***Biphosphonates

Question 1

Describe the phases of bone remodelling

Answer 1

1. Activation phase:
   Can be stimulated by:
   · Low levels of calcium stimulates the release of parathyroid hormone (PTH) from the parathyroid gland
   · An alteration of mechanical loading sensed by the osteocytes or factors like tumour necrosis factor-α (TNF-α) and interleukin-6 (IL-6)

· These stimulants activate lining cells which are dormant osteoblasts
· Lining cells increase their own surface expression of RANKL (a ligand), that binds to its receptor RANK present on pre-osteoclasts
· RANKL binding to RANK stimulates osteoclast differentiation

2. Resorption phase:
   · Once differentiated, osteoclasts adhere to the bone surface and begin to dissolve bone
   · They acidify the bone matrix to dissolve the inorganic component
   · They release of lysosomal enzymes, such as cathepsin K, and of MMP9 to degrade the organic component of bon
   · Once accomplished their function, osteoclasts undergo to apoptosis
3. Formation phase:
   · Bone matrix resorption leads to the release of several growth factors like bone morphogenetic proteins (BMPs), fibroblast growth factors (FGFs) and transforming growth factor β (TGF β)
   · They recruit osteoblasts which will produce the new bone matrix, PTH increases the number and activity of osteoclasts

Question 2

List examples of bone remodelling pathologies (4)

Answer 2

• Osteolytic lesions
• Paget’s disease
• Secondary osteoporosis due to glucocorticoids
• Osteoporosis

Question 3

Describe osteolytic lesions

Answer 3

· Is a softened section of a patients bones, with sparse trabeculae
· Is characterized by increased resorption of old bone tissue and a decrease in bone rebuilding
· This imbalance is due tumours releasing regulatory factors like MCP-1 which attracts monocytes (even though the body doesn’t need remodelling)
· Tumour cells will also pump a PTH related protein that will initiate bone resorption EVEN THOUGH IT ISN’T needed. In addition they produce IL 1, IL6 and IL 11 which stimulates osteoclasts even more

Question 4

Describe Paget’s disease

Answer 4

A condition where there is rapid and excessive bone breakdown followed by disorganized new bone formation which has low fracture resistance

Question 5

Describe secondary osteoporosis due to glucocorticoids

Answer 5

· Glucocorticoid go through phospholipid bilayer and into cytoplasm which activates the glucocorticoid receptor
· When two glucocorticoid joins with its receptors and they join together, they form a dimer
· This “dimer” now has the ability to mess with transcription. It’ll attach to DNA and as a result this attracts other proteins to join
· The conjugation of these proteins forms histone acetyl transferase.
· This weakens DNA’s histone rapping ability
· Thus, this allows other things to come in and upregulate (initiate osteoclasts) and downregulate genes (apoptosis of osteocytes/ blasts)

Question 6

Describe osteoporosis

Answer 6

· A hormone called estradiol directly acts on osteoclasts and inactivates them
· Meanwhile, they interfere with an enzyme called caspase -3 that initiates apoptosis of osteoblasts
· This, along with androgens (testosterone or DHT) stimulates osteoblast differentiation

Question 7

Describe two types of biphosphonates

Answer 7

· The difference between the two types of Bisphosphonates is due to residues, residue 1 (R1) and residue 2 (R2)
· If there is no nitrogen present in R1 or R2 = non-nitrogen containing bisphosphonates
· If nitrogen presents in either R1 or R2 = nitrogen-containing bisphosphonates

Question 8

Describe the mode of action of N-BPs

Answer 8

· Bisphosphonates, when attached to bone tissue, are released by osteoclaststhe bone cells that break down bone tissue. Bisphosphonate molecules then attach to and enter osteoclasts where they disrupt intracellular enzymatic functions needed for bone resorption
· Work via interrupting the mevalonate pathway occuring in the osteoclasts
· They inhibit prenyltransferase which is essential for allowing the mevalonate pathway to continue
· Also inhibits prenylate proteins (which causes the formation of a ruffled border and sealant for osteoclasts), which is otherwise actually needed for osteoclast functioning
· N-BPs do not cause apoptosis of osteoclasts directly. N-BPs cause the loss of function of osteoclasts

Question 9

Describe the mode of action of NN-BPs

Answer 9

· Bisphosphonates, when attached to bone tissue, are released by osteoclaststhe bone cells that break down bone tissue. Bisphosphonate molecules then attach to and enter osteoclasts where they disrupt intracellular enzymatic functions needed for bone resorption
· In all cells, there is an ongoing reaction to create ATP
· N-NBPs attach to AMP molecules which creates a more deformed ATP
· This deformed ATP with the bisphosphonate molecule attached to it will block the ADP ion- channel; stopping the formation of ATP
· Since the ion channel is blocked, it messes with the -mv of the mitochondrium membrane. This triggers the release of cytochrome C from the inner membrane of mitochondrium
· Cytochrome C will move into the cytoplasm which activates the lysis of osteoclasts

Question 10

Describe how biphosphonates cause osteonecrosis of the jaw

Answer 10

· Jaw has a higher rate of bone remodelling. As a result, a higher number of BPs accumulate there
· Because of the high rate of remodelling, the biphosphonates is released from the bone rapidly and thus, they go into other cells which is toxic for them and it causes those cells to die
· Holistically, this causes inflammation = low pH and necrosis of the jaw
· BPs also have an immunosuppressive effect, thus exposed bone is susceptible to bacterial colonisation

Question 11

Detail how the mevalonate pathway works.

Answer 11

The mevalonate pathway leads to the formation of prenylated proteins and cholesterol.

The pathway starts when two acetyl CoA molecules join together to form “Acetoacteyl- CoA”.

Then, “Acetoacteyl- CoA” gets into a reaction with acetyl- CoA to form HMG - CoA.

HMG - CoA then gets into a reaction with HMG- reductase to form MEVALONATE.

MEVALONATE goes into a series of many reactions to eventually form two important things:

• Isopentenyl pyrophosphate
• Dimethylallyl pyrophosphate

Isopentenyl pyrophosphate and Dimethylallyl pyrophosphate get into a reaction to form Geranyl transferase. This is catalysed by PRENYLTRANSFERASE.

PRENYLTRANSFERASE transforms geranyl transferase into Farnesyl transferase = prenylated proteins.

NBP’s block the function of prenyltransferase