1. Limitless replicative
Self-sufficiency
in growth signals
Sustained
angiogenesis
Insensitivity to
anti-growth signals
Limitless replicative
potential
Evading apoptosis
Tissue invasion
& metastasis
Cancer
The development of human cancer cells

2. Cdc6 is essential in the initiation of DNA replication:
A). Assembling pre-replication complex (pre-RC) with ORC,
and loading MCMs to replication origins;
B). Licensing DNA replication to ensure one round
DNA replication per cell cycle.

3. the specific genes for cell proliferation:
The novel function of GLS101/TSUKUSI FL1 is the silencing of human Cdc6,
the specific genes for cell proliferation:
Specific Cdc6 shRNAs produced from the viral vector Inhibit Cdc6 and block
Cancer cell proliferation, but not normal cells.
Cdc6
THTD6Ai
THTP
CHLA255
LA-N-2
Mock
MCF7
-actin
Cdc6 knockdown
Cdc6 shRNA

4. Apoptosis Differentiation Senescence The Cell Cycle M G1 Recombinant
Lentivirus
GLS101 G1 S G2 M
STOP
Senescence
The Cell Cycle

5. Reactivation of p16INK4a CDK inhibitory activity by Cdc6 knockdown
Phosph-Rb-C Ser780
Phosph-Rb-C Ser807/811
IgG, light chain
—    IP/β-p16INK4a
— —   THTD
 — — — IP/mouse IgG
Phosph Rb-C Ser 807/811
1:5
1:10
1:25
 IP/β-p16INK4a
— — —  THTD

6. Senescence-associated β-glactosidase (SA-β-Gla)
positive staining caused by Cdc6 knockdown
MCF7
MCF7/THTP
MCF7/THTD6Ai

7. The formation of senescence associated heterochromatic foci via
Cdc6 knockdown
Anti-H3K9me3/me2
indirect fluorescent
staining
THTD6Ai
Mock
Confocal fluorescent
microscopy