Simultaneous evaluation of treatment efficacy and toxicity for bispecific T-cell engager therapeutics in a humanized mouse model.

Immuno-oncology (IO)-based therapies such as checkpoint inhibitors, bi-specific antibodies, and CAR-T-cell therapies have shown significant success in the treatment of several cancer indications. However, these therapies can result in the development of severe adverse events, including cytokine release syndrome (CRS). Currently, there is a paucity of in vivo models that can evaluate dose-response relationships for both tumor control and CRS-related safety issues.

Serine phosphorylation of the small phosphoprotein ICAP1 inhibits its nuclear accumulation.

Nuclear accumulation of the small phosphoprotein integrin cytoplasmic domain-associated protein-1 (ICAP1) results in recruitment of its binding partner, Krev/Rap1 interaction trapped-1 (KRIT1), to the nucleus. KRIT1 loss is the most common cause of cerebral cavernous malformation, a neurovascular dysplasia resulting in dilated, thin-walled vessels that tend to rupture, increasing the risk for hemorrhagic stroke. KRIT1's nuclear roles are unknown, but it is known to function as a scaffolding or adaptor protein at cell-cell junctions and in the cytosol, supporting normal blood vessel integrity and development.

Nuclear Localization of Integrin Cytoplasmic Domain-associated Protein-1 (ICAP1) Influences β1 Integrin Activation and Recruits Krev/Interaction Trapped-1 (KRIT1) to the Nucleus.

Binding of ICAP1 (integrin cytoplasmic domain-associated protein-1) to the cytoplasmic tails of β1 integrins inhibits integrin activation. ICAP1 also binds to KRIT1 (Krev interaction trapped-1), a protein whose loss of function leads to cerebral cavernous malformation, a cerebrovascular dysplasia occurring in up to 0.5% of the population.

CCM2-CCM3 interaction stabilizes their protein expression and permits endothelial network formation.

Mutations in the essential adaptor proteins CCM2 or CCM3 lead to cerebral cavernous malformations (CCM), vascular lesions that most frequently occur in the brain and are strongly associated with hemorrhagic stroke, seizures, and other neurological disorders. CCM2 binds CCM3, but the molecular basis of this interaction, and its functional significance, have not been elucidated. Here, we used x-ray crystallography and structure-guided mutagenesis to show that an α-helical LD-like motif within CCM2 binds the highly conserved "HP1" pocket of the CCM3 focal adhesion targeting (FAT) homology domain.

Cerebral cavernous malformation proteins at a glance.

Loss-of-function mutations in genes encoding KRIT1 (also known as CCM1), CCM2 (also known as OSM and malcavernin) or PDCD10 (also known as CCM3) cause cerebral cavernous malformations (CCMs). These abnormalities are characterized by dilated leaky blood vessels, especially in the neurovasculature, that result in increased risk of stroke, focal neurological defects and seizures. The three CCM proteins can exist in a trimeric complex, and each of these essential multi-domain adaptor proteins also interacts with a range of signaling, cytoskeletal and adaptor proteins, presumably accounting for their roles in a range of basic cellular processes including cell adhesion, migration, polarity and apoptosis.

Mechanism for KRIT1 release of ICAP1-mediated suppression of integrin activation.

KRIT1 (Krev/Rap1 Interaction Trapped-1) mutations are observed in ∼40% of autosomal-dominant cerebral cavernous malformations (CCMs), a disease occurring in up to 0.5% of the population. We show that KRIT1 functions as a switch for β1 integrin activation by antagonizing ICAP1 (Integrin Cytoplasmic Associated Protein-1)-mediated modulation of "inside-out" activation.

Structural basis for paxillin binding and focal adhesion targeting of β-parvin.

β-Parvin is a cytoplasmic adaptor protein that localizes to focal adhesions where it interacts with integrin-linked kinase and is involved in linking integrin receptors to the cytoskeleton. It has been reported that despite high sequence similarity to α-parvin, β-parvin does not bind paxillin, suggesting distinct interactions and cellular functions for these two closely related parvins. Here, we reveal that β-parvin binds directly and specifically to leucine-aspartic acid repeat (LD) motifs in paxillin via its C-terminal calponin homology (CH2) domain.

Structural basis for small G protein effector interaction of Ras-related protein 1 (Rap1) and adaptor protein Krev interaction trapped 1 (KRIT1).

Cerebral cavernous malformations (CCMs) affect 0.1-0.5% of the population resulting in leaky vasculature and severe neurological defects.

Epithelial stem cells.

It is likely that adult epithelial stem cells will be useful in the treatment of diseases, such as ectodermal dysplasias, monilethrix, Netherton syndrome, Menkes disease, hereditary epidermolysis bullosa, and alopecias. Additionally, other skin problems such as burn wounds, chronic wounds, and ulcers will benefit from stem cell-related therapies. However, there are many questions that need to be answered before this goal can be realized.

Isolation and culture of adult epithelial stem cells from human skin.

The homeostasis of all self-renewing tissues is dependent on adult stem cells. As undifferentiated stem cells undergo asymmetric divisions, they generate daughter cells that retain the stem cell phenotype and transit-amplifying cells (TA cells) that migrate from the stem cell niche, undergo rapid proliferation and terminally differentiate to repopulate the tissue. Epithelial stem cells have been identified in the epidermis, hair follicle, and intestine as cells with a high in vitro proliferative potential and as slow-cycling label-retaining cells in vivo (1-3).

Targeting the Notch1 and mTOR pathways in a mouse T-ALL model.

Mutations in NOTCH1 are frequently detected in patients with T-cell acute lymphoblastic leukemia (T-ALL) and in mouse T-ALL models. Treatment of mouse or human T-ALL cell lines in vitro with gamma-secretase inhibitors (GSIs) results in growth arrest and/or apoptosis. These studies suggest GSIs as potential therapeutic agents in the treatment of T-ALL.

p19Arf inhibits the invasion of hepatocellular carcinoma cells by binding to C-terminal binding protein.

The INK4A/ARF tumor suppressor locus is frequently inactivated in hepatocellular carcinoma (HCC), yet the consequences of this remain unknown. We recently described a HCC mouse model in which loss of the Ink4a/Arf locus accelerates the development of metastasis and enhances tumor cell migration and invasion in cell culture assays. We show here that knockdown of p19Arf in an HCC cell line increases invasion in cell culture assays.

The Notch1/c-Myc pathway in T cell leukemia.

The Notch receptor family and its ligands (Delta-like and Jagged) have been found deregulated in several human cancers. We and the Aster/Pear group recently identified c-myc as a direct transcriptional target gene of the Notch1 pathway in T cell acute lymphoblastic leukemia (T-ALL). Although the oncogenic roles of c-Myc and Notch1 are established, a direct link between Notch1 and c-Myc had not been demonstrated.

Notch1 contributes to mouse T-cell leukemia by directly inducing the expression of c-myc.

Recent work with mouse models and human leukemic samples has shown that gain-of-function mutation(s) in Notch1 is a common genetic event in T-cell acute lymphoblastic leukemia (T-ALL). The Notch1 receptor signals through a gamma-secretase-dependent process that releases intracellular Notch1 from the membrane to the nucleus, where it forms part of a transcriptional activator complex. To identify Notch1 target genes in leukemia, we developed mouse T-cell leukemic lines that express intracellular Notch1 in a doxycycline-dependent manner.

NFKB1 is a direct target of the TAL1 oncoprotein in human T leukemia cells.

We recently showed that a subset of human T acute lymphoblastic leukemia (T-ALL) cell lines expresses low basal levels of p50, a nuclear factor-kappaB (NF-kappaB)/Rel family member, resulting in their capacity to activate the atypical p65:cRel complex rather than the classic p50:p65 dimer. Here, we show that the transcription factor TAL1 (also known as SCL) binds to the promoter of the NFKB1 gene that encodes p50 and represses its transcription to set up this unique response in T-ALL cells. When TAL1 expression is reduced in CEM T leukemia cells, basal NFKB1 expression is increased, and the levels of p65:cRel complex and transcription of its target gene, such as intercellular adhesion molecule-1 (ICAM-1), are reduced in response to etoposide treatment.