ANOTHER INTERNET EPP GROUP
EPPREF member Lori Givan has also started an internet group for EPP people only (not other forms of porphyria) through Yahoo. If you want more information and instructions on how to join the group, contact her at lori64064@yahoo.com.
MORE COPING TIPS
Another source of sun-protective clothing
An EPPREF member told us about a store which sells clothes treated with a UV inhibitor, blocking both the UV-B (sunburn) and UV-A (longer) wavelengths of light. We can’t guarantee that it will protect against visible light, because these clothes are made mainly to protect against skin cancer development, but if the substance does make the material impervious to light, it would work for EPP people. You can check out their web-site at www.sunsolutionsclothing.com. Their telephone number is 1-800-895-0010.
A “sun block mask”
A dermatologist invented a visor with a 5 to 6 inch plastic strip attached which shields the face from the rays of the sun - it is rated to absorb both UV-B and UV-A rays. The sun-block mask is obtainable from the George Tieman Surgical Supply Company - their phone number is 1-800-843-6266.
Dietary fish oils and EPP
You may have heard or read on the web about a report that suggested that dietary fish oils (one brand being Maxepa, sold in the US by Twin Labs) seem to lessen the symptoms of EPP. A paper describing a beneficial effect in one patient was reported in a dermatology journal, but unfortunately this patient also developed gall bladder problems. It is not clear if these were caused by the fish oil, but the doctors did not seem to think so. At this point, we cannot recommend this use of fish oils, until careful studies under medical supervision are done, and FDA approval is obtained. This would be a treatment, not a cure - the oils may work as does beta-carotene (Lumitene), by acting as anti-oxidants.
MESSAGE FROM DR. ROTH - GENE THERAPY AND EPP
In issue 26 of EPPREF NEWS I mentioned that I was starting some work on seeing if we could cure EPP in the strain of mice which have it with gene therapy aimed at the bone marrow. As I mentioned at the time, I teamed up with some scientists at the Harvard Medical School - Massachusetts Institute of Technology Health Sciences and Technology division, and to make a long story short, we did manage to cure the mice of their EPP. We published our results in a pretty good scientific journal, Nature Medicine, in July, 1999 (volume 5, page 768, for any scientist among you who wants to look up the article). I should add that the early part of this study was aided by contributions you made to EPPREF. We did get a research grant from the NIH to continue work on gene therapy for EPP using the mice. But, unfortunately, some problems have developed with gene therapy studies in human beings. You may have heard about the study in France involving the children with a disease called “severe combined immuno-deficiency”. This is a disease of the bone marrow, in which its victims are unable to fight off infections. These children received bone marrow-based gene therapy using a viral vector (gene carrier) which contained the normal gene. This was done by removing some of their bone-marrow, isolating the stem cells and mixing the stem cells in culture with the viral vector, and then re-injecting the “cured” stem cells back into the children. In the majority of the children treated, this cured their disease - but, unfortunately two of the cured children also developed a form of leukemia. It was found that the viral vector inserted itself into the middle of a gene in the DNA of the marrow stem cell, which causes leukemia when activated. As a result, human studies on viral-based gene therapy targeting the bone marrow have been put on hold by the FDA until it can be figured out exactly what caused this problem. So, it puts on hold for now any attempt for human EPP gene studies with the viral vector we used successfully in the mice. My plan now is to do studies on ways to do gene therapy to bone marrow stem cells without the use of viral vectors.
MESSAGE FROM DR. ROTH - WHAT YOU NEED TO KNOW ABOUT STEM CELLS
Stem cells are a pretty popular subject of discussion these days. It is important for people to know the basics about them because these cells are potentially important in the cure of EPP and other genetic diseases. Stem cells are unspecialized cells which have the ability to reproduce themselves without limit, and also to give rise to specialized (differentiated) cells, such as heart or blood or muscle or brain cells, which could be used to make replacements for diseased tissues or organs, when the stem cells are grown in the presence of specific chemical nutrients.
Stem cells normally are present in very young embryos, older embryos and fetuses, umbilical cords at birth, and in children and adults.
Embryonic stem cells
It is a fact of embryology that people start their lives as one cell, called the zygote - this is the cell which forms when the oocyte (egg cell) of the mother joins with the sperm cell of the father. This is the first cell of each of us, because it contains our unique set of chromosomes, half of which were inherited from our moms and half from our dads. The zygote starts to divide into two, then four cells. By the third day of life, 12 to 15 cells have formed, making a solid ball of cells: at this stage, the growing young human is called a morula. On the fourth day of life, the morula enters the mother’s uterus. Soon a fluid-filled space forms in the morula - this separates the little ball of cells into two parts: 1) a thin outer layer of cells called the trophoblast, which gives rise to the baby’s part of the placenta and its associated membranes, and 2) a little ball of cells located inside the trophoblast and surrounded by the fluid, called the inner cell mass - these cells are the embryonic stem cells. The embryonic stem cells go on to form the entire embryo - every kind of cell and tissue in the growing little human (and eventually, with further growth, in the fetus, child and adult). At this stage the growing human is called a blastocyst.
This is how embryonic stem cells are formed in the “usual” way. You have heard of “in vitro fertilization” (IVF), a technique which helps couples who are having problems conceiving a baby. In IVF, oocytes and sperm are mixed in a petri dish which contains a special nutrient solution, and they join in the dish to form a zygote, just as they would join in the mother’s uterus. The zygote starts to divide as it would in the uterus: the early development of the embryo (the generic term for a growing human from fertilization until the end of the 8th week of life - from then until birth we are called a fetus) is the same in a petri dish as it is in the mom’s uterus. Usually, the embryos are allowed to grow for about 3 days, checked to make sure they are healthy and then either implanted in the prospective mom’s uterus, or frozen in a special protective solution for future use, in case the first implantation does not result in pregnancy. Occasionally the embryos are cultured to the blastocyst stage, and then are frozen. Implantation of thawed 3 day old embryos and blastocysts have resulted in successful pregnancies.
People who want to do research on embryonic stem cells want to obtain "extra" frozen embryos and blastocysts from IVF clinics and either culture the 3-day old ones to the blastocyst stage, or work directly with the blastocysts which had been frozen. To obtain the inner cell mass cells, i.e. embryonic stem cells, the blastocysts are put in a special solution containing specific antibodies, which cause the trophoblast cells to dissolve, and the inner cell mass cells are released. The stem cells are then washed and cultured in appropriate media for various studies, hopefully to get them to form specialized cells and tissues. It is important for people to realize that the process of opening up the trophoblast and obtaining the embryonic stem cells kills the growing young human being. Unfortunately, in addition to this serious ethical problem, of killing one very young member of our own human species, there are also some medical problems associated with the use of embryonic stem cells.
The same problems of immunological rejection, graft-versus-host disease, etc., that a patient gets from receiving an organ from an organ donor who is not one's identical twin will occur with cells or tissues made from embryonic stem cells, because the parents of the embryos are not related to the patient who will receive the cells and tissues made from those embryonic stem cells. Additionally, embryonic stem cells have the tendency of forming teratomas, which are tumors made up of several kinds of differentiated cells, which often form when the stem cells are implanted into different organs, as has been found in animal studies.
Embryonic stem cells from cloned embryos
Scientists thought that the problems of immunological rejection could be avoided by making cloned embryos and harvesting their stem cells and using these to make the cells and tissues a patient needs to treat hi/her disease - they call this “therapeutic cloning”. Cloning is done by taking an oocyte from a female donor, and removing its nucleus. Then, a somatic cell (a body cell, not an oocyte or sperm cell) is obtained from the patient to be treated (or cloned), and its nucleus is removed and is placed into that oocyte. The oocyte with its new nucleus, which has all 46 chromosomes, is the first cell, the zygote, of the cloned individual. This zygote is then stimulated to start its growth and development. The cloned zygote's development is the same as that of a zygote produced by the union of egg and sperm by either sexual reproduction (the usual way) or by in-vitro fertilization (IVF) However, stem cells from cloned embryos would still lead to some immunological rejection problems, because a crucial part of all cloned cells, the mitochondria, which direct certain aspects of cell metabolism, are all derived from the mitochondria from the oocyte, not from the genes of the donor nucleus, and thus could trigger rejection. Also, teratoma formation could occur. Additionally, since many mutations occur in the early embryo in the first few days of life, a cloned embryo would not be exempt from developing these mutations, which would be transmitted to the inner mass cells (i.e. the embryonic stem cells). In normal development in the uterus, the majority of these defective embryos would be eliminated because they can't develop to implantation into the mother’s uterus and develop beyond the early stages of life, but when development is stopped at 5 to 7 days by the cloning process, these early mutations are not eliminated. There is the possibility that these mutations may cause problems in the differentiated cells developed from the defective clone's embryonic stem cells. Also, reprogramming and imprinting errors of the patient’s chromosomes developed in the early embryo would probably remain in the stem cells developed from that embryo, and may lead to future problems, perhaps malignancies, in the cells and tissues developed from them. It is important to remember that destroying a cloned embryo to get its stem cells would also be killing a little growing human being.
Ethical problems aside, we have to remember that there is no guarantee that we will be able to master the process of directing embryonic stem cells from either cloned embryos or IVF embryos into developing into the kinds of differentiated cells or tissues we need for therapy without causing harm to the recipient of these cells or tissues: we are years away from achieving the goal of safe and effective embryonic stem cell therapy.
Older embryo and fetal stem cells
Stem cells are also obtained by isolating cells from an area of the body of 5-8 week embryos and 9 week-old fetuses called the germinal ridge: these cells are called primordial germ cells and are the cells which would eventually differentiate into oocytes or sperm, depending on the sex of the embryo from which they are obtained.
These cells are diploid (i.e. have 46 chromosomes), as they have not yet undergone the “meiotic” division which they need to undergo to become mature haploid (i.e. have 23 chromosomes) oocytes or sperm. The embryos and young fetuses, which are usually obtained from abortions, are autopsied, and their germinal ridges are removed and the cells from there are isolated and grown in special tissue culture media. It is also possible to obtain such stem cells from embryos and fetuses who have died due to miscarriage or premature birth. However, specialized cells and tissues formed from these stem cells would lead to the same immunological rejection problems as would embryonic stem cells. They also form teratomas, as was found in a study with human patients where these cells were injected into the brain - teratomas formed and had to be surgically removed. So, these kinds of stem cells also present problems.
Umbilical cord stem cells
These are harvested from umbilical cords and placentas at the time of birth. Many parents are having their newborn’s umbilical cord stem cells harvested and stored in cell banks (similar to blood banks) for future use, if the need for a bone marrow transplant ever arises. These cells will not form teratomas, but would cause rejection problems if used by others besides the child from whose umbilical cord they come. Much work is being done on them, but it is not yet known how many types of organs or tissues they could form.
Adult (i.e. post-natal) stem cells
Children and adults have stem cells in their bodies. Adult stem cells are obtained from bone marrow donations or organ biopsies: these procedures do not permanently harm the donor, though they may cause a bit of local pain when they are performed. Several organs, such as pancreas, liver and brain, have been found to have stem cells specific to them and there are also certain other kinds of stem cells which can make a variety of different tissues, which would be very useful for treating disease: there are many reports about this in medical and scientific journals. One of the more versatile adult stem cells is the bone marrow stromal cell, which can give rise to many different kinds of specialized cells, including liver cells (hepatocytes), brain cells (neurons), dendritic cells (immune system cells), bone cells, fat cells and cartilage cells. Another very exciting example of an adult stem cell is one also found in the bone marrow and called multipotent adult progenitor cells (MAPCs) which can be made to differentiate into cells of all three embryonic layers - endoderm, mesoderm and ectoderm, including the various kinds of blood cells: the ability to do this makes these cells as versatile as embryonic stem cells. The MAPCs are found in human bone marrow, as well as in mouse bone marrow. They do not form teratomas, and would not cause immunological rejection, as they would be isolated from the bone marrow of the patient who would receive the cells or tissues made from them. Another potentially exciting new stem cell has been found in peripheral blood, which can be differentiated into endothelial cells, nerve cells and liver cells and certain kinds of blood cells, but it is not yet known if these can develop into red blood cells, which they would have to do to be useful for EPP treatments. The advantage of these cells would be that patients would not have to undergo bone marrow punctures, which can be somewhat painful - drawing blood from a vein is much less so.
From our survey of stem cells, it would appear that the best and safest ones to use would be adult stem cells, specifically those able to make hematopoietic (blood) cells. The word “erythropoietic” in EPP’s name means red blood cells. So, my plan now is to investigate the traditional bone marrow stem cells, as well as the MAPCs and the new peripheral blood stem cell I mentioned, and see if we can get the normal gene for ferrochelatase (the enzyme which is defective in EPP and causing all the problems) into these stem cells without using viral vectors. So, keep in touch with EPPREF - we’ll keep you posted. And wish me luck - prayers won’t hurt either! If any of you have any questions about stem cells or gene therapy, please don’t hesitate to contact me.
Contact EPPREF:
By phone: 617-525-8249
By e-mail: mmmathroth@rics.bwh.harvard.edu.
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