Human Tumor Models Research Articles

CD47 predominates over CD24 as a macrophage immune checkpoint in cancer.

Macrophages hold tremendous promise as effectors of cancer immunotherapy, but the best strategies to provoke these cells to attack tumors remain unknown. Here, we evaluated the therapeutic potential of targeting two distinct macrophage immune checkpoints: CD47 and CD24. We found that antibodies targeting these antigens could elicit maximal levels of phagocytosis when combined together in vitro. However, to our surprise, via unbiased genome-wide CRISPR screens, we found that CD24 primarily acts as a target of opsonization rather than an immune checkpoint. In a series of in vitro and in vivo genetic validation studies, we found that CD24 was neither necessary nor sufficient to protect cancer cells from macrophage phagocytosis in most mouse and human tumor models. Instead, anti-CD24 antibodies exhibit robust Fc-dependent activity, and as a consequence, they cause significant on-target hematologic toxicity in mice. To overcome these challenges and leverage our findings for therapeutic purposes, we engineered a collection of 77 novel bispecific antibodies that bind to a tumor antigen with one arm and engage macrophages with the second arm. We discovered multiple novel bispecifics that maximally activate macrophage-mediated cytotoxicity and reduce binding to healthy blood cells, including bispecifics targeting macrophage immune checkpoint molecules in combination with EGFR, TROP2, and CD71. Overall, our findings indicate that CD47 predominates over CD24 as a macrophage immune checkpoint in cancer, and that the novel bispecifics we created may be optimal immunotherapies to direct myeloid cells to eradicate solid tumors.

Tumour-intrinsic PDL1 signals regulate the Chk2 DNA damage response in cancer cells and mediate resistance to Chk1 inhibitors

BackgroundAside from the canonical role of PDL1 as a tumour surface-expressed immune checkpoint molecule, tumour-intrinsic PDL1 signals regulate non-canonical immunopathological pathways mediating treatment resistance whose significance, mechanisms, and therapeutic targeting remain incompletely understood. Recent reports implicate tumour-intrinsic PDL1 signals in the DNA damage response (DDR), including promoting homologous recombination DNA damage repair and mRNA stability of DDR proteins, but many mechanistic details remain undefined.MethodsWe genetically depleted PDL1 from transplantable mouse and human cancer cell lines to understand consequences of tumour-intrinsic PDL1 signals in the DNA damage response. We complemented this work with studies of primary human tumours and inducible mouse tumours. We developed novel approaches to show tumour-intrinsic PDL1 signals in specific subcellular locations. We pharmacologically depleted tumour PDL1 in vivo in mouse models with repurposed FDA-approved drugs for proof-of-concept clinical translation studies.ResultsWe show that tumour-intrinsic PDL1 promotes the checkpoint kinase-2 (Chk2)-mediated DNA damage response. Intracellular but not surface-expressed PDL1 controlled Chk2 protein content post-translationally and independently of PD1 by antagonising PIRH2 E3 ligase-mediated Chk2 polyubiquitination and protein degradation. Genetic tumour PDL1 depletion specifically reduced tumour Chk2 content but not ATM, ATR, or Chk1 DDR proteins, enhanced Chk1 inhibitor (Chk1i) synthetic lethality in vitro in diverse human and murine tumour models, and improved Chk1i efficacy in vivo. Pharmacologic tumour PDL1 depletion with cefepime or ceftazidime replicated genetic tumour PDL1 depletion by reducing tumour Chk2, inducing Chk1i synthetic lethality in a tumour PDL1-dependent manner, and reducing in vivo tumour growth when combined with Chk1i.ConclusionsOur data challenge the prevailing surface PDL1 paradigm, elucidate important and previously unappreciated roles for tumour-intrinsic PDL1 in regulating the ATM/Chk2 DNA damage response axis and E3 ligase-mediated protein degradation, suggest tumour PDL1 as a biomarker for Chk1i efficacy, and support the rapid clinical potential of pharmacologic tumour PDL1 depletion to treat selected cancers.

Bojungikki-Tang Augments Pembrolizumab Efficacy in Human PBMC-Injected H460 Tumor-Bearing Mice

Bojungikki-Tang (BJIKT) is traditionally used to enhance digestive function and immunity. It has gained attention as a supplement to chemotherapy or targeted therapy owing to its immune-boosting properties. This study aimed to evaluate the synergistic anti-tumor effects of BJIKT in combination with pembrolizumab in a preclinical model. MHC I/II double knockout NSG mice were humanized with peripheral blood mononuclear cells (PBMCs) and injected subcutaneously with H460 lung tumor cells to establish a humanized tumor model. Both agents were administered to evaluate their impact on tumor growth and immune cell behavior. Immunohistochemistry showed decreased exhaustion markers in CD8(+) and CD4(+) T cells within the tumor, indicating enhanced T cell activity. Additionally, RNA sequencing, transcriptome analysis, and quantitative PCR analysis were performed on tumor tissues to investigate the molecular mechanisms underlying the observed effects. The results confirmed that BJIKT improved T cell function and tumor necrosis factor signaling while suppressing transforming growth factor-β signaling. This modulation led to cell cycle arrest and apoptosis. These findings demonstrate that BJIKT, when combined with pembrolizumab, produces significant anti-tumor effects by altering immune pathways and enhancing the anti-tumor immune response. This study provides valuable insights into the role of BJIKT in the tumor microenvironment and its potential to improve therapeutic outcomes.

Therapeutic vaccine targeting dual immune checkpoints induces potent multifunctional CD8+ T cell anti-tumor immunity

BackgroundVaccines targeting immune checkpoints represent a promising immunotherapeutic approach for solid tumors. However, the therapeutic efficacy of dual targeting immune checkpoints is still unclear in renal carcinoma. MethodsAn adenovirus (Ad) vaccine targeting B7H1 and B7H3 was developed and evaluated for its therapeutic efficacy in subcutaneous, lung metastasis or orthotopic renal carcinoma mouse and humanized models using flow cytometry, Enzyme-linked immunosorbent spot (ELISPOT), cytotoxic T lymphocyte (CTL) killing, cell deletion, hematoxylin and eosin (HE) staining, and immunohistochemistry (IHC) assays. ResultsThe Ad-B7H1/B7H3 immunization effectively inhibited tumor growth and increased the induction and percentages of CD8+ T cells in subcutaneous tumor models. The vaccine enhanced the induction and maturation of CD11c+ or CD8+CD11c+ cells, promoting tumor-specific CD8+ T cell immune responses. This was evidenced by increased proliferation of CD8+ T cells and enhanced CTL killing activity. Deletion of CD8+ T cells in vivo abolished the anti-tumor effect of the Ad-B7H1/B7H3 vaccine, highlighting the pivotal role of functional CD8+ T cell immune responses. Moreover, significant therapeutic efficacy of the Ad-B7H1/B7H3 vaccine was observed in lung metastasis, orthotopic, and humanized tumor models through multifunctional CD8+ T cell immune responses. ConclusionsThe Ad vaccine targeting dual immune checkpoints B7H1 and B7H3 exerts a potent therapeutic effect for renal carcinoma and holds promise for solid tumor treatment.

Adenovirus vaccine targeting kinases induces potent antitumor immunity in solid tumors

BackgroundTargeting kinases presents a potential strategy for treating solid tumors; however, the therapeutic potential of vaccines targeting kinases remains uncertain.MethodsAdenovirus (Ad) vaccines encoding Aurora kinase A (AURKA) or cyclin-dependent kinase...

DPPA4 increases aggressiveness of pituitary neuroendocrine tumors by enhancing cell stemness.

Pituitary neuroendocrine tumors, PitNETs, are often aggressive and precipitate in distant metastases that are refractory to current therapies. However, the molecular mechanism in PitNETs' aggressiveness is not well understood. Developmental pluripotency-associated 4 (DPPA4) is known as a stem cell regulatory gene and overexpressed in certain cancers, but its function in the context of PitNETs' aggressiveness is not known. We employed both rat and human models of PitNETs. In the rat pituitary tumor model (RPT), we used prenatal-alcohol-exposed (PAE) female Fischer rats which developed aggressive PitNETs following estrogen treatment, while in the human pituitary tumor (HPT) model, we used aggressively proliferative cells from pituitary tumors of patients undergone surgery. Various molecular, cellular, and epigenetic techniques were used to determine the role of DPPA4 in PitNETs' aggressiveness. We show that DPPA4 is overexpressed in association with increased cell stemness factors in aggressive PitNETs of PAE rats and of human patients. Gene-editing experiments demonstrate that DPPA4 increases the expression of cell stemness and tumor aggressiveness genes and promotes proliferation, colonization, migration, and tumorigenic potential of PitNET cells. ChIP assays and receptor antagonism studies reveal that DPPA4 binds to canonical WINTs promoters and increases directly or indirectly the Wnt/β-catenin control of cell stemness, tumor growth, and aggressiveness of PitNETs. Epigenetic studies show involvement of histone methyltransferase in alcohol activation of DPPA4. These findings support a role of DPPA4 in tumor stemness and aggressiveness and provide a preclinical rationale for modulating this stemness regulator for the treatment of PitNETs.

Extracellular Matrix Sulfation in the Tumor Microenvironment Stimulates Cancer Stemness and Invasiveness.

Tumor extracellular matrices (ECM) exhibit aberrant changes in composition and mechanics compared to normal tissues. Proteoglycans (PG) are vital regulators of cellular signaling in the ECM with the ability to modulate receptor tyrosine kinase (RTK) activation via their sulfated glycosaminoglycan (sGAG) side chains. However, their role on tumor cell behavior is controversial. Here, it is demonstrated that PGs are heavily expressed in lung adenocarcinoma (LUAD) patients in correlation with invasive phenotype and poor prognosis. A bioengineered human lung tumor model that recapitulates the increase of sGAGs in tumors in an organotypic matrix with independent control of stiffness, viscoelasticity, ligand density, and porosity, is developed. This model reveals that increased sulfation stimulates extensive proliferation, epithelial-mesenchymal transition (EMT), and stemness in cancer cells. The focal adhesion kinase (FAK)-phosphatidylinositol 3-kinase (PI3K) signaling axis is identified as a mediator of sulfation-induced molecular changes in cells upon activation of a distinct set of RTKs within tumor-mimetic hydrogels. The study shows that the transcriptomic landscape of tumor cells in response to increased sulfation resembles native PG-rich patient tumors by employing integrative omics and network modeling approaches.

H-1 Parvovirus-Induced Oncolysis and Tumor Microenvironment Immune Modulation in a Novel Heterotypic Spheroid Model of Cutaneous T-Cell Lymphoma.

The rat protoparvovirus H-1 (H-1PV) is an oncolytic virus known for its anticancer properties in laboratory models of various human tumors, including non-Hodgkin lymphomas (NHL) of B-cell origin. However, H-1PV therapeutic potential against hematological malignancies of T-cell origin remains underexplored. The aim of the present study was to conduct a pilot preclinical investigation of H-1PV-mediated oncolytic effects in cutaneous T-cell lymphoma (CTCL), a type of NHL that is urgently calling for innovative therapies. We demonstrated H-1PV productive infection and induction of oncolysis in both classically grown CTCL suspension cultures and in a novel, in vivo-relevant, heterotypic spheroid model, but not in healthy donor controls, including peripheral blood mononuclear cells (PBMCs). H-1PV-mediated oncolysis of CTCL cells was not prevented by Bcl-2 overexpression and was accompanied by increased extracellular ATP release. In CTCL spheroid co-cultures with PBMCs, increased spheroid infiltration with immune cells was detected upon co-culture treatment with the virus. In conclusion, our preclinical data show that H-1PV may hold significant potential as an ingenious viroimmunotherapeutic drug candidate against CTCL.

Downregulation of ATP5F1D inhibits mtROS/NLRP3/caspase-1/GSDMD axis to suppress pyroptosis-mediated malignant progression of endometrial cancer

PurposeIn developed countries, endometrial cancer (EC) is the most prevalent gynecological cancer and its occurrence is associated with chronic inflammation. ATP5F1D is a subunit of ATP synthase (complex V), as well as the important component of mitochondrial electron transport chain (ETC). ETC play compelling roles in carcinogenesis. To date, little is known about the role of ATP5F1D in EC. MethodsATP5F1D expression was identified in EC tissues and EC cell lines. We evaluated the influence of ATP5F1D on clinical features and prognosis based on TCGA database. The effects of ATP5F1D in EC malignant progression by applying loss-of-function assays in KLE and Ishikawa cell lines were detected by EdU, CCK-8, wound healing, Transwell, and flow cytometry assays. Additionally, electron microscope, LDH release, ELISA, mitochondrial ROS measurement, and Immunofluorescence were performed to demonstrate ATP5F1D can affect the pyroptosis of EC. To observe the anti-tumor effect on ATP5F1D silencing, we established an in vivo human endometrial tumor model using nude mice. ResultsATP5F1D expression was significantly upregulated in EC and was associated with favorable prognosis. ATP5F1D knockdown inhibited the proliferation, invasion, and migration of EC cells. Similarly, in nude mice, ATP5F1D knockdown suppressed the growth EC cells. Knocking down ATP5F1D lead to decrease the production of mitochondrial ROS (mtROS) and inhibited pyroptosis of EC cells. ConclusionDownregulation of ATP5F1D as a new therapeutic strategy that could mediate pyroptosis via suppressing mtROS/NLRP3/caspase-1/GSDMD pathway to inhibit EC progression.

Targeting Tumor-Associated Sialic Acids Using Chimeric Switch Receptors Based on Siglec-9 Enhances the Antitumor Efficacy of Engineered T Cells.

Cancer exploits different mechanisms to escape T-cell immunosurveillance, including overexpression of checkpoint ligands, secretion of immunosuppressive molecules, and aberrant glycosylation. Herein, we report that IFNγ, a potent immunomodulator secreted in the tumor microenvironment, can induce α2,6 hypersialylation in cancer cell lines derived from various histologies. We focused on Siglec-9, a receptor for sialic acid moieties, and demonstrated that the Siglec-9+ T-cell population displayed reduced effector function. We speculated that Siglec-9 in primary human T cells can act as a checkpoint molecule and demonstrated that knocking out Siglec-9 using a clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 system enhanced the functionality of primary human T cells. Finally, we aimed to augment cancer-specific T-cell activity by taking advantage of tumor hypersialylation. Thus, we designed several Siglec-9-based chimeric switch receptors (CSR), which included an intracellular moiety derived from costimulatory molecules (CD28/41BB) and different hinge regions. In an antigen-specific context, T cells transduced with Siglec-9 CSRs demonstrated increased cytokine secretions and upregulation of activation markers. Moreover, T cells equipped with specific Siglec-9 CSRs mediated robust antitumor activity in a xenograft model of human tumors. Overall, this work sheds light on tumor evasion mechanisms mediated by sialylated residues and exemplifies an approach to improve engineered T cell-based cancer treatment. See related Spotlight by Abken, p. 1310.

Probe-guided endoscopic system for 5-aminolevulinic acid-based photodynamic diagnosis in cholangiocarcinoma

Background and AimThe diagnostic accuracy for cholangiocarcinoma (CCA) is inadequate, necessitating the exploration of novel diagnostic approaches. Protoporphyrin IX (Pp IX), a metabolic product of 5-aminolevulinic acid (5-ALA), emits red fluorescence upon blue light exposure. Because it accumulates selectively in cancer cells, photodynamic diagnosis using 5-ALA (5-ALA-PDD) has been integrated into clinical practice for diverse cancer types. Nevertheless, there is currently no device capable of capturing Pp IX-derived fluorescence for real-time 5-ALA-PDD within the biliary tract, largely due to challenges in device miniaturization. MethodsTo investigate the feasibility of real-time 5ALA-PDD in CCA, we developed two essential components of the cholangioscopy system: a small-diameter flexible camera and a light guide for emitting blue light. We evaluated the detectability of Pp IX fluorescence using these devices in experimental gels and animal models. ResultsOur camera and light guide were smoothly inserted into the lumen of existing cholangioscopes. Incorporating a long-pass filter at the camera tip enabled efficient detection of red fluorescence without significantly impacting white-light observation. The integration of these devices facilitated clear visualization of red fluorescence from gels containing Pp IX at concentrations of 5 μM or higher. Additionally, when observing subcutaneous human CCA tumor models in nude mice treated with 5-ALA, we successfully demonstrated distinct red fluorescence from Pp IX accumulation in tumors compared to peritumoral subcutaneous areas. ConclusionThe integration of our device combination holds promise for real-time 5-ALA-PDD in human CCA, potentially enhancing the diagnostic accuracy for this complex condition.

Physical activities aid in tumor prevention: A finite element study of bio-heat transfer in healthy and malignant breast tissues

The objective of the present research is to explore the temperature diffusion in healthy and cancerous tissues, with a specific focus on how physical activity impacts on the weakening of breast tumors. Previous research lacked numerical analysis regarding the effectiveness of physical activity in tumor prevention or attenuation, prompting an investigation into the mechanism behind physical activity and tumor prevention from a bio-heat transfer perspective. The study employs a realistic model of human breasts and tumors in COMSOL Multiphysics® to analyze temperature distribution by utilizing Penne's bio-heat equation. The research examines their influence on tissue temperature by varying tumor diameter (10–20 mm) and exercise intensities (such as walking speeds and other activities like carpentry, swimming, and marathon running). Results demonstrate that cancerous tissues generate notably more heat than normal tissues at rest and during physical activity. Smaller tumors exhibit higher temperatures during exercise, emphasizing the significance of tumor size in treatment effectiveness. Tumor temperatures range between 40 and 43.2 °C, while healthy tissue temperatures remain below 41 °C during physical activity. High-intensity exercises, particularly swimming, walking at 1.8 m/s, and marathon running, display a therapeutic effect on tumors, increasing effectiveness with intensity. The temperatures of healthy and malignant tissues rise noticeably due to constant metabolic heat and decreased blood flow. The study also identifies the optimal duration of high-intensity exercise, recommending at least 20 min for optimal therapeutic outcomes. The outcomes of this research would help individuals, doctors, and cancer researchers understand and weaken malignant tissues.

A bi-adjuvant nanovaccine amplifying STING activation for cancer immunotherapy

The insufficient immune activation and immunosuppressive microenvironment restrict the therapeutic efficacy of cancer vaccines. Herein, we exploited a bi-adjuvant nanovaccine to amplify the activation of the stimulator of interferon genes (STING) for potent and durable anti-tumor immunity. STING agonist (MnO2 nanoparticles), toll-like receptor 9 agonist (CpG), and antigenic proteins or peptides were strategically formulated into nanovaccines (MnC NVs) by a one-step biomineralization method. The nanovaccine efficiently targeted dendritic cells of draining lymph nodes, in which CpG-mediated the stimulation of nuclear factor-kappa B pathway augmented STING signaling. RNA sequencing analysis validated the enhanced expression of relevant genes of type-I interferon signaling pathways. The priming of antigen-specific CD8+ T cells and humoral immune responses led to potent tumor growth inhibition and prolonged survival time in melanoma and human papilloma virus-derived tumor models. MnC NVs were able to synergize with immune checkpoint blockade therapy, with 80 % survival over 40 days in TC-1 tumor model. Moreover, rechallenging of tumor-free mice with homotypic cells resulted in complete tumor growth inhibition, suggesting the generation of a long-term memory effect. The simplified nanovaccine platform provides a safe and robust strategy to boost STING-mediated anti-tumor immunity for personalized cancer immunotherapy, showing significant potential for clinical translation.

Targeting of SUMOylation leads to cBAF complex stabilization and disruption of the SS18::SSX transcriptome in Synovial Sarcoma.

Synovial Sarcoma (SS) is driven by the SS18::SSX fusion oncoprotein and is ultimately refractory to therapeutic approaches. SS18::SSX alters ATP-dependent chromatin remodeling BAF (mammalian SWI/SNF) complexes, leading to the degradation of canonical (cBAF) complex and amplified presence of an SS18::SSX-containing non-canonical BAF (ncBAF or GBAF) that drives an SS-specific transcription program and tumorigenesis. We demonstrate that SS18::SSX activates the SUMOylation program and SSs are sensitive to the small molecule SAE1/2 inhibitor, TAK-981. Mechanistically, TAK-981 de-SUMOylates the cBAF subunit SMARCE1, stabilizing and restoring cBAF on chromatin, shifting away from SS18::SSX-ncBAF-driven transcription, associated with DNA damage and cell death and resulting in tumor inhibition across both human and mouse SS tumor models. TAK-981 synergized with cytotoxic chemotherapy through increased DNA damage, leading to tumor regression. Targeting the SUMOylation pathway in SS restores cBAF complexes and blocks the SS18::SSX-ncBAF transcriptome, identifying a therapeutic vulnerability in SS, positioning the in-clinic TAK-981 to treat SS.

Multimodal Mass Spectrometry Imaging of an Osteosarcoma Multicellular Tumour Spheroid Model to Investigate Drug-Induced Response.

A multimodal mass spectrometry imaging (MSI) approach was used to investigate the chemotherapy drug-induced response of a Multicellular Tumour Spheroid (MCTS) 3D cell culture model of osteosarcoma (OS). The work addresses the critical demand for enhanced translatable early drug discovery approaches by demonstrating a robust spatially resolved molecular distribution analysis in tumour models following chemotherapeutic intervention. Advanced high-resolution techniques were employed, including desorption electrospray ionisation (DESI) mass spectrometry imaging (MSI), to assess the interplay between metabolic and cellular pathways in response to chemotherapeutic intervention. Endogenous metabolite distributions of the human OS tumour models were complemented with subcellularly resolved protein localisation by the detection of metal-tagged antibodies using Imaging Mass Cytometry (IMC). The first application of matrix-assisted laser desorption ionization-immunohistochemistry (MALDI-IHC) of 3D cell culture models is reported here. Protein localisation and expression following an acute dosage of the chemotherapy drug doxorubicin demonstrated novel indications for mechanisms of region-specific tumour survival and cell-cycle-specific drug-induced responses. Previously unknown doxorubicin-induced metabolite upregulation was revealed by DESI-MSI of MCTSs, which may be used to inform mechanisms of chemotherapeutic resistance. The demonstration of specific tumour survival mechanisms that are characteristic of those reported for in vivo tumours has underscored the increasing value of this approach as a tool to investigate drug resistance.

Transcriptomic analysis-guided assessment of precision-cut tumor slices (PCTS) as an ex-vivo tool in cancer research

With cancer immunotherapy and precision medicine dynamically evolving, there is greater need for pre-clinical models that can better replicate the intact tumor and its complex tumor microenvironment (TME). Precision-cut tumor slices (PCTS) have recently emerged as an ex vivo human tumor model, offering the opportunity to study individual patient responses to targeted therapies, including immunotherapies. However, little is known about the physiologic status of PCTS and how culture conditions alter gene expression. In this study, we generated PCTS from head and neck cancers (HNC) and mesothelioma tumors (Meso) and undertook transcriptomic analyses to understand the changes that occur in the timeframe between PCTS generation and up to 72 h (hrs) in culture. Our findings showed major changes occurring during the first 24 h culture period of PCTS, involving genes related to wound healing, extracellular matrix, hypoxia, and IFNγ-dependent pathways in both tumor types, as well as tumor-specific changes. Collectively, our data provides an insight into PCTS physiology, which should be taken into consideration when designing PCTS studies, especially in the context of immunology and immunotherapy.

The FLRT3-UNC5B Pathway is a Novel Regulator of T Immunosurveillance

Abstract Our understanding of molecular cell-surface interactions that control T cell responses in cancer remains limited despite many advances in the field. We hypothesized that tumor cells take advantage of developmental protein-protein interactions such as axon guidance interactions that could inhibit or repel tumor-immune interactions in the TME to evade T cell anti-tumor immunity. We used gain-of-function screening to evaluate a gene-set of axon guidance proteins for inhibitory function in human T cells and identified and validated Fibronectin and Leucine-Rich Transmembrane protein-3 (FLRT3) as an inhibitor of T cell activity. FLRT3 inhibited human T cell function and promoted tumor growth in humanized tumor cell-derived xenograft (CDX) models. FLRT3 inhibited T cells through interaction with Unc-5 Netrin Receptor B (UNC5B) expressed on activated T cells. A FLRT3 monoclonal antibody designed to specifically block FLRT3 interactions with UNC5 family proteins reduced tumor growth in CDX models that was dependent on T cells. FLRT3 blockade also enhanced CAR-T and BiTE + T cell activity and infiltration in humanized zebrafish tumor models. These studies support the notion that aberrant axon guidance protein expression in the TME act as T cell checkpoints, and the FLRT3:UNC5B pathway can be targeted therapeutically to improve T cell-based immunotherapy activity and infiltration.

Half-life extension of single-domain antibody-drug conjugates by albumin binding moiety enhances antitumor efficacy.

Single-domain antibody-drug conjugates (sdADCs) have been proven to have deeper solid tumor penetration and intratumor accumulation capabilities due to their smaller size compared with traditional IgG format ADCs. However, one of the key challenges for improving clinical outcomes of sdADCs is their abbreviated in vivo half-life. In this study, we innovatively fused an antihuman serum albumin (αHSA) nanobody to a sdADCs targeting oncofetal antigen 5T4, conferring serum albumin binding to enhance the pharmacokinetic profiles of sdADCs. The fusion protein was conjugated with monomethyl auristatin E (MMAE) at s224c site mutation. The conjugate exhibited potent cytotoxicity against various tumor cells. Compared with the nonalbumin-binding counterparts, the conjugate exhibited a 10-fold extended half-life in wild-type mice and fivefold prolonged serum half-life in BxPC-3 xenograft tumor models as well as enhanced tumor accumulation and retention in mice. Consequently, n501-αHSA-MMAE showed potent antitumor effects, which were comparable to n501-MMAE in pancreatic cancer BxPC-3 xenograft tumor models; however, in human ovarian teratoma PA-1 xenograft tumor models, n501-αHSA-MMAE significantly improved antitumor efficacy. Moreover, the conjugate showed mitigated hepatotoxicity. In summary, our results suggested that fusion to albumin-binding moiety as a viable strategy can enhance the therapeutic potential of sdADCs through optimized pharmacokinetics.

Inhibition of protein translational machinery in triple-negative breast cancer as a promising therapeutic strategy

Y-box binding protein-1 (YB-1) is a proto-oncogenic protein associated with protein translation regulation. It plays a crucial role in the development and progression of triple-negative breast cancer (TNBC). In this study, we describe a promising approach to inhibit YB-1 using SU056, a small-molecule inhibitor. SU056 physically interacts with YB-1 and reduces its expression, which helps to restrain the progression of TNBC. Proteome profiling analysis indicates that the inhibition of YB-1 by SU056 can alter the proteins that regulate protein translation, an essential process for cancer cell growth. Preclinical studies on human cells, mice, and patient-derived xenograft tumor models show the effectiveness of SU056. Moreover, toxicological studies have shown that SU056 treatment and dosing are well tolerated without any adverse effects. Overall, our study provides a strong foundation for the further development of SU056 as a potential treatment option for patients with TNBC by targeting YB-1.

Tumor expressed BCAM impedes anti-tumor T cell immunity and can be targeted therapeutically

Abstract The significant albeit limited clinical benefit of therapeutic targeting of checkpoint inhibitors highlights the need for immunological understanding of cancer escape from anti-tumor immunity to develop novel and better therapeutics. We surmise that many protein-protein interactions that regulate T cell immunity have yet to be identified. Using human gain-of-function genetic screens, thousands of proteins were screened for their ability to inhibit primary human T cell function. B Cell Adhesion Molecule (BCAM) was identified as a potent inhibitor of T cell proliferation and cytotoxic T cell (CTL) ability to kill human tumors in vitro. Tumor cell surface BCAM inhibited anti-tumor immunity promoted tumor growth in vivo. Blockade with decoy soluble BCAM protein fused to a either a silenced or functional Fc domain (BCAM Fc fusion protein) was able to promote T cell anti-tumor immunity and reduce tumor growth in syngeneic and humanized CDX tumor models. Moreover, tumor BCAM interaction with known ligand Laminin-a5 (LAMA5) resulted in increased production of LAMA5 that was associated with ECM modifications and potent exclusion of T cells from the tumor microenvironment (TME). BCAM Fc disruption of BCAM-LAMA5 interactions resulted in decreased LAMA5 production, increased T cell infiltration and decreased tumor growth. These data support a multi-pronged role for BCAM in anti-tumor T cell suppression and exclusion that can be targeted therapeutically.