Lack of intratumoral heterogeneity in DNA ploidy pattern of hepatocellular carcinoma

doi:10.1016/0016-5085(93)90150-B

Gastroenterology

Volume 105, Issue 5, November 1993, Pages 1449-1454

https://doi.org/10.1016/0016-5085(93)90150-B Get rights and content

Abstract

$Background$ : From biological and clinical perspectives, it is important to clarify tumor heterogeneity. This study was aimed to investigate whether or not intratumoral heterogeneity of DNA ploidy pattern exists in hepatocellular carcinoma (HCC). $Methods$ : Using fresh materials resected from 31 untreated patients, DNA ploidy was analyzed at different sites of the same HCC by means of flow cytometry. The tumor size ranged from 1.7 to 25.0 cm. $Results$ : There was no case in which euploid and aneuploid HCCs coexisted in the same tumor. The DNA ploidy pattern was euploid in 15 and aneuploid in 16 instances. Of 15 euploid tumors, the areas analyzed were all diploid in 12 and tetraploid in 2 but diploid/tetraploid in 1. Among 16 aneuploid tumors, the DNA indices (DI) at different sites were similar in 9, but apparently, different aneuploid subclones coexisted in 7 cases. The incidence of DI heterogeneity in aneuploid HCCs was similar between small (<5 cm) and large (≧5 cm) tumors; 3 of 7 (42.9%) versus 4 of 9 (44.4%). $Conclusions$ : It is assumed that euploid and aneuploid HCCs develop in their own ploidy pattern and that the evolution of aneuploid subpopulations from euploid HCC is rare, but new aneuploid subclones can evolve from aneuploid HCC due to increased instability of its karyotype.

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Cited by (31)

Mice With Increased Numbers of Polyploid Hepatocytes Maintain Regenerative Capacity But Develop Fewer Hepatocellular Carcinomas Following Chronic Liver Injury
2020, Gastroenterology
Citation Excerpt :
However, we cannot assert that polyploid cells never transform or become cancer. The increased likelihood that HCCs have a diploid origin is supported by the fact that most human HCCs are diploid.15,46–49 In our experiments, super-polyploid livers are protected from both DEN + CCl4 and CCl4-induced liver cancers.
Thirty to 90% of hepatocytes contain whole-genome duplications, but little is known about the fates or functions of these polyploid cells or how they affect development of liver disease. We investigated the effects of continuous proliferative pressure, observed in chronically damaged liver tissues, on polyploid cells.
We studied Rosa-rtTa mice (controls) and Rosa-rtTa;TRE-short hairpin RNA mice, which have reversible knockdown of anillin, actin binding protein (ANLN). Transient administration of doxycycline increases the frequency and degree of hepatocyte polyploidy without permanently altering levels of ANLN. Mice were then given diethylnitrosamine and carbon tetrachloride (CCl₄) to induce mutations, chronic liver damage, and carcinogenesis. We performed partial hepatectomies to test liver regeneration and then RNA-sequencing to identify changes in gene expression. Lineage tracing was used to rule out repopulation from non-hepatocyte sources. We imaged dividing hepatocytes to estimate the frequency of mitotic errors during regeneration. We also performed whole-exome sequencing of 54 liver nodules from patients with cirrhosis to quantify aneuploidy, a possible outcome of polyploid cell divisions.
Liver tissues from control mice given CCl₄ had significant increases in ploidy compared with livers from uninjured mice. Mice with knockdown of ANLN had hepatocyte ploidy above physiologic levels and developed significantly fewer liver tumors after administration of diethylnitrosamine and CCl₄ compared with control mice. Increased hepatocyte polyploidy was not associated with altered regenerative capacity or tissue fitness, changes in gene expression, or more mitotic errors. Based on lineage-tracing experiments, non-hepatocytes did not contribute to liver regeneration in mice with increased polyploidy. Despite an equivalent rate of mitosis in hepatocytes of differing ploidies, we found no lagging chromosomes or micronuclei in mitotic polyploid cells. In nodules of human cirrhotic liver tissue, there was no evidence of chromosome-level copy number variations.
Mice with increased polyploid hepatocytes develop fewer liver tumors following chronic liver damage. Remarkably, polyploid hepatocytes maintain the ability to regenerate liver tissues during chronic damage without generating mitotic errors, and aneuploidy is not commonly observed in cirrhotic livers. Strategies to increase numbers of polypoid hepatocytes might be effective in preventing liver cancer.
Surviving Acute Organ Failure: Cell Polyploidization and Progenitor Proliferation
2019, Trends in Molecular Medicine
Citation Excerpt :
Solid organs utilizing progenitor-driven repair can develop cancer, for example, the liver and possibly the renal tubules. Indeed, in mutagen- and high-fat-induced mouse models, tumorigenesis was accelerated by proliferation of diploid hepatocytes [92], and accordingly, human data show that, for example, hepatocarcinomas are predominantly diploid [93–95]. Consistently, in mouse models of a stable polyploidy knockout, hepatocyte progenitors proliferate faster than in wild-type mice and become highly tumorigenic when challenged with tumor-promoting stimuli, suggesting that tumors are driven by rapid progenitor proliferation [20].
In acute organ failure, rapid compensation of function loss assures survival. Dedifferentiation and/or proliferation of surviving parenchymal cells could imply a transient (and potentially fatal) impairment of residual functional performance. However, evolution has selected two flexible life-saving mechanisms acting synergistically on organ function recovery. Sustaining residual performance is possible when the remnant differentiated parenchymal cells avoid cell division, but increase function by undergoing hypertrophy via endoreplication, leading to polyploid cells. In addition, tissue progenitors, representing a subset of less-differentiated and/or self-renewing parenchymal cells completing cytokinesis, proliferate and differentiate to regenerate lost parenchymal cells. Here, we review the evolving evidence on polyploidization and progenitor-driven regeneration in acute liver, heart, and kidney failure with evolutionary advantages and trade-offs in organ repair.
The Polyploid State Plays a Tumor-Suppressive Role in the Liver
2018, Developmental Cell
Citation Excerpt :
Consistent with this, DEN tumors were most often diploid, suggesting a diploid origin. This is consistent with human data showing that HCCs are predominantly diploid (Anti et al., 1994; Fujimoto et al., 1991; Nagasue et al., 1993). We also showed that MYC-induced oncogenesis in the liver was agnostic to ploidy states.
Most cells in the liver are polyploid, but the functional role of polyploidy is unknown. Polyploidization occurs through cytokinesis failure and endoreduplication around the time of weaning. To interrogate polyploidy while avoiding irreversible manipulations of essential cell-cycle genes, we developed orthogonal mouse models to transiently and potently alter liver ploidy. Premature weaning, as well as knockdown of E2f8 or Anln, allowed us to toggle between diploid and polyploid states. While there was no detectable impact of ploidy alterations on liver function, metabolism, or regeneration, mice with more polyploid hepatocytes suppressed tumorigenesis and mice with more diploid hepatocytes accelerated tumorigenesis in mutagen- and high-fat-induced models. Mechanistically, the diploid state was more susceptible to Cas9-mediated tumor-suppressor loss but was similarly susceptible to MYC oncogene activation, indicating that polyploidy differentially protected the liver from distinct genomic aberrations. This suggests that polyploidy evolved in part to prevent malignant outcomes of liver injury.
Intratumoral genomic heterogeneity in human hepatocellular carcinoma detected by restriction landmark genomic scanning
2000, Journal of Hepatology
Background/Aims: One of the unique features of advanced hepatocellular carcinoma (HCC) is the morphological heterogeneity in a single tumor nodule. In order to investigate the intratumoral genomic heterogeneity of HCC, we performed Restriction Landmark Genomic Scanning (RLGS), which allows genomic DNAs to be surveyed at approximately 2000 landmark sites in a single 2-dimensional gel electrophoresis.
Methods: RLGS profiles of two regions from a single HCC nodule in six patients were compared with nontumorous liver tissue. Four HCCs consisting of moderately-differentiated cells were separated into several small parts by thin fibrous septa, but not encapsulated. DNA samples were obtained from both parts of these so-called “nodule-in-nodule” HCC. Two HCCs consisting of well-differentiated cells which did not have a definite partition appeared pathologically homogeneous, and two independent regions of the HCC were used for the analysis.
Results: All six HCCs demonstrated different RLGS profiles (in total about 160 different spots) from the corresponding non-tumorous liver, and the number of different spots was greater in the 4 moderately-differentiated nodule-in-nodule HCCs (39–68 spots) than in the 2 well-differentiated homogeneous HCCs (6 and 3 spots). RLGS profiles of the two parts were different to each other in all 4 nodule-in-nodule HCCs. On the other hand, two other homogeneous HCCs showed the same RLGS profiles in the two regions.
Conclusion: Thus, intratumoral genomic heterogeneity was demonstrated in the advanced HCC samples, and the genomic alterations may relate to the progression of HCC.
Progression of hepatocellular carcinoma as reflected by nuclear DNA ploidy and cellular differentiation
1998, Journal of Hepatology
Background/Aims: Intratumor heterogeneity of DNA ploidy within a single hepatocellular carcinoma is not well understood. The present study was designed to examine the histologic distribution of intratumor DNA ploidy in hepatocellular carcinomas of different growth types in relation to cell differentiation.
Methods: Twenty patients (16 men and four women; mean age, 60.2 years) with hepatocellular carcinoma (mean diameter, 4.3 cm) were studied. One hundred and twenty-seven samples from different sites of each tumor were analyzed by determination of the nuclear DNA content and histological examination.
Results: The DNA ploidy was heterogeneous in nine (45%) of the 20 tumors. Five tumors had a mixture of diploid and aneuploid regions, and the remaining four consisted of aneuploid regions with different DNA indices. There was no significant difference in patient characteristics between the heterogeneous and homogeneous groups. A significant correlation was found between tumor growth type and the incidence of heterogeneity. Only 16% of single nodular carcinomas without intratumor septal formation exhibited heterogeneity, while single nodular tumors with septal formation or confluent multinodular tumors were associated with high incidences of different DNA ploidy patterns or DNA indices. There was no aneuploidy in well-differentiated foci, while aneuploidy was frequently found in moderately or poorly differentiated foci (incidences of 67% and 74%, respectively).
Conclusions: Heterogeneity of DNA ploidy may develop along with changes in growth pattern and cell dedifferentiation or by confluence of nodules originating from different tumor cell clones.
Flow cytometric dna-ploidy and proliferative activity of diethylnitrosamine-induced hepatocellular carcinoma and pulmonary metastases in monkeys
1995, Hepatology
Flow cytometric DNA analysis was carried out on diethylnitrosamine (DEN) induced primary hepatocellular carcinomas (HCC) and lung metastases in monkeys. In analyzing one sample from each of 113 HCC cases, 76 (67.2%) were diploid and 37 (32.7%) aneuploid. When more samples were analyzed from the same tumorous liver, all of the 76 diploid cases maintained their pattern, whereas 5 (13.5%) of the aneuploid cases displayed both diploid and aneuploid DNA. Studies of lung metastases from 44 (28 diploid, 16 aneuploid) HCC cases showed that the DNA-ploidy pattern characterizing the primary HCC was preserved in the metastases in 78.6% of the diploid and 93.7% of the aneuploid cases. The average synthetic phase fraction (SPF) value for the diploid tumors was 7.7% and the aneuploid tumors 14.9%. The difference is highly significant (P < .01). Highly significant correlation was found between the DNA ploidy and the SPF values, both in the primary HCC (P = .0001) and the metastases (P = .0266). Of different tumor and host features examined, statistically significant correlation was only found between DNA-ploidy/SPF and the cytological tumor grade. This study represents the first DNAploidy analysis of HCC in monkeys. The data showed that diploid and aneuploid tumors displayed comparable metastatic potential. The DNA-ploidy pattern was preserved in the metastases in the majority of the cases.

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Lack of intratumoral heterogeneity in DNA ploidy pattern of hepatocellular carcinoma

Abstract

Biochim Biophys Acta

Isolation, karyotype, and clonal growth of heterogeneous subpopulations of human malignant gliomas

Cancer Res

Tumor heterogeneity

Cancer Res

Flow cytometry in clinical cancer research

Cancer Res

Clinical and biological significance of aneuploidy in human tumors

J Clin Pathol

Flow cytometry of breast carcinoma: I Relation of DNA ploidy level to histology and estrogen receptor

Cancer

Intratumoral variations in DNA distribution patterns in mammary adenocarcinoma

Cytometry

Cellular DNA content—a stable feature in epithelial ovarian cancer

Br J Cancer

Comparison of nuclear DNA content in primary and metastatic papillary thyroid carcinoma

Cancer Res

Flow-cytometric demonstration of tumor-cell subpopulations with different DNA contents in human colorectal carcinomas

Eur J Cancer

Heterogeneity of colorectal adenocarcinomas evaluated by flow cytometry and histopathology

Br J Cancer

DNA stemline heterogeneity in colorectal cancer

Cancer

Intratumoral regional differences in DNA ploidy of gastrointestinal carcinomas

Cancer

Regional ploidy variations in signet ring cell carcinomas of the stomach

Cancer

Intratumoral heterogeneity in DNA ploidy of esophageal squamous cell carcinomas

Cancer

Clonal heterogeneity of small-cell anaplastic carcinoma of the lung demonstrated by flow-cytometric DNA analysis

Cancer Res