Broad-spectrum therapeutic suppression of metastatic melanoma through nuclear hormone receptor activation.

Melanoma metastasis is a devastating outcome lacking an effective preventative therapeutic. We provide pharmacologic, molecular, and genetic evidence establishing the liver-X nuclear hormone receptor (LXR) as a therapeutic target in melanoma. Oral administration of multiple LXR agonists suppressed melanoma invasion, angiogenesis, tumor progression, and metastasis. Molecular and genetic experiments revealed these effects to be mediated by LXRß, which elicits these outcomes through transcriptional induction of tumoral and stromal apolipoprotein-E (ApoE). LXRß agonism robustly suppressed tumor growth and metastasis across a diverse mutational spectrum of melanoma lines. LXRß targeting significantly prolonged animal survival, suppressed the progression of established metastases, and inhibited brain metastatic colonization. Importantly, LXRß activation displayed melanoma-suppressive cooperativity with the frontline regimens dacarbazine, B-Raf inhibition, and the anti-CTLA-4 antibody and robustly inhibited melanomas that had acquired resistance to B-Raf inhibition or dacarbazine. We present a promising therapeutic approach that uniquely acts by transcriptionally activating a metastasis suppressor gene.

Convergent multi-miRNA targeting of ApoE drives LRP1/LRP8-dependent melanoma metastasis and angiogenesis.

Through in vivo selection of human cancer cell populations, we uncover a convergent and cooperative miRNA network that drives melanoma metastasis. We identify miR-1908, miR-199a-5p, and miR-199a-3p as endogenous promoters of metastatic invasion, angiogenesis, and colonization in melanoma. These miRNAs convergently target apolipoprotein E (ApoE) and the heat shock factor DNAJA4. Cancer-secreted ApoE suppresses invasion and metastatic endothelial recruitment (MER) by engaging melanoma cell LRP1 and endothelial cell LRP8 receptors, respectively, while DNAJA4 promotes ApoE expression. Expression levels of these miRNAs and ApoE correlate with human metastatic progression outcomes. Treatment of cells with locked nucleic acids (LNAs) targeting these miRNAs inhibits metastasis to multiple organs, and therapeutic delivery of these LNAs strongly suppresses melanoma metastasis. We thus identify miRNAs with dual cell-intrinsic/cell-extrinsic roles in cancer, reveal convergent cooperativity in a metastatic miRNA network, identify ApoE as an anti-angiogenic and metastasis-suppressive factor, and uncover multiple prognostic miRNAs with synergistic combinatorial therapeutic potential in melanoma.

Nanoparticle delivery of immunostimulatory oligonucleotides enhances response to checkpoint inhibitor therapeutics.

The recent advent of immune checkpoint inhibitor (CPI) antibodies has revolutionized many aspects of cancer therapy, but the efficacy of these breakthrough therapeutics remains limited, as many patients fail to respond for reasons that still largely evade understanding. An array of studies in human patients and animal models has demonstrated that local signaling can generate strongly immunosuppressive microenvironments within tumors, and emerging evidence suggests that delivery of immunostimulatory molecules into tumors can have therapeutic effects. Nanoparticle formulations of these cargoes offer a promising way to maximize their delivery and to enhance the efficacy of checkpoint inhibitors. We developed a modular nanoparticle system capable of encapsulating an array of immunostimulatory oligonucleotides that, in some cases, greatly increase their potency to activate inflammatory signaling within immune cells in vitro. We hypothesized that these immunostimulatory nanoparticles could suppress tumor growth by activating similar signaling in vivo, and thereby also improve responsiveness to immune checkpoint inhibitor antibody therapies. We found that our engineered nanoparticles carrying a CpG DNA ligand of TLR9 can suppress tumor growth in several animal models of various cancers, resulting in an abscopal effect on distant tumors, and improving responsiveness to anti-CTLA4 treatment with combinatorial effects after intratumoral administration. Moreover, by incorporating tumor-homing peptides, immunostimulatory nucleotide-bearing nanoparticles facilitate antitumor efficacy after systemic intravenous (i.v.) administration.

Metastasis-suppressor transcript destabilization through TARBP2 binding of mRNA hairpins.

Aberrant regulation of RNA stability has an important role in many disease states. Deregulated post-transcriptional modulation, such as that governed by microRNAs targeting linear sequence elements in messenger RNAs, has been implicated in the progression of many cancer types. A defining feature of RNA is its ability to fold into structures. However, the roles of structural mRNA elements in cancer progression remain unexplored. Here we performed an unbiased search for post-transcriptional modulators of mRNA stability in breast cancer by conducting whole-genome transcript stability measurements in poorly and highly metastatic isogenic human breast cancer lines. Using a computational framework that searches RNA sequence and structure space, we discovered a family of GC-rich structural cis-regulatory RNA elements, termed sRSEs for structural RNA stability elements, which are significantly overrepresented in transcripts displaying reduced stability in highly metastatic cells. By integrating computational and biochemical approaches, we identified TARBP2, a double-stranded RNA-binding protein implicated in microRNA processing, as the trans factor that binds the sRSE family and similar structural elements--collectively termed TARBP2-binding structural elements (TBSEs)--in transcripts. TARBP2 is overexpressed in metastatic cells and metastatic human breast tumours and destabilizes transcripts containing TBSEs. Endogenous TARBP2 promotes metastatic cell invasion and colonization by destabilizing amyloid precursor protein (APP) and ZNF395 transcripts, two genes previously associated with Alzheimer's and Huntington's disease, respectively. We reveal these genes to be novel metastasis suppressor genes in breast cancer. The cleavage product of APP, extracellular amyloid-α peptide, directly suppresses invasion while ZNF395 transcriptionally represses a pro-metastatic gene expression program. The expression levels of TARBP2, APP and ZNF395 in human breast carcinomas support their experimentally uncovered roles in metastasis. Our findings establish a non-canonical and direct role for TARBP2 in mammalian gene expression regulation and reveal that regulated RNA destabilization through protein-mediated binding of mRNA structural elements can govern cancer progression.

Sustained-release synthetic biomarkers for monitoring thrombosis and inflammation using point-of-care compatible readouts.

Postoperative infection and thromboembolism represent significant sources of morbidity and mortality but cannot be easily tracked after hospital discharge. Therefore, a molecular test that could be performed at home would significantly impact disease management. Our lab has previously developed intravenously delivered 'synthetic biomarkers' that respond to dysregulated proteases to produce a urinary signal. These assays, however, have been limited to chronic diseases or acute diseases initiated at the time of diagnostic administration. Here, we formulate a subcutaneously administered sustained release system by using small PEG scaffolds (<10 nm) to promote diffusion into the bloodstream over a day. We demonstrate the utility of a thrombin sensor to identify thrombosis and an MMP sensor to measure inflammation. Finally, we developed a companion paper ELISA using printed wax barriers with nanomolar sensitivity for urinary reporters for point-of-care detection. Our approach for subcutaneous delivery of nanosensors combined with urinary paper analysis may enable facile monitoring of at-risk patients.

Protease activity sensors noninvasively classify bacterial infections and antibiotic responses.

BACKGROUND: Respiratory tract infections represent a significant public health risk, and timely and accurate detection of bacterial infections facilitates rapid therapeutic intervention. Furthermore, monitoring the progression of infections after intervention enables 'course correction' in cases where initial treatments are ineffective, avoiding unnecessary drug dosing that can contribute to antibiotic resistance. However, current diagnostic and monitoring techniques rely on non-specific or slow readouts, such as radiographic imaging and sputum cultures, which fail to specifically identify bacterial infections and take several days to identify optimal antibiotic treatments. METHODS: Here we describe a nanoparticle system that detects P. aeruginosa lung infections by sensing host and bacterial protease activity in vivo, and that delivers a urinary detection readout. One protease sensor is comprised of a peptide substrate for the P. aeruginosa protease LasA. A second sensor designed to detect elastases is responsive to recombinant neutrophil elastase and secreted proteases from bacterial strains. FINDINGS: In mice infected with P. aeruginosa, nanoparticle formulations of these protease sensors-termed activity-based nanosensors (ABNs)-detect infections and monitor bacterial clearance from the lungs over time. Additionally, ABNs differentiate between appropriate and ineffective antibiotic treatments acutely, within hours after the initiation of therapy. INTERPRETATION: These findings demonstrate how activity measurements of disease-associated proteases can provide a noninvasive window into the dynamic process of bacterial infection and resolution, offering an opportunity for detecting, monitoring, and characterizing lung infections. FUND: National Cancer Institute, National Institute of Environmental Health Sciences, National Institutes of Health, National Science Foundation Graduate Research Fellowship Program, and Howard Hughes Medical Institute.

Genome-scale analysis identifies paralog lethality as a vulnerability of chromosome 1p loss in cancer.

Functional redundancy shared by paralog genes may afford protection against genetic perturbations, but it can also result in genetic vulnerabilities due to mutual interdependency1-5. Here, we surveyed genome-scale short hairpin RNA and CRISPR screening data on hundreds of cancer cell lines and identified MAGOH and MAGOHB, core members of the splicing-dependent exon junction complex, as top-ranked paralog dependencies6-8. MAGOHB is the top gene dependency in cells with hemizygous MAGOH deletion, a pervasive genetic event that frequently occurs due to chromosome 1p loss. Inhibition of MAGOHB in a MAGOH-deleted context compromises viability by globally perturbing alternative splicing and RNA surveillance. Dependency on IPO13, an importin-ß receptor that mediates nuclear import of the MAGOH/B-Y14 heterodimer9, is highly correlated with dependency on both MAGOH and MAGOHB. Both MAGOHB and IPO13 represent dependencies in murine xenografts with hemizygous MAGOH deletion. Our results identify MAGOH and MAGOHB as reciprocal paralog dependencies across cancer types and suggest a rationale for targeting the MAGOHB-IPO13 axis in cancers with chromosome 1p deletion.