FAK suppresses antigen processing and presentation to promote immune evasion in pancreatic cancer.

OBJECTIVE: Immunotherapy for the treatment of pancreatic ductal adenocarcinoma (PDAC) has shown limited efficacy. Poor CD8 T-cell infiltration, low neoantigen load and a highly immunosuppressive tumour microenvironment contribute to this lack of response. Here, we aimed to further investigate the immunoregulatory function of focal adhesion kinase (FAK) in PDAC, with specific emphasis on regulation of the type-II interferon response that is critical in promoting T-cell tumour recognition and effective immunosurveillance. DESIGN: We combined CRISPR, proteogenomics and transcriptomics with mechanistic experiments using a KrasG12Dp53R172H mouse model of pancreatic cancer and validated findings using proteomic analysis of human patient-derived PDAC cell lines and analysis of publicly available human PDAC transcriptomics datasets. RESULTS: Loss of PDAC cell-intrinsic FAK signalling promotes expression of the immunoproteasome and Major Histocompatibility Complex class-I (MHC-I), resulting in increased antigen diversity and antigen presentation by FAK-/- PDAC cells. Regulation of the immunoproteasome by FAK is a critical determinant of this response, optimising the physicochemical properties of the peptide repertoire for high affinity binding to MHC-I. Expression of these pathways can be further amplified in a STAT1-dependent manner via co-depletion of FAK and STAT3, resulting in extensive infiltration of tumour-reactive CD8 T-cells and further restraint of tumour growth. FAK-dependent regulation of antigen processing and presentation is conserved between mouse and human PDAC, but is lost in cells/tumours with an extreme squamous phenotype. CONCLUSION: Therapies aimed at FAK degradation may unlock additional therapeutic benefit for the treatment of PDAC through increasing antigen diversity and promoting antigen presentation.

FAK promotes stromal PD-L2 expression associated with poor survival in pancreatic cancer.

BACKGROUND: Pancreatic Cancer is one of the most lethal cancers, with less than 8% of patients surviving 5 years following diagnosis. The last 40 years have seen only small incremental improvements in treatment options, highlighting the continued need to better define the cellular and molecular pathways contributing to therapy response and patient prognosis. METHODS: We combined CRISPR, shRNA and flow cytometry with mechanistic experiments using a KrasG12Dp53R172H mouse model of pancreatic cancer and analysis of publicly available human PDAC transcriptomic datasets. RESULTS: Here, we identify that expression of the immune checkpoint, Programmed Death Ligand 2 (PD-L2), is associated with poor prognosis, tumour grade, clinical stage and molecular subtype in patients with Pancreatic Ductal Adenocarcinoma (PDAC). We further show that PD-L2 is predominantly expressed in the stroma and, using an orthotopic murine model of PDAC, identify cancer cell-intrinsic Focal Adhesion Kinase (FAK) signalling as a regulator of PD-L2 stromal expression. Mechanistically, we find that FAK regulates interleukin-6, which can act in concert with interleukin-4 secreted by CD4 T-cells to drive elevated expression of PD-L2 on tumour-associated macrophages, dendritic cells and endothelial cells. CONCLUSIONS: These findings identify further complex heterocellular signalling networks contributing to FAK-mediated immune suppression in pancreatic cancer.

T-cell co-stimulation in combination with targeting FAK drives enhanced anti-tumor immunity.

Focal Adhesion Kinase (FAK) inhibitors are currently undergoing clinical testing in combination with anti-PD-1 immune checkpoint inhibitors. However, which patients are most likely to benefit from FAK inhibitors, and what the optimal FAK/immunotherapy combinations are, is currently unknown. We identify that cancer cell expression of the T-cell co-stimulatory ligand CD80 sensitizes murine tumors to a FAK inhibitor and show that CD80 is expressed by human cancer cells originating from both solid epithelial cancers and some hematological malignancies in which FAK inhibitors have not been tested clinically. In the absence of CD80, we identify that targeting alternative T-cell co-stimulatory receptors, in particular OX-40 and 4-1BB in combination with FAK, can drive enhanced anti-tumor immunity and even complete regression of murine tumors. Our findings provide rationale supporting the clinical development of FAK inhibitors in combination with patient selection based on cancer cell CD80 expression, and alternatively with therapies targeting T-cell co-stimulatory pathways.

Identification of 4-(Aminomethyl)-6-(trifluoromethyl)-2-(phenoxy)pyridine Derivatives as Potent, Selective, and Orally Efficacious Inhibitors of the Copper-Dependent Amine Oxidase, Lysyl Oxidase-Like 2 (LOXL2).

LOXL2 catalyzes the oxidative deamination of ε-amines of lysine and hydroxylysine residues within collagen and elastin, generating reactive aldehydes (allysine). Condensation with other allysines or lysines drives the formation of inter- and intramolecular cross-linkages, a process critical for the remodeling of the ECM. Dysregulation of this process can lead to fibrosis, and LOXL2 is known to be upregulated in fibrotic tissue. Small-molecules that directly inhibit LOXL2 catalytic activity represent a useful option for the treatment of fibrosis. Herein, we describe optimization of an initial hit 2, resulting in identification of racemic-trans-(3-((4-(aminomethyl)-6-(trifluoromethyl)pyridin-2-yl)oxy)phenyl)(3-fluoro-4-hydroxypyrrolidin-1-yl)methanone 28, a potent irreversible inhibitor of LOXL2 that is highly selective over LOX and other amine oxidases. Oral administration of 28 significantly reduced fibrosis in a 14-day mouse lung bleomycin model. The (R,R)-enantiomer 43 (PAT-1251) was selected as the clinical compound which has progressed into healthy volunteer Phase 1 trials, making it the "first-in-class" small-molecule LOXL2 inhibitor to enter clinical development.

Small Molecule Lysyl Oxidase-like 2 (LOXL2) Inhibitors: The Identification of an Inhibitor Selective for LOXL2 over LOX.

Two series of novel LOXL2 enzyme inhibitors are described: benzylamines substituted with electron withdrawing groups at the para-position and 2-substituted pyridine-4-ylmethanamines. The most potent compound, (2-chloropyridin-4-yl)methanamine 20 (hLOXL2 IC50 = 126 nM), was shown to be selective for LOXL2 over LOX and three other amine oxidases (MAO-A, MAO-B, and SSAO). Compound 20 is the first published small molecule inhibitor selective for LOXL2 over LOX.

Efficient parallel synthesis of macrocyclic peptidomimetics.

A new method for solid phase parallel synthesis of chemically and conformationally diverse macrocyclic peptidomimetics is reported. A key feature of the method is access to broad chemical and conformational diversity. Synthesis and mechanistic studies on the macrocyclization step are reported.