Protective role of cytoplasmic p21Cip1/Waf1 in apoptosis of CDK4/6 inhibitor‐induced senescence in breast cancer cells

Abstract

Inhibition of CDK4/6 slows the cell cycle and induces senescence in breast cancer cells. However, senescent cancer cells promote invasion and metastasis. Several drugs reportedly target senescent cells, including . We examined the effects of the CDK4/6 inhibitor abemaciclib and ABT-263 on two human breast cancer cell lines. The abemaciclib and ABT-263 combination additively decreased the viability of MDA-MB-231 cells, but not MCF-7 cells. Also, the combination therapy-induced caspase-dependent apoptosis in MDA-MB-231 cells. Combination therapy with abemaciclib and ABT-737, an ABT-263 analog, significantly suppressed the in vivo growth of MDA-MB-231 with transient bodyweight loss. Given that p16 Ink4a and p21 Cip1/Waf1 are key factors in senescence and that both cell lines were negative for p16, the role of p21 in apoptosis of treated breast cancer cells was investigated. Although abemaciclib increased the cytoplasmic p21 level in both cell lines as a hallmark of senescence, the abemaciclib and ABT-263 combination decreased it only in MDA-MB-231 cells. This decrease of p21 expression was relieved by caspase inhibition, and p21 was colocalized with caspase-3 in the cytoplasm of MDA-MB-231 cells. Alternatively, small interfering RNA-mediated knockdown of p21 rendered caspase-3-negative MCF-7 cells susceptible to abemaciclib and ABT-263, as well as TNF-related apoptosisinducing ligand. Furthermore, a clinical database analysis showed that p21 high breast cancer patients had a poorer prognosis compared to p21 low patients. These results suggest that cytoplasmic p21 plays a protective role in apoptosis of CDK4/6 inhibitor-induced senescent breast cancer cells.

Protective roles of cytoplasmic p21Cip1/Waf1 in senolysis and ferroptosis of lung cancer cells

“…Given that TIS cells increase cytoplasmic p21 expression, p21 may have inhibitory effects on senolysis. Likewise, we previously reported the protective role of cytoplasmic p21 in the apoptosis of CDK4/6 inhibitor‐induced senescent breast cancer cells 17 . In this study, we utilized PEM, an anti‐folate drug, which has been widely used for the treatment of NSCLC patients 27,41 and can induce senescence in NSCLC cell lines, including A549 cells 24,25 .…”

“…Immunoblot was performed, as previously reported 17 . The following primary antibodies were used: anti‐p21 Cip1/Waf1 (#2947; Cell Signaling Technology), anti‐p16 Ink4a (SPC‐1280; StressMarq Biosciences), anti‐p53 (p8999; Sigma‐Aldrich), anti‐phospho‐p53 (#9284; Cell Signaling Technology) anti‐glutathione peroxidase 4 (GPx4) antibody (Cayman), anti‐TATA‐binding protein (TBP; #22006‐I‐AP; Proteintech), anti‐GAPDH (#015‐25473; Fujifilm Wako Pure Chemical) and anti‐β actin (#622102; BioLegend).…”

“…Cell viability was measured using the Cell Counting Kit‐8 (CCK‐8) (Dojindo Molecular Technologies), as previously reported 17 …”

“…Despite its role in promoting apoptosis, p21 prevents apoptosis in cancer cells in response to anticancer therapies 15,16 . Previously, we reported the anti‐apoptotic role of cytoplasmic p21 in CDK4/6 inhibitor‐induced senescent breast cancer cells 17 …”

A review on the role of cyclin dependent kinases in cancers

“…The combination teraphy was found to be more effective than single treatments also under hypoxia [266] circ_0001588/miR-874/CDK4 axis SK-Hep- Female BALB nude mice abemaciclib and ABT-263 combination: ↓ tumor growgh [218] 4-week-old BALB/c nude mice ↑↑ miR-124: ↓ tumor growth [219] 4-week-old nude mice ∆ MALAT1: ↑ inhibitory effect of miR-124 on the tumor growth [222] Cervical cancer 4-5-week-old male BALB/c nude mice ∆ circ_0000326: ↓ tumor growth [227] Clear cell renal cell carcinoma 4-5-week-old male BALB/c nude mice ↑↑ miR-1: ↓ tumor growth [228] NOD/SCID/IL2Rγ-null (NSG) mice ∆ DMDRMR: ↓ tumor growth [229] 4-5-week-old male BALB/c nude mice ↑↑ miR-206: ↓ tumor size and weigh [230] Colon cancer Male athymic BALB/c nude mice ∆ HAGLR: ↓ tumor growth [231] 6-week-old female Balb/c nude mice ↑↑ miRNA-20b-5p: ↓ tumor growth [232] Colorectal cancer 6-week-old BALB/c athymic nude mice ↑↑ MCM3AP-AS1: ↑ tumor growth [233] 5-6-week-old male BALB/c nude mice ↑↑ miR-142-3p: ↓ tumor growth [234] 4-6-week-old male BALB/c athymic nude mice ↑↑ miR-875-5p: ↓ tumor growth [236] Esophageal squamous cell carcinoma 4-week-old male BALB/c nude mice ∆ hsa_circ_0016788: ↓ tumor growth [268] 6-week male Bl6/Rag2/GammaC double knockout nude mice ∆ CCDC144NL-AS1/WDR5 or ↑↑ miR-940: ↓ tumor growth [270] 4-week-old female BALB/c nude mice ∆ circSP3: ↓ tumor volume and weight [273] BALB/c nude mice ∆ VPS9D1-AS1: ↓ tumor growth [24] Kaposi's sarcoma-associated herpesvirus 4-6-week-old female BALB/c nude mice ↑↑ miR-34a-5p: ↓ tumor volume and weight [274] Lung cancer female athymic BALB/c nude mice ∆ LINC01194: ↓ tumor volume and weight [278] 6-week-old male BALB/c nude mice ↑↑ hsa_circ_0014235: ↑ DDP chemoresistance [279] 5-6-week-old male BALB/c nude mice ↑↑ miR-613: ↓ tumor growth [280] 4-week-old female BALB/c nude mice ↑↑ miR-340: ↓ tumor growth [283] 4-6-week-old male BALB/c athymic nude mice ↑↑ miR-326: ↑ tumor volume and weight [285] 4-6-week-old male BALB/c athymic nude mice ↑↑ miR-134: ↓ tumor growth [285] male athymic BALB/c nude mice ∆ lncSENCR: ↓ tumor growth [291] 5…”