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DIFFERENTIATION AGENTS IN NEUROFIBROMATOSIS
Peter Adamson, M.D.
National Cancer Institute
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I will certainly do my best to try to explain what differentiation agents, and
differtiation therapy, are and to talk briefly about how these agents may find a role
for the treatment of neurofibromatosis. Before I go into some of these details, I would
like to tell a little bit about who I am and who we are, right across the street in
building 10 at the national cancer Institute (NCI), how we ended up beginning to look
at some of these drugs, and how they may apply particularly for children with
neurofibromatosis.
I am in the pharmacology and experimental therapeutic section of the pediatric
oncology branch of NCI. Our mission is to develop novel anti-cancer drugs for children
with refractory cancer, and to study the clinical pharmacology of anti-cancer drugs
in children. In other words, once we give a drug, new or old, to a child, we do studies
to determine what happens to that drug when in the body. Children handle drugs
differently than adults, they are not just small adults. We perform studies after we
administer new drugs to determine what is happening to the drug once it is in the child.
In the past three to four years, we have focused our efforts into developing new
treatments for children with tumors of the nervous system.
Now, what is differentiation therapy? Differentiation therapy says that the
genes required for normal growth and differentiation are not necessarily lost during
malignant transformation, but the regulation ofthese genes is often lost. What does
all ofthat mean? Cells have "machinery" that allows them to grow and divide, and they
also have machinery that allow them to stop growing and become mature cells. Mature
cells usually neither grow nor divide. Certain mature cells, in fact, if they stop
growing and dividing, go on to die spontaneously. Cancer, or with some
similarities benign tumors, have genes that have been altered that tell the cell to
keep growing, The "machinery" to stop growing is still working very well, but what is
broken is the on/off switch. The cells are growing, but the off switch for cell growth,
or the on switch to become mature cells has broken. What differentiation therapy says
is even though there is a genetic alteration that is pushing the machinery in one
direction, perhaps there are other switches that we can use to nun it off. We will
not fix the genetic defects, but what we will do is try to find a different on/off
switch. That sounds very interesting. The question is, does that happen in human tumors,
malignant or benign, and if so, are there drugs that we can use that would work in
patients.
Perhaps the best study of this type ofapproach comes from a tumor of childhood
called neuroblatstoma. Neuroblastoma is a tumor of the nervous system. It typically
occurs in young children and, unfortunately, our current treatments for it are not
very good for the majority of patients. However, we know that in infants when this
tumor occurs, it can spontaneously differentiate into a mature tissue and go on to
die without any therapy. So, what we and many other investigators have been looking
at are ways to take the rumors that do not do well and see if we can turn off the
growth switch. One drug that does this in the laboratory is called a retinoic acid,
which you may have heard of. It is actually a derivative of vitamin A. In this slide
the neuroblasts that have been growing without any retinoic acid tend to be round,
grow rather quickly, and tend to clump up. In the laboratory when we add retinoic
acid, the cell number is greatly decreased, and the cells themselves begin to take
on an appearance of mature neurons by extending these thin lines called axons. We are
currently studying this approach in patients with neuroblastoma. We do not yet know
if retinoic acid is going to be effective in patients, but the idea here is that we
are not correcting what is the known genetic defect, but rather we are applying something
else that turns the growth switch offand the maturation switch on.
I am going to talk about a clinical trial that we have done, a phase-1 trial of
a drug called phenylacetate. Phenylacetate is another differentiation agent. Phase 1
trials, historically, have only been done in children with refractory cancer. In good
part the reason for this is that the risks one is taking with a new drug must outweigh
the potential benefits, and for children with cancer whose disease is refractory or
will not respond to any kmown effective treatment, often times the potential benefits
of experimental treatment outweigh the risks. Phenylacetate is not a typical anti-cancer
agent. It is a deaminated metabolite of Phenylanine. What that means is Phenylanine
is a naturally occurring amino acid within the body, and its breakdown product is
phenylacetate. However, phenyiacetate only exists in the body at very, very low
concentrations. ln the laboratory, phenylacetate effects a range of neural tumor cell
lines, including brain tumor cell lines, and neuroblastoma cell lines. One finds that
in high concentration phenylacetate can induce tumor cytostasis, which means it stops
the tumors from growing. It also induces differentiation which means it takes an
immature cell and transforms it into a mature appearing cell, which stops growing and
goes on to die.
We began a phase- 1 trial of phenylacatate in children that I would like to talk
to you about today, but before doing that, I think it is important to put it in
perspective what a phase-1 trial is how it may or may not apply for patients with
neurofibromatosis. There are four primary goals of it phase- 1 trial. The first is, we
wanted to determine what the maximum tolerated dose of a new drug is. In other words,
for most drugs, especially, anti-cancer drugs, the more one gives, the more effective
it seems to be. That is not true for all drugs, but many drugs. In a phase-i trial, we
want to determine in humans, in this case the children, what is the highest dose that we can
safely give where side effects are acceptable and the likelihood for benefit is
maximum. As part of this, we need to learn what the side effects are. There are many,
many models that we can use in the laboratory that help us predict this. But until we
test it in patients we are not certain what all of the potential side effects are. We
also want to see what is happening to this drug in the body. We do what are called
pharmacokinetic studies. After giving the drug, we take numerous small blood samples
to see what is happening to the drug in the body. This helps us to better understand how
to give the drug in future studies. Finally, in the phase- 1 trial we look for signs of
activity in what will turn out to be a wide spectrum of different refractory cancers.
One thing that we do not learn from a phase- 1 trial is, if this drug is truly
effective and if so, how effective is it. We only begin to learn that in a phase-II
trial where we treat children with the same type of malignancy with the maximum
tolerated dose. So phase-1 trials can sometimes give us a clue as to what tumors we
may want to loot in phase II trial, but it does not answer the question, "is it
really an effective drug?" So in general, especially in pediatric trials, who are
the patients who should participate in a phase-1 trial? The patients have to have
progressive cancer, no nmown curative treatment, and for safety reasons, since all
drugs that we give are eliminated from the body by the kidney, liver or combination
they must have to have adequate kidney and liver function. This has been the standard
approach for phase-1 drug trials in pediatric oncology. However, for Phenylacetate,
because of its laboratory activity in a number of different tumors of the nervous
system, and because its side effect profile was expected to be quite a bit different
than the normal side effect profile that we see with anti-cancer drugs, that is
significantly safer than typical anti-cancer drugs, we allowed the patients who
had neurofibromatosis and severe complications from their disease where there were
no other interventions to participate in this trial. On the order of 30 or so patients
with a wide variety of diagnoses entered on to the trial. Because of the pre-clinical
work, most ofthe diagnoses where, in fact, tumors ofthe nervous syseem. As you can see
here we treated three children with neurofibromatosis, two had NF-I and one had NF-2.
There were also other tumors ofthe nervous system in the study.
How do we do a phase-1 trial? In children, we usually rely on the initial
experience in adult phase-1 trials which start at very, very low doses and slowly work
their way up to high doses. There are far fewer children with cancer, fortunately, than
there are adults. So we will usually wait for information from the adult phase-1 trial
and estimate a safe starting dose. We started Phenylacetate in children at a dose of
9 grams per meter square, that is based on their size, given every day and they
received it for 28 consecutive days.
In phase-I trials we treat small groups of children and follow them to see if
they develop signs of side effects or toxicity. At the very first dose level, we
found out that our dose limiting toxicity was somnolence, The children became very,
very sleepy. Fortunately, this side effect rapidly went away when we turned off the
infusion of the drug. To leave anyone on a drug that makes them sleepy through the
day, if you are going to do it every day, is simply not acceptable. So we had to go
down on our dose and try it in another group of patients. In those groups of patients
we had six children and in only one did we have to adjust the dosage.
We were also doing pharmacologic studies that told us that the drug in this
body started at high levels but would rapidly decrease. So we changed our study
design to start children at a lower dose and, after a week, see if tey would tolerate
a higher dose. That tuned out to be a case. By just starting them at a lower dose and
then increasing the dose, we treated a group of seven children from seven to nine
grams/m(2) and then seven more children from nine to twelve g/m(2). When we got up to
a dose twelve to fifteen g/m(2) we started to have other types of toxicity which told
us that we could go no higher. We went back and studied the nine to twelve gram per
metered squared dose and then recommended this as a safe starting dose for larger
trials. Those trials will begin within the next two months.
We measured the drug concentration in the blood in different patients. The
vast majority of patients had no side effects, though a small number of patients did
have side effects. We found that patients who had very high levels of the drug in
their blood were the ones most likely to develop side effects. Blood tests may be a
way for us to monitor patients who are treated with this drug and adjust their dose
to leave them in a range that we believe is effective without running into a risk of
side effects.
As I said, we never answer the question, "is this drug effective" on a phase-1
trial but there are times when we get clues to say that we should look at this in more
detail in a phase-II trial. We had one child who participated on this trial with a
rare type of brain tumor called a germinoma which had grown back after surgery,
radiation, and chemotherapy. An MRI scan of the brain showed that the tumor had
infiltrated a significant portion ofthe brain. After the first month of treatment
there was a clear change on the MRI scan with a marked decrease in the tumor area.
This, to us, was a clue that perhaps Phenylacetate is a drug that should be explored
further in a phase-II trial.
Now how does this apply to neurofibromatosis? The answer is, we do not yet
know, but let me share with you just one of the observations. One of the children with
neurofibromatosis that was entered into the trial was a 15-year old young man with
NF-2 who had severe complications of his disease. When he came to us he had
quadraparasis secondary to cervical cord compression. In other words, his
neurofibromas and meningiomas were choking off the spinal cord high in the neck and
he had lost the use of his arms and legs. He no longer had any type of surgical option
that was going to reverse this process. We felt it would be reasonable for this child
to participate in the phase-1 trial. Over several months time while on Phenylacetate,
he regained strength in his upper extremities, not normal strength, but he clearly
regained strength and movement of his upper arms. However, on an MRI we could not
measure a significant change in his tumors. As you know, these tumors tend to be
quite slow growing and it may take quite some time to see changes, but despite his
clinical improvement, we could not find the evidence on his MRI. He remained on
treatment for one year and then unfortunately began developing a worsening of this
disease and symptoms. Again, this was at least a clue that Phenylacetate deserves
further exploration both in the laboratory as well as in the clinic.
What are our future directions? We need to do additional laboratory evaluation of
Phenylacetate in tumors of the nervous system. I specifically say neuron tumors because
even though I am a pediatric oncologist, it has not taken too many children with
neurofibromatosis to convince me that we need to be doing more for this disease. We
are going back to the laboratory with drugs such as Phenylacetate, to see what may
help in the treatment of tumors of the nervous system.
We are beginning anotber phase-1 trial with a drug that is related to Phenylacetate
called phenylbutyrate. As we did for the Phenylacetate, we will allow children whom
there is no alternative for treatment who are suffering severe or life threatening
consequences of their disease to be enrolled in the study if it is safe to do so.
The last direction is really not related to differentiation therapy, but in some
ways is related to the talk that proceeded mine, and that is a phase- 1 trial of a
drug that is called farnesyl-transferase inhibitor. Now I had better explain why I
think this may be an important type of drug to look at in patients particularly in NF-1.
It is being developed as an anti-cancer agent, but it may have features that make it
applicable to non-cancerous tumors. You may have heard something about this, and I am
sure there are people in the room that know more about this than I do. There is
something call Ras, a critical protein for the growth and control of growth of tumors,
both benign and malignant. Ras can exist in two forms, an inactive form as well as an
active form -- active cells go on to grow and divide. Now, part of the gene product
from NF- 1 seems to be closely related to another group of protein called GAP protein's
that keep Ras in its active form. Whether this is in fact the mechanism that
neurofibromas grow, we do not know, but there is certainly suggestions that the NF- 1
gene product is mimicking the actions of GAP, keeping Ras in an active form and leading
to mitosis. The farnesyl-transferase inhibitors do block the activation of Ras, keeping
it in an inactive form. This may be important in a number of cancers, and may be
important in neurofiromas. This is a brand new class of drugs that are in early stages
of testing for cancer. We think, because of its mechanism of action, if its side
effects profile turns out to be acceptable, it may well be a class of drugs that is
important to study in patients suffering with severe complications of NF-l.
I thank you for your attention and I would welcome any questions that you may have.
Questions:
Q: Is this Ras also in NF-2, what is the difference?
A: No, although it may have a role in the growth of any type of tumor, but as far as
the underlying mechanism for the growth of that tumor, it is more likely to be
related to NF-1 simply because of how Ras is activated.
Q: How did you get turned on to the Phenylafetate for this purpose?
A. We have found focusing our efforts on developing drugs for children with brain
tumors and other neural tumors. We have many drugs affective in many different
types of cancer. The side effects of manmy of those drugs have a lot in common and
in fact, the major side effects are the effects on the bone marrow and the blood
counts. By the time children have tried ah of the standard therapy, they usually
cannot tolerate cytotoxic chemotherapy. So although we develop those type of
agents as well, we are always on the lookout for drugs that do not have that side
effect. Phenylacetate has been looked at in the laboratory, not in my laboratory,
but in other laboratories in the NCI, and showed potential -- initially in prostate
cancer and then in brain tumors. Activities in brain tumors and its side effects
profile prompted us into looking at its effects in children with cancer.
Q: Are you accepting patients for this new study of Phenylbutyrate?
A. Right now the phase-1 trial ofphenylbutyrate is open and yes, we will be taking
children with NF. I think it is only going to apply, I hope, to a very, very small
fraction of children with NF; but we certainly would consider children who have
severe complications oftheir disease where there is really no standard approach to
relief of conmplications. The farnesyl transfease inhibitor is probably six months
away, but we are in discussion with the drug company already to allow us to treat
children with NF on their trial.
Q: When do you consider a drug to be successful - tumor shrinkage, no growth
of the tumor?
A: I think ultimately for a drug to be successfull it has to relieve symptoms and ideally
make the tumors get smaller. Stopping tumors from growing would certainly be a
benefit to many patients. It is harder to convince people because of the nature of
NF and the long time course that it takes for tumors to grow. If the tumor get
smaller, it is a lot more convincing. If the tumor stays the same, it takes a larger
trial and longer trial to show that in fact, it was the drug that make the difference.
Q: Is there age or weight restrictions?
A: I believe that the age cut off for Phenylbutyrate is three years. There is no weight
restriction.
Q: You mentioned retinelic acid. Is there a difference between retinelic acid and
retinoic acid?
A: There are many different types of retinoic acid and the one that was out there, I
think there was in fact a trial of 13-cis-retonoic acid also known as Accutane or
isotretinoin, and is the one used for severe acne. Accutane was the first
commercially available retinoic acid. The more recent retinoic acid to come along
is All-Trans retinoic acid and that is the one that is likely to be more potent. The
retinoids are the class of Vitamin A compounds that include all of the retinoic acid
isomers as well as other Vitamin A derivatives. Right now there are two
commercially available forms of retinoic acid, the 13-cis which is used in acne and
the All-Trans which is used in a disease called an acute promyelocytic leukemia.
Q: There was a report just this week that I saw on television with the use of
retinoic acid which has been effective in shrinking tumors.
A: That would probably be All-Trans retinoic acid, which has been used in a rare form
of leukemia and which we are testing in neuroblastoma, but it has been looked at in
a number of other tumors. 13-cis retinoic acid has also been used in combination
with interferons for other types of cancers.
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