Riyao Yang - Academia.edu (original) (raw)

Papers by Riyao Yang

Scientific Reports

The landscape of current cancer immunotherapy is dominated by antibodies targeting PD-1/PD-L1 and... more The landscape of current cancer immunotherapy is dominated by antibodies targeting PD-1/PD-L1 and CTLA-4 that have transformed cancer therapy, yet their efficacy is limited by primary and acquired resistance. The blockade of additional immune checkpoints, especially TIGIT and LAG-3, has been extensively explored, but so far only a LAG-3 antibody has been approved for combination with nivolumab to treat unresectable or metastatic melanoma. Here we report the development of a PDL1 × TIGIT bi-specific antibody (bsAb) GB265, a PDL1 × LAG3 bsAb GB266, and a PDL1 × TIGIT × LAG3 tri-specific antibody (tsAb) GB266T, all with intact Fc function. In in vitro cell-based assays, these antibodies promote greater T cell expansion and tumor cell killing than benchmark antibodies and antibody combinations in an Fc-dependent manner, likely by facilitating T cell interactions (bridging) with cancer cells and monocytes, in addition to blocking immune checkpoints. In animal models, GB265 and GB266T ant...

Scientific Reports, Jun 18, 2023

The landscape of current cancer immunotherapy is dominated by antibodies targeting PD-1/ PD-L1 an... more The landscape of current cancer immunotherapy is dominated by antibodies targeting PD-1/ PD-L1 and CTLA-4 that have transformed cancer therapy, yet their efficacy is limited by primary and acquired resistance. The blockade of additional immune checkpoints, especially TIGIT and LAG-3, has been extensively explored, but so far only a LAG-3 antibody has been approved for combination with nivolumab to treat unresectable or metastatic melanoma. Here we report the development of a PDL1 × TIGIT bi-specific antibody (bsAb) GB265, a PDL1 × LAG3 bsAb GB266, and a PDL1 × TIGIT × LAG3 tri-specific antibody (tsAb) GB266T, all with intact Fc function. In in vitro cell-based assays, these antibodies promote greater T cell expansion and tumor cell killing than benchmark antibodies and antibody combinations in an Fc-dependent manner, likely by facilitating T cell interactions (bridging) with cancer cells and monocytes, in addition to blocking immune checkpoints. In animal models, GB265 and GB266T antibodies outperformed benchmarks in tumor suppression. This study demonstrates the potential of a new generation of multispecific checkpoint inhibitors to overcome resistance to current monospecific checkpoint antibodies or their combinations for the treatment of human cancers. Checkpoint inhibitors reactivate T cells by blockade checkpoint receptor/ligand interactions that restrict T cell activity 1. Several checkpoint inhibitor antibodies targeting CTLA-4 and PD-1/PD-L1 have been approved and have shown efficacy for a variety of tumor indications 2. Responders to such agents often exhibit durable remission, which is rarely seen for other therapies 3. However, many cancer patients do not benefit from such treatments due to primary or acquired resistance 4,5. The landscape of checkpoint inhibitors is still dominated by antibodies that target PD-1 or PD-L1, although the potential of targeting other immune checkpoints are also being actively investigated 2,6,7. Among those TIGIT and LAG-3 are two distinct immune checkpoints that contribute to T cell exhaustion 1,8,9. TIGIT is expressed in T cells and NK cells, and inhibits T cell activation mainly by competing for CD155 binding with CD226, a T cell stimulatory receptor 8,10-13. LAG-3 is expressed on CD4 and CD8 T cells, and suppresses T cells once it is engaged by one of its ligands including MHC-II 14,15 , FGL-1 16 , or galectin-3 17,18. These two immune checkpoints are now emerging as promising targets in the post-PD-1/CTLA-4 era in cancer immunotherapy. Strong preclinical evidence supports blocking the TIGIT pathway for cancer immunotherapy 12,19-22 , although a recent Phase III clinical trial of anti-TIGIT tiragolumab combined with anti-PD-L1 (Tecentriq) failed. Much remains to be learned about the LAG-3 pathway, including the relative importance of its three potential ligands. However, the clinical efficacy of LAG-3 inhibition has been attested, and an anti-LAG-3 antibody (relatlimab) has been approved by the FDA in combination therapy with anti-PD-1 (nivolumab) for the management of patients with unresectable or metastatic melanoma 23. These recent developments highlight the promise of simultaneously targeting PD-1/PD-L1 and TIGIT or LAG-3 pathways for cancer immunotherapy. Although combination immune checkpoint therapy is currently the prevailing strategy to overcome resistance to anti-PD-1/PD-L1 therapy, new technology platforms now allow for the development of a different class of therapeutic antibodies, i.e., multispecific antibodies, which can target different epitopes on two or more checkpoint molecules 24-28. In addition to blocking multiple inhibitory pathways, such multispecific agents can have additional MOAs, including cell-cell bridging via the crosslinking of antigens on neighboring cells to achieve greater T cell activation and/or cancer cell killing 24-27. Another factor to consider when designing therapeutic

International Journal of Biological Sciences, 2023

Although current cancer immunotherapies that target PD-1/PD-L1 immune checkpoint to reinvigorate ... more Although current cancer immunotherapies that target PD-1/PD-L1 immune checkpoint to reinvigorate exhausted T cells have achieved impressive clinical outcomes, only a small proportion of patients respond. New therapeutic targets are therefore needed to be identified to further unleash the anti-tumor potential of T cells and benefit more patients. Galectin-9 (Gal-9), initially identified as a ligand for TIM-3 to induce T cell death, acts as an immunosuppressive regulator in the tumor microenvironment (TME) but its potential as a therapeutic target remains largely elusive. Here we show that antibody neutralization of Gal-9, in combination with inhibition of Ataxia telangiectasia mutated (ATM), a kinase essential for DNA damage response (DDR), is a promising modality for cancer immunotherapy. Genetic depletion of ATM in tumors markedly potentiated anti-Gal-9 therapy in a syngeneic mouse model. Mechanistically, ATM inhibition greatly upregulated Gal-9 expression and secretion in a variety of human and murine tumor cells via the cGAS-STING-interferon β (IFNβ) innate immune pathway. Combination of Gal-9 inhibition with AZD1390, a selective ATM inhibitor currently evaluated in clinical trials, significantly suppressed tumor growth and prolonged survival in multiple syngeneic mouse models, including the poorly-immunogenic LLC lung tumors that do not respond to PD-1/PD-L1 blockade, concomitant with increased T cell infiltration. These results reveal Gal-9 induction via STING/IFNβ signaling as an important mechanism mediating tumor immune escape that could be targeted for cancer immunotherapies, and unveil a novel anti-Gal-9-based combination strategy for cancer immunotherapies in a wide variety of malignancies, including those resistant to PD-1/PD-L1 blockade.

3T3-L1 fibroblasts engineered with the doxycycline-induced LacZ or VPS13C knockdown syst... more 3T3-L1 fibroblasts engineered with the doxycycline-induced LacZ or VPS13C knockdown system were treated for three days without or with doxycycline in the presence of chloroquine, MG-132, or 3-MA. Cell viability was then analyzed using MTS assay[<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0153534#pone.0153534.ref058" target="_blank">58</a>]. Results are representative of three experiments. Asterisks denote statistical significance.</p

Cancer Research

Background: Cancer immunotherapy has transformed cancer therapy. In particular, blockade of the P... more Background: Cancer immunotherapy has transformed cancer therapy. In particular, blockade of the PD-L1/PD-1 axis with PD-1 or PD-L1 antibodies has achieved successful and durable treatment outcomes in some patients across a variety of cancer types. However, primary or acquired resistance to immunotherapy and adverse effects have been observed in cancer patients, due to a hostile TME and the immunosuppressive activity of various inhibitory checkpoint receptors and their ligands on immune cells and cancer cells. It was recently reported that dual blockade of PD-1/PD-L1 and TIGIT/CD155 could improve treatment efficacy in NSCLC patients. We therefore develop bispecific antibody against both PD-L1 and TIGIT with the aim of enhancing the efficacy of therapy and mitigating potential adverse effects. Methods: We obtained TIGIT binders by VHH library panning, and constructed PD-L1/TIGIT BsAbs of two different structures either by fusing the TIGIT VHH to a PD-L1 IgG or by fusing the TIGIT VHH ...

Expression was assayed by quantitative real-time RT-PCR for mRNA levels (A and ... more Expression was assayed by quantitative real-time RT-PCR for mRNA levels (A and B) and by immunoblotting for protein levels (C and D) in subconfluent 3T3-L1 fibroblasts, or at different time points of adipocyte differentiation. Adipocyte differentiation was induced at day 0, when cells were three days post confluence, following an established adipogenic regimen [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0153534#pone.0153534.ref022" target="_blank">22</a>]. Bar graphs present data (means ± s.e.) from three experiments.</p

Both human VPS13C (A) and ATG2 (B) carry a Chorein... more Both human VPS13C (A) and ATG2 (B) carry a Chorein_N and a ATG_C domains. cc, coiled coil. Data from the Pfam protein families database [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0153534#pone.0153534.ref057" target="_blank">57</a>]. (C) An ATG-C-nearby sequence in ATG2A (1723–1829), essential for autophagy and required for ATG2A localization to both the autophagic membrane and lipid droplets [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0153534#pone.0153534.ref033" target="_blank">33</a>], is also conserved in VPS13C. (D) 3T3-L1 fibroblasts engineered with a doxycycline-regulated knockdown system for control (LacZ) or Vps13c gene were treated for 3 days with or without doxycycline. Cells were then cultured 3 h under basal conditions, or in a medium depleted of amino acid and serum (starvation), in the absence or presence of chloroquine. Cells were lysed and analyzed by immunoblotting for indicated proteins. Results are representative of three experiments.</p

Journal of Biological Chemistry, 2022

Antibodies that target immune checkpoint proteins such as programmed cell death protein 1, progra... more Antibodies that target immune checkpoint proteins such as programmed cell death protein 1, programmed death ligand 1, and cytotoxic T-lymphocyte–associated antigen 4 in human cancers have achieved impressive clinical success; however, a significant proportion of patients fail to respond to these treatments. Galectin-9 (Gal-9), a β-galactoside-binding protein, has been shown to induce T-cell death and facilitate immunosuppression in the tumor microenvironment by binding to immunomodulatory receptors such as T-cell immunoglobulin and mucin domain–containing molecule 3 and the innate immune receptor dectin-1, suggesting that it may have potential as a target for cancer immunotherapy. Here, we report the development of two novel Gal-9-neutralizing antibodies that specifically react with the N-carbohydrate-recognition domain of human Gal-9 with high affinity. We also show using cell-based functional assays that these antibodies efficiently protected human T cells from Gal-9-induced cell death. Notably, in a T-cell/tumor cell coculture assay of cytotoxicity, these antibodies significantly promoted T cell-mediated killing of tumor cells. Taken together, our findings demonstrate potent inhibition of human Gal-9 by neutralizing antibodies, which may open new avenues for cancer immunotherapy.

Cancer Research

Targeting immune checkpoints such as programmed cell death 1 (PD-1) and programmed cell death lig... more Targeting immune checkpoints such as programmed cell death 1 (PD-1) and programmed cell death ligand 1 (PD-L1) has transformed cancer treatment, with durable clinical responses across a wide range of tumor types. However, a high percentage of patients fail to respond to anti–PD-1/PD-L1 treatment. A greater understanding of PD-L1 regulation is critical to improving the clinical response rate of PD-1/PD-L1 blockade. Here, we demonstrate that PD-L1 is phosphorylated and stabilized by casein kinase 2 (CK2) in cancer and dendritic cells (DC). Phosphorylation of PD-L1 at Thr285 and Thr290 by CK2 disrupted PD-L1 binding with speckle-type POZ protein, an adaptor protein of the cullin 3 (CUL3) ubiquitin E3 ligase complex, protecting PD-L1 from CUL3-mediated proteasomal degradation. Inhibition of CK2 decreased PD-L1 protein levels by promoting its degradation and resulted in the release of CD80 from DC to reactivate T-cell function. In a syngeneic mouse model, combined treatment with a CK2 in...

Science China Life Sciences, 2017

Cancer treatment in the past few years has been transformed by a new kind of therapy that targets... more Cancer treatment in the past few years has been transformed by a new kind of therapy that targets the immune system instead of the cancer itself to reinvigorate antitumor immunity with astonishing results. However, primary and acquired resistance to this type of treatment, namely immune checkpoint blockade (ICB), continue to counter treatment efficacy. In many cases, resistance has been attributed to defective or chronically enhanced interferon signaling and/or upregulation of alternative immune checkpoints, including T-cell immunoglobulin mucin-3 (Tim-3) and its ligand galactin-9 (Gal-9). In this article, we briefly describe the current knowledge of common checkpoint resistance mechanisms, focusing on the Tim-3/Gal-9 pathway as an alternative checkpoint that holds great promise as another target for ICB.

Oncogene

Programmed cell death 1 (PD-1) is widely expressed in tumor-infiltrating lymphocytes (TILs) of tr... more Programmed cell death 1 (PD-1) is widely expressed in tumor-infiltrating lymphocytes (TILs) of triple-negative breast cancer (TNBC). As a dominant inhibitory immune checkpoint (ICP) receptor, cell surface PD-1 is well-known to transduce negative signaling of effector T cell activity during cell–cell contact. However, despite its well-documented inhibitory effects, higher PD-1 expression in TILs is significantly associated with longer survival in TNBC patients. This phenomenon raises an interesting question whether PD-1 harbors positive activity to enhance anti-tumor immunity. Here, we show that PD-1 is secreted in an exosomal form by activated T cells and can remotely interact with either cell surface or exosomal programmed death-ligand 1 (PD-L1), induce PD-L1 internalization via clathrin-mediated endocytosis, and thereby prevent subsequent cellular PD-L1: PD-1 interaction, restoring tumor surveillance through attenuating PD-L1-induced suppression of tumor-specific cytotoxic T cell ...

Nature Communications

The two T cell inhibitory receptors PD-1 and TIM-3 are co-expressed during exhausted T cell diffe... more The two T cell inhibitory receptors PD-1 and TIM-3 are co-expressed during exhausted T cell differentiation, and recent evidence suggests that their crosstalk regulates T cell exhaustion and immunotherapy efficacy; however, the molecular mechanism is unclear. Here we show that PD-1 contributes to the persistence of PD-1+TIM-3+ T cells by binding to the TIM-3 ligand galectin-9 (Gal-9) and attenuates Gal-9/TIM-3-induced cell death. Anti-Gal-9 therapy selectively expands intratumoral TIM-3+ cytotoxic CD8 T cells and immunosuppressive regulatory T cells (Treg cells). The combination of anti-Gal-9 and an agonistic antibody to the co-stimulatory receptor GITR (glucocorticoid-induced tumor necrosis factor receptor-related protein) that depletes Treg cells induces synergistic antitumor activity. Gal-9 expression and secretion are promoted by interferon β and γ, and high Gal-9 expression correlates with poor prognosis in multiple human cancers. Our work uncovers a function for PD-1 in exhaus...

International Journal of Molecular Sciences

Galectin-12 is a member of a family of mammalian lectins characterized by their affinity for β-ga... more Galectin-12 is a member of a family of mammalian lectins characterized by their affinity for β-galactosides and consensus amino acid sequences. The protein structure consists of a single polypeptide chain containing two carbohydrate-recognition domains joined by a linker region. Galectin-12 is predominantly expressed in adipose tissue, but is also detected in macrophages and other leukocytes. Downregulation of galectin-12 in mouse 3T3-L1 cells impairs their differentiation into adipocytes. Conversely, overexpression of galectin-12 in vitro induces cell cycle arrest in G1 and apoptosis. Upregulation of galectin-12 and initiation of G1 cell cycle arrest are associated with driving pre-adipocytes toward terminal differentiation. Galectin-12 deficiency increases insulin sensitivity and glucose tolerance in obese animals. Galectin-12 inhibits macrophage polarization to the M2 population, enhancing inflammation and decreasing insulin sensitivity in adipocytes. Galectin-12 also affects myeloid differentiation, which is associated with chemotherapy resistance. In addition to highlighting the above-mentioned aspects, this review also discusses the potential clinical applications of modulating the function of galectin-12.

American journal of cancer research, 2018

The ataxia telangiectasia and Rad3-related (ATR) kinase plays a crucial role in maintaining genom... more The ataxia telangiectasia and Rad3-related (ATR) kinase plays a crucial role in maintaining genome stability in response to DNA damage. Once activated, ATR acts via its downstream target to arrest the cell cycle, promote DNA repair, and enhance cell survival. Therefore, ATR has become an attractive therapeutic target in cancer therapy. Multiple clinical studies have demonstrated that ATR inhibitors can sensitize cancer cells to conventional DNA damaging agents. However, the potential effects of ATR inhibitors on immune response in the tumor microenvironment, especially on the expression of immune checkpoint-related proteins, remain elusive. Here we show that DNA damaging agents, such as ionizing radiation and cisplatin, significantly induce cell surface PD-L1 expression in various cancer cell types. This effect is blocked by depletion or pharmacological inhibition of ATR, suggesting the essential role of ATR in DNA damage-induced PD-L1 expression. Mechanistically, we show that disru...

Journal of leukocyte biology, 2016

As a member of the galectin family of animal lectins, galectin-12 is preferentially expressed in ... more As a member of the galectin family of animal lectins, galectin-12 is preferentially expressed in adipocytes and leukocytes. In adipocytes, galectin-12 is associated with lipid droplets and regulates lipid metabolism and energy balance, whereas its role in leukocytes is not clear. Analysis of galectin-12 expression in a public data set of acute myeloid leukemia (AML) samples revealed that it is selectively overexpressed in the M3 subtype, which is also known as acute promyelocytic leukemia (APL). To investigate the role of galectin-12 in APL cells, we manipulated its expression in the APL cell line, NB4, and measured resultant effects on all-trans-retinoic acid (ATRA)-induced granulocytic differentiation. With a doxycycline-inducible gene knockdown system, we found that suppression of galectin-12 promoted ATRA-induced neutrophil differentiation but inhibited lipid droplet formation. Our results indicate that overexpression of galectin-12 contributes to a differentiation block in APL ...

PloS one, 2016

Galectin-12, a member of the galectin family of β-galactoside-binding animal lectins, is preferen... more Galectin-12, a member of the galectin family of β-galactoside-binding animal lectins, is preferentially expressed in adipocytes and required for adipocyte differentiation in vitro. This protein was recently found to regulate lipolysis, whole body adiposity, and glucose homeostasis in vivo. Here we identify VPS13C, a member of the VPS13 family of vacuolar protein sorting-associated proteins highly conserved throughout eukaryotic evolution, as a major galectin-12-binding protein. VPS13C is upregulated during adipocyte differentiation, and is required for galectin-12 protein stability. Knockdown of Vps13c markedly reduces the steady-state levels of galectin-12 by promoting its degradation through primarily the lysosomal pathway, and impairs adipocyte differentiation. Our studies also suggest that VPS13C may have a broader role in protein quality control. The regulation of galectin-12 stability by VPS13C could potentially be exploited for therapeutic intervention of obesity and related ...

Journal of Investigative Dermatology, 2012

The epidermal growth factor receptor (EGFR)-mediated signaling pathways are important in a variet... more The epidermal growth factor receptor (EGFR)-mediated signaling pathways are important in a variety of cellular processes, including cell migration and wound re-epithelialization. Intracellular trafficking of EGFR is critical for maintaining EGFR surface expression. Galectin-3, a member of an animal lectin family, has been implicated in a number of physiological and pathological processes. Through studies of galectin-3-deficient mice and cells isolated from these mice, we demonstrated that absence of galectin-3 impairs keratinocyte migration and skin wound reepithelialization. We have linked this pro-migratory function to a crucial role of cytosolic galectin-3 in controlling intracellular trafficking and cell surface expression of EGFR after EGF stimulation. Without galectin-3, the surface levels of EGFR are dramatically reduced and the receptor accumulates diffusely in the cytoplasm. This is associated with reduced rates of both endocytosis and recycling of the receptor. We have provided evidence that this novel function of galectin-3 may be mediated through interaction with its binding partner Alix, which is a protein component of the endosomal sorting complex required for transport (ESCRT) machinery. Our results suggest that galectin-3 is potentially a critical regulator of a number of important cellular responses through its intracellular control of trafficking of cell surface receptors.

Scientific Reports

Scientific Reports, Jun 18, 2023

International Journal of Biological Sciences, 2023

Cancer Research

Journal of Biological Chemistry, 2022

Cancer Research

Science China Life Sciences, 2017

Oncogene

Nature Communications

International Journal of Molecular Sciences

American journal of cancer research, 2018

Journal of leukocyte biology, 2016

PloS one, 2016

Journal of Investigative Dermatology, 2012