Zhiqiang An

HER3/ErbB3 has emerged as a new therapeutic target for cancer. Currently, more than a dozen anti-HER3 antibodies are in clinical trials for treatment of various cancers. However, limited understanding of the complex HER3 signaling in cancer and lack of established biomarkers have made it challenging to stratify cancer patients who can benefit from HER3 targeted therapies. In this study, we identified DJ-1/PARK7 (Parkinson Protein 7) as a novel interaction partner of HER3 and demonstrated the potential of DJ-1 as a biomarker for anti-HER3 cancer therapy. DJ-1 association with HER3 protects HER3 from ubiquitination and degradation through the proteasomal pathway in breast cancer cells. However, neuregulin 1 (NRG-1) mediated HER3 activation results in a reduced association of DJ-1 with HER3. DJ-1 shRNA knockdown in cancer cells resulted in decreased levels of HER3 and its downstream signaling through the PI3K/AKT and Ras/Raf/ERK pathways. DJ-1 shRNA knockdown cancer cells significantly...

Antibodies are an important component in host immune responses to viral pathogens. Because of their unique maturation process, antibodies can evolve to be highly specific to viral antigens. Physicians and researchers have been relying on such high specificity in their quest to understand host–viral interaction and viral pathogenesis mechanisms and to find potential cures for viral infection and disease. With more than 60 recombinant monoclonal antibodies developed for human use in the last 20 years, monoclonal antibodies are now considered a viable therapeutic modality for infectious disease targets, including newly emerging viral pathogens such as Ebola representing heightened public health concerns, as well as pathogens that have long been known, such as human cytomegalovirus. Here, we summarize some recent advances in identification and characterization of monoclonal antibodies suitable as drug candidates for clinical evaluation, and review some promising candidates in the development pipeline.

Studies have demonstrated that cancer-associated matrix metalloproteinases (MMPs) can generate single peptide bond cleavages in the hinge region of immunoglobulin G1 (IgG1). This study investigated the cleavage of endogenous IgGs by MMPs in the tumor microenvironment and the consequences of the IgG hinge cleavage for humoral immunity. We investigated the occurrence of single peptide bond cleaved IgGs (scIgGs) in tumor tissues and plasma samples collected from a cohort of breast cancer patients (n=60). Samples from healthy people (n=20) were used as the control. Antibody hinge cleavage was detected by multiple assays including IHC, ELISA, and flow cytometry. A correlation analysis was conducted between scIgG levels and patient clinical parameters. Levels of scIgGs in tumors were significantly higher than in normal tissues. In addition, scIgG levels in tumors were enriched compared to that in the plasma of the same patients. The appearance of scIgGs in tumor tissues was associated wit...

Nonhuman primates (NHPs) are used as a preclinical model for vaccine development, and the antibody profiles to experimental vaccines in NHPs can provide critical information for both vaccine design and translation to clinical efficacy. However, an efficient protocol for generating monoclonal antibodies from single antibody secreting cells of NHPs is currently lacking. In this study we established a robust protocol for cloning immunoglobulin (IG) variable domain genes from single rhesus macaque (Macaca mulatta) antibody secreting cells. A sorting strategy was developed using a panel of molecular markers (CD3, CD19, CD20, surface IgG, intracellular IgG, CD27, Ki67 and CD38) to identify the kinetics of B cell response after vaccination. Specific primers for the rhesus macaque IG genes were designed and validated using cDNA isolated from macaque peripheral blood mononuclear cells. Cloning efficiency was averaged at 90% for variable heavy (VH) and light (VL) domains, and 78.5% of the clo...

Trastuzumab has been used for the treatment of HER2-overexpressing breast cancer for more than a decade, but the mechanisms of action for the therapy are still being actively investigated. Ab-dependent cell-mediated cytotoxicity mediated by NK cells is well recognized as one of the key mechanisms of action for trastuzumab, but trastuzumab-mediated Ab-dependent cellular phagocytosis (ADCP) has not been established. In this study, we demonstrate that macrophages, by way of phagocytic engulfment, can mediate ADCP and cancer cell killing in the presence of trastuzumab. Increased infiltration of macrophages in the tumor tissue was associated with enhanced efficacy of trastuzumab whereas depletion of macrophages resulted in reduced antitumor efficacy in mouse xenograft tumor models. Among the four mouse FcγRs, FcγRIV exhibits the strongest binding affinity to trastuzumab. Knockdown of FcγRIV in mouse macrophages reduced cancer cell killing and ADCP activity triggered by trastuzumab. Consi...

ERBB3/HER3 is emerging as a molecular target for various cancers. HER3 is overexpressed and activated in a number of cancer types under the conditions of acquired resistance to other HER family therapeutic interventions such as tyrosine kinase inhibitors and antibody therapies. Regulation of the HER3 expression and signaling involves numerous HER3 interacting proteins. These proteins include PI3K, Shc, and E3 ubiquitin ligases NEDD4 and Nrdp1. Furthermore, recent identification of a number of HER3 oncogenic mutations in colon and gastric cancers elucidate the role of HER3 in cancer development. Despite the strong evidence regarding the role of HER3 in cancer, the current understanding of the regulation of HER3 expression and activation requires additional research. Moreover, the lack of biomarkers for HER3-driven cancer poses a big challenge for the clinical development of HER3 targeting antibodies. Therefore, a better understanding of HER3 regulation should improve the strategies t...

Aberrant expression and activation of EGFR and ERBB2 (HER2) have been successfully targeted for cancer therapeutics. Recent evidence from both basic and clinical studies suggests that ERBB3 (HER3) serves as a key activator of downstream signaling through dimerization with other ERBB proteins and plays a critical role in the widespread clinical resistance to EGFR and HER2 targeting cancer therapies. As a result, HER3 is actively pursued as an antibody therapeutic target for cancer. Ligand binding is thought to be a prerequisite for dimerization of HER3 with other ERBB proteins, which results in phosphorylation of its c-terminal tyrosine residues and activation of downstream AKT and MAPK signaling pathways. In this study, we report that an anti-HER2 monoclonal antibody (HER2Mab), which blocks HER2 dimerization with HER3, induces HER3 dimerization with EGFR in both low and high HER2 expressing cancer cells. Treatment of the low HER2 expressing MCF7 cancer cells with HER2Mab promoted ce...

Dimerization among the EGFR family of tyrosine kinase receptors leads to allosteric activation of the kinase domains of the partners. Unlike other members in the family, the kinase domain of HER3 lacks key amino acid residues for catalytic activity. As a result, HER3 is suggested to serve as an allosteric activator of other EGFR family members which include EGFR, HER2 and HER4. To study the role of intracellular domains in HER3 dimerization and activation of downstream signaling pathways, we constructed HER3/HER2 chimeric receptors by replacing the HER3 kinase domain (HER3-2-3) or both the kinase domain and the C-terminal tail (HER3-2-2) with the HER2 counterparts and expressed the chimeric receptors in Chinese hamster ovary (CHO) cells. While over expression of the intact human HER3 transformed CHO cells with oncogenic properties such as AKT/ERK activation and increased proliferation and migration, CHO cells expressing the HER3-2-3 chimeric receptor showed significantly reduced HER...

Primary and acquired resistance to anti-cancer antibody immunotherapies presents significant clinical challenges. Here we demonstrate that proteolytic inactivation of cancer-targeting antibodies is an unappreciated contributor to cancer immune evasion and the finding presents novel opportunities for therapeutic intervention. A single peptide bond cleavage in the immunoglobulin G1 (IgG1) hinge impairs cancer cell killing due to structural derangement of the Fc region. Hinge-cleaved trastuzumab gradually accumulated on the surfaces of HER2-expressing cancer cell lines in vitro, and was greatly accelerated when the cells were engineered to express the potent bacterial IgG-degrading proteinase (IdeS). Similar to cancer-related matrix metalloproteinases (MMPs), IdeS exposes a hinge neo-epitope that we have developed an antibody, mAb2095-2, to specifically target the epitope. In in vitro studies, mAb2095-2 restored the lost ADCC functionality of cell-bound single-cleaved trastuzumab (scIg...

HER3 is a member of the HER (EGFR/ErbB) receptors family consisting of four closely related type 1 transmembrane receptors (EGFR, HER2, HER3, and HER4). HER receptors are part of a complex signaling network intertwined with the Ras/Raf/MAPK, PI3K/AKT, JAK/STAT, and PKC signaling pathways. Aberrant activation of the HER receptors and downstream signaling molecules tips the balance on cellular events, leading to various types of cancers. Monoclonal antibodies (mAbs) and small molecule inhibitors targeting EGFR and HER2 tyrosine kinase activities exhibit clinical benefits in the treatment of several types of cancers, but their clinical efficacy is limited by the occurrence of drug resistance. HER3 is the preferred dimerization partner of HER2 and it is well established that HER3 plays an important role in drug resistance to EGFR- and HER2-targeting therapies. Since HER3 has limited kinase activity, mAbs are being explored to target HER3 for cancer therapy. Currently, approximately a dozen of anti-HER3 mAbs are at different stages of clinical development. However, the lack of established biomarkers has made it more challenging to stratify cancer patients to whom HER3-targeting therapies can be more effective. In this review, we focus on the validation of HER3 as a cancer drug target, the recent development in biomarker discovery for anti-HER3 therapies, and the progress made in the clinical development of HER3-targeting mAbs.