Can DNA Methylation Profiling Classify Histologic Subtypes and Grades in Soft Tissue Sarcoma?

BACKGROUND: A clear classification of the subtype and grade of soft tissue sarcoma is important for predicting prognosis and establishing treatment strategies. However, the rarity and heterogeneity of these tumors often make diagnosis difficult. In addition, it remains challenging to predict the response to chemotherapy and prognosis. Thus, we need a new method to help diagnose soft tissue sarcomas and determine treatment strategies in conjunction with traditional methods. Genetic alterations can be found in some subtypes of soft tissue sarcoma, but many other types show dysregulated gene expression attributed to epigenetic changes, such as DNA methylation status. However, research on DNA methylation profiles in soft tissue sarcoma is still insufficient to provide information to assist in diagnosis and therapeutic decisions. QUESTIONS/PURPOSES: (1) Do DNA methylation profiles differ between normal tissue and soft tissue sarcoma? (2) Do DNA methylation profiles vary between different histologic subtypes of soft tissue sarcoma? (3) Do DNA methylation profiles differ based on tumor grade? METHODS: Between January 2019 and December 2022, we treated 85 patients for soft tissue sarcomas. We considered patients whose specimens were approved for pilot research by the Human Biobank of St. Vincent's Hospital, The Catholic University of Korea, as potentially eligible. Based on this, 41% (35 patients) were eligible; 1% (one patient) was excluded because of gender mismatch between clinical and genetic data after controlling for data quality. Finally, 39 specimens (34 soft tissue sarcomas and five normal samples) were included from 34 patients who had clinical data. All tissue samples were collected intraoperatively. The five normal tissue samples were from muscle tissues. There were 20 female patients and 14 male patients, with a median age of 58 years (range 19 to 82 years). Genomic DNA was extracted from frozen tissue, and DNA methylation profiles were obtained. Genomic annotation of DNA methylation sites and hierarchical cluster analysis were performed to interpret results from DNA methylation profiling. A t-test was used to analyze different methylation probes. Benjamini-Hochberg-adjusted p value calculations were used to account for bias resulting from evaluating thousands of methylation sites. RESULTS: The most common histologic subtypes were liposarcoma (n = 10) and leiomyosarcoma (n = 9). The tumor grade was Fédération Nationale des Centres de Lutte Contre Le Cancer Grades 1, 2, and 3 in 3, 15, and 16 patients, respectively. DNA methylation profiling demonstrated differences between soft tissue sarcoma and normal tissue as 21,188 cytosine-phosphate-guanine sites. Despite the small number of samples, 72 of these sites showed an adjusted p value of < 0.000001, suggesting a low probability of statistical errors. Among the 72 sites, 70 exhibited a hypermethylation pattern in soft tissue sarcoma, with only two sites showing a hypomethylation pattern. Thirty of 34 soft tissue sarcomas were distinguished from normal samples using hierarchical cluster analysis. There was a different methylation pattern between leiomyosarcoma and liposarcoma at 7445 sites. Using the data, hierarchical clustering analysis showed that liposarcoma was distinguished from leiomyosarcoma. When we used the same approach and included other subtypes with three or more samples, only leiomyosarcoma and myxofibrosarcoma were separated from the other subtypes, while liposarcoma and alveolar soft-part sarcoma were mixed with the others. When comparing DNA methylation profiles between low-grade (Grade 1) and high-grade (Grades 2 and 3) soft tissue sarcomas, a difference in methylation pattern was observed at 144 cytosine-phosphate-guanine sites. Among these, 132 cytosine-phosphate-guanine sites exhibited hypermethylation in the high-grade group compared with the low-grade group. Hierarchical clustering analysis showed a division into two groups, with most high-grade sarcomas (28 of 31) separated from the low-grade group and few (3 out of 31) clustered together with the low-grade group. However, three high-grade soft tissue sarcomas were grouped with the Grade 1 cluster, and all of these sarcomas were Grade 2. When comparing Grades 1 and 2 to Grade 3, Grade 3 tumors were separated from Grades 1 and 2. CONCLUSION: We observed a different DNA methylation pattern between soft tissue sarcomas and normal tissues. Liposarcoma was distinguished from leiomyosarcoma using methylation profiling. High-grade soft tissue sarcoma samples showed a hypermethylation pattern compared with low-grade ones. Our findings indicate the need for research using methylation profiling to better understand the diverse biological characteristics of soft tissue sarcoma. Such research should include studies with sufficient samples and a variety of subtypes, as well as analyses of the expression and function of related genes. Additionally, efforts to link this research with clinical data related to treatment and prognosis are necessary. LEVEL OF EVIDENCE: Level III, diagnostic study.

Dorsal rhizotomy for pain from neoplastic lumbosacral plexopathy in advanced pelvic cancer.

BACKGROUND: Pain from neoplastic lumbosacral plexopathy is resistant to conventional pain treatment. According to a recent review of destructive procedures for cancer pain, only cordotomy has been reported to play an important role in the treatment of cancer pain. To date, the effectiveness of dorsal rhizotomy, which selectively interrupts pain transmission, has not been shown in neoplastic lumbosacral plexopathy. OBJECTIVES: The present study seeks to find out the effectiveness of selective dorsal rhizotomies for intractable pain from neoplastic lumbosacral plexopathy in terminal pelvic cancer patients. METHODS: Dorsal rhizotomies of the involved segments were performed on 6 cancer patients in whom neuropathic pain from lumbosacral plexus involvement in terminal pelvic cancer had been refractory to other therapies. Clinical efficacy of the procedure was assessed by comparing patient pain ratings and narcotic usage before and after dorsal rhizotomy. RESULTS: Examination of the results indicated a significant reduction in pain ratings as well as a significant reduction in daily narcotic use. No adverse neurological effects were observed and no recurrence of pain from neoplastic lumbosacral plexopathy was noted. CONCLUSIONS: These findings provide corroborating clinical evidence for the effectiveness of selective dorsal root rhizotomy for the intractable pain from lumbosacral plexopathy in terminal pelvic cancer patients.

Clinicopathological implications of human papilloma virus (HPV) L1 capsid protein immunoreactivity in HPV16-positive cervical cytology.

BACKGROUND: The objective of this study was to investigate the expression of human papilloma virus (HPV) L1 capsid protein in abnormal cervical cytology with HPV16 infection and analyze its association with cervical histopathology in Korean women. MATERIAL AND METHODS: We performed immunocytochemistry for HPV L1 in 475 abnormal cervical cytology samples from patients with HPV16 infections using the Cytoactiv(®) HPV L1 screening set. We investigated the expression of HPV L1 in cervical cytology samples and compared it with the results of histopathological examination of surgical specimens. RESULTS: Of a total of 475 cases, 188 (39.6%) were immunocytochemically positive and 287 (60.4%) negative for HPV L1. The immunocytochemical expression rates of HPV L1 in atypical squamous cells of unknown significance (ASCUS), low-grade squamous intraepithelial lesions (LSIL), high-grade squamous intraepithelial lesions (HSIL), and cancer were 21.8%, 59.7%, 19.1%, and 0.0%, respectively. LSIL exhibited the highest rate of HPV L1 positivity. Of a total of 475 cases, the multiple-type HPV infection rate, including HPV16, in HPV L1-negative cytology samples was 27.5%, which was significantly higher than that in HPV L1-positive cytology samples (p = 0.037). The absence of HPV L1 expression in ASCUS and LSIL was significantly associated with high-grade (≥ cervical intraepithelial neoplasia [CIN] 2) than low-grade (≤ CIN1) histopathology diagnoses (p < 0.05), but was not significantly different between HPV16 single and multiple-type HPV infections (p > 0.05). On the other hand, among 188 HPV L1-positive cases, 30.6% of multiple-type HPV infections showed high-grade histopathology diagnoses (≥ CIN3), significantly higher than the percentage of HPV16 single infections (8.6%) (p = 0.0004) CONCLUSIONS: Our study demonstrates that the expression of HPV L1 is low in advanced dysplasia. Furthermore, the absence of HPV L1 in HPV16-positive low-grade cytology (i.e., ASCUS and LSIL) is strongly associated with high-grade histopathology diagnoses. The multiplicity of HPV infections may have an important role in high-grade histopathology diagnoses (≥ CIN3) in HPV L1-positive cases.

Treatment outcome in patients with vulvar cancer: comparison of concurrent radiotherapy to postoperative radiotherapy.

PURPOSE: To evaluate outcome and morbidity in patients with vulvar cancer treated with radiotherapy, concurrent chemoradiotherapy or postoperative radiotherapy. MATERIALS AND METHODS: The records of 24 patients treated with radiotherapy for vulvar cancer between July 1993 and September 2009 were retrospectively reviewed. All patients received once daily 1.8-4 Gy fractions external beam radiotherapy to median 51.2 Gy (range, 19.8 to 81.6 Gy) on pelvis and inguinal nodes. Seven patients were treated with primary concurrent chemoradiotherapy, one patient was treated with primary radiotherapy alone, four patients received palliative radiotherapy, and twelve patients were treated with postoperative radiotherapy. RESULTS: Twenty patients were eligible for response evaluation. Response rate was 55% (11/20). The 5-year disease free survival was 42.2% and 5-year overall survival was 46.2%, respectively. Fifty percent (12/24) experienced with acute skin complications of grade III or more during radiotherapy. Late complications were found in 8 patients. 50% (6/12) of patients treated with lymph node dissection experienced severe late complications. One patient died of sepsis from lymphedema. However, only 16.6% (2/12) of patients treated with primary radiotherapy developed late complications. CONCLUSION: Outcome of patients with vulvar cancer treated with radiotherapy showed relatively good local control and low recurrence. Severe late toxicities remained higher in patients treated with both node dissection and radiotherapy.

Correlation between the digital cervicography and pathological diagnosis performed at private clinics in Korea.

OBJECTIVE: To evaluate the correlation tendency between abnormal findings of digital cervicography and cervical pathology at private clinics in Korea. METHODS: Abnormal finding of digital cervicography performed at private clinics in Korea between January 1, 2010 and May 31, 2012 were analysed retrospectively. The patient's age, abnormal findings of digital cervicography, cervical cytology, human papillomaviru (HPV) test and cervical pathology were investigated and the rate of agreement between abnormal finding of digital cervicography and cervical pathology results was calculated. Abnormal findings of digital cervicography were divided into 4 categories: atypical, compatible with CIN1, compatible with CIN2/3 and compatible with cancer. RESULTS: The study group was composed of 1547 women with a mean (range) age of 37.4 (14-91 years). The agreement rate between abnormal findings of digital cervicography and cervical pathology was 52.0% in "compatible with CIN1", 78.9% in "compatible with CIN2/3", and 90.2% in "compatible with cancer". CONCLUSIONS: Abnormal findings of digital cervicography were highly concordant with cervical intraepithelial neoplasia (CIN) and cancer examined at outpatient clinics in Korea. Therefore, abnormal interpretations of digital cervicography can be used as an excellent auxiliary technique with cervical cytology for CIN and cancer.

Apoptosis-related mRNA expression profiles of ovarian cancer cell lines following cisplatin treatment.

OBJECTIVE: The aim of this study was to identify apoptosis-related genes of ovarian cancer cell lines following cisplatin treatment. METHODS: We used IC(50) values and fluorescence-activated cell sorting analysis to compare cell death in 2 ovarian cancer cell lines, namely, SKOV-3 and OVCAR-3, upon treatment with cisplatin. Moreover, the change in transcriptional levels of apoptosis-associated genes was measured with a dendron-modified DNA microarray. RESULTS: The protein levels for the up-regulated genes in each cell line were validated to identify the molecules that may determine the cellular behavior of cisplatin resistance. Eight genes were over-expressed in the 2 cell lines. The cisplatin-induced up-regulation of DAD1 in transcriptional and protein levels contributed to the cisplatin resistance of OVCAR-3, and the up-regulation of FASTK and TNFRSF11A in SKOV-3 resulted in its higher sensitivity to cisplatin than that of OVCAR-3. CONCLUSION: In the present study, we have identified a set of genes responsible for apoptosis following cisplatin treatment in ovarian cancer cell lines. These genes may give information about the understanding of cisplatin-induced apoptosis in ovarian cancer.

HCCR-1, a novel oncogene, encodes a mitochondrial outer membrane protein and suppresses the UVC-induced apoptosis.

BACKGROUND: The Human cervical cancer oncogene (HCCR-1) has been isolated as a human oncoprotein, and has shown strong tumorigenic features. Its potential role in tumorigenesis may result from a negative regulation of the p53 tumor suppressor gene. RESULTS: To investigate the biological function of HCCR-1 in the cell, we predicted biological features using bioinformatic tools, and have identified a LETM1 homologous domain at position 75 to 346 of HCCR-1. This domain contains proteins identified from diverse species predicted to be mitochondrial proteins. Fluorescence microscopy and fractionation experiments showed that HCCR-1 is located in mitochondria in the COS-7, MCF-7 and HEK/293 cell lines, and subcompartamentally at the outer membrane in the HEK/293 cell line. The topological structure was revealed as the NH2-terminus of HCCR-1 oriented toward the cytoplasm. We also observed that the D1-2 region, at position 1 to 110 of HCCR-1, was required and sufficient for posttranslational mitochondrial import. The function of HCCR-1 on mitochondrial membrane is to retard the intrinsic apoptosis induced by UVC and staurosporine, respectively. CONCLUSION: Our experiments show the biological features of HCCR-1 in the cell, and suggest that uncontrolled expression of HCCR-1 may cause mitochondrial dysfunction that can result in resisting the UVC or staurosporine-induced apoptosis and progressing in the tumor formation.