Targeting the tumor vasculature with antibody-drug conjugates (ADCs) is a promising anti-cancer strategy that in order to be realized must overcome several obstacles, including identification of ...suitable targets and optimal warheads. Here, we demonstrate that the cell-surface protein CD276/B7-H3 is broadly overexpressed by multiple tumor types on both cancer cells and tumor-infiltrating blood vessels, making it a potentially ideal dual-compartment therapeutic target. In preclinical studies CD276 ADCs armed with a conventional MMAE warhead destroyed CD276-positive cancer cells, but were ineffective against tumor vasculature. In contrast, pyrrolobenzodiazepine-conjugated CD276 ADCs killed both cancer cells and tumor vasculature, eradicating large established tumors and metastases, and improving long-term overall survival. CD276-targeted dual-compartment ablation could aid in the development of highly selective broad-acting anti-cancer therapies.
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•CD276/B7-H3 is broadly overexpressed in both cancer cells and tumor vasculature•Both angiogenic and non-angiogenic tumor vasculature express CD276•Anti-CD276-drug conjugates display potent anti-tumor and anti-metastatic activity•Pyrrolobenzodiazepine dimers are optimal warheads for targeting tumor vasculature
Seaman et al. show that CD276 is broadly overexpressed in cancer cells and tumor vascular cells and demonstrate anti-CD276-drug conjugates as promising anti-cancer reagents. The drug selected for conjugation is important because tumor vascular cells can be resistant to a drug to which tumor cells are sensitive.
Defining mechanisms that maintain tissue stem cells during homeostasis, stress, and aging is important for improving tissue regeneration and repair and enhancing cancer therapies. Here, we show that ...Id1 is induced in hematopoietic stem cells (HSCs) by cytokines that promote HSC proliferation and differentiation, suggesting that it functions in stress hematopoiesis. Genetic ablation of Id1 increases HSC self-renewal in serial bone marrow transplantation (BMT) assays, correlating with decreases in HSC proliferation, mitochondrial biogenesis, and reactive oxygen species (ROS) production. Id1−/− HSCs have a quiescent molecular signature and harbor less DNA damage than control HSCs. Cytokines produced in the hematopoietic microenvironment after γ-irradiation induce Id1 expression. Id1−/− HSCs display a blunted proliferative response to such cytokines and other inducers of chronic proliferation including genotoxic and inflammatory stress and aging, protecting them from chronic stress and exhaustion. Thus, targeting Id1 may be therapeutically useful for improving HSC survival and function during BMT, chronic stress, and aging.
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•Id1−/− HSCs have enhanced self-renewal potential and are maintained during BMT•Id1−/− HSCs show reduced oxidative stress and increased quiescence after BMT•Id1−/− HSCs are protected from inflammatory cytokine-induced proliferative stress•Id1−/− HSCs are protected from exhaustion by chronic inflammatory stress and aging
Singh et al. show that Id1 is an important mediator of stress hematopoiesis. They found that Id1 deletion protects HSCs from exhaustion in multiple paradigms of chronic and physiologically relevant stress and promotes their quiescence, suggesting that targeting Id1 may improve HSC survival and function during chronic stress and aging.
Defining mechanism(s) that maintain tissue stem quiescence is important for improving tissue regeneration, cell therapies, aging, and cancer. We report here that genetic ablation of Id2 in adult ...hematopoietic stem cells (HSCs) promotes increased HSC activation and differentiation, which results in HSC exhaustion and bone marrow failure over time. Id2J/J HSCs showed increased cycling, ROS production, mitochondrial activation, ATP production, and DNA damage compared with Id2+'+ HSCs, supporting the conclusion that Id2J/J HSCs are less quiescent. Mechanistically, HIF-1a expression was decreased in Id2J/J HSCs, and stabilization of HIF-1a in Id2J/J HSCs restored HSC quiescence and rescued HSC exhaustion. Inhibitor of DNA binding 2 (ID2) promoted HIF-1a expression by binding to the von Hippel-Lindau (VHL) protein and interfering with proteasomal degradation of HIF-1a. HIF-1a promoted Id2 expression and enforced a positive feedback loop between ID2 and HIF-1a to maintain HSC quiescence. Thus, sustained ID2 expression could protect HSCs during stress and improve HSC expansion for gene editing and cell therapies.
Poly (ADP-ribose) polymerase (PARP) inhibitor (PARPi) olaparib has been approved for treatment of advanced ovarian cancer associated with BRCA1 and BRCA2 mutations. BRCA1- and BRCA2-mutated cells, ...which are homologous recombination (HR) deficient, are hypersensitive to PARPi through the mechanism of synthetic lethality. Here we examine the effect of PARPi on HR-proficient cells. Olaparib pretreatment, PARP1 knockdown or Parp1 heterozygosity of Brca2(cko/ko) mouse embryonic stem cells (mESCs), carrying a null (ko) and a conditional (cko) allele of Brca2, results in viable Brca2(ko/ko) cells. PARP1 deficiency does not restore HR in Brca2(ko/ko) cells, but protects stalled replication forks from MRE11-mediated degradation through its impaired recruitment. The functional consequence of Parp1 heterozygosity on BRCA2 loss is demonstrated by a significant increase in tumorigenesis in Brca2(cko/cko) mice. Thus, while olaparib efficiently kills BRCA2-deficient cells, we demonstrate that it can also contribute to the synthetic viability if PARP is inhibited before BRCA2 loss.
Investigating mechanisms that regulate endothelial cell (EC) growth and survival is important for understanding EC homeostasis and how ECs maintain stem cell niches. We report here that targeted loss ...of Id genes in adult ECs results in dilated, leaky sinusoids and a pro-inflammatory state that increases in severity over time. Disruption in sinusoidal integrity leads to increased hematopoietic stem cell (HSC) proliferation, differentiation, migration, and exhaustion. Mechanistically, sinusoidal ECs (SECs) show increased apoptosis because of reduced Bcl2-family gene expression following Id gene ablation. Furthermore, Id1−/−Id3−/− SECs and upstream type H vessels show increased expression of cyclin-dependent kinase inhibitors p21 and p27 and impaired ability to proliferate, which is rescued by reducing E2-2 expression. Id1−/−Id3−/− mice do not survive sublethal irradiation because of impaired vessel regeneration and hematopoietic failure. Thus, Id genes are required for the survival and regeneration of BM SECs during homeostasis and stress to maintain HSC development.
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•Bone marrow sinusoids lacking Id genes become dilated, leaky, and pro-inflammatory•Sinusoidal ECs and type H vessels depend on Id genes for survival and proliferation•Id1 and Id3 restrain E proteins and CDKis and promote Bcl2-family gene expression•Loss of Id1 and Id3 in ECs promotes HSC activation, proliferation, and exhaustion
Gadomski et al. show that Id genes are critical regulators of bone marrow endothelial cell proliferation and survival. Selective loss of Id genes in endothelial cells promotes vessel dilation, apoptosis, and increased permeability, leading to the functional decline of hematopoietic stem cells under steady-state and stress conditions.
Growth factor independence 1 (Gfi-1) is a part of the transcriptional network that regulates the development of adult hematopoietic stem and progenitor cells. Gfi-1-null (Gfi-1−/−) mice have reduced ...numbers of hematopoietic stem cells (HSCs), impaired radioprotective function of hematopoietic progenitor cells (HPCs), and myeloid and erythroid hyperplasia. We found that the development of HPCs and erythropoiesis, but not HSC function, was rescued by reducing the expression of inhibitor of DNA-binding protein 2 (Id2) in Gfi-1−/− mice. Analysis of Gfi-1−/−;Id2+/− mice revealed that short-term HSCs, common myeloid progenitors (CMPs), erythroid burst-forming units, colony-forming units in spleen, and more differentiated red cells were partially restored by reducing Id2 levels in Gfi-1−/− mice. Moreover, short-term reconstituting cells, and, to a greater extent, CMP and megakaryocyte-erythroid progenitor development, and red blood cell production (anemia) were rescued in mice transplanted with Gfi-1−/−;Id2+/− bone marrow cells (BMCs) in comparison with Gfi-1−/− BMCs. Reduction of Id2 expression in Gfi-1−/− mice increased the expression of Gata1, Eklf, and EpoR, which are required for proper erythropoiesis. Reducing the levels of other Id family members (Id1 and Id3) in Gfi-1−/− mice did not rescue impaired HPC function or erythropoiesis. These data provide new evidence that Gfi-1 is linked to the erythroid gene regulatory network by repressing Id2 expression.
•Reducing Id2 in Gfi-1−/− mice restores radioprotective function of hematopoietic progenitors and partially rescues erythroid development.•Rescue of erythroid development in Gfi-1−/− mice by Id2 reduction directly correlates with an increase of Gata1, Eklf, and EpoR expression.
Pancreatic cancer is the fourth leading cause of cancer‐related mortality in the world. Pancreatic cancer can be localized, locally advanced, or metastatic. The median 1‐ and 5‐year survival rates ...are 25% and 6%, respectively. Epigenetic modifications such as DNA methylation play a significant role during both normal human development and cancer progression. To investigate epigenetic regulation of genes in the tumor‐initiating population of pancreatic cancer cells, which are also termed cancer stem cells (CSCs), we conducted epigenetic arrays in PANC1 and HPAC pancreatic cancer cell lines and compared the global DNA methylation status of CpG promoters in invasive cells, demonstrated to be CSCs, to their noninvasive counterparts, or non‐CSCs. Our results suggested that the NF‐κB pathway is one of the most activated pathways in pancreatic CSCs. In agreement with this, we determined that upon treatment with NF‐κB pathway inhibitors, the stem cell‐like properties of cells are significantly disrupted. Moreover, SOX9, demethylated in CSCs, is shown to play a crucial role in the invasion process. Additionally, we found a potential NF‐κB binding site located in the SOX9 promoter and determined that the NF‐κB subunit p65 positively regulates SOX9 expression by binding to its promoter directly. This interaction can be efficiently blocked by NF‐κB inhibitors. Thus, our work establishes a link between the classic NF‐κB signaling transduction pathway and the invasiveness of pancreatic CSCs, which may result in the identification of novel signals and molecules that function at an epigenetic level, and could potentially be targeted for pharmaceutical investigations and clinical trials. STEM Cells
2013;31:1454–1466
Fetal globin genes are transcriptionally silenced during embryogenesis through hemoglobin switching. Strategies to derepress fetal globin expression in the adult could alleviate symptoms in sickle ...cell disease and β-thalassemia. We identified a zinc-finger protein, pogo transposable element with zinc-finger domain (POGZ), expressed in hematopoietic progenitor cells. Targeted deletion of Pogz in adult hematopoietic cells in vivo results in persistence of embryonic β-like globin expression without affecting erythroid development. POGZ binds to the Bcl11a promoter and erythroid-specific intragenic regulatory regions. Pogz+/− mice show elevated embryonic β-like globin expression, suggesting that partial reduction of Pogz expression results in persistence of embryonic β-like globin expression. Knockdown of POGZ in primary human CD34+ progenitor cell-derived erythroblasts reduces BCL11A expression, a known repressor of embryonic β-like globin expression, and increases fetal hemoglobin expression. These findings are significant, since new therapeutic targets and strategies are needed to treat β-globin disorders.
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•Pogz is highly expressed in mouse megakaryocyte erythroid progenitors•POGZ is required to repress murine embryonic β-like globin during erythropoiesis•Pogz+/− mice develop normally and show elevated embryonic β-like globin expression•POGZ knockdown decreases BCL11A and increases fetal globin expression in human cells
Gudmundsdottir et al. show that POGZ represses embryonic globin gene expression in mouse and human erythroid cells, in part by regulating Bcl11a expression in vitro and in vivo. The molecular pathways regulated by POGZ may represent potential therapeutic targets to increase fetal globin expression in patients with sickle cell disease and β-thalassemia.
Toll-like receptor 4 (Tlr4) has a pivotal role in innate immune responses, and the transcription factor CCAAT/enhancer binding protein delta (C/EBPδ, Cebpd) is a Tlr4-induced gene. Here we identify a ...positive feedback loop in which C/EBPδ activates Tlr4 gene expression in macrophages and tumour cells. In addition, we discovered a negative feedback loop whereby the tumour suppressor FBXW7α (FBW7, Cdc4), whose gene expression is inhibited by C/EBPδ, targets C/EBPδ for degradation when C/EBPδ is phosphorylated by GSK-3β. Consequently, FBXW7α suppresses Tlr4 expression and responses to the ligand lipopolysaccharide. FBXW7α depletion alone is sufficient to augment pro-inflammatory signalling in vivo. Moreover, as inflammatory pathways are known to modulate tumour biology, Cebpd null mammary tumours, which have reduced metastatic potential, show altered expression of inflammation-associated genes. Together, these findings reveal a role for C/EBPδ upstream of Tlr4 signalling and uncover a function for FBXW7α as an attenuator of inflammatory signalling.
The relationship between cancer and autoimmunity is complex. However, the incidence of solid tumors such as melanoma has increased significantly among patients with previous or newly diagnosed ...systemic autoimmune disease (AID). At the same time, immune checkpoint blockade (ICB) therapy of cancer induces
autoinflammation and exacerbates underlying AID, even without evident antitumor responses. Recently, systemic lupus erythematosus (SLE) activity was found to drive myeloid-derived suppressor cell (MDSC) formation in patients, a known barrier to healthy immune surveillance and successful cancer immunotherapy. Cross-talk between MDSCs and macrophages generally drives immune suppressive activity in the tumor microenvironment. However, it remains unclear how peripheral pregenerated MDSC under chronic inflammatory conditions modulates global macrophage immune functions and the impact it could have on existing tumors and underlying lupus nephritis. Here we show that pathogenic expansion of SLE-generated MDSCs by melanoma drives global macrophage polarization and simultaneously impacts the severity of lupus nephritis and tumor progression in SLE-prone mice. Molecular and functional data showed that MDSCs interact with autoimmune macrophages and inhibit cell surface expression of CD40 and the production of IL27. Moreover, low CD40/IL27 signaling in tumors correlated with high tumor-associated macrophage infiltration and ICB therapy resistance both in murine and human melanoma exhibiting active IFNγ signatures. These results suggest that preventing global macrophage reprogramming induced by MDSC-mediated inhibition of CD40/IL27 signaling provides a precision melanoma immunotherapy strategy, supporting an original and advantageous approach to treat solid tumors within established autoimmune landscapes. SIGNIFICANCE: Myeloid-derived suppressor cells induce macrophage reprogramming by suppressing CD40/IL27 signaling to drive melanoma progression, simultaneously affecting underlying autoimmune disease and facilitating resistance to immunotherapy within preexisting autoimmune landscapes.
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