatory molecules CD80 and CD86, and induction of IL-12. In addition, ADC-1013 induces direct tumor killing of CD40+ tumors, e.g. via antibody-dependent cellular cytotoxicity (ADCC). The anti-tumor effects of ADC-1013 were first assessed in a bladder cancer model (EJ) in immunodeficient NSG mice. Significant anti-tumor responses were demonstrated, and further augmented in mice repopulated with human moDCs/T cells. To study the anti-tumor effects related to the immune activating properties of ADC-1013 in more detail, a human CD40 positive transgenic mouse (hCD40tg) in C57/BL-6 background was used. This transgenic mouse strain has an intact immune system and fully functional dendritic cells that are activated upon ADC-1013 treatment. Furthermore, the dendritic cells obtained from this strain are able to induce antigen specific T cell activation in vitro upon stimulation with ADC-1013. Importantly, treatment with ADC-1013 in a syngeneic bladder cancer (MB49) model, which is hCD40 negative, demonstrated that ADC-1013 induce significant tumor protection and long term immunity independent of direct tumor targeting. In addition, the anti-tumor immunity was shown to be T-cell dependent. To our knowledge, ADC-1013 represents the first immunomodulatory antibody optimized for local immunotherapy of cancer. It is currently in late pre-clinical development and will enter clinical trials in 2014.

Bispecific antibodies in cancer immunotherapy

Following the clinical success of immune checkpoint antibodies targeting CTLA-4, PD-1 or PD-L1 in cancer treatment, bispecific antibodies are now emerging as a growing class of immunotherapies with potential to further improve clinical efficacy and safety. We describe three classes of immunotherapeutic bispecific antibodies: (a) cytotoxic effector cell redirectors; (b) tumor-targeted immunomodulators; and (c) dual immunomodulators. Cytotoxic effector cell redirectors are dominated by T-cell redirecting compounds, bispecific compounds engaging a tumor-associated antigen and the T-cell receptor/CD3 complex, thereby redirecting T-cell cytotoxicity to malignant cells. This is the most established class of bispecific immunotherapies, with two compounds having reached the market and numerous compounds in clinical development. Tumor-targeted immunomodulators are bispecific compounds binding to a tumor-associated antigen and an immunomodulating receptor, such as CD40 or 4-1BB. Such compounds are usually designed to be inactive until binding the tumor antigen, thereby localizing immune stimulation to the tumor environment, while minimizing immune activation elsewhere. This is expected to induce powerful activation of tumor-specific T cells with reduced risk of immune-related adverse events. Finally, dual immunomodulators are bispecific compounds that bind two distinct immunomodulating targets, often combining targeting of PD-1 or PD-L1 with that of LAG-3 or TIM-3. The rationale is to induce superior tumor immunity compared to monospecific antibodies to the same targets. In this review, we describe each of these classes of bispecific antibodies, and present examples of compounds in development.

Pre-clinical development of the human CD40 agonistic antibody ADC-1013.

Activation of CD40 on dendritic cells increases cross-presentation of tumor antigens and as a result the number of activated tumor directed T effector cells. CD40 agonists have been shown to generate powerful systemic anti-tumor responses when administered in situ. The pre-clinical proof of concept data has been generated mainly using an hCD40 transgenic mouse (hCD40tg) strain. Anti-tumor effects has been demonstrated in multiple syngeneic tumor models including bladder tumor (MB49), melanoma (B16) and lymphoma (A20) models. The anti-tumor effects has been shown to induce long term, T cell dependent, tumor immunity. As expected, the anti-tumor effects depends on the immune status of the tumors as evaluated by immunohistochemistry and flow cytometry. Since CD40 agonistic antibodies mainly exert their effects upstream of the checkpoint inhibitors they have the potential to be ideal candidates for combination regimens including e.g. PD-1 antagonists and OX40 antibodies. Combining ADC-1013 with antibodies against these targets has been shown to improve the anti-tumor effects in B16.F10 melanoma. Toxicology studies were performed in cynomolgus monkeys using subcutaneous or intravenous administration. Doses of 1 to 10 mg/kg were evaluated in both studies. ADC-1013 was well tolerated at all dose levels and no major safety concerns were identified. Receptor saturation of approximately 95% was achieved and pharmacodynamic activity, notably B-cell depletion, was observed at all dose levels. In cynomolgus monkey, serum concentrations of ADC-1013 declined in a multi-phasic manner following a single intravenous dose. The mean terminal half-life was approximately 30 hours. Following subcutaneous administration the mean elimination half-life was found to be between 0.6 and 3.2 days. The clearance (CL_F) decreased and the half-life increased with increasing dose indicating target-mediated clearance. Alligator Bioscience has recently started dosing of the first patients in a clinical trial of the CD40 agonistic antibody ADC-1013. The study is a first-in-human, multicenter, open-label, multiple ascending dose Phase I study in patients with advanced solid tumors to determine the safety, pharmacokinetics and pharmacodynamics of intratumorally administered ADC-1013.

The human CD40 agonistic antibody ADC-1013 generates immune mediated anti-tumor effects in syngeneic tumor models in hCD40 transgenic mice

Local activation of costimulatory pathways by e.g. CD40 activation has been shown to generate powerful systemic anti-tumor responses. Here we report significant anti-tumor effects obtained with an optimized fully human agonistic CD40 antibody, ADC-1013, in two syngeneic tumor models. An hCD40 transgenic mouse (hCD40tg) strain was used to evaluate the immune mediated anti-tumor effects of ADC-1013. Dendritic cells obtained from hCD40tg mice were hCD40 positive and could be activated by stimulation with ADC-1013 to a similar extent as human monocyte derived dendritic cells. Furthermore, stimulation of dendritic cells from hCD40tg mice in vitro, with ADC-1013, induced antigen specific T cell activation. Two different syngeneic tumor models, hCD40 negative MB49 bladder cancer and hCD40 positive B16.F10.hCD40+ melanoma, was used to demonstrate anti-tumor effects. Subcutaneous tumors from both models were characterized by flow cytometry and immunohistochemistry in hCD40tg mice. Treatment of the bladder cancer model (MB49) with ADC-1013 resulted in significant anti-tumor response and long term tumor immunity. The anti-tumor immunity was shown to be T cell dependent and naive mice were protected from tumor challenge by transplantation of splenocytes from cured hCD40tg mice. In addition, significant anti-tumor effect was demonstrated in a subcutaneous B16.F10.hCD40+ melanoma model. The human CD40 agonistic antibody ADC-1013 is the first costimulatory antibody optimized for local immunotherapy of cancer. Strong immune mediated anti-tumor effects were demonstrated. ADC-1013 is currently in late pre-clinical development.