Lrig1 / 5’-GGGGACAGAAAGGAGGAAGT-3’
5’-TAAACGACGAAGACTCTCCCG-3’ / [1]
Sox2 / 5’-CGAGATAAACATGGCAATCAAATG-3’
5’-CACCAGAACAAATTCCGTTTGCAA-3’ / [2]
HPV16E6 / 5’-GTTACTGCGACGTGAGGTATATG-3’
5’-CATTTATCACATACAGCATATGGATTC-3’ / [3]
HPV16E7 / 5’-CCGGACAGAGCCCATTACAAT-3’
5’-ACGTGTGTGCTTTGTACGCAC-3’ / [4]
Notch1 / 5’-CACTGTGGGCGGGTCC-3’
5’-GTTGTATTGGTTCGGCACCAT-3’ / [5]
Hes-1 / 5’-TGGAAATGACAGTGAAGCACCT-3’
5’-GTTCATGCACTCGCTGAAGC-3’ / [6]
Nanog / 5’-CGATCTCCTGACCTTGT-3’
5’-CACGCCTGTAAATCCCA-3’ / [7]
ABCG2 / 5’-GCAGATGCCTTCTTCGTTATG-3’
5’- TCTTCGCCAGTACATGTTGC-3’ / [8]
CD133 / 5’-ACATGAAAAGACCTGGGGG-3’
5’-GATCTGGTGTCCCAGCATG-3’ / [9]
Supplementary Table 1: List of real-time RT-PCR primers used in the study
Supplementary Table2: Result of cervical cancer cell line or primary culture derived xenografting in nude mice
Cell Types* / Cell no. implanted / No. of Mice / Mice with Tumor**Parental (Low dose) / 20x103 / 6 / 0
Parental (High dose) / 0.5x106 / 6 / 3
Non-CaCxSLC
(NSP→CD49f-veCD71+ve→CD133-ve) / 20x103 / 6 / 0
CaCxSLC
(SP→CD49f+veCD71-ve→CD133+ve) / 20x103 / 6 / 5
pCaCxSLC
(SP→CD49f+veCD71-ve→CD133+ve) / 0.1x106 / 6 / 4
*SP - Side population, NSP - Non-side population, CaCxSLC – Cervical cancer stem-like cells, Non-CaCxSLC – Non-stem cervical cancer cells, pCaCxSLC – Primary tumor derived cervical cancer stem-like cells.
**Tumor formation after 4-weeks post injection
Supplementary Table 3: Physical state of HPV16 DNA in CINI/II, CINIII/IV cervical cancer lesionsand primary xenograft
Lesion grade / HPV16 physical stateEpisomal
(E2:E6 = 1.0) / Mixed
(0<E2:E6<1.0) / Integrated
(E2:E6 = 0)
CINI/II (n=5) / 4/5(80%) / 1/5(20%) / 0
CINIII/IV (n=10) / 1/10(10%) / 2/10(20%) / 7/10 (70%)
Primary Xenograft (n=4)a, b / 0 / 1/4(25%) / 3/4(75%)
P<0.005, CINI/II vs CINIII/IV, aCINI/II vs Primary xenograft, bCINIII/IVvs Primary xenograft; CIN – Cervical intraepithelial neoplasia
Supplementary References:
1.Jensen KB, Watt FM: Single-cell expression profiling of human epidermal stem and transit-amplifying cells: Lrig1 is a regulator of stem cell quiescence. Proc Natl Acad Sci U S A 2006, 103:11958-11963.
2.Ji J, Werbowetski-Ogilvie TE, Zhong B, Hong SH, Bhatia M: Pluripotent transcription factors possess distinct roles in normal versus transformed human stem cells. PLoS One 2009, 4:e8065.
3.de Boer MA, Jordanova ES, Kenter GG, Peters AA, Corver WE, Trimbos JB, Fleuren GJ: High human papillomavirus oncogene mRNA expression and not viral DNA load is associated with poor prognosis in cervical cancer patients. Clin Cancer Res 2007, 13:132-138.
4.Darnell GA, Schroder WA, Antalis TM, Lambley E, Major L, Gardner J, Birrell G, Cid-Arregui A, Suhrbier A: Human papillomavirus E7 requires the protease calpain to degrade the retinoblastoma protein. J Biol Chem 2007, 282:37492-37500.
5.Martin CH, Woll PS, Ni Z, Zuniga-Pflucker JC, Kaufman DS: Differences in lymphocyte developmental potential between human embryonic stem cell and umbilical cord blood-derived hematopoietic progenitor cells. Blood 2008, 112:2730-2737.
6.Li YP, Paczesny S, Lauret E, Poirault S, Bordigoni P, Mekhloufi F, Hequet O, Bertrand Y, Ou-Yang JP, Stoltz JF, et al: Human mesenchymal stem cells license adult CD34+ hemopoietic progenitor cells to differentiate into regulatory dendritic cells through activation of the Notch pathway. J Immunol 2008, 180:1598-1608.
7.Chiou SH, Wang ML, Chou YT, Chen CJ, Hong CF, Hsieh WJ, Chang HT, Chen YS, Lin TW, Hsu HS, Wu CW: Coexpression of Oct4 and Nanog enhances malignancy in lung adenocarcinoma by inducing cancer stem cell-like properties and epithelial-mesenchymal transdifferentiation. Cancer Res 2010, 70:10433-10444.
8.Saxena M, Stephens MA, Pathak H, Rangarajan A: Transcription factors that mediate epithelial-mesenchymal transition lead to multidrug resistance by upregulating ABC transporters. Cell Death Dis 2011, 2:e179.
9.Clement V, Marino D, Cudalbu C, Hamou MF, Mlynarik V, de Tribolet N, Dietrich PY, Gruetter R, Hegi ME, Radovanovic I: Marker-independent identification of glioma-initiating cells. Nat Methods 2010, 7:224-228.