Transcript Immunological Bioinformatics Ole Lund Center for Biological Sequence Analysis BioCentrum-DTU Technical University of Denmark [email protected].

Immunological Bioinformatics
Ole Lund
Center for Biological Sequence Analysis
BioCentrum-DTU
Technical University of Denmark
[email protected]
Challenges and failures of the immune
system
Outside
Infection
with
microbe A
Infection
with
microbe B
Allergen ->
allergy
Peptide
drugs
Time
Creation Creation
of self
of an
immune
system/
Tolerance
to self
Autoimmunity
(break of
tolerance to
self)
Inside
Cancer
How does the immune system “see” a virus?
Immune system overview
•Innate – fast, unspecific
•Addaptive – specific, remembers…
•Cellular
•Cytotoxic T lymphocytes (CTL)
•Helper T lymphocytes (HTL)
•Humoral
•B lymphocytes
Figures by Eric A.J. Reits
MHC Class I pathway
Response to
1:(5x20x200)
= 1:2000
9mers
1:5
1:200
Figure by Eric A.J. Reits
1:2
Genomes to vaccines
Lauemøller et al., 2000
Cytotoxic T
Lymphocyte
T Cell receptor (TCR)
Peptide
MHC
b2m
CD8
Fugure by Thomas Blicher
Antigen Presenting cell
Figure by Anne Mølgaard, peptide (KVDDTFYYV) used as vaccine by Snyder et al. J Virol 78, 7052-60 (2004).
Influenza A virus (A/Goose/Guangdong/1/96(H5N1))
Genome
>Segment 1
agcaaaagcaggtcaattatattcaatatggaaagaataaaagaactaagagatctaatg
tcgcagtcccgcactcgcgagatactaacaaaaaccactgtggatcatatggccataatc
aagaaatacacatcaggaagacaagagaagaaccctgctctcagaatgaaatggatgatg
gcaatgaaatatccaatcacagcagacaagagaataatggagatgattcctgaaaggaat
and 13350 other nucleotides on 8 segments
Proteins
9mer peptides
>polymerase“
MERIKELRD
MERIKELRDLMSQSRTREILTKTTVDHMAIIKKYTSGRQEKNPALRMKWMMAMKYPITAD
ERIKELRDL
KRIMEMIPERNEQGQTLWSKTNDAGSDRVMVSPLAVTWWNRNGPTTSTVHYPKVYKTYFE
RIKELRDLM
KVERLKHGTFGPVHFRNQVKIRRRVDINPGHADLSAKEAQDVIMEVVFPNEVGARILTSE
IKELRDLMS
SQLTITKEKKEELQDCKIAPLMVAYMLERELVRKTRFLPVAGGTSSVYIEVLHLTQGTCW
KELRDLMSQ
EQMYTPGGEVRNDDVDQSLIIAARNIVRRATVSADPLASLLEMCHSTQIGGIRMVDILRQ
ELRDLMSQS
NPTEEQAVDICKAAMGLRISSSFSFGGFTFKRTNGSSVKKEEEVLTGNLQTLKIKVHEGY
LRDLMSQSR
EEFTMVGRRATAILRKATRRLIQLIVSGRDEQSIAEAIIVAMVFSQEDCMIKAVRGDLNF
RDLMSQSRT
...
DLMSQSRTR
LMSQSRTRE
and 9 other proteins
and 4376 other 9mers
Weight matrices
(Hidden Markov models)
YMNGTMSQV
GILGFVFTL
ALWGFFPVV
ILKEPVHGV
ILGFVFTLT
LLFGYPVYV
GLSPTVWLS
WLSLLVPFV
FLPSDFFPS
CVGGLLTMV
FIAGNSAYE
A2 Logo
G
F
C
A
Lauemøller et al., 2000
Human Leukocyte antigen (HLA=MHC in humans)
polymorphism - alleles
http://www.anthonynolan.com/HIG/index.html
HLA polymorphism - supertypes
•Each HLA molecule within a supertype
essentially binds the same peptides
•Nine major HLA class I supertypes have been
defined
•HLA-A1, A2, A3, A24,B7, B27, B44, B58, B62
Sette et al, Immunogenetics (1999) 50:201-212
HLA polymorphism - frequencies
Supertypes
Phenotype frequencies
Caucasian
Black
Japanese
A2,A3, B7
83 %
86 %
88 %
88 %
86 %
86%
+A1, A24, B44
100 %
98 %
100 %
100 %
99 %
99 %
+B27, B58, B62
100 %
100 %
100 %
100 %
100 %
100 %
A Sette et al, Immunogenetics (1999) 50:201-212
Chinese Hispanic
Average
O Lund et al., Immunogenetics. 2004 55:797-810
O Lund et al., Immunogenetics. 2004 55:797-810
O Lund et al., Immunogenetics. 2004 55:797-810
Combined method
•Combining predicted MHC-I affinity with prediction of Cterminal proteasomal cleavage and TAP transport efficiency
improves the ability to identify known CTL epitopes
MV Larsen et al., Accepted for publication in European Journal of Immunology
Infectious Diseases
•More than 400 microbial agents are associated with disease in healthy
adult humans
•There are only licensed vaccines in the United states for 22 microbial
agents (vaccines for 34 pathogens have been developed)
•Immunological Bioinformatics may be used to
•Identify immunogenic regions in pathogen
•These regions may be used as in rational vaccine design
•Which pathogens to focus on? Infectious diseases may be ranked
based on
•Impact on health
•Dangerousness
•Economic impact
Infectious Diseases in the World
•11 million (19%) of the 57 million people who died in the world in 2002
were killed by infectious or parasitic infection [WHO, 2004]
•The three main single infectious diseases are HIV/AIDS, tuberculosis,
and malaria, each of which causes more than 1 million deaths
Deaths from
infectious diseases
in the world in 2002
www.who.int/entity/whr/2004/annex/topic/en/annex_2_en.pdf
Dodo
Pathogenic Viruses
1st column: log10 of the number of deaths caused by the
pathogen per year
2nd column: DNA Advisory Committee (RAC)
classification
DNA Advisory Committee guidelines [RAC, 2002] which
includes those biological agents known to infect humans, as
well as selected animal agents that may pose theoretical risks
if inoculated into humans. RAC divides pathogens into
four classes.
Risk group 1 (RG1). Agents that are not associated with disease in
healthy adult humans
Risk group 2 (RG2). Agents that are associated with human disease
which is rarely serious and for which preventive or therapeutic interventions
are often available
Risk group 3 (RG3). Agents that are associated with serious or lethal
human disease for which preventive or therapeutic interventions may be
available (high individual risk but low community risk)
Risk group 4 (RG4). Agents that are likely to cause serious or lethal
human disease for which preventive or therapeutic interventions are not
usually available (high individual risk and high community risk)
3rd column: CDC/NIAID bioterror classification
classification of the pathogens according to the Centers for
Disease Control and Prevention (CDC) bioterror categories
A–C, where category A pathogens are considered the worst
bioterror threats
4th column: Vaccines available
A letter indicating the type of vaccine if one is available (A:
acellular/adsorbet; C: conjugate; I: inactivated; L: live; P:
polysaccharide; R: recombinant; S staphage lysate; T: toxoid).
Lower case indicates that the vaccine is released as an
investigational new drug (IND)).
5th column: G: Complete genome is sequenced
Data derived from /www.cbs.dtu.dk/databases/Dodo.
Biodefence
Targets
www2.niaid.nih.gov/Biodefense/
bandc_priority.htm
NIH project
Pathogen
HLA binding
Elispot
Influenza
X
X
Variola major (smallpox) vaccine strain
X
X
Yersinia pestis
X
Francisella tularensis (tularemia)
X
LCM
X
Lassa Fever
X
Hantaan virus (Korean hemorrhagic fever virus)
X
Rift Valley Fever
X
Dengue
X
Ebola
X
Marburg
X
Multi-drug resistant TB (BCG vaccine)
X
Yellow fever
X
Typhus fever (Rickettsia prowazekii)
X
West Nile Virus
X
CBS/panum
X
Strategy for determination of peptide-HLA binding
Step I: Folding of MHC class I molecules in solution
b2m
Heavy chain
peptide
Incubation
Peptide-MHC
complex
Step II: Detection of de novo folded MHC class I molecules by ELISA
Development
C Sylvester-Hvid et al., Tissue Antigens. 2002 59:251-8
ELISPOT assay
•Measure number of white blood cells that in vitro produce
interferon-g in response to a peptide
•A positive result means that the immune system have
earlier reacted to the peptide (during a response ot a
vaccine/natural infection)
SLFNTVATL
SLFNTVATL
SLFNTVATL
SLFNTVATL
SLFNTVATL
SLFNTVATL
Two spots
Preliminary results
•167 peptides have so far been tested for binding to a HLA
molecule
•113 of these (67%) have been shown to bind to the relevant
HLA allele with a affinity better than 500nM
•180 predicted epitopes from influenza A virus were tested in
an ELISPOT assay
•12 were so far found to be epitopes (recognized by donors
previously exposed to Influenza)
•14% of peptides binding with an affinity better than 500nM
were found to be epitopes
•1:2000 randomly chosen peptides are epitopes
Vaccination
•Vaccination
•Administration of a substance to a person with the purpose of
preventing a disease
•Traditionally composed of a killed or weakened microorganism
•Vaccination works by creating a type of immune response that enables
the memory cells to later respond to a similar organism before it can
cause disease
Early History of Vaccination
•Pioneered India and China in the 17th century
•The tradition of vaccination may have originated in India in AD 1000
•Powdered scabs from people infected with smallpox was used to
protect against the disease
•Smallpox was responsible for 8 to 20% of all deaths in several
European countries in the 18th century
•In 1721 Lady Mary Wortley Montagu brought the knowledge of these
techniques from Constantinople (now Istanbul) to England
•Two to three percent of the smallpox vaccinees, however, died from the
vaccination itself
•Benjamin Jesty and, later, Edward Jenner could show that vaccination
with the less dangerous cowpox could protect against infection with
smallpox
•The word vaccination, which is derived from vacca, the Latin word for
cow.
Vaccination Today
•Vaccines have been made for only 34 of the more than 400 known
pathogens that are harmful to man.
•Immunization saves the lives of 3 million children each year, but that 2
million more lives could be saved if existing vaccines were applied on a
full-scale worldwide
Human Vaccines
against pathogens
Immunological Bioinformatics, The MIT press.
Categories of Vaccines
•Live vaccines
•Are able to replicate in the host
•Attenuated (weakened) so they do not cause disease
•Subunit vaccines
•Part of organism
•Genetic Vaccines
•Part of genes from organism
Polytope construction
Linker
NH2 M
Epitope
COOH
C-terminal cleavage
Cleavage within epitopes
cleavage
New epitopes
Helper responses
Figures by Eric A.J. Reits
Figure by Anne Mølgaard
MHC class II prediction
Complexity of problem
– Peptides of different
length
– Weak motif signal
Alignment crucial
Gibbs Monte Carlo
sampler
M Nielsen et al., Bioinformatics. 2004 20:1388-97
RFFGGDRGAPKRG
YLDPLIRGLLARPAKLQV
KPGQPPRLLIYDASNRATGIPA
GSLFVYNITTNKYKAFLDKQ
SALLSSDITASVNCAK
PKYVHQNTLKLAT
GFKGEQGPKGEP
DVFKELKVHHANENI
SRYWAIRTRSGGI
TYSTNEIDLQLSQEDGQTIE
Class II binding motif
Alignment by Gibbs sampler
RFFGGDRGAPKRG
YLDPLIRGLLARPAKLQV
KPGQPPRLLIYDASNRATGIPA
GSLFVYNITTNKYKAFLDKQ
SALLSSDITASVNCAK
PKYVHQNTLKLAT
GFKGEQGPKGEP
DVFKELKVHHANENI
SRYWAIRTRSGGI
TYSTNEIDLQLSQEDGQTI
M Nielsen et al., Bioinformatics. 2004 20:1388-97
Random
ClustalW
Gibbs sampler
Antibody responses
Figures by Eric A.J. Reits
Prediction of Antibody epitopes
Linear
– Hydrophilicity scales (average in ~7 window)
• Hoop and Woods (1981)
• Kyte and Doolittle (1982)
• Parker et al. (1986)
– Other scales & combinations
• Pellequer and van Regenmortel
• Alix
– New improved method (Pontoppidan et al. in preparation)
• http://www.cbs.dtu.dk/services/BepiPred/
Discontinuous
– Protrusion (Novotny, Thornton, 1986)
• Pernille Haste Andersen, 2005, in preparation
Immunological bioinformatics
Classical experimental research
– Few data points
– Data recorded by pencil and
paper/spreadsheet
New experimental methods
– Sequencing
– DNA arrays
– Proteomics
Need to develop new methods for
handling these large data sets
• Immunological Bioinformatics/Immunoinformatics
Sune
Frankild
Jens
Pontoppidan
Morten
Nielsen
Pernille
Haste
Andersen
Thomas
Blicher
Claus
Lundegaard
Anne
Mølgaard
Xiuxiu
Ye