nf1: tumor suppressor gene

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NF1: Tumor Suppressor Gene

Presentation by

Hana Masood

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Overview of Presentation

Disease – Neurofibromatosis Type I NF1 Gene Protein - Neurofibromin Protein Function – RasGAP Biological Role – Active in Ras Pathway NF1 gene and Neurofibromin Role in Cancer

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Neurofibromatosis Type I Among most common neurogenetic disorders-It is heritable; all affected members of family have same form of NF Affects 1 in 3,500- Whereas NF2 affects only 1 in 40,000 Diagnosis based on any 2 of the following clinical criteria:- Greater than or equal to 6 café-au-lait spots- Freckles underarm or groin- Greater than or equal to 2 neurofibromas, or 1 plexiform

neurofibroma- Iris Lisch nodules identified by ophthalmologist- Tumor of optic nerve called optic pathway glioma (in astrocytes)- Bone deformities (i.e. around eye, tibia)- First degree relative (parent, sibling, or child)

http://www.understandingnf1.org/id/int_id_win.html

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Neurofibromatosis Type I

Affects peripheral nerves brain, and spinal cord

More active as get older In adolescence, also find

learning disabilities in reading, math, trouble following directions, and paying attention

All racial and ethnic backgrounds

Symptoms vary person to person

http://www.understandingnf1.org/exp/int_exp_wcn.html

European Genetics Foundationhttp://www.charite.de/ch/medgen/eumedis/embryology04/neurocutan-disorders.html

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NF1 Gene

NF1 is a tumor suppressor gene

NF1 is located on Chromosome 17 long arm- NF2 on other hand is located on Chromosome 22

NF1 gene encodes the protein Neurofibromin

http://www.charite.de/ch/medgen/eumedis/embryology04/neurocutan-disorders.html

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NF1 Gene

Cloned in 1990 Over 300kb of genomic DNA, 50 exons Very large and complicated, and genetic

changes can be very small and subtle A missense mutation in GAP-Related Domain

is one way NF1 is inactivated- usually a mutation that causes a charge inversion; mostly change from Lysine to something like Glutamine in protein

cc.oulu.fi/~anatwww/ NF/Neurofibromin/Ras.jpg

Scheffzek et al, The Embo Journal, 1998, Vol. 17

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NF1 Gene Inactivation (Loss of Function)

50% of cases inherited from parent, other 50% of cases start out as a new NF1 mutation in sperm or egg that makes the child

All cell types are initially NF+/- until a subpopulation of Schwann cells undergo LOH and become NF-/-

Bi-allelic inactivation in Schwann cells causes the lack of expression of Neurofibromin protein

- Schwann cells are the initiating cells in neurofibroma formation

So, NF1 behaves as a dominant trait phenotypically, but is still recessive genotypically

http://www.understandingnf1.org/id/int_id_win.htmlHarrisingh and Lloyd, Cell Cycle, 2004, Vol. 3, p. e69

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Biochemical and Cell Biological Role of Neurofibromin

Neurofibromin acts as a RasGAP that acts mostly in Schwann cells

- What is a RasGAP?

Lodish et al. Fig. 20-22

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Neurofibromin Molecule

Has homology to p120GAP Expressed in all tissues, highest levels in

brain, kidney, and spleen Associates with cytokeratin bundles in human

keratinocytes and also binds tubulin Thus, exerts its effects in part by controlling

organization of cytoskeleton during the formation of cellular contacts


RasGAP(250-400 AA)

2818

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Neurofibromin GAP-Related Domain

Scheffzek et al, The Embo Journal, 1998, Vol. 17http://www.nature.com/emboj/journal/v17/n15/full/7591132a.html

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Neurofibromin GAP-Related Domain and Ras Interaction

Scheffzek et al, The Embo Journal, 1998, Vol. 17

http://www.nature.com/emboj/journal/v17/n15/full/7591132a.html

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Neurofibromin in Ras Pathway


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Neurofibromin involvement in Schwann cells – Biological Role

Neurofibromin involved in Schwann cell differentiation

Schwann cells are a regenerative cell type that has no known stem-cell population

Part of peripheral nerves

http://fig.cox.miami.edu/~cmallery/150/neuro/sf42x10b.jpg

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Schwann Cells

http://www.unis.org/UNIScienceNet/Schwann%20cell.jpg http://www.bu.edu/histology/i/21301loa.jpg

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Biological Role of Neurofibromin

Following nerve damage, myelinating Schwann cells produce new cells by de-differentiating, proliferating, and then re-differentiating

Axons in vivo provide signals that regulate Schwann cell differentiation

Ras causes de-differentiation and leads to proliferation, and then Neurofibromin RasGAP causes re-differentiation of Schwann cells

Harrisingh and Lloyd, Cell Cycle, 2004, Vol. 3, p. e69, 2004

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Biological Role of Neurofibromin NF1 inactivation causes

constitutive Ras activity in Schwann cells

Ras signalling drives the de-differentiation of myelinating Schwann cells that lead to tumors if uncontrolled

Thus, the role of Neurofibromin is to act as a tumor suppressor and control Ras activity by catalyzing its inactivation when necessary

Harrisingh and Lloyd, Cell Cycle, 2004, Vol. 3, p. e69-70, 2004

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Biological Role of Neurofibromin – Studies

NF1 bi-allelic inactivation is embryonically lethal in mice

NF1-/- Schwann cells are angiogenic and invasive in vitro, and have:- elevated levels of Ras-GTP (active) compared to wild-type cells- altered morphology elongated and hyper-refractile, long processes

• NF1+/- Schwann cells have intermediate phenotype (some Neurofibromin activity)


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Summary of Biological Role of Neurofibromin

Ras pathway in Schwann cell acts by driving the de-differentiation process itself, not by blocking a differentiation signal

The Raf/ERK signal can act dominantly over differentiation signals emanating from axons

Neurofibromin indirectly allows re-differentiation to happen by controlling the level of de-differentiation


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NF1 Inactivation Role in Cancer – De-Differentiation

• NF1+/- Schwann cells might not have sufficiently increased Ras signalling to de-differentiate

• But, state may make Schwann cells more susceptible to de-differentiation and facilitate loss of second allele (LOH) to shift equilibrium in favor of the de-differentiated state

Harrisingh and Lloyd, Cell Cycle, 2004, Vol. 3, p. e70-71

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NF1 Inactivation Role in Cancer – Nonautonomous Process

NF1-/- Schwann cells really induce neurofibroma formation in the presence of abnormal heterozygous NF1 environment that drive proliferation

Haploinsufficient state of somatic tissue surrounding NF1 tumors initiates or progresses tumor formation

When cellular environment has both functional NF1 alleles, the onset, growth potential, and multicullular nature of NF1-/-

neurofibromas is suppressed

Zhu et al, Science, 2002, Vol. 296, p. 920, 922

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“Tumor formation is a coordinated process in which incipient tumor cells recruit collaborating cells (i.e. mast, fibroblast cells) from the environment”

Through heterozygous inactivation mouse study, believed that heterozygous mast cells interacting with nullizygous Schwann cells creates a cytokine-rich microenvironment permissive for tumor growth

Zhu et al, Science, 2002, Vol. 296, p. 921-22

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• NF1+/- mice have abnormal responses to skin wounding, with increased collagen deposition and fibroblast proliferation, also observed in neurofibroma formation

NF+/- neurons, mast cells, and fibroblasts in mice do not undergo NF1 LOH, promote abnormal cell environment

NF+/- fibroblasts have defects in proliferation and perineurial sheath formation, allowing inappropriate exposure to circulating growth factors and hormone

NF-/- Schwann cells secrete increased levels of stem cell factor, thought to stimulate mast cell chemotaxis, which induces collagen deposition, cell proliferation, and angiogenesis


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NF1 Inactivation Role in Cancer - Overall• Nerve damage could trigger

the production of proliferative signals by NF1+/- fibroblasts, nerve cells, and mast cells that could result in the unregulated proliferation of de-differentiated NF1-/- Schwann cells that have lost Neurofibromin activity

• NF1 patients undergo an altered response to normal nerve repair signal with repeated cycles of attempted repair resulting in neurofibroma formation


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Role of the Inactivation of NF1 in Cancer (Neurofibromatosis) - Animation

Inactivation of NF1 in Cancer

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Neurofibromatosis Type I and Cancer Neurofibromas – benign Plexiform neurofibromas – subtype of benign

tumors, can undergo transformation by CDKN2A/p16 or INK4A inactivation (causes loss of p16 cdk inhibitor) into malignant peripheral nerve sheath tumors (MPNSTs)

- Also, MPNSTs aberrantly express EGFR- MPNSTs are highly aggressive cancers- 8-13% of NF1 patients develop it 15-20% of NF1 children develop low-grade

astrocytomas

Arun and Gutmann, Current Opinion in Neurology, 2004, Vol. 17, p. 101http://www.cancerindex.org/geneweb/NF1.htm#malig

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Neurofibromatosis Type I and Cancer “NF1 patients developing

malignant neoplasms can have any type of NF1 germline mutation such as a total gene deletion, a frameshift mutation, an in-frame deletion, or a missense mutation. We conclude that in our series no specific type of NF1 germline mutation was found in NF1 individuals with malignancies, but that large NF1 gene deletions were more frequently found in this group than reported for the general population of NF1 individuals.”

Genes Chromosomes Cancer, Vol. 26, p. 376-80, 1999

European Genetics Foundationhttp://www.charite.de/ch/medgen/eumedis/embryology04/neurocutan-disorders.html

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What did the Elephant Man have?

Joseph Merrick did not have NF1 He really had another disease Proteus Syndrome

(sine had mutation on another gene)

Genetic Science Learning Center at the University of Utahhttp://gslc.genetics.utah.edu/units/disorders/nf1/elephantman.cfm

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Treatment for NF1 Primary option is surgery for both benign and

malignant tumors Noncomparative phase II evaluation of cis-retinoic

acid (promaturation agent) and interferon-alpha (anti-inflammatory and antiangiogenic effects)

Phase I studies for farnesyl protein transferase inhibitors that inhibit Ras activity

Pirfenidone, an antibiotic drug that regulates cytokine action, in clinical trial

EGFR inhibitors may be effective in MPNSTs Possibly therapies that neutralize the effects of

haploinsufficiency before the onset of tumorogenesis

Arun and Gutmann, Current Opinion in Neurology, 2004, Vol. 17, p. 102, 104

nf1: tumor suppressor gene

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