© 2005 John D. Furber

Glycation Crosslink Breakers:Glycation Crosslink Breakers: Necessity and Prospects Necessity and Prospects

John D. FurberJohn D. Furber

Legendary PharmaceuticalsLegendary Pharmaceuticals

Gainesville, FloridaGainesville, Florida

www.LegendaryPharma.comwww.LegendaryPharma.com

© 2005 John D. Furber

Extracellular AgingExtracellular Agingof Extracellular Matrix (ECM)of Extracellular Matrix (ECM)Glycation, Crosslinking, StiffnessGlycation, Crosslinking, Stiffness

Cardiovascular hemodynamics, capillary circulationCardiovascular hemodynamics, capillary circulation

Lens accommodation, cataracts, glaucomaLens accommodation, cataracts, glaucoma

ArthritisArthritis

Bladder distensibilityBladder distensibility

Erectile dysfunctionErectile dysfunction

Interactions of Cells and ECM: Interactions of Cells and ECM:

Differentiation, ApoptosisDifferentiation, Apoptosis

InflammationInflammation

© 2005 John D. Furber

Alberts, et.al. Molecular Biology of the Cell. (4th Ed. New York: Garland Publishing, Inc. 2002.)

Elastin surrounds the AortaElastin surrounds the Aorta

Alberts, et.al. Molecular Biology of the Cell. (4th Ed. Alberts, et.al. Molecular Biology of the Cell. (4th Ed. New York: Garland Publishing, Inc. 2002.New York: Garland Publishing, Inc. 2002.))

alpha-diketone Crosslinkalpha-diketone Crosslink

•Forms from Glucose

•Perhaps Links Lysine to (Lysine or Cysteine or Histidine)

Ulrich P, Cerami A. Protein glycation, diabetes, and aging. Recent Prog Horm Res. 2001; 56:1-21.

Beneficial Crosslinks in CollagenBeneficial Crosslinks in Collagen

Alberts, et.al. Molecular Biology of the Cell. (4th Ed. Alberts, et.al. Molecular Biology of the Cell. (4th Ed. New York: Garland Publishing, Inc. 2002.New York: Garland Publishing, Inc. 2002.))

•Formed in young collagen

•Give strength to mature fibers

•Formed under enzymatic control

•Lysyl oxidase mediates Lys-Lys bond

•No sugar linker

Fibroblast LineageFibroblast Lineage

Alberts, et.al. Molecular Biology of the Cell. (4th Ed. Alberts, et.al. Molecular Biology of the Cell. (4th Ed. New York: Garland Publishing, Inc. 2002.New York: Garland Publishing, Inc. 2002.))

•Synthesize new Collagen and Elastin

•Arrange fibers properly in place

•Secrete enzymes to digest old ECM

© 2005 John D. Furber

Vicious Cycle Accelerates Vicious Cycle Accelerates ECM Crosslinking ECM Crosslinking

• Crosslinked collagen has very small spaces.Crosslinked collagen has very small spaces.

• Restricts access to Fibroblasts and EnzymesRestricts access to Fibroblasts and Enzymes

• Glycation crosslinking accumulates over timeGlycation crosslinking accumulates over time

• Collagen fibers drawn even more closely togetherCollagen fibers drawn even more closely together

• Further slowing the natural turnover processesFurther slowing the natural turnover processes

Glucosepane CrosslinkGlucosepane Crosslink

•Forms from Glucose

•Links Lysine to Arginine

Sell DR, Biemel KM, Reihl O, Lederer MO, Strauch CM, Monnier VM. Glucosepane is a major protein cross-link of the senescent human extracellular matrix. Relationship with diabetes. J Biol Chem. 2005 Apr 1;280(13):12310-5.

alpha-diketone Crosslinkalpha-diketone Crosslink

•Forms from Glucose

•Perhaps Links Lysine to (Lysine or Cysteine or Histidine)

Ulrich P, Cerami A. Protein glycation, diabetes, and aging. Recent Prog Horm Res. 2001; 56:1-21.

© 2005 John D. Furber

K2P CrosslinkK2P Crosslink

Lysine-

dihydropyridinium-

Lysine

Cheng R, Feng Q, Argirov OK, Ortwerth BJ. Structure elucidation of a novel yellow chromophore from human lens protein. J Biol Chem. 2004 Oct 29;279(44):45441-9.

Two Nucleophilic CentersTwo Nucleophilic CentersAttack two Carbonyl CarbonsAttack two Carbonyl Carbons

© 2005 John D. Furber

Crosslink Breaker Crosslink Breaker Alagebrium Alagebrium

S

N+

CH3

CH3O

Cl

3-(2-phenyl-2-oxoethyl)-4,5-dimethyl thiazolium chloride

ALT-711

Drawing © John D. Furber 2005

© 2005 John D. Furber

Research We NeedResearch We Need

• High-Throughput Screening (HTS) for High-Throughput Screening (HTS) for small-molecule breakers of glucosepane small-molecule breakers of glucosepane and K2P.and K2P.

© 2005 John D. Furber

Research We NeedResearch We Need

• High-Throughput Screening (HTS) for High-Throughput Screening (HTS) for small-molecule breakers of glucosepane small-molecule breakers of glucosepane and K2P.and K2P.

• But... What if new small-molecule breakers But... What if new small-molecule breakers are not quickly found?are not quickly found?

© 2005 John D. Furber

Research We NeedResearch We Need

Plan B: Stimulate Natural TurnoverPlan B: Stimulate Natural Turnover

– Stimulate Fibroblasts or mesenchymal-Stimulate Fibroblasts or mesenchymal-lineage cells to do more of what they lineage cells to do more of what they already do.already do.

– ExtracorporealExtracorporeal

– In vivo injectionsIn vivo injections

© 2005 John D. Furber

Research We NeedResearch We Need

Plan C: Combination Breaker TherapyPlan C: Combination Breaker Therapy

– Develop or discover large enzyme Develop or discover large enzyme breakers for Glucosepane and K2P.breakers for Glucosepane and K2P.

– Use them in combination with AlagebriumUse them in combination with Alagebrium

© 2005 John D. Furber

Research We NeedResearch We Need

Plan D: Nanobots to the Rescue!Plan D: Nanobots to the Rescue!

– Design and build billions of little machines Design and build billions of little machines to do what fibroblast and mesenchymal to do what fibroblast and mesenchymal lineage cells do, only better.lineage cells do, only better.

– Digest old collagen and synthesize new… Digest old collagen and synthesize new… in exactly the right pattern. in exactly the right pattern.

© 2005 John D. Furber

In the Meantime…In the Meantime…

Exercise, fasting, and low blood sugar levels Exercise, fasting, and low blood sugar levels are good for your collagen.are good for your collagen.

[M Kjaer. "Metabolic activity and collagen turnover in [M Kjaer. "Metabolic activity and collagen turnover in human tendon in response to physical activity.” J human tendon in response to physical activity.” J Musculoskelet Neuronal Interact. 2005 Mar;5(1):41-52.]Musculoskelet Neuronal Interact. 2005 Mar;5(1):41-52.]

© 2005 John D. Furber

Glycation Inhibitors in Food, Herbs,Glycation Inhibitors in Food, Herbs, and Supplements and Supplements

• Vitamins B1, C, and EVitamins B1, C, and E

• CurcuminCurcumin• N-acetyl-cysteineN-acetyl-cysteine• EGCG (black tea, green tea, white tea)EGCG (black tea, green tea, white tea)• PABAPABA• QuercetinQuercetin• ResveratrolResveratrol• RutinRutin• thymethyme

© 2005 John D. Furber

Extracellular Rejuvenation Extracellular Rejuvenation is the answer to is the answer to Extracellular Aging Extracellular Aging

•

© 2005 John D. Furber

Parallel Development to Parallel Development to Maximize Early Success Maximize Early Success

1. HTS for small-molecule breakers of glucosepane 1. HTS for small-molecule breakers of glucosepane and K2Pand K2P

2. Stimulate natural ECM turnover processes, such 2. Stimulate natural ECM turnover processes, such as fibroblastsas fibroblasts

3. Rational Design of enzymatic breakers of 3. Rational Design of enzymatic breakers of glucosepane and K2P - Use in combination with glucosepane and K2P - Use in combination with diketone breakers.diketone breakers.

4. Nanobots (to be invented) come to the rescue4. Nanobots (to be invented) come to the rescue

© 2005 John D. Furber

Legendary PharmaceuticalsLegendary Pharmaceuticals Scientific Advisory BoardScientific Advisory Board

• Ulf T. Brunk, MD, Ph.D.

• Judith Campisi, Ph.D.

• Richard M. Cawthon, MD, Ph.D.

• Ana Maria Cuervo, MD, Ph.D.

• Aubrey D.N.J. de Grey, Ph.D.

• Ron Garren, MD

• John C. Guerin

• Kenneth Hensley, Ph.D.

• Alan R. Hipkiss, Ph.D.

• Shalesh Kaushal, Ph.D.

• Jiankang Liu, Ph.D.

• Charles Ostman

• Kamala Rose, MD

• Mark A. Smith, Ph.D.

• Alexei Terman, MD, Ph.D.

• John H. Weiss, MD, Ph.D.

www.LegendaryPharma.com