Evaluation of Loss of Heterozygosity Before and After Interferon Therapy in Patients With Hepatitis C Virus Infection Who Developed Hepatocellular Carcinoma During Follow Up

Seiji Suzuki; Yutaka Kondo; Noboru Hirashima; Hideaki Kato; Fuminaka Sugauchi; Yasuhito Tanaka; Etsuro Orito; Ying Yang; Yu Shen; Kenji Sakakibara; Ryuzo Ueda; Masashi Mizokami

Disclosures

J Gastroenterol Hepatol. 2003;18(12) 

In This Article

Results

The results of LOH analysis are shown in Table 2 . Two microdissected samples were examined in each patient using 12 microsatellite markers, and it was found that 12 (9%) of 138 and 32 (22%) of 143 informative samples showed LOH in the pre-IFN-N and pre-IFN-H samples, respectively. When LOH was present in at least one tissue specimen for each marker in each patient, then LOH was considered as positive. The proportion of positive LOH in all informative markers was calculated for each patient, and this proportion was termed the LOH score (for example, LOH score of N-1 in pre-IFN-N samples is 2/7 = 0.29).

The LOH scores for each patient in the pre-IFN-N and pre-IFN-H samples are illustrated in Figure 3. According to this data, the LOH score is a significant risk factor for predicting HCC (likelihood ratio test from the logistic regression, P = 0.01). The medians of the LOH scores in the pre-IFN-N and pre-IFN-H samples were 0.14 and 0.25, respectively. The LOH score in the pre-IFN-H samples was significantly higher than that in the pre-IFN-N samples (exact Wilcoxon test, P = 0.04). There was no significant difference in the LOH score between CR and NR in both samples.

Comparison of loss of heterozygosity (LOH) score distributions between pre-IFN-N and pre-IFN-H samples. Each blot indicates LOH score for each patient. CR and NR indicate the complete responder and non-responder to IFN therapy, respectively. The median of LOH score for the pre-IFN-N and pre-IFN-H samples are 0.14 and 0.25, respectively. The LOH score for the pre-IFN-H samples is significantly higher than that in the pre-IFN-N samples (P = 0.04). However, when comparing the response to IFN therapy, there was no difference in LOH score between CR and NR for both samples. IFN, interferon.

To evaluate the LOH incidence in different steps in cancer patients, LOH was examined in cancerous (CT samples) and adjacent non-CT (ANCT samples) derived from surgically resected specimens in seven of the nine cases in the HCC group ( Table 2 ). Loss of heterozygosity was detected in 44/186 (24%) and 47/119 (40%) of informative samples of ANCT samples and of CT samples, respectively.

The LOH scores for each patient from theHCC group are plotted in Figure 4. The medians of the LOH scores for the pre-IFN-H, ANCT, and CT samples from the patients were 0.25, 0.44, and 0.44, respectively. Intriguingly, in the HCC group, the LOH score for the pre-IFN-H, ANCT, and CT samples was not significantly different (exact Wilcoxon test P = 0.22), although some patients, such as H-1, H-4, and H-7 showed a trend of stepwise increase in the incidence of LOH, suggesting that IFN therapy had little or no effect on the incidence of LOH in patients who developed cancers. The authors could not observe a distinct tendency in the LOH score among pre-IFN-H, ANCT and CT samples between the CR (H-1, H-3, H-4, H-7, H-8) and NR (H-6, H-9) patients.

Comparison of loss of heterozygosity (LOH) score for each patient of the hepatocellular carcinoma (HCC) group. H-1-H-9 indicate patient numbers. CR and NR indicate the complete responder and non-responder to interferon (IFN) therapy, respectively. Loss of heterozygosity score for the (white bar) pre-IFN-H samples, (grey bar) adjacent non-cancerous tissue (ANCT) samples and (black bar) cancerous tissue (CT) samples from the HCC group patients. Although H-1, H-4, and H-7 showed a trend of stepwise increase in LOH, the LOH scores for pre-IFN-H, ANCT and CT samples were not significantly different (P = 0.22). IFN, interferon.

The percentages of LOH-positive patients for each marker in each group are illustrated in Figure 5. Loss of heterozygosity was not detected on the markers D2S336, D8S258, D13S153, D13S171, and D18S1118 in the non-HCC group. In contrast, a high frequency of LOH was detected on every marker in the HCC group. The proportions of positive LOH in markers D1S199, D6S305, D7S493, D8S277, D8S258, and D18S1118 tended to sequentially increase during the development of HCC, while those in D1S235, D4S426, D13S153, D13S171, and D17S786 were almost the same before and after the cancer had arisen.

Percentages of loss of heterozygosity (LOH)-positive patients for each marker in each group. (speckled bar) pre-interferon (IFN)-N samples, (white bar) pre-IFN-H samples, (grey bar) adjacent non-cancerous tissue (ANCT) samples and (black bar) cancerous tissue (CT) samples. On markers D2S336, D8S258, D13S153, D13S171 and D18S1118, no LOH was detected in non-hepatocellular carcinoma (HCC) samples. In contrast, LOH was detected on every marker in the HCC group (pre-IFN-H, ANCT and CT samples).

The incidences of MSI were 0/195 (0%), 3/199 (2%), 3/247(1%), and 6/162 (4%) for the pre-IFN-N, pre-IFN-H, ANCT, and CT samples, respectively.

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