# Presentation: 205907

## rhBMP-2:origin, biology and preclinical safety

- Gerard E. Riedel, Ph.D.
- Wyeth-Genetics Institute, Inc.
- Cambridge, Massachusetts  USA

## 	
	rhBMP-2 =
	recombinant human bone morphogenetic protein-2 

**	****rhBMP-2 =**
**	**_**r**_**ecombinant **_**h**_**uman **_**b**_**one **_**m**_**orphogenetic **_**p**_**rotein-2**** **

## Origin of rhBMP-2 

- The gene for human BMP-2 was isolated
- This gene was inserted into a chromosome of a mammalian cell
- This cell line grows in large culture vessels and secretes rhBMP-2 into the media
- rhBMP-2 is purified, vialed and freeze-dried

## Properties of rhBMP-2

- Member of “growth and differentiation” protein family
- Endogenous BMP-2 is active in bone repair and in embryonic development
- Homodimeric, glycosylated protein that is highly conserved across species
- Osteoinductive in animals and humans

## rhBMP-2 induces bonerat ectopic implant assay 

**14 days **
**post-implantation**

## Mechanism of bone induction

**Mechanism of bone induction**

- Chemotaxis/ proliferation
- Differentiation of mesenchymal cells
- Bone induction
- trabecular, woven bone
- remodels appropriately
- Increased vascularity

## Mechanism of rhBMP-2 action

- Cell biology
- Responsive cell types, receptors and signal transduction pathways identified
- Bone induction biology
- Local administration is required

## Implantation of rhBMP-2 requires a matrix

- Delivers rhBMP-2 to surgical site
- Augments rhBMP-2 retention at site
- Provides a compatible environment for bone induction

## Matrix = ACS(absorbable collagen sponge)

- Commercially available in US since 1981
- Approved PMA (Integra LifeSciences)
- Implantable hemostatic agent
- Extensive experience of safe use
- Bovine tendon Type I collagen
- Manufacturing process meets/exceeds all US guidance

## ACS = Helistat®

## rhBMP-2/ACS preparation

**ACS**

**rhBMP-2**

**WFI**

**rhBMP-2 solution**

## rhBMP-2/ACS & LT- CAGETM device 

## ACS augments rhBMP-2 retention at implantation site

- Orthotopic model (rabbit ulna)
- Radiolabelled rhBMP-2
- Gamma camera scintigraphy
- Validated method

**	****  ****1         4 ****	****        7                         14**
**                                 ****days**
** **

**% initial dose**

** **

## Safety information 

- rhBMP-2/ACS studies
- implant safety studies
- ADME studies
- rhBMP-2 studies
- Tumor biology studies
- Systemic toxicity studies
- Reproductive toxicity studies

## rhBMP-2/ACS safety studies

- Three anatomic sites
- craniofacial, long bone, spine
- Meyer et al. 1999. Spine 24: 747-754.
- Dosing >>> therapeutic range(species-adjusted)
- Results: no systemic effect
- local effects = rhBMP-2 bioactivity

## Biodistribution studies of rhBMP-2/ACS 

- Two species (rats, rabbits)
- Two implant sites
- ectopic or long-bone  (+/- osteotomy)
- Results:
- rhBMP-2 is released slowly from implant site
- low systemic availability (rat long-bone model)
- maximum rhBMP-2 detected = 0.1% of dose

## rhBMP-2 is slowly released from implantation site

## ADME studies of rhBMP-2

- Pharmacokinetics/biodistribution
- rats (adults and juveniles) and NHP
- IV administration
- Results: rhBMP-2 is cleared rapidly from the circulation, primarily through the liver, and rapidly degraded/excreted into urine

**rhBMP-2 is rapidly cleared ****from the systemic circulation**

## slow release from implant site+ rapid clearance from circulation

**low rhBMP-2 systemic exposure**

## Tumor biology studies 

- Screening studies
- Some tumors express BMP-2, BMP receptors
- No evidence that BMP-2 initiates tumors
- No cytotoxic/mutagenic activity in vitro
- No evidence of abnormal cell biology in implant toxicity studies
- Pharmacology studies (in vitro)

## Effect on tumor growth (in vitro) 

- 51 tumor cell lines tested
-   3: growth promotion (in serum-free media)       [pancreas (2), prostate]
- 48: no effect or inhibition
- 71 primary tumor isolates tested
- 71: no effect or inhibition
- Inhibition is not sufficient for therapy

## Additional tumor studies (FDA consultation)

- 1. Screen tumor cells
-     for BMP receptor
-     messenger RNA

- 2. Assess effect on
-     tumor cell growth
-     in vitro (cell culture)

- 3. Assess effect on
-     tumor cell growth
-     in vivo (xenograft)

## rhBMP-2 toxicity studies 

- Systemic toxicity of rhBMP-2
- single or repeat IV dosing, two species
- exposure > 1000x expected in patients
- Reproductive toxicity of rhBMP-2
- repeat IV dosing, two species
- fertility and teratology
- exposure > 1000x expected in patients
- Results: no systemic effect

## Additional safety studies 

- Tripartite guideline safety studies
- in vivo, in vitro
- cytotoxicity, mutagenicity, others
- General safety studies
- in vivo, IV administration
- behavior, locomotion, other body functions
- Results: no effect

## New studies (FDA consultation) 

- Focus: better understanding of an immune response to rhBMP-2
- New clinical assay
- measure all human antibody isotypes(current assay measures human IgG)
- Neutralization assay
- ability of antisera to block rhBMP-2 activity

## Preclinical safety findings

- No systemic effects
- attributed to low systemic exposure
- local effects = osteoinduction by rhBMP-2
- No toxicity detected
- Safety assessment supports use of InFUSETM Bone Graft