Immunogenetica della LLC: implicazionui patogenetiche e prognostiche mediante analisi del gene IGHV1-69

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Immunogenetica della LLC implicazionui
patogenetiche e prognostiche mediante analisi del
gene IGHV1-69
Orvieto, Palazzo Coelli 21 Novembre 2009

• Francesco Forconi
• Ematologia e Trapianti
• Università di Siena

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IGH Variable region
IGHV-D-J rearrangement
Hypervariable Region
HCDR3
HCDR1
HCDR2
HFR1
HFR2
HFR3
N Region
IGHV
IGHJ
IGHD
51 IGHV genes
27 IGHD genes
6 IGHJ genes
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Midollo Osseo
Organi linfoidi secondari/ Marginal Zone
IgD
IgM
IgG
UM-CLL 40
M-CLL 60
MUTAZIONI SOMATICHE
IgM
IgM
Ig
IgD
Ig
Cellula B immatura
Cellula B naive
Cellula B memoria
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Sopravvivenza e stato mutazionale dei geni IGHV
17 anni
293 mesi
9 anni
95 mesi
Hamblin, T. J. et al. Blood 1999941848-1854
Damle, R. N. et al. Blood 1999941840-1847
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BCR
IgD
IgM
IgG
UM-CLL 40
M-CLL 60
ZAP70
ZAP70 -

• phosphorylation of p72Syk
• intracellular Ca(2)(i)
• Rapid disease progression

• Slow disease progression

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Shared sequence stereotypic characteristics of
the HCDR3 suggest antigen selection of the
leukemic clones
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Top 10 in CLL
Murray et al BLOOD, 2008 (111).
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• Selective stimulation of the B cell of origin (?)
• Antigenic drive continuing following
  transformation (?)
• Are Stereotypes CLL-specific?

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IGHV1-69

• 14 alleli di cui i più frequenti IGHV1-6901,
  02, e 12 (riconosciuti dallanticorpo anti-51p1
  G6)
• Infrequente nella popolazione B del sangue
  periferico da analisi molecolari (Lipsky lt1)
• 13 di tutte le CLL
• 30 delle UM-CLL
• 227/259 (88) cases gt98 homology to germline
  alleles
• Dal 47 al 55 delle CLL stereotipate
• Nella CLL mediana dei casi 1-69 è 69 anni

N214
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51p1-IGHJ6 rearrangements expressed in the normal
B cell repertoire
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Comparison of the HCDR3 sequences of CLL and
normal B cells in the 51p1-IGHJ6-derived subset 5.
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G6-positive (IGHV1-69 51p1-expressing) B- cells
are part of the conventional resting naïve B-cell
population.

• 4.8 of all B-cells.
• CD27-negative, indicative of naïve B cells.
• IgM IgD CD23 CD5- CD38 (as in G6-ve naïve
  B-cells).
• A small percentage of CD5 B cells, not found in
  the memory B-cell subset.
• CD38 expression was similarly high in naïve and
  G6-positive populations.
• IgK (65) IgL (35) comparable to normal B
  cells and 51p1ve CLL (data not shown).
• Absence of activation markers (CD25 and CD69).

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Are Stereotypes CLL-specific?

• by focusing only on the IGHV1-69-derived
  sequences combined to IGHJ6 in age-matched normal
  subjects, we have found Stereotypic sequences
  of several of the major subsets described in CLL
  and of new potential subsets in gt 33 sequences
  cloned from normal donors.
• it is possible that this conserved sequences are
  a likely source of transformation to U-CLL and
  that they derive from the naïve B-cell
  repertoire.
• Little similarity in the HCDR3 junctional amino
  acids between cases of CLL and little similarity
  within normal B cells

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How does antigenic stimulation would continue
following transformation?

• HCDR3 driven clustering to identify prognostic
  subsets

Stamatopoulos, K. et al. Blood 2007109259-270
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Subset 1
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IGHV4-39 and transformation to Richter Sydrome
plt.001
No IGHV4-39
IGHV4-39
IGHV4-39/stereotypic HCDR3
IGHV4-39/no stereotypic HCDR3
No IGHV4-39/stereotypic HCDR3
No IGHV4-39/no stereotypic HCDR3
Events/N 5-year risk SE
IGHV4-39 6/20 35.4 13.5
No IGHV4-39 33/733 5.6 1.1
5-year risk p
IGHV4-39/stereotypic HCDR3 68.7 .003
IGHV4-39/no stereotypic HCDR3 0 .003
No IGHV4-39/stereotypic HCDR3 9.9 .005
No IGHV4-39/no stereotypic HCDR3 4.2 .005
Rossi, Clinical Cancer Research 2009
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IGHV1-69 progression
Stamatopoulos, K. et al. Blood 2007109259-270
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HCDR3 length in CLL
HCDR3 length in 1-69
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Summary

• By investigating the IGHV1-69-J6 repertoire we
  can observe that CLL-specific HCDR3 are present
  in the normal individuals
• The subsets with different clinical behavior may
  rely on (super)antigen stimulation. However, it
  remains to be demonstrated that stimulation
  occurs through specific CDR3 interaction.
• Lack of different behavior (CLL progression and
  overall survival) between stereotyped and non
  stereotyped UM-CLL using IGHV1-69 point to
  antigen stimulation via CDR3-independent antigen.
  
• Clinically, mutational status keeps being
  confirmed as the relevant tool to stratify
  progression risk in CLL

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p66Shc levels and clinical behavior of U-CLL and
M-CLL
Capitani et al, submitted 2009
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Siena Emanuele Cencini Elisa Sozzi Nagaja
Capitani Cosima Baldari Novara Davide
Rossi Gianluca Gaidano Aviano Riccardo
Bomben Valter Gattei Bellinzona Andrea
Rinaldi Francesco Bertoni Niguarda Milano Silvio
Veronese Marco Montillo Roma Dimitar
Efremov Luca Laurenti Giovanni Del
Poeta Modena Roberto Marasca Torino Marta
Coscia Massimo Massaia Southampton Kathy
Potter Freda K. Stevenson Salonicco Kostas
Stamatopoulos