Recent Patents on Biotechnology (v.6, #3)

Ryanodine Receptor Patents by Alexander Kushnir, Andrew R. Marks (157-166).
Research over the past two decades has implicated dysfunction of the ryanodine receptor (RyR), a Ca2+ releasechannel on the sarcoplasmic reticulum (SR) required for excitation-contraction (EC) coupling, in the pathogenesis of cardiacand skeletal myopathies. These discoveries have led to the development of novel drugs, screening tools, and researchmethods. The patents associated with these advances tell the story of the initial discovery of RyRs as a target for plant alkaloids,to their central role in cardiac and skeletal muscle excitation-contraction coupling, and ongoing clinical trials witha novel class of drugs called RycalsTM that inhibit pathological intracellular Ca2+ leak. Additionally, these patents highlightquestions, controversies, and future directions of the RyR field.

Myokines in Myogenesis and Health by Tora Henriksen, Charlotte Green, Bente Klarlund Pedersen (167-171).
Although some myokines exert their actions on other organs in a hormone-like fashion, many of them operatelocally on skeletal muscle themselves. Myokines may thereby provide a feedback loop for the muscle to regulate its owngrowth and regeneration allowing for adaptation to exercise training. The myokine concept provides a conceptual basisand a whole new paradigm for understanding how muscles communicate with other organs within the muscle itself. Newbiotechnological patents are published based on the identification of new myokines, and these myokines and their receptorswill potentially serve as pharmacological targets for treating muscle diseases, metabolic disorders and other diseasesassociated with muscle disuse.

Hyperthermia: From Diagnostic and Treatments to New Discoveries by Sandra Romero-Suarez, Chenglin Mo, Chad Touchberry, Nuria Lara, Kendra Baker, Robin Craig, Leticia Brotto, Jon Andresen, Michael Wacker, Simon Kaja, Eduardo Abreu, Wolfgang Dillmann, Ruben Mestril, Marco Brotto, Thomas Nosek (172-183).
Hyperthermia is an important approach for the treatment of several diseases. Hyperthermia is also thought toinduce hypertrophy of skeletal muscles in vitro and in vivo, and has been used as a therapeutic tool for millennia. In thefirst part of our work, we revise several relevant patents related to the utilization of hyperthermia for the treatment and diagnosticof human diseases. In the second part, we present exciting new data on the effects of forced and natural overexpressionof HSP72, using murine in vitro (muscle cells) and ex vivo (primary skeletal muscles) models. These studies helpto demonstrate that hyperthermia effects are orchestrated by tight coupling between gene expression, protein function, andintracellular Ca2+ signaling pathways with a key role for calcium-induced calcium release. We hope that the review of currentpatents along with previous unknown information on molecular signaling pathways that underlie the hypertrophy responseto hyperthermia in skeletal muscles may trigger the curiosity of scientists worldwide to explore new inventionsthat fully utilize hyperthermia for the treatment of muscle diseases.

Hyperkalemic periodic paralysis (HyperKPP) is a disease characterized by periods of myotonic discharges andparalytic attacks causing weakness, the latter associated with increases in plasma [K+]. The myotonic discharge is due toincreased Na+ influx through defective Na+ channels that triggers generation of several action potentials. The subsequentincrease in extracellular K+ concentration causes excessive membrane depolarization that inactivates Na+ channels triggeringthe paralysis. None of the available treatments is fully effective. This paper reviews the capacity of Na+ K+ ATPasepumps, KATP and ClC-1 Cl- channels in improving membrane excitability during muscle activity and how using thesethree membrane components we can study future and more effective treatments for HyperKPP patients. The review ofcurrent patents related to HyperKPP reinforces the need of novel approaches for the treatment of this channelopathy.

Jatropha curcas: From Biodiesel Generation to Medicinal Applications by Luis Cesar Paulillo, ChengLin Mo, Janalee Isaacson, Luciene Lessa, Edjacy Lopes, Sandra Romero-Suarez, Leticia Brotto, Eduardo Abreu, William Gutheil, Marco Brotto (192-199).
Jatropha curcas (JC) is a multipurpose perennial plant that belongs to the Euphorbiaceae family and is nativeto arid and semiarid tropical regions worldwide. It has many attributes and considerable potential for renewable energy,fish and livestock feeding. Despite its rich application as a renewable source and for animal feeding, JC has barely beenexplored for its medicinal potential. Here we review several patents related to JC that show it has been underused for medicinalpurposes. For example, only one invention disclosure to date utilizes JC, combined with three other plants, in apreparation for wound healing. Motivated by support from the Brazilian funding agencies and anecdotal accounts in Brazilof the medicinal value of JC, we performed a series of pilot studies that demonstrate that JC is able to protect skeletalmuscle cells in vitro against the deleterious effects of ethanol. We were able to determine that JC's effects are mediated bythe up regulation of HSP60, a critical mitochondrial heat shock related protein that is essential for intracellular REDOXregulation. Given the fact that ethanol myopathy accounts for more than 50% of all cases of myopathy worldwide, wehope that our studies will sparkle new interest from the scientific community to explore the medicinal properties of Jatrophacurcas, including the development of new patents leading to new drugs and new targets for the treatment of musclediseases and other human diseases.

Poloxamer 188 (P188) as a Membrane Resealing Reagent in Biomedical Applications by Joseph G. Moloughney, Noah Weisleder (200-211).
Maintenance of the integrity of the plasma membrane is essential for maintenance of cellular function and preventionof cell death. Since the plasma membrane is frequently exposed to a variety of mechanical and chemical insultsthe cell has evolved active processes to defend against these injuries by resealing disruptions in the plasma membrane.Cell membrane repair is a conserved process observed in nearly every cell type where intracellular vesicles are recruitedto sites of membrane disruption where they can fuse with themselves or the plasma membrane to create a repair patch.When disruptions are extensive or there is an underlying pathology that reduces the membrane repair capacity of a cellthis defense mechanism may prove insufficient and the cell could die due to breakdown of the plasma membrane. Extensiveloss of cells can compromise the integrity and function of tissues and leading to disease. Thus, methods to increasemembrane resealing capacity could have broad utility in a number of disease states. Efforts to find reagents that canmodulate plasma membrane reseal found that specific tri-block copolymers, such as poloxamer 188 (P188, or PluronicF68), can increase the structural stability and resealing of the plasma membrane. Here we review several current patentsand patent applications that present inventions making use of P188 and other copolymers to treat specific disease statessuch as muscular dystrophy, heart failure, neurodegenerative disorders and electrical injuries, or to facilitate biomedicalapplications such as transplantation. There appears to be promise for the application of poloxamers in the treatment ofvarious diseases, however there are potential concerns with toxicity with long term application and bioavailability in somecases.

Magnesium (Mg2+) is used pharmacologically to sedate specific forms of arrhythmias. Administration of pharmacologicaldoses of catecholamine or adrenergic receptor agonists often results in arrhythmias onset. Results from thepresent study indicate that stimulation of cardiac adrenergic receptors elicits an extrusion of cellular Mg2+ into the extracellularspace. This effect occurs in both perfused hearts and isolated cells within 5-6 min following either ?- or ?1-adrenergic receptor stimulation, and is prevented by specific adrenergic receptors antagonists. Sequential stimulation ofthe two classes of adrenergic receptor results in a larger mobilization of cellular Mg2+ provided that the two agonists areadministered together or within 1-2 min from each other. A longer delay in administering the second stimulus results inthe abolishment of Mg2+ extrusion. Hence, these data suggest that the stimulation of ?- and ?1-adrenergic receptors mobilizesMg2+ from two distinct cellular pools, and that Mg2+ loss from either pool triggers a Mg2+ redistribution within thecardiac myocyte. At the sarcolemmal level, Mg2+ extrusion occurs through a Na+/Mg2+ exchange mechanism phosphorylatedby cAMP. Administration of quinidine, a patent anti-arrhythmic agent, blocks Na+ transport in a non-specific mannerand prevents Mg2+ extrusion. Taken together, these data indicate that catecholamine administration induces dynamicchanges in total and compartmentalized Mg2+ pools within the cardiac myocytes, and suggest that prevention of Mg2+ extrusionand redistribution may be an integral component of the effectiveness of quinidine and possibly other cardiac antiarrhythmicagents. Confirmation of this possibility by future experimental and clinical studies might result in new patentsof these compounds as Mg2+ preserving agents.

Prostaglandin E2: From Clinical Applications to Its Potential Role in Bone- Muscle Crosstalk and Myogenic Differentiation by Chenglin Mo, Sandra Romero-Suarez, Lynda Bonewald, Mark Johnson, Marco Brotto (223-229).
Prostaglandin E2 (PGE2), a prostanoid synthesized from arachidonic acid via the cyclooxygenase pathway, is amodulator of physiological responses including inflammation, fever, and muscle regeneration. Several patents have beenfiled that are related to PGE2, one of them being directly related to skeletal muscles. In this report, we first summarize thekey patents describing inventions for the utilization of PGE2 for either diagnostic or therapeutic purposes, including skeletalmuscle. In the second part of our work we present new and exciting data that demonstrates that PGE2 accelerates skeletalmuscle myogenic differentiation. Our discovery resulted from our recent and novel concept of bone-muscle crosstalk.Bone and muscle are anatomically intimate endocrine organs and we aimed to determine whether this anatomical intimacyalso translates into a biochemical communication from bone cells to muscle cells at the in vitro level. The effects of MLOY4osteocyte-like cell conditioned medium (CM) and three osteocyte-secreted factors, PGE2, sclerostin and monocytechemotactic protein (MCP-3), on C2C12 myogenic differentiation were evaluated using morphological analyses, a customized96-gene PCR array, and measurements of intracellular calcium levels. MLO-Y4 CM and PGE2, but not sclerostinand MCP-3, induced acceleration of myogenesis of C2C12 myoblasts that was linked with significant modifications in intracellularcalcium homeostasis. This finding should further stimulate the pursuit of new patents to explore the use ofPGE2 and the new concept of bone-muscle crosstalk for the development and application of inventions designed to treatmuscle diseases characterized by enhanced muscle wasting, such as sarcopenia.

Genome-wide association study (GWAS) has become a commonly adopted approach for revealing the geneticarchitecture of complex diseases, with respect to uncovering the unknown genetic variants involved in the disease, theirvariations in the population and the magnitude of their effects. Though a substantial number of disease-susceptibility variantshave been identified, the genetic architecture of complex diseases has remained elusive. It is unclear how many geneticvariants in the human genome are associated with diseases, and how the genetic variants interact with one another tocause diseases. This challenge is partly due to the pervasive gene-gene interactions that underlie complex human diseases.Whereas a number of statistical methods have been developed for detecting gene-gene interactions, they are designed forvarious purposes, such as a particular study design, the order of the interactions being examined, and the measurement ofdisease phenotypes. This paper provides a survey of the currently available statistical methods and patents from the perspectiveof their application to various types of phenotypic traits. We also discuss the strength of each method as well asthe biological interpretation of results.