Friday, September 7, 2012

Parathyroid hormone to treat achondroplasia? Part 3: exploring the boundaries

What we have learned so far

In this series of articles we have been reviewing the available information regarding the parathyroid hormone (PTH) actions in the bone and cartilage to understand if PTH could turn to be an appropriate therapeutic option for achondroplasia. We have briefly visited this hormone’s metabolic properties and learned that it is part of a small family, as there is a very close correlate protein produced by the body, called PTH-related protein (PTHrP). Both molecules produce quite similar effects acting through the same cell membrane receptor, the PTHR.
We also learned that while PTH circulates in the blood, released by the parathyroid glands, PTHrP is found within the tissues which produce it; in other words, in normal situations, PTHrP is a local agent, it doesn’t circulate. PTHrP is important for us (and by extension PTH) in the context of achondroplasia, because this protein exerts a crucial role in bone growth.
If you have been following this blog, you might have already read about the complexities of the cartilage growth plate and know that PTHrP is part of a very intricate symphony where many different agents control bone growth in children. While PTHrP is a growth promoter, fibroblast growth factor receptor type 3 (FGFR3), the protein which is mutated (altered) in achondroplasia, works like a growth brake. As in achondroplasia FGFR3 is working too much, bone growth is severely impaired. We saw that there is evidence that the growth arrest caused by the mutated FGFR3 may cause reduction of the availability of PTHrP within the growth plate, and this in turn may account for part of the FGFR3 effects in the cartilage (a vicious cycle).
We have also seen that PTH is not like a simple on/off button, with just one single activity. Its molecule contains segments that when cleaved (sliced) may have distinct metabolic functions. The two examples given in the last article are worth to repeat here because they may help us to choose the right PTH form for potential tests in achondroplasia:
  • The N-terminal part of PTH, constituted by the last 34 amino acids of the hormone chain seems to be responsible for the strong anabolic action of this hormone in bone (and is called PTH 1-34);
  • The C-terminal part of PTH seems to be responsible for a modulation of this anabolic activity, and probably is responsible for the differences of the anabolic effects of PTH 1-34 compared to the integral hormone, which is also called PTH 1-84 (1).
Furthermore, we saw that PTH as a medicine has been used for the treatment of osteoporosis in two different forms, PTH 1-84 and as a PTH 1-34 analogue called teriparatide. Meanwhile, it is also being tested in other conditions, with clinical studies already carried on in adults and children with hypoparathyroidism. The evidence obtained in those studies about using PTH in a hormone replacement therapy for hypoparathyroidism is compelling. Then, why PTH is still not part of the treatment of hypoparathyroidism? And why this topic may affect the decision to use PTH in achondroplasia?
To answer these questions we will need to examine the results of the toxicity studies performed with PTH when it was being evaluated as an experimental drug for osteoporosis.
 Is PTH risky?
For any experimental drug to become a medicine it not only need to prove it works (efficacy) but also must pass a series of exhaustive studies both in vitro (in the lab) and in vivo (in animal and human) to test if it is safe. So, let’s check what happened with one of the PTH analogues, teriparatide, during the obligatory tests made to assess its safety. The findings of those tests, which we will be reviewing below brought concern about its long term use in osteoporosis. You see, osteoporosis is a chronic, slow evolving bone disorder and therapies for this condition must be also long term. In summary, tests made in animals, looking for a carcinogen effect (capability of causing cancer), showed that mice submitted to lifelong high dose exposure to teriparatide developed osteosarcomas and other kinds of bone cancer (2).
What does explain those findings? PTH has a physiological pro-proliferative role (it stimulates the multiplication of cells) so the results of those tests were interpreted as the cancer observed in the animals being a consequence of the continuous pro-proliferative stimulation of PTH (2).
These findings caused the Food and Drug Administration (FDA) to impose a black box warning in the teriparatide label with two main restrictions to its use in the clinic (3):
1. Prohibition of its administration in patients with open epiphyses (a synonym of growth plates; growing children and adolescents);
2. Continuous use up to 2 years only.
FDA determined that children and adolescents could not use teriparatide because they are under a natural growing pace and cells are multiplying in great speed, so there is a fear about the risk of having a booster (PTH) working in these fast developing organisms that could cause a cancer transformation in the exposed cells and tissues.
These restrictions reflect concerns over risks not certainties and currently there is no clear evidence that long term use of PTH analogues in fact induce osteosarcoma or other kinds of cancer in man. On the contrary, the subsequent studies made in animals (mice and monkeys) to confirm the cancer increase failed to demonstrate so. These subsequent studies concluded that the oncogenic potential (property of causing cancer) of PTH is linked to the dose used, several times higher than the approved dose in humans (4-6).
Furthermore, as part of an agreement with the manufacturer, FDA requested also a long term follow up of teriparatide use after is approval. The epidemiological surveillance about the risk of osteosarcoma in patients treated with teriparatide showed that it is similar to what is found in the general population. For instance, in 2007 there were already about 300,000 patients treated with teriparatide and there was no clear evidence that in the recommended doses it increased the rate of bone cancer compared to the unexposed population (7). In summary, pre-clinical and surveillance studies with PTH (the full form) and teriparatide have not been showing any tendency of increased risk of cancer in treated patients in defined therapeutic doses. Furthermore, as we have already reviewed, PTH analogues have been tested in adults and children with hypoparathyroidism and have been showing to be safe.
Is there a place for PTH therapy in achondroplasia?
This is the key question. Now it’s time to address it.
First thing we need to know is that PTH and/or PTHrP are not FGFR3 antagonists. They work quite independently one from each other, although we already know that FGFR3 could reduce PTHrP availability in the growth plate (8).
And then, could these two proteins be used to treat achondroplasia? You see, as PTH or PTHrP could not block FGFR3, they would theoretically only overcome one of the main consequences of the FGFR3 actions in the growth plate, which is the chondrocyte proliferation arrest.
This is an interesting concept, treating a disorder without working directly in the causative agent. However, this is exactly the idea being applied in the CNP (BMN-111) development. The goal here would be to rescue the proliferation of chondrocytes, which is impaired because of the FGFR3 excessive activity.
The idea of using PTH in achondroplasia is not new. Back on 2004, the group of Amizuka published an interesting work, where they tested mouse models to study the consequences of switching off FGFR3 or PTHrP or both at the same time. You should look at the published Figures 1 and 2 in their paper, which I don’t reproduce here to respect the copyright (access should be free, after registering at the editor’s website).
Figure 1 shows the differences among the animal models they created. Pay attention to the size of the long bones in the figures 1A (normal mouse, called wild type), 1B (no FGFR3, PTHrP positive) and 1C (FGFR3 positive, no PTHrP). Figure 1C resembles figures published in other studies showing mice models of achondroplasia. Figure 1B highlights the proliferative effect of PTHrP: the long bones are longer than those of the wild type mouse.
Figure 2 shows microscopic cuts of the growth plates corresponding to those of the Figure 1. Pay attention to the same sequence of figures 1A, 1B and 1C. Figure 2B shows a larger proliferative chondrocyte layer compared to the wild type in 2A, while Figure 2C highlights the reduction of the thickness of this layer.
The study by Amizuka et al. is important because it shows the natural effects of both FGFR3 and PTHrP. And then, is there anything done with PTH? The answer is yes.
In 2007, Koso Ueda and cols. (10) published a study where they tested the use of a recombinant (synthetic) form of PTH in bone explants of an achondroplasia mouse model (these were mouse long bones excised from the animal and kept in a culture medium). The administration of PTH rescued the proliferative layer of chondrocytes despite the presence of the mutant, overactive FGFR3. PTH works. Look at the figures showing the differences perceived among the different chondrocyte layers. Ueda’s group also published a small work in a medical meeting in 2009, showing further experiments with the use of PTH in a mouse model of achondroplasia, again with positive results (Ueda K et al. 2009).
Recently, the group of Chen, who has also been very interested in the growth plate cartilage and achondroplasia, published a large study where they tested long term therapy of achondroplastic mice with an intermittent injection of a PTH analogue (12). Results are striking in terms of chondrocyte proliferation and in the growth plate proliferative layer, although authors pointed out that the rescue was not complete. Importantly, they also found that FGFR3 was downregulated (had its production inhibited or reduced) in PTH-treated animals. You see, the cycle is closing, isn’t it? Excessive FGFR3 inhibits PTHrP production; introduction of PTH in achondroplasia reduces FGFR3 production.
Let’s see a bit more about the study by the Chen group (12), which brings several very interesting insights in terms of design, findings and conclusions. The authors used an intermittent scheme of PTH administration, resembling the strategy used for the treatment of osteoporosis in humans with the commercial available PTH analogues. Intermittent use of PTH has been showed to build bone in osteoporosis. When used continuously, PTH causes osteoporosis among other metabolic disturbances and we surely wouldn´t want it to happen if we were treating a child with achondroplasia. So, by using intermittent administration the authors very likely simulated the way PTH could be used in humans to treat the bone growth arrest in achondroplasia.
In summary
In summary, PTH and its analogues represent a true potential therapy for achondroplasia. There are several steps to be performed in terms of understanding its mechanism of action and the effects in the animal model and also to decide which analogue should be safer and to explore the potential undesired effects before it could be used in clinic. For instance, in children with hypoparathyroidism, PTH has been showed to be safe in long term, with few and manageable side effects. However, they have low PTH from the beginning, which would not be true about a child with achondroplasia. PTH use could cause renal stones, an issue we would not like to cause in the patient. How can we manage these effects? What we need are diligent tests, focused researchers and resources.
A final note
In this series of articles about PTH we approached a true boundary. To decide to work on a known molecule in a new indication, especially if it is a rare condition affecting children is not an easy step to do. There is a lot of risks involved, from the affected child’s health to the money to be invested. However, if we just sit down and wait for the lower hanging fruit – as this is the common behavior within the industry – little will be achieved. In the last two years I have made contact with four different pharma industries or biotechs which are developing PTH or PTHrP analogues. I received polite answers explaining that it would be difficult to address the indication at that time or simply received no answer at all. Well, I don’t think achondroplasia is untreatable, but it is the kind of condition that won’t have true attention from major health sponsors easily. It is up to those directly or indirectly affected to change the future.
I realize I have extended a lot this article series. The goal is always to inform, to share knowledge while trying not to be excessively technical. It is the scientific language which prevents many interested people to understand what is really going on with achondroplasia. The lack of scientific background could be a kind of barrier which makes people in front of the inevitable to just sit and wait. I wish these articles I am publishing help the achondroplasia community to get more conscience about the science ongoing and become stronger. It will be this strength that can make the difference for our children. We can do more.
Well, this is far from the end. There are other potential therapeutic solutions waiting for review. I will be bringing a new one in the next article. Not really new since I have already addressed it in one of the first reviews in this blog.
1. Divieti P et al. Receptors specific for the carboxyl-terminal region of parathyroid hormone on bone-derived cells: determinants of ligand binding and bioactivity. Endocrinology 2005;146(4): 1863–70.
2. Vahle JL et al. Skeletal Changes in Rats Given Daily Subcutaneous Injections of Recombinant Human Parathyroid Hormone (1-34) for 2 Years and Relevance to Human Safety. Toxicol Pathol 2002; 30: 3312-21.
4. Vahle JL et al. Bone neoplasms in F344 rats given teriparatide [rhPTH(1-34)] are dependent on duration of treatment and dose. Toxicol Pathol 2004;32(4):426-38.
5. Vahle JL et al. Lack of bone neoplasms and persistence of bone efficacy in cynomolgus macaques after long-term treatment with teriparatide [rhPTH(1-34)]. J Bone Miner Res 2008; 23:2033–9.
6. Jolette J. et al. Defining a noncarcinogenic dose of recombinant human parathyroid hormone 1–84 in a 2-year study in Fischer 344 rats.Toxicol Pathol 2006; 34: 929-40.
7. Harper KD et al. Osteosarcoma and teriparatide? (letter) J Bone Miner Res 2007; 22:334.
8. Chen L et al.  A Ser(365)-->Cys mutation of fibroblast growth factor receptor 3 in mouse downregulates Ihh/PTHrP signals and causes severe achondroplasia. Hum Mol Genet 2001; 10(5): 457-65. 
9. Amizuka et al. Signalling by fibroblast growth factor receptor 3 and parathyroid hormone-related peptide coordinate cartilage and bone development. Bone 2004; 34(1): 13-25.
10. Ueda et al. PTH has the potential to rescue disturbed bone growth in achondroplasia. Bone 2007; 41: 13–18.  
11. Ueda K et al. Effect of rhPTH on bone growth disturbance in achondroplastic mouse. Bone 2009; 45 (Suppl 2): S57-8. doi:10.1016/j.bone.2009.04.041. 
12. Xie et al. Intermittent PTH (1-34) injection rescues the retarded skeletal development and postnatal lethality of mice mimicking human achondroplasia and thanatophoric dysplasia. Hum Mol Genet 2012; 21(18): 3941-55.

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