© The American Society of Gene & Cell Therapy
references
1. Aiuti, A, Cattaneo, F, Galimberti, S, Benninghoff, U, Cassani, B, Callegaro, L et al. (2009). Gene therapy for immunodeficiency due to adenosine deaminase deficiency. N Engl J Med 360: 447–458. 2. Biffi, A, Montini, E, Lorioli, L, Cesani, M, Fumagalli, F, Plati, T et al. (2013). Lentiviral hematopoietic stem cell gene therapy benefits metachromatic leukodystrophy. Science 341: 1233158. 3. Naldini, L (2011). Ex vivo gene transfer and correction for cell-based therapies. Nat Rev Genet 12: 301–315. 4. Chuah, MK, Evens, H and VandenDriessche, T (2013). Gene therapy for hemophilia. J Thromb Haemost 11 (suppl. 1): 99–110. 5. Moayeri, M, Ramezani, A, Morgan, RA, Hawley, TS and Hawley, RG (2004). Sustained phenotypic correc-
commentary
tion of hemophilia A mice following oncoretroviralmediated expression of a bioengineered human factor VIII gene in long-term hematopoietic repopulating cells. Mol Ther 10: 892–902. 6. Follenzi, A, Raut, S, Merlin, S, Sarkar, R and Gupta, S (2012). Role of bone marrow transplantation for correcting hemophilia A in mice. Blood 119: 5532–5542. 7. Shi, Q, Wilcox, DA, Fahs, SA, Fang, J, Johnson, BD, Du, LM et al. (2007). Lentivirus-mediated platelet-derived factor VIII gene therapy in murine haemophilia A. J Thromb Haemost 5: 352–361. 8. Kuether, EL, Schroeder, JA, Fahs, SA, Cooley, BC, Chen, Y, Montgomery, RR et al. (2012). Lentivirusmediated platelet gene therapy of murine hemophilia A with pre-existing anti-factor VIII immunity. J Thromb Haemost 10: 1570–1580.
9. Wang, X, Shin, SC, Chiang, AFJ, Khan, I, Pan, D, Rawlings, DJ et al. (2015). Intraosseous delivery of lentiviral vectors targeting factor VIII expression in platelets corrects murine hemophilia A. Mol Ther 23: 617–626 10. Agudo, J, Ruzo, A, Kitur, K, Sachidanandam, R, Blander, JM and Brown, BD (2012). A TLR and non-TLR mediated innate response to lentiviruses restricts hepatocyte entry and can be ameliorated by pharmacological blockade. Mol Ther 20: 2257–2267. 11. Wang, CX, Sather, BD, Wang, X, Adair, J, Khan, I, Singh, S et al. (2013). Rapamycin relieves lentiviral vector transduction resistance in human and mouse hematopoietic stem cells. Blood 124: 913–923.
See page 707
Antiobesity Strategy Targets Energy Economy Safeguards Michel Vivaudou1–3 and André Terzic4 doi:10.1038/mt.2015.39
A
fundamental barrier in obesity management is that caloric restriction triggers energy-conserving responses that evolved to prevent body weight loss. ATPsensitive potassium (KATP) channels have been identified as safeguards controlling energy expenditure in skeletal muscles and thereby key factors determining body weight.1 In this issue of Molecular Therapy, Koganti and colleagues report the successful reduction of muscle energy efficiency through targeted intramuscular injections of cell-penetrating vivo-morpholinos to prevent translation of the channel poreforming Kir6.2 subunit.2 In this elegant proof-of-concept study, the authors demonstrate localized reduction of KATP channel expression and function, leading in turn to an increase in activity-related energy consumption, without compromising exercise tolerance. This report opens a new avenue of investigation in targeted 1 Université Grenoble Alpes, Institut de Biologie Structurale, Grenoble, France; 2Centre National de la Recherche Scientifique, Institut de Biologie Structurale, Grenoble, France; 3CEA, Institut de Biologie Structurale, Grenoble, France; 4Center for Regenerative Medicine, Mayo Clinic, Rochester, Minnesota, USA Correspondence: Michel Vivaudou, Institut de Biologie Structurale, 71 Avenue des Martyrs, 38044 Grenoble, France. E-mail: vivaudou@ ibs.fr or André Terzic, Center for Regenerative Medicine, Mayo Clinic, 200 First Street SW, Rochester, Minnesota 55905, USA. E-mail: [email protected]
Molecular Therapy vol. 23 no. 4 april 2015
therapies aiming to control weight management. Obesity reflects an imbalance between calorie intake and expenditure. At present, more than 1 billion adults worldwide are considered overweight, underscoring a rampant epidemic.3 The global prevalence of obesity has precipitated a major escalation in comorbidities associated with an increase in overall mortality.4 Beyond intensive counseling and change in lifestyle, diverse strategies targeting weight loss are being pursued.5 Progress is reflected by US Food and Drug Administration approval of new medications for chronic weight management in obese patients, such as lorcaserin and phentermine/topiramate. Yet antiobesity drug therapy has been largely unsuccessful because of lack of efficacy, poor adherence, and adverse effects.6 Elucidating the molecular pathways that underlie the caloric intake–expenditure equilibrium is necessary to inform the selection of promising therapeutic targets. Indeed, a deeper insight into the innate mechanisms regulating appetite, nutrient exposure, and energy balance is warranted to aid in the discovery and future development of next-generation therapies. Stimulation of energy expenditure may be a potent strategy for obesity treatment. KATP channels, expressed at high density in striated muscles and other excitable tissues, are established membrane sensors of ATP/ADP.7–9 It has been postulated that
under conditions of energy deficit, activation of channel complexes would result in protective energy economy, whereas under energy surplus, downregulation of KATP channels would increase thermogenesis.10 As such, KATP channels provide a low-fuel warning that signals muscle fibers to slow down and avoid irreversible energy depletion. Through tight regulation by adenine nucleotides and integration with metabolic pathways, muscle KATP channels seem to sense both static metabolic levels and the dynamics of energy consumption, thus maintaining an optimal balance between lost heat production and useful mechanical work.11 Without functional KATP channels, energy efficiency decreases and muscles burn more calories than normal.1 Thus, weight loss could be achieved without additional exercise, by “simply” reducing the activity of skeletal muscle KATP channels. A pharmacological approach is conceivable, as numerous molecules are known to modulate KATP channels by binding to their regulatory SUR subunit.12 Unfortunately, blockers of KATP channels specific for skeletal muscle do not yet exist and are unlikely to be discovered soon, given that comparable channels (incorporating isoform SUR2) are also found in cardiac and smooth muscle.13 In the new work, Koganti and colleagues2 used an alternative approach of reducing protein levels by leveraging antisense oligonucleotides packaged as a 615
© The American Society of Gene & Cell Therapy
commentary morpholino for stability and coupled to a guanidine dendrimer for intracellular delivery (“vivo-morpholinos”).14 Using mice, the authors demonstrate that it is feasible to selectively knock down KATP channels in designated skeletal muscles by intramuscular injections of vivo-morpholinos targeting the Kir6.2 pore subunit of the channels.2 At the molecular level, application of vivo-morpholinos decreased Kir6.2 protein levels threefold (measured with western blot) and decreased channel functional expression to the same extent (measured by the patch-clamp technique). Importantly, these effects persisted for at least a week, and nearby muscles were not affected. These measurements demonstrate the efficacy of vivo-morpholinos in achieving a significant and long-lasting reduction of targeted KATP channel activity.2 The response of the fibers to exercise was assessed by monitoring membrane potential, action potential characteristics, and force. At rest, where it is thought that KATP channels are mostly inactive, there was little difference between treated and untreated fibers. Under prolonged stimulation, skeletal muscle adaptation is seen by a steady decline in action potential amplitude and force. Experiments on whole animals showed that exercising treated subjects produced more heat and consumed more oxygen than did the same treatment of controls. Based on these results, the authors suggest that antiKir6.2 vivo-morpholinos could be used to treat obesity, as they document that it is possible to reduce the expression of KATP channels in a specific muscle, thus directing that muscle to burn more calories than usual without disabling the muscle’s mechanical function.2 Selective knockdown in designated muscle areas seems particularly important, in that previous studies using Kir6.2-knockout models have implicated Kir6.2-dependent KATP channel activity required for attainment of the physiological benefits of exercise without injury.15 The study by Koganti et al. thus provides an original approach to target KATP channel-dependent energy economy, introducing a fit-for-purpose experimental tool and opening a spectrum of new con-
616
siderations. Which and how many muscles should be targeted to achieve significant weight loss? How predictable, how persistent, and how stable are the effects of vivomorpholinos on channel density and ultimate outcome? Is there a concomitant loss of myoprotection conferred by KATP channels that would thereby limit treatment to nonexercising subjects? To what extent are genetic manipulation underpinning therapy and the invasive mode of intramuscular delivery, though feasible, safe and effective in the long term? Thus, the translational value of this promising approach, as a means to manage obesity, is predicated on further testing with demonstration of spatial and temporal effectiveness, and of a safe therapeutic index. references
1. Alekseev, AE, Reyes, S, Yamada, S, Hodgson-Zingman, DM, Sattiraju, S, Zhu, Z et al. (2010). Sarcolemmal ATP-sensitive K+ channels control energy expenditure determining body weight. Cell Metab 11: 58–69. 2. Koganti, SRK, Zhu, Z, Subbotina, E, Gao, Z, Sierra, A, Proenza, M et al. (2015). Disruption of KATP channel expression in skeletal muscle by targeted oligonucleotide delivery promotes activity-linked thermogenesis. Mol Ther 23: 707–716 3. Valentino, MA, Terzic, A and Waldman, SA (2010). Sizing up pharmacotherapy for obesity. Clin Transl Sci 3: 123–125. 4. Terzic, A and Waldman, S (2011). Chronic diseases: the emerging pandemic. Clin Transl Sci 4: 225–226. 5. Rueda-Clausen, CF, Padwal, RS and Sharma, AM (2013). New pharmacological approaches for obesity management. Nat Rev Endocrinol 9: 467–478. 6. Kim, GW, Lin, JE, Blomain, ES and Waldman, SA (2014). Antiobesity pharmacotherapy: new drugs and emerging targets. Clin Pharmacol Ther 95: 53–66. 7. Flagg, TP, Enkvetchakul, D, Koster, JC and Nichols, CG (2010). Muscle KATP channels: recent insights to energy sensing and myoprotection. Physiol Rev 90: 799–829. 8. Olson, TM and Terzic, A (2010). Human KATP channelopathies: diseases of metabolic homeostasis. Pflugers Arch 460: 295–306. 9. Terzic, A, Alekseev, AE, Yamada, S, Reyes, S and Olson, TM (2011). Advances in cardiac ATP-sensitive K+ channelopathies from molecules to populations. Circ Arrhythm Electrophysiol 4: 577–585. 10. Reyes, S, Park, S, Terzic, A and Alekseev, AE (2010). KATP channels process nucleotide signals in muscle thermogenic response. Crit Rev Biochem Mol Biol 45: 506–519. 11. Alekseev, AE, Hodgson, DM, Karger, AB, Park, S, Zingman, LV and Terzic, A (2005). ATP-sensitive K+ channel channel/enzyme multimer: metabolic gating in the heart. J Mol Cell Cardiol 38: 895–905. 12. Jahangir, A and Terzic, A (2005). KATP channel therapeutics at the bedside. J Mol Cell Cardiol 39: 99–112. 13. Moreau, C, Prost, AL, Dérand, R and Vivaudou, M (2005). SUR, ABC proteins targeted by KATP channel openers. J Mol Cell Cardiol 38: 951–963. 14. Morcos, PA, Li, Y and Jiang, S (2008). Vivo-Morpholinos: a non-peptide transporter delivers Morpholinos into a wide array of mouse tissues. Biotechniques 45: 613–623. 15. Kane, GC, Behfar, A, Yamada, S, Perez-Terzic, C, O’Cochlain, F, Reyes, S et al. (2004). ATP-sensitive K+ channel knockout compromises the metabolic benefit of exercise training, resulting in cardiac deficits. Diabetes 53 (suppl. 3): S169–S175.
www.moleculartherapy.org vol. 23 no. 4 april 2015
references
1. Aiuti, A, Cattaneo, F, Galimberti, S, Benninghoff, U, Cassani, B, Callegaro, L et al. (2009). Gene therapy for immunodeficiency due to adenosine deaminase deficiency. N Engl J Med 360: 447–458. 2. Biffi, A, Montini, E, Lorioli, L, Cesani, M, Fumagalli, F, Plati, T et al. (2013). Lentiviral hematopoietic stem cell gene therapy benefits metachromatic leukodystrophy. Science 341: 1233158. 3. Naldini, L (2011). Ex vivo gene transfer and correction for cell-based therapies. Nat Rev Genet 12: 301–315. 4. Chuah, MK, Evens, H and VandenDriessche, T (2013). Gene therapy for hemophilia. J Thromb Haemost 11 (suppl. 1): 99–110. 5. Moayeri, M, Ramezani, A, Morgan, RA, Hawley, TS and Hawley, RG (2004). Sustained phenotypic correc-
commentary
tion of hemophilia A mice following oncoretroviralmediated expression of a bioengineered human factor VIII gene in long-term hematopoietic repopulating cells. Mol Ther 10: 892–902. 6. Follenzi, A, Raut, S, Merlin, S, Sarkar, R and Gupta, S (2012). Role of bone marrow transplantation for correcting hemophilia A in mice. Blood 119: 5532–5542. 7. Shi, Q, Wilcox, DA, Fahs, SA, Fang, J, Johnson, BD, Du, LM et al. (2007). Lentivirus-mediated platelet-derived factor VIII gene therapy in murine haemophilia A. J Thromb Haemost 5: 352–361. 8. Kuether, EL, Schroeder, JA, Fahs, SA, Cooley, BC, Chen, Y, Montgomery, RR et al. (2012). Lentivirusmediated platelet gene therapy of murine hemophilia A with pre-existing anti-factor VIII immunity. J Thromb Haemost 10: 1570–1580.
9. Wang, X, Shin, SC, Chiang, AFJ, Khan, I, Pan, D, Rawlings, DJ et al. (2015). Intraosseous delivery of lentiviral vectors targeting factor VIII expression in platelets corrects murine hemophilia A. Mol Ther 23: 617–626 10. Agudo, J, Ruzo, A, Kitur, K, Sachidanandam, R, Blander, JM and Brown, BD (2012). A TLR and non-TLR mediated innate response to lentiviruses restricts hepatocyte entry and can be ameliorated by pharmacological blockade. Mol Ther 20: 2257–2267. 11. Wang, CX, Sather, BD, Wang, X, Adair, J, Khan, I, Singh, S et al. (2013). Rapamycin relieves lentiviral vector transduction resistance in human and mouse hematopoietic stem cells. Blood 124: 913–923.
See page 707
Antiobesity Strategy Targets Energy Economy Safeguards Michel Vivaudou1–3 and André Terzic4 doi:10.1038/mt.2015.39
A
fundamental barrier in obesity management is that caloric restriction triggers energy-conserving responses that evolved to prevent body weight loss. ATPsensitive potassium (KATP) channels have been identified as safeguards controlling energy expenditure in skeletal muscles and thereby key factors determining body weight.1 In this issue of Molecular Therapy, Koganti and colleagues report the successful reduction of muscle energy efficiency through targeted intramuscular injections of cell-penetrating vivo-morpholinos to prevent translation of the channel poreforming Kir6.2 subunit.2 In this elegant proof-of-concept study, the authors demonstrate localized reduction of KATP channel expression and function, leading in turn to an increase in activity-related energy consumption, without compromising exercise tolerance. This report opens a new avenue of investigation in targeted 1 Université Grenoble Alpes, Institut de Biologie Structurale, Grenoble, France; 2Centre National de la Recherche Scientifique, Institut de Biologie Structurale, Grenoble, France; 3CEA, Institut de Biologie Structurale, Grenoble, France; 4Center for Regenerative Medicine, Mayo Clinic, Rochester, Minnesota, USA Correspondence: Michel Vivaudou, Institut de Biologie Structurale, 71 Avenue des Martyrs, 38044 Grenoble, France. E-mail: vivaudou@ ibs.fr or André Terzic, Center for Regenerative Medicine, Mayo Clinic, 200 First Street SW, Rochester, Minnesota 55905, USA. E-mail: [email protected]
Molecular Therapy vol. 23 no. 4 april 2015
therapies aiming to control weight management. Obesity reflects an imbalance between calorie intake and expenditure. At present, more than 1 billion adults worldwide are considered overweight, underscoring a rampant epidemic.3 The global prevalence of obesity has precipitated a major escalation in comorbidities associated with an increase in overall mortality.4 Beyond intensive counseling and change in lifestyle, diverse strategies targeting weight loss are being pursued.5 Progress is reflected by US Food and Drug Administration approval of new medications for chronic weight management in obese patients, such as lorcaserin and phentermine/topiramate. Yet antiobesity drug therapy has been largely unsuccessful because of lack of efficacy, poor adherence, and adverse effects.6 Elucidating the molecular pathways that underlie the caloric intake–expenditure equilibrium is necessary to inform the selection of promising therapeutic targets. Indeed, a deeper insight into the innate mechanisms regulating appetite, nutrient exposure, and energy balance is warranted to aid in the discovery and future development of next-generation therapies. Stimulation of energy expenditure may be a potent strategy for obesity treatment. KATP channels, expressed at high density in striated muscles and other excitable tissues, are established membrane sensors of ATP/ADP.7–9 It has been postulated that
under conditions of energy deficit, activation of channel complexes would result in protective energy economy, whereas under energy surplus, downregulation of KATP channels would increase thermogenesis.10 As such, KATP channels provide a low-fuel warning that signals muscle fibers to slow down and avoid irreversible energy depletion. Through tight regulation by adenine nucleotides and integration with metabolic pathways, muscle KATP channels seem to sense both static metabolic levels and the dynamics of energy consumption, thus maintaining an optimal balance between lost heat production and useful mechanical work.11 Without functional KATP channels, energy efficiency decreases and muscles burn more calories than normal.1 Thus, weight loss could be achieved without additional exercise, by “simply” reducing the activity of skeletal muscle KATP channels. A pharmacological approach is conceivable, as numerous molecules are known to modulate KATP channels by binding to their regulatory SUR subunit.12 Unfortunately, blockers of KATP channels specific for skeletal muscle do not yet exist and are unlikely to be discovered soon, given that comparable channels (incorporating isoform SUR2) are also found in cardiac and smooth muscle.13 In the new work, Koganti and colleagues2 used an alternative approach of reducing protein levels by leveraging antisense oligonucleotides packaged as a 615
© The American Society of Gene & Cell Therapy
commentary morpholino for stability and coupled to a guanidine dendrimer for intracellular delivery (“vivo-morpholinos”).14 Using mice, the authors demonstrate that it is feasible to selectively knock down KATP channels in designated skeletal muscles by intramuscular injections of vivo-morpholinos targeting the Kir6.2 pore subunit of the channels.2 At the molecular level, application of vivo-morpholinos decreased Kir6.2 protein levels threefold (measured with western blot) and decreased channel functional expression to the same extent (measured by the patch-clamp technique). Importantly, these effects persisted for at least a week, and nearby muscles were not affected. These measurements demonstrate the efficacy of vivo-morpholinos in achieving a significant and long-lasting reduction of targeted KATP channel activity.2 The response of the fibers to exercise was assessed by monitoring membrane potential, action potential characteristics, and force. At rest, where it is thought that KATP channels are mostly inactive, there was little difference between treated and untreated fibers. Under prolonged stimulation, skeletal muscle adaptation is seen by a steady decline in action potential amplitude and force. Experiments on whole animals showed that exercising treated subjects produced more heat and consumed more oxygen than did the same treatment of controls. Based on these results, the authors suggest that antiKir6.2 vivo-morpholinos could be used to treat obesity, as they document that it is possible to reduce the expression of KATP channels in a specific muscle, thus directing that muscle to burn more calories than usual without disabling the muscle’s mechanical function.2 Selective knockdown in designated muscle areas seems particularly important, in that previous studies using Kir6.2-knockout models have implicated Kir6.2-dependent KATP channel activity required for attainment of the physiological benefits of exercise without injury.15 The study by Koganti et al. thus provides an original approach to target KATP channel-dependent energy economy, introducing a fit-for-purpose experimental tool and opening a spectrum of new con-
616
siderations. Which and how many muscles should be targeted to achieve significant weight loss? How predictable, how persistent, and how stable are the effects of vivomorpholinos on channel density and ultimate outcome? Is there a concomitant loss of myoprotection conferred by KATP channels that would thereby limit treatment to nonexercising subjects? To what extent are genetic manipulation underpinning therapy and the invasive mode of intramuscular delivery, though feasible, safe and effective in the long term? Thus, the translational value of this promising approach, as a means to manage obesity, is predicated on further testing with demonstration of spatial and temporal effectiveness, and of a safe therapeutic index. references
1. Alekseev, AE, Reyes, S, Yamada, S, Hodgson-Zingman, DM, Sattiraju, S, Zhu, Z et al. (2010). Sarcolemmal ATP-sensitive K+ channels control energy expenditure determining body weight. Cell Metab 11: 58–69. 2. Koganti, SRK, Zhu, Z, Subbotina, E, Gao, Z, Sierra, A, Proenza, M et al. (2015). Disruption of KATP channel expression in skeletal muscle by targeted oligonucleotide delivery promotes activity-linked thermogenesis. Mol Ther 23: 707–716 3. Valentino, MA, Terzic, A and Waldman, SA (2010). Sizing up pharmacotherapy for obesity. Clin Transl Sci 3: 123–125. 4. Terzic, A and Waldman, S (2011). Chronic diseases: the emerging pandemic. Clin Transl Sci 4: 225–226. 5. Rueda-Clausen, CF, Padwal, RS and Sharma, AM (2013). New pharmacological approaches for obesity management. Nat Rev Endocrinol 9: 467–478. 6. Kim, GW, Lin, JE, Blomain, ES and Waldman, SA (2014). Antiobesity pharmacotherapy: new drugs and emerging targets. Clin Pharmacol Ther 95: 53–66. 7. Flagg, TP, Enkvetchakul, D, Koster, JC and Nichols, CG (2010). Muscle KATP channels: recent insights to energy sensing and myoprotection. Physiol Rev 90: 799–829. 8. Olson, TM and Terzic, A (2010). Human KATP channelopathies: diseases of metabolic homeostasis. Pflugers Arch 460: 295–306. 9. Terzic, A, Alekseev, AE, Yamada, S, Reyes, S and Olson, TM (2011). Advances in cardiac ATP-sensitive K+ channelopathies from molecules to populations. Circ Arrhythm Electrophysiol 4: 577–585. 10. Reyes, S, Park, S, Terzic, A and Alekseev, AE (2010). KATP channels process nucleotide signals in muscle thermogenic response. Crit Rev Biochem Mol Biol 45: 506–519. 11. Alekseev, AE, Hodgson, DM, Karger, AB, Park, S, Zingman, LV and Terzic, A (2005). ATP-sensitive K+ channel channel/enzyme multimer: metabolic gating in the heart. J Mol Cell Cardiol 38: 895–905. 12. Jahangir, A and Terzic, A (2005). KATP channel therapeutics at the bedside. J Mol Cell Cardiol 39: 99–112. 13. Moreau, C, Prost, AL, Dérand, R and Vivaudou, M (2005). SUR, ABC proteins targeted by KATP channel openers. J Mol Cell Cardiol 38: 951–963. 14. Morcos, PA, Li, Y and Jiang, S (2008). Vivo-Morpholinos: a non-peptide transporter delivers Morpholinos into a wide array of mouse tissues. Biotechniques 45: 613–623. 15. Kane, GC, Behfar, A, Yamada, S, Perez-Terzic, C, O’Cochlain, F, Reyes, S et al. (2004). ATP-sensitive K+ channel knockout compromises the metabolic benefit of exercise training, resulting in cardiac deficits. Diabetes 53 (suppl. 3): S169–S175.
www.moleculartherapy.org vol. 23 no. 4 april 2015
