Longevity Gene Therapy – Updated Projects


While discussing the longevity gene therapy project we encountered various questions and observations that prompted us to broaden the project and slightly change it. Generally, all the comments can be reduced into 5 main points:

  1. You need to enlarge the list of therapeutic genes by adding to it this and that.
  2. You want to use too many genes; therefore you need to make the project simpler by keeping only the most effective genes
  3. If you apply all the genes at the same time, some of them may cancel out the effects of other genes.
  4. Will it be safe to use viral vectors to deliver genetic constructs?
  5. How safe are therapeutic genes for the body?

Some of the observations were of completely opposite nature, so we decided to do 2 versions of the project. One of them is for aging geneticists. In it we almost double the list of the genes extending lifespan. This project will allow testing many poorly studied genes, but promising in terms of aging. Besides, some unexpected results can be obtained, which is always valuable.

The socond version of the project, on the contrary, has only 5 best studies and most effective genes. This project description focuses on the safety issues. We will use adeno-associated viruses for gene delivery that don’t integrate into the genome, which means that the possibility of insertion mutagenesis is eliminated. Plus, adeno-associated viruses cause really low immune response. To reduce potential side effects of delivered genes we will make their expression inducible. This means that the genes will only be activated upon adding a specific drug. Therefore, it will be possible to control the time during which the therapeutic gene is active. Moreover, in the case of delivering telomerase gene in order to decrease the probability of oncogenesis we will not only optimise the duration of its expression, but will also deliver it together with p53 gene, which is a tumorsuppressor. The second version of the project can be seen as a preliminary step before the clinical studies.

One of the comments mentioned the possibility of genes having neutrolizing effects on one onther. In order to aviod it and also to identify genes that may have a cumulative effect on longevity we will study the genes in different combinations.

On another note, we previous version of the project suggested using both aging modifying genes and those affecting a single age-related pathology, for example preventing age-related deline in cardio-vascular function, or immune system activity, or aging conitive decline, etc. However, a number of publication states that life extension is accompanied by slowing down the development of various age-related diseases. For example, delivering the telomerase gene not only extends lifespan, but also prevents aging-associated decline in bone density, subcutaneous fat, deficits in motor activity, etc. Therefore, to eliminate decrepitude, it may be more promising to use one gene that extends longevity than a whole set dealing with various pathologies. Besides, delivering a large number of genes is tied to higher strain on the body and may yield side-effects. That’s why we removed the genes that prevent aging changes in separate tissues.

Longevity Gene Therapy – Longer Version

Project description

Gene engineering is the most powerful existing tool for life extension. Mutations in certain genes result in up to 10-fold increase in nematode lifespan and in up to 2-fold increase in a mouse life expectancy. Gene therapy represents a unique tool to transfer achievements of gene engineering into medicine. This approach has already been proven successful for treatment of numerous diseases, in particular those of genetic and multigenic nature. More than 2000 clinical trials have been launched to date.

We propose developing a gene therapy that will radically extend lifespan. Genes that promote longevity of model animals will be used as therapeutic agents. We will manipulate not a single gene, but several aging mechanisms simultaneously. A combination of different approaches may lead to an additive or even a synergistic effect, resulting in a very long life expectancy. For this purpose, an animal will be affected by a set of genes that contribute to longevity and prevent age-related pathologies.

As a result, we will develop a comprehensive treatment that will dramatically extend lifespan and prevent the decrepitude of the body. Experiments will be conducted in old mice. Thus, in case of success, the developed method of aging treatment can be quickly moved to clinical trials.

The goal of the project is to develop a complex gene therapy that will drastically increase mouse lifespan and prevent tissue pathology in old age, coupled with the safety assessment of the treatment.

Project description

20 genes that are most promising in terms of life extension (table 1) will be used as targets for gene therapy. We will affect both the biological aging mechanisms, common to the majority of the cells, as well as the primary neuroendocrine center, that regulates the whole organism’s longevity – the hypothalamus. The expression increase or decrease of these genes in animal models was shown to result in boosted longevity. If the increase in expression of a particular gene is necessary for longevity, we will deliver this gene into the body. If, on the other hand, longevity depends on the inhibition of a certain gene’s expression, we will introduce a genetic construct that encodes small RNAs that inhibit the expression of the target gene. Two out of 20 genes have previously been used for gene therapy of aging: the lifespan of mice was increased by 20% (Zhang et al., 2013, Bernardes de Jesus et al., 2012). Moreover, delivery of these genes resulted in prevention of numerous age-related pathologies.

Therapeutic genes will be introduced into the body using viral vectors – the most powerful method of delivering genetic constructs. In case of gene delivery to hypothalamus therapeutic construct will be injected directly into this part of the brain. For other genes we will perform systemic gene delivery into the bloodstream.

  • At first, we will reveal the most effective longevity promoting genes. For this purpose, each of the therapeutic genes will be tested individually and increase in lifespan will be evaluated. All the experiments will be conducted in 2-year old mice. We will use untreated old and young animals as a control.
  1. The efficiency of therapeutic gene delivery will be tested.
  2. All groups of mice will be regularly tested for aging markers, behavioral test will be performed, average and maximum lifespan of mice will be determined. In addition, a detailed study of side effects will be performed. Mice will be compared with old and young mice of the control groups.
  3. We will select 5 most effective and safe longevity-promoting genes.
  • Then we will reveal the most effective combinations of the selected longevity promoting genes. We will study all genes in combinations of 2, 3 and 4. We will also utilize all 5 genes simultaneously. The impact of gene combinations on mouse lifespan will be compared to the impact of these genes applied individually as well as to untreated old and young mice.
  1. The efficiency of therapeutic gene delivery will be tested.
  2. All groups of mice will be regularly tested for aging markers, behavioral test will be performed, average and maximum lifespan of mice will be determined. In addition, a detailed study of side effects will be performed. Mice will be compared with old and young mice of the control groups.
  3. Thus, we will reveal the most efficient and safe combination of genes, which promote longevity and prevent age-related pathologies.

Table 1. Target genes for life extending gene therapy.

Target gene The impact on gene expression Therapeutic effect Research links
Gene delivery to hypothalamus
NF-кВ Expression inhibition The inhibition of NF-KB transcription factor reduces inflammation in hypothalamus during aging and increases mouse lifespan by 23%. Zhang et al., 2013
UCP2 Overexpression Uncoupling protein 2 elevates the temperature of the hypothalamus, which is accompanied by a slight decrease in the overall body temperature and increased lifespan (20%) in mice Conti et al., 2006
Systemic gene delivery
TERT Overexpression The catalytic subunit of the telomerase extends the telomeres, which increase the replicative potential of cells and longevity of the body (13-20%) Bernardes de Jesus et al., 2012
CRTC1 Expression inhibition Inhibiting TOR-kinase, which promotes cell growth and proliferation, leads to increased lifespan (20%) in mice Wu et al., 2013
GHR Expression inhibition Growth hormone receptor, which inhibition results in dwarf phenotype with significantly extended lifespan (40-50%) Coschigano et al., 2003
Igf1r Expression inhibition insulin-like growth factor 1 receptor mediates growth. Igf1r +/- mice live 26% longer than controls. Holzenberger et al., 2003
Adcy5 Expression inhibition Adenylate cyclase 5 gene knockout in mice results in resistance to cardiac stress and an increased median lifespan of 30% Yan et al., 2007
NUDT1 Overexpression NUDT1 participates in oxidative damage repair and its overexpression results in 16% increase of mouse lifespan. De Luca et al., 2013
LINE retrotransposons and endogenous retroviruses Expression inhibition Inhibiting retrotransposon expression leads to reduced genetic instability in old age Wang et al., 2011
Suv39h1 Overexpression Suv39h methyltransferase directs H3K9me3 at heterochromatin and opposes age-dependent loss of gene silencing. Helfand et al., 2014
FOXO3 Overexpression A transcription factor that triggers stress response and promotes longevity Brunet et al., 2004
TFEB Overexpression A transcription factor that activates autophagy and leads to longevity Lapierre et al., 2013
ELAVL1 Overexpression RNA – binding protein HuR stabilizes mRNA of factors regulating the cell cycle. The overexpression of HuR leads to rejuvenation of senescent cells Wang et al., 2001
SIRT6 Overexpression Overexpression of sirtuin 6 – a NAD + -dependent deacetylase, leads to an increase in life expectancy Kanfi et al., 2012
AMPK genes Overexpression AMPK overexpression triggers stress response and promotes longevity Greer et al., 2007
Nrf2 Overexpression NFE2-related factor 2 is a stress-sensing transcription factor, which mediates longevity Sykiotis et al., 2008
Gdf11 Overexpression Growth differentiation factor 11 is a rejuvenating factor for skeletal muscle Sinha et al., 2014
RAGE Expression inhibition RAGE – receptors for advanced glycation endproducts (AGEs) contribute to oxidative stress, inflammatory reactions and are involved in age-related diseases and aging process. Bierhaus et al., 2009
KL Overexpression Peptide hormone α-klotho reduces phosphate blood level. klotho gene mutation cause multiple aging-related disorders and shortens lifespan. Kuro-o et al., 1997
Gene delivery to senescent cells
HSV-TK Overexpression HSV-TK promotes the transformation of a non-toxic prodrug into a toxic product. HSV-TK gene is under p16 promoter, which is activated in senescent cells. Thus, exposure to the prodrug induces death of senescent cells Campisi, 2014

Literature

  1. Bernardes de Jesus B, Vera E, Schneeberger K, Tejera AM, Ayuso E, Bosch F, Blasco MA. Telomerase gene therapy in adult and old mice delays aging and increases longevity without increasing cancer. EMBO Mol Med. 2012. v4(8). P691-704.
  2. Bierhaus A, Nawroth PP. Multiple levels of regulation determine the role of the receptor for AGE (RAGE) as common soil in inflammation, immune responses and diabetes mellitus and its complications. Diabetologia. 2009. v(11). P.2251-63.
  3. Brunet A, Sweeney LB, Sturgill JF, Chua KF, Greer PL, Lin Y, Tran H, Ross SE, Mostoslavsky R, Cohen HY, Hu LS, Cheng HL, Jedrychowski MP, Gygi SP, Sinclair DA, Alt FW, Greenberg ME. Stress-dependent regulation of FOXO transcription factors by the SIRT1 deacetylase. Science. 2004. v303(5666). P2011-5.
  4. Campisi J, Report on Rejuvenation biotechnology conference, August 21-23, 2014, Santa Clara, USA
  5. Conti B, Sanchez-Alavez M, Winsky-Sommerer R, Morale MC, Lucero J, Brownell S, Fabre V, Huitron-Resendiz S, Henriksen S, Zorrilla EP, de Lecea L, Bartfai T. Transgenic mice with a reduced core body temperature have an increased life span. Science. 2006. v314(5800). P825-8.
  6. Coschigano KT, Holland AN, Riders ME, List EO, Flyvbjerg A, Kopchick JJ. Deletion, but not antagonism, of the mouse growth hormone receptor results in severely decreased body weights, insulin, and insulin-like growth factor I levels and increased life span. Endocrinology. 2003. v144(9). P3799-810.
  7. De Luca G, Ventura I, Sanghez V, Russo MT, Ajmone-Cat MA, Cacci E, Martire A, Popoli P, Falcone G, Michelini F, Crescenzi M, Degan P, Minghetti L, Bignami M, Calamandrei G. Prolonged lifespan with enhanced exploratory behavior in mice overexpressing the oxidized nucleoside triphosphatase hMTH1. Aging Cell. 2013. v12(4). P695-705.
  8. Greer EL, Dowlatshahi D, Banko MR, Villen J, Hoang K, Blanchard D, Gygi SP, Brunet A. An AMPK-FOXO pathway mediates longevity induced by a novel method of dietary restriction in C. elegans. Curr Biol. 2007. v17(19). P1646-56.
  9. Holzenberger M, Dupont J, Ducos B, Leneuve P, Géloën A, Even PC, Cervera P, Le Bouc Y. IGF-1 receptor regulates lifespan and resistance to oxidative stress in mice. Nature. 2003. v421(6919). P182-7.
  10. Kanfi Y, Naiman S, Amir G, Peshti V, Zinman G, Nahum L, Bar-Joseph Z, Cohen HY. The sirtuin SIRT6 regulates lifespan in male mice. Nature. 2012. v483(7388). P218-21.
  11. Kuro-o M, Matsumura Y, Aizawa H, Kawaguchi H, Suga T, Utsugi T, Ohyama Y, Kurabayashi M, Kaname T, Kume E, Iwasaki H, Iida A, Shiraki-Iida T, Nishikawa S, Nagai R, Nabeshima YI. Mutation of the mouse klotho gene leads to a syndrome resembling ageing. Nature. 1997. v390(6655). P45-51.
  12. Lapierre LR, De Magalhaes Filho CD, McQuary PR, Chu CC, Visvikis O, Chang JT, Gelino S, Ong B, Davis AE, Irazoqui JE, Dillin A, Hansen M. The TFEB orthologue HLH-30 regulates autophagy and modulates longevity in Caenorhabditis elegans. Nat Commun. 2013. v2267.
  13. Sinha M, Jang YC, Oh J, Khong D, Wu EY, Manohar R, Miller C, Regalado SG, Loffredo FS, Pancoast JR, Hirshman MF, Lebowitz J, Shadrach JL, Cerletti M, Kim MJ, Serwold T, Goodyear LJ, Rosner B, Lee RT, Wagers AJ. Restoring systemic GDF11 levels reverses age-related dysfunction in mouse skeletal muscle. Science. 2014. V344(6184).
  14. Sykiotis GP, Bohmann D. Keap1/Nrf2 signaling regulates oxidative stress tolerance and lifespan in Drosophila. Dev Cell. 2008. v14(1). P76–85.
  15. Wang J, Geesman GJ, Hostikka SL, Atallah M, Blackwell B, Lee E, Cook PJ, Pasaniuc B, Shariat G, Halperin E, Dobke M, Rosenfeld MG, Jordan IK, Lunyak VV. Inhibition of activated pericentromeric SINE/Alu repeat transcription in senescent human adult stem cells reinstates self-renewal. Cell Cycle. 2011. v10(17). P3016-30.
  16. Wang W, Yang X, Cristofalo VJ, Holbrook NJ, Gorospe M. Loss of HuR is linked to reduced expression of proliferative genes during replicative senescence. Mol Cell Biol. 2001. v21(17). P5889-98.
  17. Wood J, Nan Jiang, Brian C Jones, Stephen L Helfand Maintaining repressive heterochromatin extends lifespan in drosophila. Abstracts of papers presented at the 2014 meeting on Molecular genetics of aging, September 29 – October 3, 2014, Cold Spring Harbor, USA.
  18. Wu JJ., Liu J, Chen EB, Wang JJ, Cao L, Narayan N, Fergusson MM, Rovira II, Allen M, Springer DA, Lago CU, Zhang S, DuBois W, Ward T, deCabo R, Gavrilova O, Mock B, Finkel T. Increased mammalian lifespan and a segmental and tissue-specific slowing of aging after genetic reduction of mTOR expression. Cell Rep. 2013. v(5). P913-20.
  19. Yan L, Vatner DE, O’Connor JP, Ivessa A, Ge H, Chen W, Hirotani S, Ishikawa Y, Sadoshima J, Vatner SF. Type 5 adenylyl cyclase disruption increases longevity and protects against stress. Cell. 2007.v130(2). P247-58.
  20. Zhang G, Li J, Purkayastha S, Tang Y, Zhang H, Yin Y, Li B, Liu G, Cai D. Hypothalamic programming of systemic ageing involving IKK-β, NF-κB and GnRH. Nature. 2013. v.497(7448). P.211-6.

Longevity Gene Therapy – Shorter Version

Project description

Gene engineering is the most powerful existing tool for life extension. Mutations in certain genes result in up to 10-fold increase in nematode lifespan and in up to 2-fold increase in a mouse life expectancy. Gene therapy represents a unique tool to transfer achievements of gene engineering into medicine. This approach has already been proven successful for treatment of numerous diseases, in particular those of genetic and multigenic nature. More than 2000 clinical trials have been launched to date.

We propose developing a gene therapy that will radically extend lifespan. Genes that promote longevity of model animals will be used as therapeutic agents. We will manipulate not a single gene, but several aging mechanisms simultaneously. A combination of different approaches may lead to an additive or even a synergistic effect, resulting in a very long life expectancy. For this purpose, an animal will be affected by a set of genes that contribute to longevity and prevent age-related pathologies. We are going to find an optimal combination of longevity promoting genes, which results in the most significant increase in lifespan and is not associated with severe side effects.

As a result, we will develop a comprehensive treatment that will dramatically extend lifespan and prevent the decrepitude of the body. Experiments will be conducted in old mice. Thus, in case of success, the developed method of aging treatment can be quickly moved to clinical trials.

The goal of the project is to develop a complex gene therapy that will drastically increase mouse lifespan and prevent tissue pathology in old age, coupled with the safety assessment of the treatment.

Research plan

5 genes that are most promising in terms of life extension (table 1) will be used as targets for gene therapy. We will affect both the biological aging mechanisms, common to the majority of the cells, as well as the primary neuroendocrine center, that regulates the whole organism’s longevity – the hypothalamus. The expression increase or decrease of these genes in animal models was shown to result in boosted longevity. If the increase in expression of a particular gene is necessary for longevity, we will deliver this gene into the body. If, on the other hand, longevity depends on the inhibition of a certain gene’s expression, we will introduce a genetic construct that encodes small RNAs that inhibit the expression of the target gene. Two out of 5 genes have previously been used for gene therapy of aging: the lifespan of mice was increased by 20% (Zhang et al., 2013, Bernardes de Jesus et al., 2012). Moreover, delivery of these genes resulted in prevention of numerous age-related pathologies.

Therapeutic genes will be introduced into the body using viral vectors – the most powerful method of delivering genetic constructs. To avoid side effects caused by random integration of genetic material into genome we will use adeno-associated viral vectors. To improve safety we will also restrict the duration of treatment. Therapeutic genes will be placed under the promoters, which are activated by specific drugs. This means that therapeutic gene is not expressed until the drug is applied. Thus, the duration of treatment can be regulated.

In case of gene delivery to hypothalamus therapeutic construct will be injected directly into this part of the brain. For other genes we will perform systemic gene delivery into the bloodstream.

We will reveal the most effective combinations of the longevity promoting genes (table 1). All the experiments will be conducted in 2-year old mice. We will study longevity promoting genes in combinations of 2, 3 and 4. We will also utilize all 5 genes simultaneously. In addition, the impact of gene combinations on mouse lifespan will be compared to the impact of these genes applied individually. Old and young untreated mice will be used as controls.

  • The efficiency of therapeutic gene delivery will be tested. We will also study whether gene delivery by the viral vectors does not result in integration of genetic constructs into cell genome.
  • All groups of mice will be regularly tested for aging markers. The blood and adipose tissue transcriptome, proteome and metabolome will be analyzed. All age-related histological and physiological changes will be studied. Behavioral test will be performed to analyze cognitive ability and locomotor activity in mice. The average and maximum lifespan of mice will be determined. In addition, a detailed study of side effects will be performed. Mice will be compared with old mice of the control group as well as with young mice. We will also optimize the duration of therapeutic gene expression.
  • Thus, we will reveal the most efficient and safe combination of genes and optimal dynamics of treatment, which promote longevity and prevent age-related pathologies. The developed method of aging treatment can be quickly moved to clinical trials.

Table 1. Target genes for longevity gene therapy.

Target gene The impact on gene expression Therapeutic effect Research links
Gene delivery to hypothalamus
NF-кВ Expression inhibition The inhibition of NF-кВ transcription factor reduces inflammation in hypothalamus during aging, causes an increase in hypothalamus hormone production and increases mouse lifespan by 23%. Zhang et al., 2013
Systemic gene delivery
TERT Overexpression The catalytic subunit of the telomerase extends the telomeres, which increase the replicative potential of cells and promotes longevity of the body (13-20% in mice). To avoid risk of cancer associated with TERT overexpression we will restrict duration of gene expression and find an optimal period which promotes longevity but does not cause cancer. We will also study the effects of simultaneous delivery of TERT gene and proapoptotic and anti-cancer p53 gene (TP53). Bernardes de Jesus et al., 2012
CRTC1 Expression inhibition Inhibiting TOR-kinase, which promotes cell growth and proliferation, activates autophagy and leads to increased lifespan (20% in mice). Wu et al., 2013
FOXO3 Overexpression A transcription factor that triggers stress response and promotes longevity. Brunet et al., 2004
Gene delivery to senescent cells
HSV-TK Overexpression HSV-TK promotes the transformation of a non-toxic prodrug into a toxic product. HSV-TK gene is under p16 promoter, which is activated in senescent cells. Thus, exposure to the prodrug induces death of senescent cells. Selective senescent cell elimination delays aging in mice. Campisi, 2014

Literature

  1. Bernardes de Jesus B, Vera E, Schneeberger K, Tejera AM, Ayuso E, Bosch F, Blasco MA. Telomerase gene therapy in adult and old mice delays aging and increases longevity without increasing cancer. EMBO Mol Med. 2012. v4(8). P691-704.
  2. Brunet A, Sweeney LB, Sturgill JF, Chua KF, Greer PL, Lin Y, Tran H, Ross SE, Mostoslavsky R, Cohen HY, Hu LS, Cheng HL, Jedrychowski MP, Gygi SP, Sinclair DA, Alt FW, Greenberg ME. Stress-dependent regulation of FOXO transcription factors by the SIRT1 deacetylase. Science. 2004. v303(5666). P2011-5.
  3. Campisi J, Report on Rejuvenation biotechnology conference, August 21-23, 2014, Santa Clara, USA
  4. Wu JJ., Liu J, Chen EB, Wang JJ, Cao L, Narayan N, Fergusson MM, Rovira II, Allen M, Springer DA, Lago CU, Zhang S, DuBois W, Ward T, deCabo R, Gavrilova O, Mock B, Finkel T. Increased mammalian lifespan and a segmental and tissue-specific slowing of aging after genetic reduction of mTOR expression. Cell Rep. 2013. v(5). P913-20.
  5. Zhang G, Li J, Purkayastha S, Tang Y, Zhang H, Yin Y, Li B, Liu G, Cai D. Hypothalamic programming of systemic ageing involving IKK-β, NF-κB and GnRH. Nature. 2013. v.497(7448). P.211-6.

1 Comment

Filed under Life Extension

One response to “Longevity Gene Therapy – Updated Projects

  1. George

    Based on the funding required for the Major Mouse Testing Project, I estimate your project will cost around a million dollars. That is perhaps too much for crowdfunding. Have you thought about contacting the Institute for Longevity Studies in Cuba? http://instituciones.sld.cu/cited/

Leave a Reply

Fill in your details below or click an icon to log in:

WordPress.com Logo

You are commenting using your WordPress.com account. Log Out / Change )

Twitter picture

You are commenting using your Twitter account. Log Out / Change )

Facebook photo

You are commenting using your Facebook account. Log Out / Change )

Google+ photo

You are commenting using your Google+ account. Log Out / Change )

Connecting to %s