1. Genetics of Langerhance Cell Histiocytosis
Dr Mohammad Keramatipour MD, PhD
Tehran University of Medical Sciences

2. Overview Langerhance Cell Histiocytosis (LCH) ?? Genetics of LCH
Genetic testing
Role in targeted therapy and prognosis
Role of new sequencing technologies in LCH management

3. Langerhans Cell Histiocytosis
Langerhans Cell Histiocytosis (LCH): a systemic disease associated with the proliferation and accumulation (granulomas) of Langerhans cells in various tissues
Langerhans cells play an immunologic role in protecting against environmental antigens

4. Langerhans Cell Histiocytosis
Epidemiology
1-2/100,000
Age of diagnosis: ~ 1-3 years
Etiology
Inflammatory, immunologic, or even neoplastic ??
Inflammatory myeloid neoplasia
Genetics??
Familial clustering
Concordance rate in monzygotic twins

5. LCH Pathogenesis RAS/RAF/ERK signaling pathway:
Key role in cell growth and proliferation

6. RAS/RAF/ERK Pathway

7. BRAF gene BRAF gene A proto-oncogene located on 7q34 21 exons
Encodes B-RAF protein
766-amino acid protein
Member of the RAF kinase family
Substantial role in regulating the RAS/RAF/ERK signaling pathway
B-RAF: three conserved domains: conserved regions 1-3

8. B-Raf protein CR1 CR2 CR3 B-Raf activation RAS interaction
A flexible linker that connects CR1 and CR3, acts as a hinge
CR3
B-Raf enzymatic kinase domain
: N-lobe, responsible for ATP-binding
: C-lobe, binds substrate proteins
B-Raf activation

9. BRAF Mutations BRAF germline mutations / AD conditions
Cardio-facio-cutaneous syndrome, type 1
Leopard syndrome, type 3
Noonan syndrome, type 7
BRAF somatic mutations
Melanoma
Lung cancers
Colorectal cancers ……

10. BRAF mutations BRAF somatic missense mutations
A majority of mutations within the kinase domain
p.V600E The most common mutation (90%) of the BRAF gene
The most common mutation tested for in clinical laboratories
66% of malignant melanomas and at lower frequency in a wide range of human cancers
non-Hodgkin lymphoma
colorectal cancer
thyroid carcinoma
non-small cell lung carcinoma
hairy cell leukemia
adenocarcinoma of lung

11. BRAF mutations Other more common pathogenic mutations:
R461I, I462S, G463E, G463V, G465A, G465E, G465V, G468A, G468E, N580S, E585K, D593V, F594L, G595R, L596V, T598I, V599D, V599E, V599K, V599R, V600K, A727V.
Most of these mutations are clustered to two regions:
Glycine-rich P loop of the N lobe
Activation segment and flanking regions
These mutations change the activation segment from inactive state to active state.

12. BRAF mutations Mutated BRAF proteins elevated kinase activity
cancer cell lines with the V600E mutation have a
RAS-independent growth
Vemurafenib (PLX4032) A highly selective and potent inhibitor of BRAF V600E.
It has marked antitumor effects against melanoma cell lines with the BRAF V600E mutation but not against cells with wild-type (non-mutated) BRAF.
Vemurafenib improves overall and progression-free survival in patients with advanced melanoma with the V600E mutation.

13. Redefining LCH 1982: RAS mutations causing tumor !
Targeted therapy prevents tumor growth
Mutations in others genes has the same effect
RAF genes: BRAF, ARAF
MEK genes: MAP2K1, MAP3K1
Genetic testing in cancer
Personalized Cancer Management

14. LCH Pathogenesis

15. RAS/RAF/MEK Pathway

16. Genetic Testing in Cancer
Genetic testing in cancer management
Screening
Hereditary cancers
Assisted diagnosis
Targeted therapy
Molecular classification of tumors
Monitoring treatment / recurrence

17. Genetic Testing in LCH First / Second lines: NGS for all related genes
Tumor sample / blood sample ???
First / Second lines:
Testing for V600E (c.1799T>A), and T599A
Analysis of exons 2 and 3 of MAP2K1
NGS for all related genes
Panel: BRAF, ARAF, MAP2K1, MAP3K1, ERBB3
Whole Exome Sequencing

18. Challenges in Genetic Testing
Detecting somatic mutations
Solutions:
Sampling
Analysis: NGS depth / Sanger sequencing

19. Thank you for listening, comments?