The TP53 tumour suppressor gene in colorectal carcinomas. I. Genetic alterations on chromosome 17

Abstract Genetic alterations play a key role in the leukemogenesis of childhood ALL. Inactivation of CDKN2A (p16), a tumour suppressor gene located at 9p21, can occur by deletion, methylation or mutation. Published reports are inconsistent in terms of incidence and mode of inactivation. We report a comprehensive analysis of CDKN2A inactivation in 1230 diagnostic and 101 relapse samples, including 46 matched diagnostic and relapse pairs, from 1285 children with ALL. Using data from cytogenetics (CC) (n=1088), FISH (n=1209), SNP arrays (SNPA) (n=106), CGH arrays (aCGH) (n=106), dHPLC (n=48) and methylation specific-PCR (MSP) (n=96) we have assessed the mode and frequency of CDKN2A inactivation. Mutation or methylation of CDKN2A was rare occurring in 1 patient each (2% and 1% respectively). In contrast, CDKN2A deletion was highly prevalent. The frequencies of deletion detected by the different methodologies were: CC 166 (15%), FISH 335 (28%), SNPA 17 (16%) and aCGH 35 (33%). The proportion of biallelic deletions also varied: CC 15 (9%), FISH 174 (52%) and aCGH 15 (65%). This variation was directly related to the resolution of each technique with a high degree of concordance across samples investigated by >1 method. Analysis of 50 deletions by aCGH showed that the size of the deletion ranged from 0.03Mb to 39.1Mb with a mean of 14.8Mb. Furthermore, analysis of 15 biallelic deletions demonstrated that they comprised one large deletion (mean size 23.3Mb) and a second much smaller deletion (mean size 1.4Mb). In addition, SNPA revealed copy number neutral LOH in 8 (8%) cases, but only once in association with a CDKN2A mutation. At diagnosis CDKN2A inactivation by any method was noted in 329 (27%) patients which was not different from that observed at relapse [25 (25%)]. However, the frequencies of CDKN2A inactivation and biallelic deletion were significantly greater in T-ALL compared with B cell precursor (BCP) ALL: 135/269 (50%) v 190/918 (21%) (p<0.001) and 83/135 (61%) v 82/190 (43%) (p=0.001), respectively. Within BCP-ALL, older patients (10+ yrs) were more likely to have CDKN2A inactivation compared to younger patients (<10 yrs) whereas the reverse was true in T-ALL: 52/196 (27%) v 138/722 (19%) (p=0.023) and 53/126 (42%) v 82/143 (57%) (p=0.012). Among 46 matched samples CDKN2A inactivation was retained (n=8), lost (n=3) or gained (n=6). The frequency of CDKN2A inactivation was strongly correlated with cytogenetics. Lower frequencies were observed among high hyperdiploid [31/302 (10%) p<0.001] and ETV6-RUNX1 patients [36/236 (15%) p=0.02] with higher frequencies among those with t(9;22) [11/19 (58%) p<0.001], t(1;19) [10/25 (40%) p=0.019] and other abnormalities [92/226 (41%) p<0.001]. In conclusion, we have confirmed the importance of CDKN2A inactivation in childhood ALL and demonstrated that by far the most prevalent method of inactivation is deletion. While it is clear that loss of CDKN2A acts as a cooperating mutation in childhood ALL it is strongly correlated with age, phenotype and genotype. The observation that it is negatively correlated with good risk cytogenetic subgroups may explain why it has been inconsistently associated with a poor outcome. The discovery of copy number neutral LOH at 9p with no evidence of CDKN2A inactivation suggests the presence of another tumour suppressor gene or oncogene in this region.

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CTCF Expression is Essential for Somatic Cell Viability and Protection Against Cancer

International Journal of Molecular Sciences ◽

10.3390/ijms19123832 ◽

2018 ◽

Vol 19 (12) ◽

pp. 3832 ◽

Cited By ~ 5

Author(s):

Charles Bailey ◽

Cynthia Metierre ◽

Yue Feng ◽

Kinsha Baidya ◽

Galina Filippova ◽

...

Keyword(s):

Gene Expression ◽

Cell Cycle ◽

Somatic Cells ◽

Suppressor Gene ◽

Genetic Alterations ◽

Tumour Suppressor Gene ◽

Tumour Suppressor ◽

Tumour Suppressor Genes ◽

Absolute Requirement ◽

The Impact

CCCTC-binding factor (CTCF) is a conserved transcription factor that performs diverse roles in transcriptional regulation and chromatin architecture. Cancer genome sequencing reveals diverse acquired mutations in CTCF, which we have shown functions as a tumour suppressor gene. While CTCF is essential for embryonic development, little is known of its absolute requirement in somatic cells and the consequences of CTCF haploinsufficiency. We examined the consequences of CTCF depletion in immortalised human and mouse cells using shRNA knockdown and CRISPR/Cas9 genome editing as well as examined the growth and development of heterozygous Ctcf (Ctcf+/−) mice. We also analysed the impact of CTCF haploinsufficiency by examining gene expression changes in CTCF-altered endometrial carcinoma. Knockdown and CRISPR/Cas9-mediated editing of CTCF reduced the cellular growth and colony-forming ability of K562 cells. CTCF knockdown also induced cell cycle arrest and a pro-survival response to apoptotic insult. However, in p53 shRNA-immortalised Ctcf+/− MEFs we observed the opposite: increased cellular proliferation, colony formation, cell cycle progression, and decreased survival after apoptotic insult compared to wild-type MEFs. CRISPR/Cas9-mediated targeting in Ctcf+/− MEFs revealed a predominance of in-frame microdeletions in Ctcf in surviving clones, however protein expression could not be ablated. Examination of CTCF mutations in endometrial cancers showed locus-specific alterations in gene expression due to CTCF haploinsufficiency, in concert with downregulation of tumour suppressor genes and upregulation of estrogen-responsive genes. Depletion of CTCF expression imparts a dramatic negative effect on normal cell function. However, CTCF haploinsufficiency can have growth-promoting effects consistent with known cancer hallmarks in the presence of additional genetic hits. Our results confirm the absolute requirement for CTCF expression in somatic cells and provide definitive evidence of CTCF’s role as a haploinsufficient tumour suppressor gene. CTCF genetic alterations in endometrial cancer indicate that gene dysregulation is a likely consequence of CTCF loss, contributing to, but not solely driving cancer growth.

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Genetic variants of the tuberous sclerosis 2 tumour suppressor gene in mouse t haplotypes

Genetics Research ◽

10.1017/s0016672399003985 ◽

1999 ◽

Vol 74 (2) ◽

pp. 139-144 ◽

Cited By ~ 1

Author(s):

ELENA V. KLEYMENOVA ◽

JEFFREY E. DECLUE ◽

CHERYL L. WALKER

Keyword(s):

Tuberous Sclerosis ◽

Germ Line ◽

Suppressor Gene ◽

Tumour Suppressor Gene ◽

Mouse Embryos ◽

Tumour Suppressor ◽

Chromosome 17 ◽

Coding Region ◽

Lethal Mutations ◽

Germ Line Mutation

The murine t complex on chromosome 17 contains a number of homozygous lethal and semi-lethal mutations that disrupt development of the mouse embryo. We recently characterized an embryonic lethality in the rat that results from a germ-line mutation in the tuberous sclerosis 2 (Tsc-2) tumour suppressor gene (the Eker mutation). Remarkably, mouse embryos homozygous for tw8 mutation display cranial defects reminiscent of those observed in rat embryos homozygous for the Eker mutation. To determine whether the Tsc-2 gene, which is in the t complex, is mutated in tw8 or other t haplotypes, we characterized this gene in a series of t haplotype mice. Four Tsc-2 polymorphisms were identified: three in the coding region and one intronic that appeared to be common to all t haplotypes analysed. No evidence was found to argue that the Tsc-2 gene is altered in tw8 haplotype mice. However, in the tw5 haplotype we found a G to T mutation in Tsc-2 that was present only in this t haplotype. In contrast to other polymorphisms within the Tsc-2 coding region which did not result in amino acid changes in Tsc-2 gene product tuberin, this mutation substituted a phenylalanine for a conserved cysteine in tw5 tuberin. Within the t complex, the Tsc-2 gene and the putative tw5 locus appeared to map to different positions, complicating identification of Tsc-2 as a candidate for the tw5 locus and suggesting that the G to T mutation in the Tsc-2 gene may have arisen independently of the tw5 functional mutation.

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The TP53 tumour suppressor gene in colorectal carcinomas. II. Relation to DNA ploidy pattern and clinicopathological variables

British Journal of Cancer ◽

10.1038/bjc.1993.15 ◽

1993 ◽

Vol 67 (1) ◽

pp. 93-98 ◽

Cited By ~ 23

Author(s):

GI Meling ◽

RA Lothe ◽

AL Børresen ◽

C Graue ◽

S Hauge ◽

...

Keyword(s):

Dna Ploidy ◽

Suppressor Gene ◽

Tumour Suppressor Gene ◽

Tumour Suppressor ◽

Colorectal Carcinomas ◽

Dna Ploidy Pattern ◽

Ploidy Pattern ◽

Clinicopathological Variables

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CTCF Expression Is Essential for Somatic Cell Viability and Protection against Cancer

10.20944/preprints201810.0451.v1 ◽

2018 ◽

Author(s):

Charles G. Bailey ◽

Cynthia Metierre ◽

Julie Feng ◽

Kinsha Baidya ◽

Galina N. Filippova ◽

...

Keyword(s):

Gene Expression ◽

Cell Cycle ◽

Somatic Cells ◽

Suppressor Gene ◽

Genetic Alterations ◽

Tumour Suppressor Gene ◽

Tumour Suppressor ◽

Tumour Suppressor Genes ◽

Absolute Requirement ◽

The Impact

CCCTC-binding factor (CTCF) is a conserved transcription factor that performs diverse roles in transcriptional regulation and chromatin architecture. Cancer genome sequencing reveals diverse acquired mutations in CTCF, which we have shown, functions as a tumour suppressor gene. While CTCF is essential for embryonic development, little is known of its absolute requirement in somatic cells and the consequences of CTCF haploinsufficiency. We examined the consequences of CTCF depletion in immortalised human and mouse cells using shRNA knockdown and CRISPR/Cas9 genome editing and examined the growth and development of heterozygous Ctcf (Ctcf+/-) mice. We also analysed the impact of CTCF haploinsufficiency by examining gene expression changes in CTCF-altered endometrial carcinoma. Knockdown and CRISPR/Cas9-mediated editing of CTCF reduced the cellular growth and colony-forming ability of K562 cells. CTCF knockdown also induced cell cycle arrest and a pro-survival response to apoptotic insult. However, in p53 shRNA-immortalised Ctcf+/- MEFs we observed the opposite: increased cellular proliferation, colony formation, cell cycle progression and decreased survival after apoptotic insult compared to wild type MEFs. CRISPR/Cas9-mediated targeting in Ctcf+/- MEFs revealed a predominance of in-frame microdeletions in Ctcf in surviving clones, however protein expression could not be ablated. Examination of CTCF mutations in endometrial cancers showed locus-specific alterations in gene expression due to CTCF haploinsufficiency, in concert with downregulation of tumour suppressor genes and upregulation of estrogen-responsive genes. Depletion of CTCF expression imparts a dramatic negative effect on normal cell function. However, CTCF haploinsufficiency can have growth-promoting effects consistent with known cancer hallmarks in the presence of additional genetic hits. Our results confirm the absolute requirement for CTCF expression in somatic cells and provide definitive evidence of CTCF’s role as a haploinsufficient tumour suppressor gene. CTCF genetic alterations in endometrial cancer indicate that gene dysregulation is a likely consequence of CTCF loss, contributing to, but not solely driving cancer growth.

Download Full-text