Multiple Pathways for Pain Control Using TRPV1 Ion Channel Agonists and Positive Allosteric Modulators

- TRPV1 agonists
a) Preclinical research b) Clinical trial in cancer pain c) Osteoarthritis

- TRPV1 positive allosteric modulators

Michael Iadarola, Ph.D. Neurobiology and Pain Therapeutics Section (NPTS) Laboratory of Sensory Biology, NIDCR National Institutes of Health, USA

Neuronal Pathways & Analgesic Targets

Central or Peripheral nervous system?
Peripheral
No CNS side effects, Can produce insensitivity to pain

Central
Possible CNS side effects Level of relief from severe pain

Cut here
Neurosurgical Lesions for Pain Relief: Antero-lateral Quadrant Cordotomy (ALQ)

Pain Control with Resiniferatoxin (RTX) a TRPV1 agonist

Central Administration Peripheral Administration
Primary afferent neuron

PERMANENT
cuts the axon or kills the cell
Terminal in spinal cord RTX

TRANSIENT
Nerve terminals die back and re-grow
Terminal In body Capsaicin

Routes:
lumbar puncture - CSF Intraganglionic - dorsal root or trigeminal ganglia

Routes:
Topical (cornea) Intra-joint Perineural Subcutaneous

RTX synthesizing Species of Euphorbia Are found in Africa (orange areas)

TRPV1 Agonists ?
If agonists activate the ion channel, wont that cause pain?

Yes: a) 8%capsaicin patch: pretreat with 4% lidocaine patch and oxycodone after.
Backonja et al., Lancet Neurol 7:110612, 2008. b) Iadarola et al., Neural activation during acute capsaicin-evoked pain and allodynia assessed with positron emission tomography. Brain 121:931-947, 1998.

Can we use this Therapeutically?

Ca++
Yes: agonist activation leads to a calcium overload. This produces a highly selective Lesion of pain fibers or neurons Literally cut the connection between the body and the spinal cord: Permanent intervention

TRPV1 Model constructed by Krisztian Kaszas, NPTS, NIDCR

Molecular Targeting of TPRV1 Using Agonists

Capsaicin Resiniferatoxin (RTX)

Karai L, Russell JT, Iadarola MJ, Olah Z: J Biol Chem. 279:16377-16387, 2004.

RTX-induced Fragmentation of ER and Mitochondria Con 0.5

TRPV1-eGFP fusion

5 eGFP

15

Laser confocal microscopy at 24 hrs after transfection; TRPV1eGFP: Green fusion protein labels ER and plasma membrane; Mitochondria: MitoTracker Red dye.

No bystander effect: cells without the receptor (red only) are unaffected
Olah Z, Karai L and Iadarola MJ: J Biol Chem 276: 31163-31170, 2001.

Agonist Selectivity
TRPV1: ENRICHED EXPRESSION IN SENSORY GANGLIA
RT-PCR (30 cycles) Rat Brain and Sensory Ganglia TRPV1 GAPDH
1 1 2 3 4 5 2

10

Cerebellum Cortex Dorsal Root Ganglion Hippocampus Hypothalamus

6 Substantia Nigra 7 Spinal Cord 8 Striatum 9 Trigeminal Ganglion 10 Blank

Animal studies of RTX

Non-clinical rodent studies: Established specificity, dose range, duration of action and route of administration Canine model that closely resembles human situation. - UPenn School of Veterinary Medicine, Dr. Dorothy Cimino Brown. - Client dogs with naturally occurring osteosarcoma - Candidates for euthanasia. Why? Poor pain control - Owner can assess higher order function

Profound analgesia, durable, resistant to disease progression, no personality change, no impact on bladder or bowel function, no impact on motor function.

Long term pain control in bone cancer

Brown DC, Iadarola MJ, et al., Anesthesiology 103:1052-1059, 2005.

Analgesic & Anti-inflammatory Medication Administered to Dogs with Bone Cancer

Dog Prior to RTX Administration Carprofen 100mg every 12 hours Prednisone 10 mg every 12 hours Piroxicam 10 mg every 24 hours Codeine 90 mg every 8 hours Piroxicam 10 mg every 24 hours Prednisone 20 mg every 12 hours Etodolac 450 mg every 12 hours Carprofen 75 mg every 12 hours Butorphanol 5 mg every 12 hours Piroxicam 20 mg every 24 hours Carprofen 100mg every 12 hours Deracoxib 100 mg every 24 hours Codeine 120 mg every 8 hours Deracoxib 100 mg every 24 hours Deracoxib 100 mg every 24 hours Codeine 30mg every 6 hours 2 Weeks Following RTX Administration Prednisone 2.5 mg every 12 hours

Dose decreased
Piroxicam 10 mg every 48 hours

2
3 4 5 6 7 8 9 10 11 14

None
Prednisone 20 mg every 12 hours Etodolac 450 mg every 12 hours

None None None

Deracoxib 100 mg every 24 hours

67%of the dogs either discontinued or decreased standard analgesic drugs

None None
Deracoxib 50 mg every 24 hours

Dose Decreased

Brown DC, Iadarola MJ, et al., Anesthesiology 103:1052-1059, 2005.

Steps to Human Clinical Trial of RTX for Cancer Pain

1) Raw Material: Latex from Euphorbia resinifera a cactus-like succulent from Morocco 4) Clinical protocol: Single administration of intrathecal resiniferatoxin for treating severe refractory pain associated with advanced disease. 5) IND to FDA Animal and cellular toxicology data, Drug master file, Stability, etc NIDA Collaboration 2a) Large scale cGMP production of RTX NIDA Collaboration 2b) Formulation of RTX for injection NIH Pharmacy 3) RTX Toxicology Two species: Complete NIDA Collaboration

6)

Phase I

Clinical trial at NIH Recruiting NOW

Protocol 09-D-0039: A Phase I Study of the Intrathecal Administration of Resiniferatoxin for Treating Severe Refractory Pain Associated With Advanced Cancer

http://clinicaltrials.gov/ct2/show/NCT00804154
Iadarola MJ and Mannes AJ: Curr Top Med Chem. 11(17):2171-2179, 2011

RTX injection procedure: lumbar puncture

Protocol 09-D-0039: A Phase I Study of the Intrathecal Administration of Resiniferatoxin for Treating Severe Refractory Pain Associated With Advanced Cancer

Eligibility Criteria: Pain in lower half of body

T12 L1 L2
Cauda Equina: site of lumbar puncture and catheter insertion

L3 L4 L5

Affected dermatomes

Dermatome map of body http://clinicaltrials.gov/ct2/show/NCT00804154

Peripheral RTX Administration

Central Administration

PERMANENT
cuts the axon or kills the cell
Terminal in spinal cord

Primary afferent neuron

Peripheral Administration TRANSIENT

Nerve terminals die back and re-grow
Terminal In body

Routes:
Topical (cornea) Intra-joint Perineural Subcutaneous

Indications:
Corneal pain Osteoarthritis Post-injury neuropathic pain with A localized site of origin or trigger zone Certain types of low back pain

TRPV1 Allosteric Modulators

Objective: new approach to TRPV1 based analgesic agents
Roadmap Grant R03 MH089480, HTS for Identifying Allosteric Modulators of the TRPV1 receptor.

Positive allosteric modulators. Concept: impose an activitydependent mode of action to enhance process of nerve terminal inactivation Lead compounds

TRPV1 PAMs should be selective for areas actively experiencing tissue damage or inflammation peripheral generators that would be sites of endovanilloid production and low pH.

MRS-1477: a TRPV1 positive allosteric modulator

Calcium imaging in primary DRG neurons

- capsaicin

+ capsasicin

Roh EJ, Keller JM, Zoltan Olah Z, Iadarola MJ, Jacobson KA: Bioorganic & Medicinal Chemistry 16:93499358, 2008

Positive Allosteric Modulation of TRPV1 with MRS1477

Positive modulator effects on orthosteric agonist and H+ ion

Ca++

Kaszas K, Keller JM, Coddu C, Mishra S, Hoon M, Stojilkovic S,Jacobson KA, Iadarola MJ: J Pharmacol Exp Ther 340: 152-160, 2012

1477 produces a long-term nerve terminal inactivation Thermal testing of the paw for A-delta or C-fiber integrity following combination Injectnion of capsaicin (the TRPV1 agonist) and MRS1477 (the TRPV1 PAM) A-delta stimulation

Lebovitz E, et al., Molecular Pain, in press

Summary
A. TRPV1 agonists can be used in many different ways. Two examples are: - Intrathecal administration of RTX to treat pain from advanced cancer - Peripheral administration into areas of inflammation or nerve damage. B. TRPV1 PAMs may also be versatile analgesic agents. - The duration of action occurs over several days in our experimental model - It may be possible to administer them systemically or by local injection - They have an activity-dependent component to their action - The role of endovanilloids requires further investigation.

Acknowledgements
Many of my colleagues were, and still are, involved in bringing the work with RTX from the bench to the bedside. The development of the PAMs is part of the next phase of TRPV1-based drugs.

RTX
NIDCR: Zoltan Olah, Laszlo Karai, Ofer Wellisch, Andrew Mannes, Jason Keller University of Pennsylvania: Dorothy Cimino Brown NIH Pharmaceutical Development Section: George Grimes NIDA: Nate Appel, Robert Walsh, Nora Chiang, Moo Park, Marta DeSantis, Jim Terrill

PAMs
NIDCR: Krisztian Kaszas, Jason Keller, Evan Lebovitz, Zoltan Olah, Ken Jacobson, NCATS, NCGC: Noel Southall, Erika Englund, Juan Marugen, Steve Titus This work was supported by the Intramural Research Program NIH, NIDCR and Molecular Libraries Screening Grant, 1 R03 MH089480, HTS for Identifying Allosteric Modulators of the TRPV1 receptor.