Table 1 Prior studies on CSCs of ccRCC

1

MACS

Human RCC nephrectomy sample (no clinical data provided)

CD105⁺

8.06 ± 3.3%

Nestin, NANOG, Musashi, OCT4, PAX2, CD29, CD44, CD73, CD90, CD146, Vimentin

Pan-CK, CD24, CD133

Clonogenic and sphere-forming ability; epithelial and endothelial differentiation in vitro; high tumorigenicity

Bussolati et. al. [66]

2

FACS

Human metastatic ccRCC biopsies (all VHL⁻) and patient-derived xenografts both VHL⁻ and VHL⁺)

CD133⁺/

CXCR4⁺

84.5% ± 2.5% in peri-necrotic area

Sphere formation; tumorigenicity; chemotherapy resistance (sunitinib)

• Varna et al. [69]

3

FACS

Human ccRCC biopsies (selected surface marker analysis of 55 patient samples, mainly pT1 and pT3; male:female = 1.8:1)

CXCR4⁺/

MET⁺/

CD44⁺

2.2%

VCAM1, E-Cadherin, KLF4, SOX9, PAX2, SALL1, PROM1 (CD133), ALDH1A1, MYST3

NOTCH and WNT signaling-dependent sphere- and organoid-formation; tumorigenicity

Fendler et al. [71]

4

MACS/

FACS

Normal and malignant biopsies from 40 ccRCC patients; pT1-pT3, male:female = 2.6:1

CD133⁺/

CD24⁺

85%

CTR2, NANOG, SOX2, CD73, SNAI1, VEGFR2, GATA-4, CD73

CD90, CD105, CD20, CD45, CD14, CD34

Self-renewal; multipotency in vitro; colony-formation in soft agar; angiogenic induction;

CD133⁺/

CD24⁺ tumor cells less differentiated than normal progenitors

• Galleggiante et al. [67]

5

FACS

Caki-1

CD105⁺; CD44⁺; CD44⁻; CD44⁻/CD105⁺; CD105⁻/CD44⁻

10.8% CD105⁺,

1.68% CD133⁺,

94.6% CD44⁺

Multiple clones expressing combinations of positive or negative CD105 and CD44 markers

Tumorigenic potency (CD105⁻/CD44⁻ cells, not CD105⁺/

CD44⁺ or single CD105⁻cells); vasculogenic promotion

Fiedorowicz et al. [72]

6

MACS

ACHN,

Caki-1

CD133⁺/

CD24⁺

ACHN, 95.8%;

Caki-1, 95.5%

CTR2, BCL-2, MDR1, OCT-4, KLF4, NOTCH1, NOTCH2, JAG1, JAG2, DLL1, DLL4

NOTCH signaling-dependent self-renewal; chemotherapy resistance (cisplatin, sorafenib); tumorigenesis in vivo

Xiao et al. [70]

7

FACS

RCC-26, RCC-53

CXCR4⁺

RCC-26, 0.8%;

RCC-53, 5%

OCT3/4, SOX2, NANOG, CD24, CD29, CD44, CD73, CD117, CD146, CXCR4

CXCR1, CD34, CD90, CD105, CD133

High ALDH1 activity; sphere-forming; tumorigenicity; RCC-53 cells, 100% incidence; resistance to RTK inhibitors

• Gassenmaier et al. [68]

8

Side population

Matched normal and malignant biopsies from ccRCC patients

Hoechst⁻

Normal kidney, 3.8 ± 0.4;

RCC, 5.9 ± 0.9%

β-catenin, NOTCH1, SHH, CD133, PAX2

Highly proliferative; retention of SP upon culture; sphere formation in 3D Matrigel

Addla et al. [81]

9

Sphere formation assay, side population

SK-RC-42

9.9 ± 0.14% (SP)

OCT3/4, BMI1, β-catenin, NANOG

MHC-II, CD80, CD86

Sphere-formation; chemo-and radiotherapy-resistance (5-FU, MMC and 2 Gy); tumorigenicity

Zhong et al. [85]

10

Side population

769-P,

786-O,

OS-RC-2, SN12C, SKRC39

Hoechst⁻

769-P, 4.82% (SP)

ABCB1

ABCG2, ABCC1

Self-renewal and proliferative potential; chemo-and radiotherapy-resistance (5-FU, MTX, 5 Gy); high tumorigenicity

Huang et al. [86]

11

Side population

ACHN, Caki1, SMKTR2, SMKTR3, RenCa

Hoechst⁻

ACHN, 2.6%; RenCa, 18% (SP)

DNAJB8

Sox2

Oct4 (Pou5f1)

Tumorigenicity

Nishizawa et al. [87]

12

Side population

ACHN, KRC/Y

ACHN, 1.4%; KRC/Y, 1.7%

ALDH1

CD105, CD133

Sphere formation, self-renewal, tumorigenicity (in ACHN but not KRC/Y)

Ueda et al. [88]

13

Sphere formation assay

ACHN,

Caki-1

OCT4, NANOG, LIN28, KLF4, CD24, CD44, ZEB1, ZEB2, TWIST1, N-cadherin, Vimentin

TGF-β-dependent sphere formation; self-renewal; tumorigenicity

Lichner et al. [90]

14

Sphere formation

Caki-1,

786-O,

(Caki-2,

769-P)

CXCR4,

SDF-1, NANOG

Sphere formation; adipogenic differentiation; tumorigenicity

(Caki-2, 769-P had none of the above)

Micucci et al. [91]