Hypoxia-inducible factors are required for
chemotherapy resistance of breast cancer stem cell
Proceedings of the National Academy of Sciences of
the united states of American (PNAS)
IF=9.8
PNAS | Published online December 1, 2014 | E5429–E5438
Background
Possible origins of cancer stem cells
肿瘤的发生是一个多基因多步骤的基因变化效应，是多阶段或多次打击后产生的。正常
细胞转化为恶性肿瘤细胞需经历一个复杂而漫长的过程，至少需要4-7次突变，这需要很长的
时间。而与已分纯细胞相比，干细胞在体内分布广泛且更新能力强，更易发生恶性转化，这
意味着突变可能更容易在干细胞中得以积累。
BCSC and Breast Cancer
1. Although many breast cancer cells enter the circulation, only BCSCs are capable of
forming a secondary tumor
2. BCSCs are also resistant to chemotherapy and the percentage of BCSCs following
chemotherapy is increased compared with before therapy
3. A reduction in cancer cell burden of >95% may result in an apparent complete
response to therapy, but may leave a population of residual BCSCs that represent the
source of subsequent disease recurrence that will eventually lead to death of the
patient.
Breast cancer therapy is based on the classification of
tumors into three groups
A: estrogen receptor/progesterone receptor [ER/PR]+ cancers (60-70%)
ER antagonists, such as tamoxifen, or aromatase inhibitors, such as letrozole
B: human epidermal growth factor receptor 2 (HER2+) cancers (15-20%)
anti-HER2 antibody, such as trastuzumab, or receptor tyrosine kinase inhibitor,
such as lapatinib
C: Triple negative breast cancers (TNBCs), which do not express ER, PR, or HER2(20%)
Cytotoxic chemotherapy, such as paclitaxel or gemcitabine
HIF and Drug resistance
Microarray data from more than 500 human breast cancers revealed that one of the
defining features of the basal molecular subtype is the increased expression of a large
battery of genes that are regulated by hypoxia-inducible factors (HIFs)
HIF-1 has been implicated in resistance to chemotherapy and in hypoxic colon cancer
cells, HIF-1 activated expression of multidrug resistance 1 (MDR1), which mediates
efflux of chemotherapy from cancer cells and represents a major mechanism
underlying treatment failure.
Increased expression of hypoxia-induced genes in breast cancer is also associated with
poor prognosis.
Methods
1. Cell Culture(MDA-MB-231+SUM-159+SUM-149 )
2. RT –qPCR
3. Immunoblot assays
4. Luciferase Assay—HIF transcriptional activity
5. Aldefluor Assay--BCSC identification
6. Mammosphere Assay
7. MitoSox Staining--ROS
Results
1. Paclitaxel Induces HIF Expression
and Transcriptional Activity in TNBC
Cells
2. Paclitaxel Treatment Increases the Percentage of BCSCs.
3. Paclitaxel-Induced
Interleukin Expression
Is Blocked by HIF
Inhibitors
4. HIF-1 α and HIF-2 α
Are Required for
Paclitaxel-Induced BCSC
nrichment
Paclitaxel-Induced SMAD2 and STAT3 Activity Is Insufficient to Induce BCSC Enrichment
TLR-4 and NF- κ B Do Not
Mediate ChemotherapyInduced BCSC Enrichment.
5. Increased ROS
Levels Mediate
Induction of HIF-1 α
and Enrichment of
BCSCs
6. Paclitaxel-Induced
MDR1 Expression Is
Blocked by HIF
Inhibitors
7. Digoxin Blocks PaclitaxelInduced BCSC Enrichment
in Vivo
8. HIF Signature Predicts Mortality of Patients Treated with Chemotherapy
DISCUSSION
We have demonstrated
that treatment of three
different TNBC cell lines
with clinically relevant
concentrations of paclitaxel
or gemcitabine increases
ROS levels, which induce
HIF-1 α and HIF-2 α mRNA
and protein expression,
leading to the HIFdependent expression of
MDR1, IL-8, and/or IL-6, and
to BCSC enrichment, both in
vitro and in vivo