A k-mer based transcriptomics approach for antisense drug discovery targeting the Ewing's family of tumors.

Ewing's sarcoma treatment failures are associated with high mortality indicating a need for new therapeutic approaches. We used a k-mer counting approach to identify cancer-specific mRNA transcripts in 3 Ewing's Family Tumor (EFT) cell lines not found in the normal human transcriptome. Phosphorodiamidate morpholino oligomers targeting six EFT-specific transcripts were evaluated for cytotoxicity in TC-32 and CHLA-10 EFT lines and in HEK293 renal epithelial control cells.

Detection of unique Ebola virus oligonucleotides using fluorescently-labeled phosphorodiamidate morpholino oligonucleotide probe pairs.

Here we identify a low-cost diagnostic platform using fluorescently-labeled phosphorodiamidate morpholino oligonucleotide (PMO) probe pairs, which upon binding target oligonucleotides undergo fluorescence resonance energy transfer (FRET). Using a target oligonucleotide derived from the Ebola virus (EBOV), we have derivatized PMO probes with either Alexa Fluor488 (donor) or tetramethylrhodamine (acceptor). Upon EBOV target oligonulceotide binding, observed changes in FRET between PMO probe pairs permit a 25 pM lower limit of detection; there is no off-target binding within a complex mixture of nucleic acids and other biomolecules present in human saliva.

Invention and Early History of Morpholinos: From Pipe Dream to Practical Products.

Beginning with my concept in 1969 to treat disease at the nucleic acid level using antisense nucleic acids, antisense has evolved to the current Morpholino oligos. Morpholinos have been the dominant gene knockdown system in developmental biology. Lack of delivery technologies has limited their use in adult animals (including humans), though alteration in muscles in Duchenne muscular dystrophy (DMD) allows delivery into adult muscle.

Tumor imaging using technetium-99m bound to pH-sensitive peptides.

Solid tumors often display metabolic abnormalities that consistently produce low pH in the extracellular space of poorly perfused tissue. These acidic regions may provide a mechanism for drug targeting. Peptides have been designed in such a manner that they exist in an anionic hydrophilic form at the pH of normal tissues, but then undergo a sharp transition to a non-ionic lipophilic form at reduced pH.

Morpholino, siRNA, and S-DNA compared: impact of structure and mechanism of action on off-target effects and sequence specificity.

Generally a gene knockdown agent should achieve high sequence specificity and should lack off-target effects (non-antisense effects due to interactions with structures other than gene transcripts). Three major gene knockdown types are compared with respect to off-target effects and sequence specificities: 1) phosphorothioate-linked DNA (S-DNA); 2) short interfering RNA (siRNA); and, 3) Morpholino. S-DNAs cause multiple off-target effects, largely because their backbone sulfurs bind to many different proteins.

Endo-Porter: a novel reagent for safe, effective delivery of substances into cells.

Delivering large molecules into the cytosol of animal cells without damaging the cells has been one of the toughest challenges in biology. Endo-Porter is a weak-base amphiphilic peptide that was designed to deliver morpholino antisense oligomers and other non-ionic substances into the cytosol/nuclear compartment of cells by an endocytosis-mediated process that avoids damaging the plasma membrane of the cell. This prevents the loss of vital cell contents and the attendant high cell toxicity typical of most delivery systems.

Custom cancer therapies: safe and effective treatments for most or all cancers.

Gene Tools, LLC has embarked on a program to develop safe and effective custom therapies for essentially all cancers. This entails: 1) identifying in each patient's cancer multiple molecular targets which are absent from that patient's normal cells, but are present in and essential to that patient's cancer; and 2) treating that patient with a custom cocktail of therapeutic agents effective for specifically inhibiting those selected targets in that patient's cancer.