Transcript Document

The chromosomal and genetic
abnormalities of cancer:
Which one is the cause?
Asilomar, Jan. 7-9, 2007
Peter Duesberg, Alice Fabarius,
Ruediger Hehlmann and Ruhong Li
1
Cancer: a genetic or
a chromosomal disease?
 It is known for over 100 y ea rs t ha t c ancer s ar e
aneuploid, ha ving too many and r ar el y to o few
chr omosomes.
 It is als o k nown for over 100 ye ars t hat r adiat ion
car cinoge ns, X-r ays, ar e an aneu ploidogen s, and for
over 80 year s t ha t t hey are al so mut ag en s.
 Fur t her , it is known fr om over 3 0 y ea rs of s eq uenc ing
t ha t c ancer s c ont ain a lot of mut at ions.
But it is s t ill de ba te d, whet her c ancer is a g enet ic or a
chr omosomal diseas e.
2
The genetic cancer theory
The deb at e o ver t he causes of ca ncer is c urre nt ly he avily
bia sed i n f avour of t he ge net ic th eory – so h eavily th at
t her e is no funding f or t he c hr omosomal t he or y.
The gen et ic ca ncer t heory p ost ula t es t ha t 2- 6 spec if ic
mutat ions ar e suff icien t to ca use ca ncer .
Chromosomal a lt er at ions, if men t ioned a t a ll, a re
considere d secondary e ven ts (se e nex t 2 slid es).
Since chr omosomal al te ra ti ons are n ot her it able and t hus
not pa rt of convent ional ge net ic condit ioning, t he y ar e
simply d isr egarded as pr imary cau ses of cancer.
3
The genetic cancer theory
According
to the NCI
2-6
mutations
cause
cancer
[NCI:
Understanding
Cancer
Series,
2007].
4
The genetic theory turns a blind
eye to cancer-specific aneuploidy
According to
the NCI and
research
supported by it,
mutations
cause cancer
by changing the
cell’s
morphology
and “color” –
but not its
karyotype.
QuickTime™ and a
TIFF (Uncompressed) decompressor
are needed to see this picture.
5
NCI website 2006: Karyotype normal = ca = 22 chromosome pairs + X,Y
Yet, the
karyotypes of
cancers are very
abnormal
Karyotype of a human colon
cancer: Highly aneuploid, with 15
rearranged marker chromosomes
Karyotype of a diploid
human cell (male)
6
Karyotype of a breast cancer cell
7
MDA231-4
Landmarks of cancer not explained
by genetic theory
1) The chr omosomes of can ce r cells ar e a ne uploid, m ost a re
hyper-d iploid s ome are hypo-d iploid.
2) Maligna ncy is p roport ional t o th e de gr ee of a neuploidy.
3) Cancers c onta in ty pe-sp ecific, me ta st asis-sp ecific, and drug
res ist ance- specif ic chromosome a lt era t ions.
4) The chr omosomes of can ce rs c onsist of clonal an d n on- clona l
aneuploidies, and a r e t hus i nher ent ly unst ab le.
8
Landmarks of carcinogenesis not
explained by genetic theory
1) The e xist e nce of n on- mut age nic car cinogens: vinyl chloride, aro mati c
hydrocarbons and as be st os.
2) Car cinogens, mut agenic a nd non- muta genic, induce ra ndom aneup loidies,
which precede and se gre gat e w it h cancer.
3) The level of ra ndom an eup loidy det er mines t he c ancer ri sk.
4) Once init iat ed, ca r cinogenesis proceeds a ut o cata lyt ical ly: First slowly
wit hout phenotypes and t hen ever fast er wit h e vermore- malignant
phenot ypes – ju st l ike a chemical chain r eaction.
Example, cancers following at omic b omb- radiations o nly decades la t e r.
Not e : Carcinogenesis is not compatib le wit h a seque nce of s pecific
mut ati ons. Mut ations ar e s tab le and not progressive.
9
The genetic cancer theory also
has genetic Achilles heels
 Despit e 30 y ear s of t r ying, t here is st ill no unambiguous
evidence t ha t 2- 6 mut an t g enes f rom c an cers t ra nsfor m
diploid c ells w ith out k ary oty pe al t er a t ions (see below).
 Numerous t ra nsgenic m ouse st ra ins c ar ry o ne or more
put at ive oncogenes in t heir ger mlines, b ut t hey p r opaga t e
nor mally a nd t heir cancer r isks ar e simila r t o t hose of ot her
la b mice.
 The genet ic t heory pr edicts high ra tes of solid c ancers i n
newborns fro m c omplement ary s ubs ets of t ra nsfor ming genes
inher it ed fr om par ent s. But , t here is p ra ct ical ly n o ca ncer in
newborns (se e next) .
10
Age-dependent incidence of cancer
Age
<1
1-4
5-9
10-14
15-19
20-24
25-29
30-34
35-39
40-44
45-49
50-54
55-59
60-64
65-69
70-74
75-79
80-84
>85
Incidence
0.24
0.22
0.12
0.13
0.21
0.31
0.44
0.59
0.88
1.48
2.70
5.37
9.47
15.41
22.64
28.29
31.23
30.83
29.77
11
Chromosomal cancer theory
 Car cinoge ns i nit iat e c ar cinogenesis by ra ndom an eu ploidies,
br ea king chromosomes o r damag ing t he sp indle ap par a t us.
 Ane uploidy t hen des ta bilizes t he ka r yot ype, m isba la ncing
t ea ms of pr ot eins t ha t s ynt hesize , se gr egat e and r epair
chr omosomes.
 Ane uploidy is t hus a st eady source of chr omosomal v ar iat ions,
fr om which , in classical Dar winian t er ms, c ancer-s pecific
aneuploidies evolve a utoc a t a lytic a lly o ver t ime.
 The r at es at w hich ca ncer - specific an euploidies ev olve
depend on t he inst a bilit y of t he r espect ive ka r yot ype.
 Cancer-s pec ific a ne uploidies gen er a t e ca ncer - specific
phenoty pes, su ch as meta st asis an d d rug res ist an ce, by
alt ering t he dosag e and ex press ions o f 1000 s of genes – jus t
like t risomy 21 genera t es Down s yndrome.
12
Carcinogenesis
as chain
reaction of
aneuploidizations
13
Despite questioning the
orthodoxy,
I expected a full absolution for
this attractive new cancer theory
–
as on previous occasions …
14
15
Instead –
The empire strikes back …
16
Coulter et al. now claim 6 mutations
are “sufficient” for tumorigenicity
Acc ording t o Coult er et al. 6 mut ant genes ar e “suff icient to
convert n ormal h uman ce lls to a t umorigen ic s ta t e.” (Ca Res 2005)
– 3 mor e th an C oult er, Weinberg e t a l. cla imed in 1999.
The 6 genes w er e ar t ificially a ct ivat ed c ellula r t el omera se, cyclin,
cyclin k inase, p 53 , myc - , an d ras genes t hat were int roduced int o
normal h uman ce lls in ret r ovir us vectors .
Since tu morige nicity was obse rved in nude mice about 2 mont hs
la t er, t he au t hors c onclude d, “t he t umors gr ew t oo f as t t o a llow
for t he s elect ion of a ddit iona l even t s”.
17
But Coulter also provides evidence
that 6 mutations are not “sufficient”
1) Only 5 out o f 100,000 “convert ed” cell s f orm 3 dimensiona lly gr owing f oci in vit r o, a s urr ogat e f or
t umor ige nicity .
2) I nocula ti on of 10 million conver t ed c ell s was u sed
(nec essar y?) to ge ner at e on e t umor in mice.
3) The s olid tu mors were re port ed t o have “large
nucle i” or “large p olymorph ic n uclei”. Bu t large
nucle i ar e t he phen oty pe s of hyper-d iploid
aneu ploidies .
18
The chromosomal theory offers
testable explanations
Coulte r’s ar t ificia ll y a cti vat ed ge nes may
dest abi liz e t he kar yot ype of “converte d
human cel ls ”.
The ce ll s would bec ome r andomly
aneuploid.
19
The chromosomes of Coulter’s
“converted cells” are unstable
Metas
(28)
Karyotyp es of the Mu6 line
1
1
1
1
1
3
1
1
1
1
1
15
42,X,-Y,-17,-19,-20[1]
45,X,-Y[1]
45,XY,-18[1]
46,XY,-2,+del(2)x2,-5 [1]
46,XY,-14,+17,-18,+19x3,-20,-22[1]
47,XY,+20[3]
47,XY,+19[1]
47,XY,+(del(1)[1]
48,XXY,+7[1]
48,XY,+del(2)x2[1]
49,XY,+del(20)x3[1]
46,XY[15]
20
Conclusion: Half of cells of Mu6 line are randomly aneuploid.
Karyotype of an aneuploid Mu6 cell
Trisomy 7,
and a
marker
derived
from
Chrom. 1
21
Chromosomal theory explains low
transformation rates of
“converted” cells
1) Foci wit h t ra nsforma ti on- spec if ic a neuploidies would
evolve spont ane ously fr om r andom a neuploidies at ra tes of
5 per 100 ,000 cell s in vit r o.
2) Tr ansformat ion- spec if ic a neuploidies of foci would be
clona l or “non-ra ndom” – a landmark of cancer .
3) Some fo ci ma y be int rinsically drug r es ist ant o wing to
r esist ance-sp ec if ic a neu ploidies – anot her l andmark of
cancer .
22
By contrast the mutation theory
predicts
Foci of t r ansf orme d c ells a re –
1) eit her diploid o r r an domly an eu ploid – j ust like
t he mot he r cell ,
2) drug s ensit ive – j ust like t he Mu6 line.
No ex plana ti on f or lo w r at es of t r an sformat ion,
beca us e 6 g enes a r e sa id t o be “s uff icie nt ”.
23
Testing spontaneous appearance of
3-dimensional foci: about 1 per
100,000 Mu6 cells
Qu i c k T i m e ™ a n d a
T I F F (Un c o m p re s s e d ) d e c o m p re s s o r
a re n e e d e d t o s e e t h i s p i c tu re .
Mu6 focus
Mu6 monolayer
24
Clonal aneuploidies of 8 Mu6 foci:
some stable others evolving
Table 1. Karyotypes and intrinsic drug resistance of 8 transfor med foci generated by the human
muscle cell line Mu6.
Cell
Met.
Aneuploid clonal or sub-clonal
Aneuploid non-clonal
Mu6
28
47,XY,+20[3]
42,X,-Y,-17,-19,-20[1]
45,X,-Y[1]
45,XY,-18[1]
46,XY,-2,+del(2)x2,-5 [1]
46,XY,-14,+17,-18,+19x3,-20,-22[1]
47,XY,+19[1]
47,XY,+(del(1)[1]
48,XXY,+7[1]
48,XY,+del(2)x2[1]
49,XY,+del(20)x3[1]
F1
34
47,XY,+20[10]
46,XY,del(17)[1]
50,XXYY,+1,+5,-8,+12,-13,+14[4]
49, XXYY,+1,+5,-8,+12,-13,+14,-18[1]
49,XXYY,+1,-8,+12,-13,+t(14;15)[1]
50,XXYY,+1,-8,-10,+del(12)x2, +14,+t(14;15),-15[1]
50,XXY,+1,+5,+12,-13,+14[1] ->[8]
47,XY,+7[4]
46,XY,+7,-21[1]
48,XXY,+7[1] ->[6]
Dipl.
[15]
ArC
S
[9]
pS/R
25
F. 2
30
F. 3
20*
F. 4
20*
F. 5
20*
48,XXY,+7[13]
47,XY,+7[2]
47,XXY,+7,-21[1]
49,XXY,+7,+del(1)[1] ->[17]
50,XXYY,+1,+5,-8,+12,-13,+14[4]
45,XXYYY,+1,+5,-8,-13,-16,-18,-20,-21[1]
47,XXYY,+1,-3,+5,-8,-11,+12,-13[1]
47,XXYY,+1,+5,-8,+12,-13,-13,+14,-16,-19[1]
49,XXY,+1,+5,-8,+12,-13,+14[1] ->[8]
47,XY,+20[3]
47,XY,+20[6]
47,XY,ctb(10q),+20[1]
47,XY,ctb(4q),+20[1]
48,XY+6,+20,del(20p)[1]
48,XY,+8,+20[1]
48,XY,+7,+20[2]
50,XXY,+8,+12,+20[1] ->[13]
48,XXY,+7[2]
49,XY,+9,+11,+13[2]
48,XXY,+7[5]
48,XXY,+7,ctb(4p),+minx2[1]
49,XXY,+7,+21[2]
49,XXY,+7,+21,ctb(2p),ctb(14q)[1] ->[9]
47,XY,+20[2]
47,XY,+20,del(20)[1] ->[3]
47,XY,+20[1]
47,XY,+20,add(2q)[2]
49,XXY,+20,+21[1]
49,XXY,+20,+22[2]
48,XY,+1,+20,+minx3[1] ->[7]
48,XXY,+7[1]
48,XXY,+7,+dmin[1] ->[2]
[2]
pS/R
45,XY,-15[1]
[3]
S
44,X,-Y,-1[1]
[7]
S
45,X,-Y[1]
46,XY,+min[1]
[9]
S
26
F. 6
40*
F. 7
20*
F. 8
20*
46,XY,+min[1]
46,XY,+dmin[1] –> [2]
49,XXY,+1,+5,-8,+12,-13,+14[1]
50,XXYY,+1,+5,-8,+12,-13,+14[14]
50,XXYY,+1,+5,-8,+12,-13,+14,+min[1]
50,XXYY,+1,+5,-8,+12,13,+14, +min,+dmin[1]
100,XXXXYYYY,+1,+1,+5,+5,-8,-8,
+12,+12, -13,-13,+14,+14[1]
100,XXXXYYYY,+1,+1,+5,+5,-8,-8
+12,+12, -13,-13,+14,+14,+min[1] –> [19]
46,XY,+min,+dmin[1]
46,XY,+minx3[1]
46,XY,ctb(1+),ctb(4p),+dmin[1]
48,XY,+1,+dmin,minx2[2]
49,XXYY,+1,+5,+min,+dmin[1]
50,XXYY,+1,t(3;7),+5,+min,+dmin[1]
52,XXYY,+1,+5,-12,+14,+min,+dmin[1]
98,XXYY,ctb(8p),+minx2[1]
98,XXYY,+minx3[1]
100,XXYY,+minx2,+dminx2,ctb(1), ctb(20)[1]
102,XXYY,+dmin[1] ->[12]
49,XXY,+1,+5[2]
46,XY,+1,+5,-8,-13,+14,-19[5]
43,X,-Y,-3,+5,-8,-19,+min[1] ->[6]
45,XY,der(3)t(3;12)(q;p),
der(12)t(3;12)(q;p)[1]
[18]
R
46,XY,t(3;7)[1]
[5]
R
[8]
R
45,XY,-18[1]
45,XY,-12[1]
46,XY,+min[1]
47,XXY,+1,-11[1]
48,XXY,+7[1]
58,XXYY,?[1]
Abbreviations: F. means focus, Met. means metaphases, Dipl. means diploid, ArC means
cytosine arabinoside, S is sensitive, pS is partially sensitive, R is resistant, and pS/R is a partially
sensitive culture becoming resistant.
Bold [numbers] indicate the sums of identical or related metaphases per clonal aneuploidy.
*Marks karyotypes analyzed by Giemsa staining instead of by hybridization with color-coded,
chromosome-specific DNA.
27
Moreover, 3 of 8 foci are
intrinsically resistant to ara-C.
Mu6 line is ara-C- sensitive.
Back to Table.
28
In sum the 6 mutations were not
“sufficient”. Instead…
 Art if icia lly acti vat ed cel lular gen es de st ab ilize t he
kar yoty pe and ge ner at e r a ndom aneu ploidy – acti ng
like c ar cinogens.
 Specif ic, c lona lly sele ct ed aneu ploidies g ener at e 3 dimensiona l gr owt h, a lias foci in vitr o.
 Owing to t he hi gh kar yoty pic insta bil it y of Mu6 c ells ,
clona lly sele ct ed, aneu ploid st emline s conti nue to
evolve dur ing focus format ion – a s in na t ur al
car cinogene sis.
 Some t r ansf or ma ti on- spe cific aneu ploidies g ener at e
int rinsic, can cer - spec if ic dru g res ist ance – ag ain a s in
nat ur al c ancers .
29
So – Which one is the
cause?
I t would a ppear t hat ch r omosomal
al t er at ions are t he gene ti c b as is of
car cinogenes is.
Mut at ions would be conseq uen ces of
canc er- spe cific a neu ploidy, which
des t abilizes t ea ms of p r ot eins t hat
syn th es ize an d r ep air gene s.
30
The chromosomal theory of cancer can be
falsified by diploid cancers.
END
31
Spontaneous focus formation of
Mu6 clone 3
Transformation
rate:
~ 1 per
10^6 cells
32