Relationship between tumor DNA ploidy and regional lymph node changes in lung cancer.

Seventy-four patients with lung cancer, resected consecutively from April 1989 to August 1990, were divided into (1) 21 with diploid tumors having a single G0/G1 peak and a coefficient of variation (CV) of 4.9 or less, (2) 18 with peridiploid tumors having a single G0/G1 peak and a CV at 5.0 or more, and (3) 35 with aneuploid tumors having multiple G0/G1 peaks. Aneuploid tumors had higher frequencies of lymphatic invasion and metastasis to the mediastinal lymph nodes. To evaluate the relationship between ploidy tumor status and immunologic competence of the regional lymph nodes, histologic findings and the proportion of killer T-lymphocytes were examined in the dissected lymph nodes. Aneuploid tumors had significantly lower proportions of paracortical hyperplasia and killer T-lymphocytes than did diploid and peridiploid ones in the nonmetastatic lymph nodes of N0 and N1 disease. These findings suggest the possibility that a decline in the antitumor competence of these lymph nodes could cause metastasis to the nodes. The recurrence rates were 19% in diploid, 33% in peridiploid, and 54% in aneuploid tumors, and the 2-year survival rates were 87%, 78%, and 44%, respectively. Peridiploid tumors showed intermediate values between diploid and aneuploid in terms of immunologic competence, recurrence rate, and survival. They were assumed to have a different proportion of aneuploid cells than the other two.     

Relationship between DNA ploidy, antigen expression and survival in renal cell carcinoma

Tumor ploidy and expression of renal differentiation antigens as recognized by a previously described series of monoclonal antibodies (MAbs) (RC2, RC3, RC4, RC38, RC69, RC154 and G250) were assessed in 48 renal-cell carcinomas. A positive association was found between aneuploidy and development of metastases. Aneuploid tumors showed a loss of antigen expression more frequently than diploid tumors. This difference was statistically significant for antigens recognized by RC3 and RC69. Loss of RC154 reactivity was associated with increased metastatic potential. Clinical stage was the most powerful single prognostic variable but ploidy and RC3 reactivity carried additional prognostic information.     

Cell dissociation techniques in human breast cancer — Variations in tumor cell viability and DNA ploidy

Summary Approximately 70% of breast cancers contain cell populations with hyperdiploid (>G₀/G₁) DNA content; however, cells cultured from breast cancers have only diploid DNA contents and karyotypes. Mechanically dissociated cells rarely, if ever, grow in culture, while enzymatically dissociated cells do grow in most cases. To determine if cell dissociation techniques used to prepare cells for culture and other laboratory procedures select for cells with specific features, and if tumor cells are killed in the process, breast cancer cells obtained by mechanical dissociation and by enzymatic dissociation were examined for DNA content and cell viability (measured by dye exclusion). Mechanical dissociation yielded more dead cells and cells with hyperdiploid (>G₀/G₁) DNA than did enzymatic dissociation. Hyperdiploid cells were also found in the dye-excluding population with each dissociation technique, suggesting that the hyperdiploid cells were not always dead.We conclude that,in vivo, tumors contain cellular subpopulations with low viability and hyperdiploid (>G₀/G₁) DNA patterns. The extent to which these subpopulations are present in a sample depends on the dissociation technique employed. That only diploid cells are found in cultures of primary breast cancers may be because enzymatic dissociation, used to prepare cells for culture, yields predominantly diploid cells. These observations also have important implications for interpreting measurements made on dispersed cells,e.g., viability, DNA content, and other cytochemical markers.     

Analysis of fibrinolytic proteins in relation to DNA ploidy in prostate cancer

The tissue concentrations of urokinase-type plasminogen activator (u-PA), urokinase-type plasminogen activator receptor (u-PAR), plasminogen activator inhibitor type 1 (PAI-1) and tissue-type plasminogen activator (t-PA) were investigated by an ELISA technique in normal and malignant samples of the prostate from 24 patients undergoing radical prostatectomy for organ-confined prostate cancer. The median concentration of u-PA was significantly higher in cancerous than in normal prostate tissue (p = 0.006). No significant increase of u-PAR, PAI-1 and t-PA was found in cancer tissue in comparison with the benign samples (p > 0.05). Assessment of the relationship between fibrinolytic proteins and DNA ploidy revealed an increased u-PA, u-PAR and PAI-1 in diploid prostate cancer as compared with the normal controls. However, in aneuploid cancer u-PA remained high but u-PAR and PAI-1 were decreased. This led to a higher local concentration of u-PA in aneuploid samples than in normal prostate and in diploid prostate cancer. No alteration of median t-PA was found in benign prostate or in diploid or aneuploid prostate cancer. The altered expression of u-PA, u-PAR and PAI-1 in diploid and aneuploid prostate cancer suggests a possible role of fibrinolytic proteins in the different biologic behavior of tumors, and may be one explanation for the higher metastatic potential of aneuploid tumors. Int. J. Cancer 78:320–325, 1998© 1998 Wiley-Liss, Inc.     

Correlation of DNA ploidy and proliferative activity in human gastric cancer

Analysis of DNA ploidy patterns was performed on 129 cases of primary gastric cancer and the results were correlated with histologic findings and in vivo bromodeoxyuridine (BrdU) labeling. Forty-nine cases were diploid (38%) and 80 cases were aneuploid (62%). There was no correlation between DNA ploidy and histologic type. In aneuploid tumors, incidence of lymphatic invasion, lymph node metastasis, and rate of advanced cases were significantly higher than those in diploid tumors. During the follow-up period of 5 to 10 years, 23 of 40 patients (55%) with aneuploid tumors died of disease within 3 to 120 months. Only 13 of 36 patients (36%) with diploid tumors died of disease. The BrdU labeling indices (BrdU LI) ranged from 2.8% to 26.7%, with a mean of 10.4%. There was no correlation between BrdU LI and histologic type or stage. The mean BrdU LI of early cancers was 8.1%. The mean BrdU LI of advanced cancers was 11.9%. The BrdU LI of cancers with lymphatic invasion or lymph node metastasis was higher than those without them. The mean BrdU LI of diploid cancers was 6.0%. The mean BrdU LI of aneuploid cancers was 11.9%. There was a good correlation between BrdU LI and DNA ploidy patterns. These results indicate that DNA ploidy patterns and BrdU LI may possibly be useful prognostic markers for gastric cancers.     

大肠癌 p27蛋白表达与 DNA倍体分析的研究

背景与目的细胞周期调控的改变存在于绝大多数肿瘤中,因此可以认为肿瘤是一类细胞周期疾病.p27蛋白是细胞周期负性调控因子的主要成员,同时也是肿瘤的重要预后因子.p27蛋白表达下降及亚细胞定位改变在大肠癌发展中的意义尚不清楚.大肠癌中 p27蛋白表达与 DNA倍体关系尚未见报道.本实验研究大肠癌 p27蛋白表达与临床病理特征及 DNA倍体分析的关系.方法应用免疫组化方法检测 p27蛋白在 40例大肠癌中的表达水平,同时应用病理图像 DNA测量系统对癌组织进行倍体分析.结果p27蛋白在大肠癌中的高表达率为 62.5%(25/40),低表达率为 37.5%(15/40),p27蛋白低表达与大肠癌分化程度低显著相关(P< 0.05).除胞核着色外,32.5%(13/40)的大肠癌出现胞浆免疫阳性反应.p27蛋白胞浆表达与 Dukes分期晚(P< 0.01)、易发生淋巴结转移显著相关(P< 0.05).p27蛋白低表达组 DNA多倍体细胞检出率(22.2± 11.3)% 明显高于 p27蛋白高表达组 [(8.0± 7.7)%,P< 0.001]. 结论p27蛋白表达的下降及亚细胞定位改变促进大肠癌的发展 ; 抑制 DNA多倍体形成可能是 p27蛋白的抑癌机理之一.     

恶性肿瘤DNA倍体异质性与细胞生物学特性的关系

目的 探讨恶性肿瘤组织DNA倍体异质性与癌细胞生物学特性的关系.方法 采用流式细胞术(FCM)检测163例恶性肿瘤组织DNA倍体异质性,并分析DNA倍体异质性与肿瘤细胞DNA指数(DI)、细胞凋亡(Apo)百分比、S期细胞百分比(SPF)以及DNA倍体类型等细胞生物学特性参数之间的相关性.结果 各种恶性肿瘤的DNA倍体异质性明显不同,在6种肿瘤中,DNA倍体异质体检出率大小分布顺序与DNA异倍体的分布十分相似.恶性肿瘤组织的DI值与DNA异质体检出率呈显著正相关(r=0.872 4,P＜0.05).异质体的DNA异倍体检出率(100.0%)和SPF[(1.51±0.67)%1均显著高于同质体[22.0%,(0.34±0.51)%](P均＜0.01),而Apo百分比[(2.08±1.25)%]显著低于同质体组织[(11.99±10.25)%](P＜0.01).结论 DNA倍体异质性是恶性肿瘤的重要生物学特性之一,且与癌细胞其他生物学特性关系十分密切,可作为肿瘤的恶性程度和预后判断的重要指标.     

Prognostic value of DNA ploidy and automated assessment of stroma fraction in prostate cancer

The combination of DNA ploidy and automatically estimated stroma fraction has been shown to correlate with recurrence and cancer death in colorectal cancer. We aimed to extend this observation and evaluate the prognostic importance of this combined marker in prostate cancer. DNA ploidy status was determined by image cytometry and the stroma fraction was estimated automatically on hematoxylin and eosin stained sections in three tumor samples from each patient to account for tumor heterogeneity. The optimal threshold for low (≤56%) and high (>56%) stroma fraction was identified in a discovery cohort (n = 253). The combined marker was validated in an independent patient cohort (n = 259) with biochemical recurrence as endpoint. The combined marker predicted biochemical recurrence independently in the validation cohort. Multivariable analysis showed that the highest risk of recurrence was observed for patients with samples that had both non-diploid ploidy status and a high stroma fraction (hazard ratio: 2.51, 95% confidence interval: 1.18–5.34). In conclusion, we suggest the combination of DNA ploidy and automatically estimated stroma fraction as a prognostic marker for the risk stratification of prostate cancer patients. It may also be a potential generic marker as concurrent results have been described in colorectal cancer.     

DNA ploidy and prostate-specific antigen as prognostic factors in clinically resectable prostate cancer.

Prostate-specific antigen (PSA) and DNA ploidy as measured by flow cytometry were compared with conventional prognostic indicators in 112 patients who underwent radical prostatectomy for clinically resectable prostate cancer. The variables examined included age, race, prostatic acid phosphatase (PAP), Gleason score of the radical prostatectomy specimen, and pathologic stage. No significant relationships were found between DNA ploidy and age, mean PAP value, and absolute PAP value. Of the 112 patients, 65 (58.0%) had disease limited to the prostate (pathologic Stages A and B); 47 (42.0%) had extraprostatic disease (pathologic Stages C and D1). The stage was related to the Gleason score (P less than 0.0001) where extraprostatic disease was associated with a Gleason score of 6 to 10. Nineteen (17.0%) patients had aneuploid tumors, and 93 (83.0%) had diploid tumors. DNA ploidy significantly correlated with pathologic stage (P = 0.04); aneuploidy was identified more frequently in patients with Stages C and D1 tumors. Aneuploid tumors occurred more frequently than diploid tumors in patients with a Gleason score of 6 to 10 (P = 0.034). Mean PSA values were higher in patients with aneuploid tumors (P = 0.078), extraprostatic neoplasms (P = 0.00001), and cancers with a Gleason score of 6 to 10 (P = 0.0004). Furthermore, PSA values greater than 10.0 ng/ml were associated with extraprostatic disease and a Gleason score of 6 to 10 (P less than 0.05 and P less than 0.001, respectively). Significant racial differences were found with respect to DNA ploidy, mean DNA indices, and mean PSA values. The 18 black patients had more DNA aneuploid tumors (P = 0.043), a higher mean DNA index (P = 0.017), and a higher mean PSA value (P = 0.043) than the 94 white patients. Both PSA and DNA ploidy analysis by flow cytometry appear to be valuable indicators in the evaluation of patients with prostatic carcinoma.     

Comparison of human prostate specific glandular kallikrein 2 and prostate specific antigen gene expression in prostate with gene amplification and overexpression of prostate specific glandular kallikrein 2 in tumor tissue.

BACKGROUND: There is a need for specific markers that can indicate the different stages of prostate carcinoma. There is ongoing speculation concerning the value of prostate specific glandular kallikrein (hK2) in this regard. METHODS: The expression levels of both hK2 and human prostate specific antigen (hPSA) were compared at the mRNA and protein levels by using in situ hybridization and immunohistochemistry techniques in tissue specimens from patients with benign prostatic hyperplasia and malignant prostate carcinoma. The respective gene copy numbers were analyzed by a competitively differential polymerase chain reaction-based method. RESULTS: In situ hybridization revealed that hK2 was expressed at significantly higher levels in malignant prostate tissue compared with benign prostate tissue (P < 0.0005), whereas hPSA expression levels were the reverse (P = 0.06). In benign tissue, the mean level of hK2 mRNA was 82% of the respective value of hPSA (P < 0.003), whereas, in tumor tissue, the mean hK2 expression level was 21% higher than that of hPSA (P < 0.01). The results at the protein level supported the mRNA findings: hPSA expression was lower in malignant tissues compared with benign tissues (17 of 20 specimens), whereas an increase in hK2 expression was detected in 17 of 19 specimens. The authors report that the hK2 gene (hKLK2) was amplified in prostate carcinoma tissue, whereas the hPSA gene was not. There was a correlation between hPSA and hK2 mRNA levels in both benign tissue (correlation coefficient [r] = 0.735; P < 0.01) and malignant tissue (r = 0.767; P < 0.01). CONCLUSIONS: Gene amplification of hKLK2 may be one of the factors leading to higher expression of hK2 in prostate carcinoma. The correlation between hK2 and hPSA expression levels indicates coordinated expression of the genes in both normal and abnormal prostate gland. The results suggest the potential value of hK2 in the diagnosis of prostate carcinoma through mRNA analyses and gene amplification.     

DNA ploidy in primary testicular cancer

The DNA stemline ploidy was measured by flow cytometry (FCM) in 129 samples from paraffin-embedded primary testicular tumours (61 seminomas, 68 non-seminomas). Only one DNA stemline was found in 38 seminomas and 44 non-seminomas. Two seminomas and one non-seminoma were DNA diploid, the other tumours being non-diploid. Twenty-three seminomas and 24 non-seminomas displayed two or three DNA stemlines. The median minimal DNA index (DI) of all seminomas was significantly higher than that of all non-seminomas (1.58 vs 1.43; P: 0.008). Three seminomas removed from two monozygotic twins within 1 week had DIs of 1.66, 1.56 and 1.59. In this limited series there was no association between DNA ploidy of the primary tumour and the metastatic status for either seminomas or non-seminomas. The results support the pathogenetic model stating that at least some (if not all) non-seminomas develop from a seminoma by additional chromosomal aberration. The clinical relevance of DNA stemline ploidy has to be further evaluated in larger series.     

Relationship between ploidy and steroid hormone receptors in primary invasive breast cancer

The relationship between ploidy, as measured by flow cytometry, and the presence of oestrogen and progesterone receptors was investigated in 145 primary invasive breast cancers. The tumours were considered as an integral group, and as subgroups of lobular and ductal carcinomas. An association was found between the presence of aneuploid stemlines and an absence of oestrogen receptors (ER), for the total tumour population (P less than 0.02), and for the ductal carcinoma group (P less than 0.05). An association between aneuploidy and an absence of progesterone receptors (PR) was observed for the total tumour group (P less than 0.05). Evaluation of a combined oestrogen and progesterone receptor status indicated that the association between aneuploidy and an absence of both receptors was highly significant. The probability of such an association was P less than 0.001 for the total tumour population, and P less than 0.01 for the ductal tumour group. Assessment of progesterone receptor expression by breast cancers containing oestrogen receptors indicated that aneuploid tumours were as likely to express PR as were diploid tumours. Hence, the biological activity of oestrogen receptors appears unmodified by the presence of aneuploid nuclei.     

Intratumor heterogeneity of DNA ploidy and correlations with clinical stage and histologic grade in prostate cancer

Paraffin blocks from 60 patients with prostate cancer were used to study the DNA ploidy patterns by flow cytometry. Nineteen patients had stage A disease, 11 had stage B, 9 had stage C, 20 had stage D, and 1 was of unknown stage. Histologically, 32 of the cancers were well differentiated, 21 were moderately differentiated, and 7 were poorly differentiated. Eighteen patients had an aneuploid or tetraploid (A/T) pattern and 42 had a diploid pattern. Seventy-one percent (5/7) of patients with poorly differentiated, 48% (10/21) with moderately differentiated, and 9% (3/32) with well-differentiated histology had A/T patterns (P< 0.01). Forty-five percent (9/20) of patients with stage D, 44% (4/9) with stage C, 27% (3/11) with stage B, and 5% (1/19) with stage A had A/T patterns (P < 0.05). Nine patients with an A/T pattern also had DNA ploidy studies done on the “benign” part of the specimen. These specimens showed diploid patterns although three of these patients had well-differentiated tumor in the “benign” designated part of the specimen. One patient with mixed histology had an aneuploid pattern on the poorly differentiated section and a diploid pattern on the well-differentiated section of the “malignant” designated part of the same specimen. We conclude that prostate cancer patients with non-diploid tumors have more advanced disease and less differentiated tumors than patients with diploid tumors and that considerable histological and ploidy heterogeneity may be present in different parts of the same paraffin-embedded specimen. © 1993 Wiley-Liss, Inc.     

DNA甲基化与肿瘤的关系

DNA甲基化在基因组的稳定性和肿瘤发生中起着重要作用。肿瘤可发生许多基因的异常甲基化，尤其是抑癌基因的高甲基化对肿瘤发生有重要作用。由于DNA甲基化的可逆转特性，因此可以作为恶性肿瘤的治疗靶点。现对甲基化在肿瘤发生中的作用和临床意义作一综述。     

Increased cell-free DNA in plasma of patients with metastatic spread in prostate cancer

We compared the plasma DNA concentrations in patients with different prostate cancer (PCa) stages and with benign prostate hyperplasia (BPH) and in healthy persons. Patients with localized cancer had DNA plasma within the reference interval. Increased plasma DNA was found in patients with lymph node and distant metastases and also in BPH. The association between plasma DNA and the survival time was similarly strong as with prostate-specific antigen. It can be concluded that plasma DNA has a limited validity as metastatic marker in PCa patients but follow-up studies combined with the investigation of tumor-related characteristics of plasma DNA would be reasonable.     

Relationship between cell ploidy and glucocorticoid induced death in human lymphoid cell lines

We have found a relationship between sensitivity to glucocorticoid induced cell death (at 10 microM glucocorticoid) and ploidy in the human lymphoid cell line CCRF/CEM-C7. Most sensitive clones are diploid, whilst resistant clones and the resistant parent line CCRF/CEM are tetraploid. Diploid sensitive clones have a tendency to become aneuploid within a few months of isolation, with alterations in their kinetic responses to glucocorticoids. This is followed by a doubling in DNA content which results in reversion to the tetraploid glucocorticoid resistant state of the parent line CCRF/CEM. A few sensitive clones have been found to be tetraploid but with different kinetic responses to glucocorticoids as compared to diploid clones. The principal difference being an extended lag period (48-72 h) prior to lethal response. The relationship between ploidy and glucocorticoid sensitivity does not appear to extend to other human lymphoid cell lines.     

DNA ploidy is closely linked to tumor invasion, lymph node metastasis, and prognosis in clinical gastric cancer

DNA ploidy microspectrophotometrically determined in 254 patients with gastric carcinoma was investigated from the standpoint of tumor invasion, lymph node metastasis, and prognosis. DNA distribution patterns were grouped into low and high ploidies. The 24.0% frequency in the high ploidy group, at the mucosal stage, increased in proportion to invasion into the deeper layers. There was a high incidence of lymph node metastasis in the high ploidy group, compared with the low ploidy group, in case of invasion beyond the mucosa. Widespread nodal involvement was frequent in the high ploidy group. The 5-year survival rate was 73.7% in patients of high ploidy, with a statistical difference compared to the 90.6% in those of low ploidy (P less than 0.01). In the multivariate analysis of 226 patients undergoing curative surgery, the DNA ploidy proved to be a major independent prognostic factor. These findings indicate a close correlation among DNA ploidy, tumor invasion and nodal involvement, and the significant clinical value of DNA analysis for predicting the prognosis in patients with gastric carcinoma.