The document discusses mutational processes in human cancer and methods for analyzing mutational signatures. It describes extracting 30 mutational signatures from the mutation catalogs of 12,023 cancer cases across 40 cancer types using non-negative matrix factorization. Certain signatures are found to dominate in specific cancer types, such as Signature 1 and Signature 5 in breast cancer. Clock-like mutational signatures accumulate with age and are present across individuals.

1. Understanding mutational processes in human
cancer
Ludmil B. Alexandrov
lba@lanl.gov

2. From fertilized egg to cancer cell

3. From fertilized egg to cancer cell

4. From fertilized egg to cancer cell

5. From fertilized egg to cancer cell

6. From fertilized egg to cancer cell

7. From fertilized egg to cancer cell

8. From fertilized egg to cancer cell

9. From fertilized egg to cancer cell

10. From fertilized egg to cancer cell

11. From fertilized egg to cancer cell

12. From fertilized egg to cancer cell

13. From fertilized egg to cancer cell

14. Mutational processes and somatic mutations

15. Mutational processes and somatic mutations
Common DNA
damages

16. Mutational processes and somatic mutations
Common DNA
damages

17. Mutational processes and somatic mutations
Common DNA
damages

18. Mutational processes and somatic mutations
Common DNA
damages

19. Mutational processes and somatic mutations
Positive Imprint
Common DNA
damages

20. Mutational processes and somatic mutations
Promoter
Methylation
Common DNA
damages

21. Mutational processes and somatic mutations
Negate Positive Imprint
Common DNA
damages
Promoter
Methylation

22. Mutational processes and somatic mutations
Mutational process: A combination of DNA damaging and repair mechanisms
that act collectively and have the ability to cause mutations in somatic cells.
Common DNA
damages

23. Using mutational signatures to distinguish between different
mutational processes

24. UV Light – Mutational Signature “Poster Child”

25. 99.9% photons
converted into
harmless heat
UV Light – Mutational Signature “Poster Child”

26. What is a mutational signature?

27. UV Light - “Poster Child” Example

28. UV Light - “Poster Child” Example
UVB mutational
signature
UVA mutational
signature

29. UV Light - “Poster Child” Example
Mutational signature: The pattern of somatic mutations imprinted by
the activity of a mutational process.
UV-A: Predominately C>T, C>A, T>G, and T>C mutations.
UV-B: Predominately C>T and CC>TT.
Mutational signature: A unique identifier of a mutational process?

30. Deciphering Mutational Signatures in Human
Cancer

31. From fertilised egg to cancer cell

32. From fertilised egg to cancer cell

33. From fertilised egg to cancer cell
A
Mutational
processes

34. From fertilised egg to cancer cell
A
B
Mutational
processes

35. From fertilised egg to cancer cell
A
B
C
Mutational
processes

36. From fertilised egg to cancer cell
A
B
C
Final cancer
genome
Mutational
processes

37. Non-negative matrix factorization (NMF)

38. Classification of base substitution mutations
C>T
C>A
C>G
T>A
T>C
T>G
6 mutation classes

39. ……ATCGGGAATCGGACCCGATG……
……ATCGGGAATTGGACCCGATG……
Classification of base substitution mutations

40. ……ATCGGGAATCGGACCCGATG……
……ATCGGGAATTGGACCCGATG……
Classification of base substitution mutations

41. ……ATCGGGAATCGGACCCGATG……
……ATCGGGAATTGGACCCGATG……
……ATCGGGAAACGGACCCGATG……
……ATCGGGAAATGGACCCGATG……
Classification of base substitution mutations

42. ……ATCGGGAATCGGACCCGATG……
……ATCGGGAATTGGACCCGATG……
……ATCGGGAAACGGACCCGATG……
……ATCGGGAAATGGACCCGATG……
……ATCGGGAAACCGACCCGATG……
……ATCGGGAAATCGACCCGATG……
Classification of base substitution mutations

43. C>T
C>A
C>G
T>A
T>C
T>G
6 mutation classes
Classification of base substitution mutations

44. C>T
C>A
C>G
T>A
T>C
T>G
ACA>ATA
ACC>ATC
ACG>ATG
ACT>ATT
CCA>CTA
CCC>CTC
CCG>CTG
CCT>CTT
GCA>GTA
GCC>GTC
GCG>GTG
GCT>GTT
TCA>TTA
TCC>TTC
TCG>TTG
TCT>TTT
6 mutation classes
Classification of base substitution mutations

45. C>T
C>A
C>G
T>A
T>C
T>G
ACA>ATA
ACC>ATC
ACG>ATG
ACT>ATT
CCA>CTA
CCC>CTC
CCG>CTG
CCT>CTT
GCA>GTA
GCC>GTC
GCG>GTG
GCT>GTT
TCA>TTA
TCC>TTC
TCG>TTG
TCT>TTT
6 mutation classes ATA>AGA
ATC>AGC
ATG>AGG
ATT>AGT
CTA>CGA
CTC>CGC
CTG>CGG
CTT>CGT
GTA>GGA
GTC>GGC
GTG>GGG
GTT>GGT
TTA>TGA
TTC>TGC
TTG>TGG
TTT>TGT
ATA>ACA
ATC>ACC
ATG>ACG
ATT>ACT
CTA>CCA
CTC>CCC
CTG>CCG
CTT>CCT
GTA>GCA
GTC>GCC
GTG>GCG
GTT>GCT
TTA>TCA
TTC>TCC
TTG>TCG
TTT>TCT
ATA>AAA
ATC>AAC
ATG>AAG
ATT>AAT
CTA>CAA
CTC>CAC
CTG>CAG
CTT>CAT
GTA>GAA
GTC>GAC
GTG>GAG
GTT>GAT
TTA>TAA
TTC>TAC
TTG>TAG
TTT>TAT
ACA>AAA
ACC>AAC
ACG>AAG
ACT>AAT
CCA>CAA
CCC>CAC
CCG>CAG
CCT>CAT
GCA>GAA
GCC>GAC
GCG>GAG
GCT>GAT
TCA>TAA
TCC>TAC
TCG>TAG
TCT>TAT
ACA>AGA
ACC>AGC
ACG>AGG
ACT>AGT
CCA>CGA
CCC>CGC
CCG>CGG
CCT>CGT
GCA>GGA
GCC>GGC
GCG>GGG
GCT>GGT
TCA>TGA
TCC>TGC
TCG>TGG
TCT>TGT
96 mutation classes
Classification of base substitution mutations

46. Extracting mutational signatures from human cancers
12,023 cancer cases of 40 cancer types
12,023 mutation catalogues = 8,024,188 somatic mutations
Apply Non-negative Matrix Factorisation (NMF)
Extract mutational signatures
Estimate the contribution of each mutational signature to the
mutational catalogue of each of the 12,023 cancer genomes

47. Extracting mutational signatures from human cancers
12,023 cancer cases of 40 cancer types
12,023 mutation catalogues = 8,024,188 somatic mutations
Apply Non-negative Matrix Factorisation (NMF)
Extract mutational signatures
Estimate the contribution of each mutational signature to the
mutational catalogue of each of the 12,023 cancer genomes

48. Extracting mutational signatures from human cancers
12,023 cancer cases of 40 cancer types
12,023 mutation catalogues = 8,024,188 somatic mutations
Apply Non-negative Matrix Factorisation (NMF)
Extract mutational signatures
Estimate the contribution of each mutational signature to the
mutational catalogue of each of the 12,023 cancer genomes

49. Extracting mutational signatures from human cancers
12,023 cancer cases of 40 cancer types
12,023 mutation catalogues = 8,024,188 somatic mutations
Apply Non-negative Matrix Factorisation (NMF)
Extract mutational signatures
Estimate the contribution of each mutational signature to the
mutational catalogue of each of the 12,023 cancer genomes

50. Extracting mutational signatures from human cancers
12,023 cancer cases of 40 cancer types
12,023 mutation catalogues = 8,024,188 somatic mutations
Apply Non-negative Matrix Factorisation (NMF)
Extract mutational signatures
Estimate the contribution of each mutational signature to the
mutational catalogue of each of the 12,023 cancer genomes

51. The landscape of mutational
signatures across human cancer

52. Mutational signatures in human cancer

53. Mutational signatures in human cancer

54. Mutational signatures in human cancer

55. Mutational signatures in human cancer
c

56. Mutational signatures in human cancer

57. Mutational signatures in human cancer

58. Mutational signatures in human cancer

59. Mutational signatures in human cancer

60. Mutational signatures in human cancer

61. Mutational signatures in human cancer

62. Mutational signatures in human cancer

63. Breast cancers
Number of
mutations
per genome
Contributions of mutational signatures to individual
cancer cases

64. Mutational signatures by cancer type

65. Mutational signatures by cancer type

66. Mutational signatures by cancer type

67. Mutational signatures by cancer type

68. Mutational signatures by cancer type

69. Mutational signatures by cancer type

70. Mutational signatures by cancer type

71. Mutational signatures by cancer type

72. Mutational signatures by cancer type

73. 73
Cancer Type Example: Breast cancer

74. 74
97.51%
94.32%
62.13% 60.14%
48.20%
29.40%
14.60%
1.81% 1.62% 0.34% 0.22% 0.14%
0.00%
30.00%
60.00%
90.00%
Signature 1 Signature 5 Signature 2 Signature 13 Signature 8 Signature 3 Signature 18 Signature 6 Signature 26 Signature 20 Signature 10 Signature 30
Percentage of samples with signature Cancer Type Example: Breast cancer

75. 75
97.51%
94.32%
62.13% 60.14%
48.20%
29.40%
14.60%
1.81% 1.62% 0.34% 0.22% 0.14%
0.00%
30.00%
60.00%
90.00%
Signature 1 Signature 5 Signature 2 Signature 13 Signature 8 Signature 3 Signature 18 Signature 6 Signature 26 Signature 20 Signature 10 Signature 30
Percentage of samples with signature Cancer Type Example: Breast cancer

76. 76
97.51%
94.32%
62.13% 60.14%
48.20%
29.40%
14.60%
1.81% 1.62% 0.34% 0.22% 0.14%
0.00%
30.00%
60.00%
90.00%
Signature 1 Signature 5 Signature 2 Signature 13 Signature 8 Signature 3 Signature 18 Signature 6 Signature 26 Signature 20 Signature 10 Signature 30
Percentage of samples with signature Cancer Type Example: Breast cancer

77. 77
97.51%
94.32%
62.13% 60.14%
48.20%
29.40%
14.60%
1.81% 1.62% 0.34% 0.22% 0.14%
0.00%
30.00%
60.00%
90.00%
Signature 1 Signature 5 Signature 2 Signature 13 Signature 8 Signature 3 Signature 18 Signature 6 Signature 26 Signature 20 Signature 10 Signature 30
Percentage of samples with signature Cancer Type Example: Breast cancer

78. 78
97.51%
94.32%
62.13% 60.14%
48.20%
29.40%
14.60%
1.81% 1.62% 0.34% 0.22% 0.14%
0.00%
30.00%
60.00%
90.00%
Signature 1 Signature 5 Signature 2 Signature 13 Signature 8 Signature 3 Signature 18 Signature 6 Signature 26 Signature 20 Signature 10 Signature 30
Percentage of samples with signature Cancer Type Example: Breast cancer

79. 79
97.51%
94.32%
62.13% 60.14%
48.20%
29.40%
14.60%
1.81% 1.62% 0.34% 0.22% 0.14%
0.00%
30.00%
60.00%
90.00%
Signature 1 Signature 5 Signature 2 Signature 13 Signature 8 Signature 3 Signature 18 Signature 6 Signature 26 Signature 20 Signature 10 Signature 30
Percentage of samples with signature Cancer Type Example: Breast cancer

80. 80
97.51%
94.32%
62.13% 60.14%
48.20%
29.40%
14.60%
1.81% 1.62% 0.34% 0.22% 0.14%
0.00%
30.00%
60.00%
90.00%
Signature 1 Signature 5 Signature 2 Signature 13 Signature 8 Signature 3 Signature 18 Signature 6 Signature 26 Signature 20 Signature 10 Signature 30
Percentage of samples with signature Cancer Type Example: Breast cancer

81. 81
97.51%
94.32%
62.13% 60.14%
48.20%
29.40%
14.60%
1.81% 1.62% 0.34% 0.22% 0.14%
0.00%
30.00%
60.00%
90.00%
Signature 1 Signature 5 Signature 2 Signature 13 Signature 8 Signature 3 Signature 18 Signature 6 Signature 26 Signature 20 Signature 10 Signature 30
Percentage of samples with signature Cancer Type Example: Breast cancer

82. Clock-like mutational signatures

83. Clock-like mutational signatures are generated through life
Time
Person A

84. Clock-like mutational signatures are generated through life
Time
Person A

85. Clock-like mutational signatures are generated through life
Time
Person A
Person B
Person C
Person D
Person E

86. Clock-like mutational signatures are generated through life
Time
Person A
Person B
Person C
Person D
Person E

87. Breast cancers
Number of
mutations
per genome
Contributions of mutational signatures to individual
cancer cases

88. Mutational signatures by cancer type

89. mber of mutations
Clock-like Signature 1 mutations
Somatic
mutations per
gigabase

90. Signature 1 is characterised by C>T substitutions
at NCG trinucleotides

91. Signature 1 is characterised by C>T substitutions
at NCG trinucleotides
ACG GCG
TCG
CCG

92. Spontaneous deamination of 5-methyl cytosine
causes C>T substitutions at NCG trinucleotides

93. Spontaneous deamination of 5-methyl cytosine
causes C>T substitutions at NCG trinucleotides

94. Spontaneous deamination of 5-methyl cytosine
causes C>T substitutions at NCG trinucleotides

95. Spontaneous deamination of 5-methyl cytosine
causes C>T substitutions at NCG trinucleotides

96. Signature 1 mutation rates in human cells

97. Signature 1 mutation rates in human cells

98. Signature 1 mutation rates in human cells

99. Signature 1 mutation rates in human cells
~90 substitutions / diploid genome / year

100. Signature 1 mutation rates in human cells

101. Signature 1 mutation rates in human cells

102. Signature 1 mutation rates in human cells

103. Signature 1 mutation rates in human cells

104. Signature 1 mutation rates in human cells

105. Signature 1 mutation rates in human cells

106. Signature 1 mutation rates in different tissues
correlate with cell turnover
Signature 1 slopes
(mutations per
gigabase per year)
P=.0033 Mann-Whitney

107. Signature 1 may be a mitotic clock

108. Signature 5 is characterised predominantly by
C>T and T>C substitutions

109. Signature 5 mutation rates in human cells

110. Signature 5 mutation rates in human cells
~90 substitutions/diploid genome/year

111. Signature 5 mutation rates in human cells

112. Signature 5 mutation rates in human cells

113. Comparing signature 1 and 5 mutation rates

114. Signature 1 and 5 mutation rates in human cells

115. Signature 1 and 5 mutation rates in human cells

116. Signature 1 and 5 mutation rates in human cells

117. Signature 1 and 5 mutation rates in human cells

118. What is the mutational process underlying
Signature 5?

119. What is the mutational process underlying
Signature 5?
Why do Signature 5 mutation rates vary between cell types?

120. c
c
de novo
germline
mutations
Signature 1
Signature 5
c
Signatures 1 and 5 together recapitulate the
germline mutation pattern

121. c
c
de novo
germline
mutations
Signature 1
Signature 5
c
Signatures 1 and 5 together recapitulate the
germline mutation pattern

122. c
c
de novo
germline
mutations
Signature 1
Signature 5
c
Signatures 1 and 5 together recapitulate the
germline mutation pattern

123. Signatures 1 and 5 together recapitulate the
germline mutation pattern

124. Signatures 1 and 5 together recapitulate the
germline mutation pattern

125. • Signatures 1 and 5 are present in all cancer types and almost all cancer samples
• Signatures 1 and 5 are clock-like
• Signature 1 may be a mitotic clock
• The mutational process underlying Signature 1 is known but not for Signature 5
• A combination of Signatures 1 and 5 recapitulates germline (inherited) variation
Clock-like Signatures Summary

126. Etiologies of mutational signatures

127. Known or speculative causes of mutational signatures

128. Known or speculative causes of mutational signatures
Deamination
of 5-methyl
cytosine;
clock-like
process

129. Known or speculative causes of mutational signatures
Unknown;
clock-like
process

130. Tobacco
smoking
Known or speculative causes of mutational signatures

131. Tobacco
chewing
Known or speculative causes of mutational signatures

132. Ultraviolet
light
Known or speculative causes of mutational signatures

133. Aflatoxin
Known or speculative causes of mutational signatures

134. Aristolochic
acid
Known or speculative causes of mutational signatures

135. Temozolomide
Known or speculative causes of mutational signatures

136. Defective DNA
mismatch repair
Known or speculative causes of mutational signatures

137. Defective
BRCA1, BRCA2,
homologous
recombination
repair
Known or speculative causes of mutational signatures

138. Defective
polymerase
epsilon activity
Known or speculative causes of mutational signatures

139. Infidelity of
polymerase eta
activity
Known or speculative causes of mutational signatures

140. Psoralen
exposure?
Known or speculative causes of mutational signatures

141. Reactive oxygen
species?
Known or speculative causes of mutational signatures

142. APOBEC
cytosine
deamination
Known or speculative causes of mutational signatures

143. Known or speculative causes of mutational signatures
Unknown

144. • There are more than 30 mutational signatures across human cancer
• Some signatures are due to known mutagenic exposures or known defects in DNA
maintenance but the cause of many is unknown
• Understanding the mutational processes underlying mutational signatures will
inform on cancer causation, prevention and treatment
Summary

145. http://cancer.sanger.ac.uk/cosmic/signatures

146. Acknowledgements – general
Signatures Collaborators
Philip Jones
Julian Sale
Steve Rozen
Teh Bin Tean
Stefan Pfister
Sean Grimmond
Peter Lichter
Jiri Zavadil
Elias Campo
Jessica Zucman-Rossi
Xose Puente
Matthew Meyerson
Lincoln Stein
Reiner Siebert
Sanger Institute
Mike Stratton
Serena Nik-Zainal
Peter Campbell
David Wedge
Los Alamos National Lab
Jennifer Green
Gnana Gnanakaran
Alan Bishop
Kim Rasmussen
William Hlavacek
Angel Garcia