Cold Spring Harbor, NY, United States
Cold Spring Harbor, NY, United States
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Robert F.,McGill University | Roman W.,McGill University | Bramoulle A.,McGill University | Fellmann C.,Mirimus Inc. | And 6 more authors.
Proceedings of the National Academy of Sciences of the United States of America | Year: 2014

Enhanced protein synthesis capacity is associated with increased tumor cell survival, proliferation, and resistance to chemotherapy. Cancers like multiple myeloma (MM), which display elevated activity in key translation regulatory nodes, such as the PI3K/ mammalian target of rapamycin and MYC-eukaryotic initiation factor (eIF) 4E pathways, are predicted to be particularly sensitive to therapeutic strategies that target this process. To identify novel vulnerabilities in MM, we undertook a focused RNAi screen in which components of the translation apparatus were targeted. Our screen was designed to identify synthetic lethal relationships between translation factors or regulators and dexamethasone (DEX), a corticosteroid used as frontline therapy in this disease. We find that suppression of all three subunits of the eIF4F cap-binding complex synergizes with DEX in MM to induce cell death. Using a suite of small molecules that target various activities of eIF4F, we observed that cell survival and DEX resistance are attenuated upon eIF4F inhibition in MM cell lines and primary human samples. Levels of MYC and myeloid cell leukemia 1, two known eIF4Fresponsive transcripts and key survival factors in MM, were reduced upon eIF4F inhibition, and their independent suppression also synergized with DEX. Inhibition of eIF4F in MM exerts pleotropic effects unraveling a unique therapeutic opportunity.


Fellmann C.,Mirimus Inc. | Hoffmann T.,Research Institute of Molecular Pathology IMP | Sridhar V.,Mirimus Inc. | Hopfgartner B.,Research Institute of Molecular Pathology IMP | And 8 more authors.
Cell Reports | Year: 2013

Short hairpin RNA (shRNA) technology enables stable and regulated gene repression. For establishing experimentally versatile RNAi tools and minimizing toxicities, synthetic shRNAs can be embedded into endogenous microRNA contexts. However, due to our incomplete understanding of microRNA biogenesis, such "shRNAmirs" often fail to trigger potent knockdown, especially when expressed from a single genomic copy. Following recent advances in design of synthetic shRNAmir stems, here we take a systematic approach to optimize the experimental miR-30 backbone. Among several favorable features, we identify a conserved element 3' of the basal stem as critically required for optimal shRNAmir processing and implement it in an optimized backbone termed "miR-E", which strongly increases mature shRNA levels and knockdown efficacy. Existing miR-30 reagents can be easily converted to miR-E, and its combination with up-to-date design rules establishes a validated and accessible platform for generating effective single-copy shRNA libraries that will facilitate the functional annotation of the genome. © 2013 The Authors.


Fellmann C.,Mirimus Inc. | Lowe S.W.,Sloan Kettering Cancer Center
Nature Cell Biology | Year: 2014

RNA interference has become an indispensable tool for loss-of-function studies across eukaryotes. By enabling stable and reversible gene silencing, shRNAs provide a means to study long-term phenotypes, perform pool-based forward genetic screens and examine the consequences of temporary target inhibition in vivo. However, efficient implementation in vertebrate systems has been hindered by technical difficulties affecting potency and specificity. Focusing on these issues, we analyse current strategies to obtain maximal knockdown with minimal off-target effects. © 2014 Macmillan Publishers Limited.


Knott S.R.V.,Howard Hughes Medical Institute | Maceli A.R.,Howard Hughes Medical Institute | Erard N.,Howard Hughes Medical Institute | Chang K.,Howard Hughes Medical Institute | And 10 more authors.
Molecular Cell | Year: 2014

The strength of conclusions drawn from RNAi-based studies is heavily influenced by the quality of tools used to elicit knockdown. Prior studies have developed algorithms to design siRNAs. However, to date, no established method has emerged to identifyeffective shRNAs, which have lower intracellular abundance than transfected siRNAs and undergo additional processing steps. We recently developed a multiplexed assay for identifying potent shRNAs and used this method to generate ~250,000 shRNA efficacy data points. Using these data, we developed shERWOOD, an algorithm capable of predicting, for any shRNA, the likelihood that it will elicit potent target knockdown. Combined with additional shRNA design strategies, shERWOOD allows the ab initio identification of potent shRNAs that specifically target the majority of each gene's multiple transcripts. We validated the performance of our shRNA designs using several orthogonal strategies and constructed genome-wide collections of shRNAs for humans and mice based on our approach. © 2014 Elsevier Inc.


Grant
Agency: Department of Health and Human Services | Branch: | Program: SBIR | Phase: Phase I | Award Amount: 189.01K | Year: 2014

DESCRIPTION (provided by applicant): An estimated 1.8 billion dollars is spent for each new successful drug developed, primarily in failed clinical trials due to lack of efficacy and safety. New approaches for rapid identification and early preclinical validation of novel therapeutic targets are crucial to make important go/no-go decisions and curb the cost of developing new cancer treatments. For decades, genetically engineered mouse models have provided a powerful platform to study disease initiation and maintenance, the tumor microenvironment and the responsiveness of cancers to known or novel therapeutics; however, the long lead times and high costs required to develop, intercross and maintain models with various cancer predisposing gene combinationshave limited their practical utility in the drug discovery process. RNA interference (RNAi), a mechanism that controls gene expression, can be exploited experimentally to silence nearly any gene target. By expressing synthetic short hairpin RNAs (shRN


M

Trademark
Mirimus Inc. | Date: 2012-08-08

Live animals, namely, genetically engineered mice for laboratory and clinical trial purposes. Medical and scientific research, namely, the design for others of genetically engineered mice for laboratory and clinical trial purposes.


Trademark
Mirimus Inc. | Date: 2012-08-08

Live animals, namely, genetically engineered mice for laboratory and clinical trial purposes. Medical and scientific research, namely, the design for others of genetically engineered mice for laboratory and clinical trial purposes.


Trademark
Mirimus Inc. | Date: 2012-10-30

Live animals, namely, genetically engineered mice for laboratory and clinical trial purposes. Medical and scientific research, namely, the design for others of genetically engineered mice for laboratory and clinical trial purposes.


Patent
Mirimus Inc. | Date: 2014-06-20

What is described is a modified miRNA molecule for producing an artificial siRNA/mature small RNA molecule that inhibits the expression of a target transcript of a host cell, comprising a stem region modified to comprise a sequence encoding the artificial siRNA molecule, consisting of a guide and a passenger strand; a conserved region having specific sequences; and a nonconserved region modified to include a recognition site for a restriction enzyme while preserving the native secondary structure of the miRNA. The modified miRNA molecule produced with these elements substantially inhibits the expression of the target transcript when expressed from an endogenous or exogenous promoter in the host cell.


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