Case report of a KIT-mutated melanoma patient with an excellent response to apatinib and temozolomide combination therapy.

Malignant melanoma is one kind of malignant disease which has high rates of mortality, metastasis, and poor prognosis. The therapeutic landscape is rapidly changing with the development of novel agents in recent decades, such as anti-PD-1 agents, anti-CTLA-4 agents, and BRAF inhibitors. However, since most of these novel agents are very expensive, not all patients can afford them. Apatinib is a novel oral small-molecule tyrosine kinase inhibitor targeting the intracellular domain of vascular endothelial growth factor receptor 2 (VEGFR-2) and may also be effective on Ret, c-KIT, and c-src. Temozolomide (TMZ) is a second-generation alkylating agent and a cytotoxic drug for melanoma treatment. In this work, we reported a case of metastatic melanoma with an excellent response to apatinib/TMZ combination therapy with progression-free survival for more than one year. This patient showed high expression of CD117, VEGFR-3, and KIT mutation in exon 11, suggesting that apatinib may induce clinical response via inhibiting VEGFR and c-KIT. Apatinib/TMZ combination therapy could be a new option for the treatment of advanced melanoma with KIT mutation.

Clear Cell Carcinoma of the Uterine Cervix: A Clinical and Pathological Analysis of 47 Patients Without Intrauterine Diethylstilbestrol Exposure.

OBJECTIVES: The aim of this study was to summarize the clinical and pathological characteristics and to conduct prognosis analysis of patients who were diagnosed with clear cell carcinoma of the uterine cervix (CCCUC) and without a history of exposure to diethylstilbestrol. METHODS: We performed a retrospective review of all the patients with CCCUC who were diagnosed and treated at Zhejiang Cancer Hospital between 1998 and 2014. Charts were reviewed for clinical and pathological characteristics, and prognosis analysis was conducted. RESULTS: A total of 47 patients were included. Median age was 52 years. No patient had a history of exposure to diethylstilbestrol. The International Federation of Gynecology and Obstetrics stage distribution was 55.3% (n = 26) stage I, 40.4% (n = 19) stage II, 2.1% (n = 1) stage III, and 2.1% (n = 1) stage IV. Forty-two patients (89.4%) underwent radical hysterectomy and pelvic lymphadenectomy. Pathological examination revealed deep cervical stromal invasion (greater than two thirds) in 20 patients (48.4%), pelvic lymph node (PLN) metastasis in 10 patients (23.8%), lymphovascular space involvement in 9 patients (21.4%), and ovarian metastasis in 1 patient (2.4%). Advanced tumor stage (IIB-IV), larger tumor size (>4 cm), and PLN metastasis had negative effects on progression-free survival (PFS) and overall survival (OS) (P < 0.05). Adjuvant radiation therapy alone or concurrent chemoradiation therapy after radical surgery did not affect PFS or OS in patients with risk factors (P > 0.05). CONCLUSIONS: International Federation of Gynecology and Obstetrics stage, tumor size, and PLN status were prognostic factors for both PFS and OS in patients with CCCUC. The long-term effects of adjuvant radiation therapy or concurrent chemoradiation therapy may be limited for CCCUC patients with risk factors. Future larger case series or clinical trials are required to confirm these findings.

Application of a cervical low incision in the functional neck dissection of thyroid papillary carcinoma.

The present study aimed to discuss the advantage of the application of a cervical low incision for functional neck dissection in patients with thyroid papillary carcinoma. The study was a retrospective analysis of 87 thyroid papillary carcinoma patients; cervical low incision in the functional neck dissection was applied for 47 cases and the classic 'L' incision was applied for 40 cases. The different integrity, surgical time, blood loss and the aesthetic property of the incision were compared between the cervical low incision and the classic 'L' incision for lateral neck dissection of thyroid cancer. The postoperative pathological diagnosis was that the average total amount and the region II lymph nodes of the unilateral neck dissection were 33 and 10 for the cervical low incision group, and 32 and 11 for the classic 'L' incision group, respectively (P>0.05). The average unilateral neck dissection times were 87 and 58 min for the cervical low incision group and the classic 'L' incision group, respectively (P<0.05). The blood loss of the cervical low incision group was 67 ml, while the loss for the classic 'L' incision group was 61 ml (P>0.05). The postoperative incision of the cervical low incision group was smaller and more concealing. Additionally, the cosmetic deformities were milder for an inconspicuous cervical scar, and the sensation was improved for the patients in comparison with the classic 'L' incision group. These results suggest that the application of cervical low incision for functional neck dissection in thyroid papillary carcinoma patients aids in reducing postoperative complications, without increasing recurrence rates. Therefore, the classic 'L' incision can be replaced by the cervical low incision.

Vascular invasion is an independent prognostic factor for distant recurrence-free survival in papillary thyroid carcinoma: a matched-case comparative study.

OBJECTIVE: It is still unclear whether the clinicopathological and outcome characteristics of vascular invasion (VI) (+) papillary thyroid carcinoma (PTC) differ from VI (-) PTC. This study aims to establish distinguishing features of patients with VI (+) and VI (-) PTC and to investigate the biological and clinical aggressiveness of the disease in these patient groups. DESIGN: A matched-case comparative study. METHODS: 412 patients (VI (+) PTC study group n=103, and VI (-) PTC control group n=309). These two patient groups were matched 1:3 for variables of age, gender, histological variants, tumour/node/metastasis (TNM) staging and approximate duration of follow-up. Clinicopathological factors and prognosis were compared across the two patient groups. RESULTS: The median age at the time of diagnosis was 42.0 years, and 68.9% were females. Across the patient groups, the incidence of tumour multifocality in patients with VI (+) PTC was slightly higher than in those with VI (-) PTC (p=0.049). In addition, when undergoing more aggressive therapy regimens patients with VI (+) PTC showed decreased distant recurrence-free survival (DRFS), but not regional recurrence-free survival (RRFS) and disease-specific survival (DSS) compared with patients who were VI (-). VI was found to be an independent predictor of DRFS, combined with tumour size >3 cm and total thyroidectomy. CONCLUSIONS: VI was an independent risk factor for DRFS, necessitating the need for appropriate postoperative treatment and careful follow-up.

[Detection of micrometastases and its clinical significance in sentinel and non-sentinel lymph nodes from early cervical carcinoma].

OBJECTIVE: To investigate the clinical significance of micrometastasis detection in sentinel lymph nodes (SLN) from patients with early cervical carcinoma. METHODS: Thirty patients with early cervical carcinoma were studied to identify SLN intraoperatively using methylene blue. One lymph node was removed randomly from palpable SLN and other pelvic lymph nodes (nSLN) in each patient, so 268 lymph nodes were collected and cut into two halves, one half of the lymph node was used to analyze the expression of cytokeratin 19 (CK19) mRNA by real-time fluorescence quantitative polymerase chain reaction to determine the presence of micrometastasis, the other half was examined by routine histology with HE staining. RESULTS: 67 SLNs were detected in 28 cases (93.3%). Pelvic lymph nodes of 6 cases were confirmed pathological metastasis. The sensitivity of SLN detection was 66.7%, the accuracy rate was 96.4%, and the false negative rate was 16.7%. Among 268 lymph nodes (including 9 lymph nodes with pathological metastasis) detected by real-time fluorescence quantitative polymerase chain reaction, 68 lymph nodes were pathological negative but had micrometastasis, accounting for 26.3% (68/259) in pathologically negative lymph nodes. Among 24 patients with pathological negative lymph nodes, 16 cases had micrometastasis, accounting for 66.7% in those patients. Among 16 patients with micrometastasis, SLN of 3 cases were negative, but nSLN were micrometastasis, so the SLN false-negative rate rose to 18.2%. There were no significant relationships between pelvic lymph nodes micrometastasis and perivascular space involvement, deep stromal invasion and tumor grade (all P > 0.05). The micrometastasis rate of nSLN in patients with SLN micrometastasis was 100%, significantly higher than that in the patients with SLN non-micrometastasis (27.3%, P < 0.01). CONCLUSIONS: Real-time fluorescence quantitative polymerase chain reaction is a sensitive method to detect SLN micrometastasis. SLN micrometastasis may be an effective complement to SLN pathology to predict nSLN metastasis. Pelvic lymph nodes micrometastases have no significant relationship with pathological risk factors in cervical cancer and prognosis of patients.

Circulating Methylated XAF1 DNA Indicates Poor Prognosis for Gastric Cancer.

BACKGROUND: Methylated DNA in fluids may be a suitable biomarker for cancer patients. XAF1 has been shown to be frequently down-regulated in human gastric cancer (GC). Here, we investigated if XAF1 methylation in GC could be a useful biomarker. METHODS: Real-time RT-PCR was used to detect XAF1 mRNA expression; immunohistochemistry and western blot were used to examine XAF1 protein expression in GC tissues (n = 202) and their corresponding para-cancerous histological normal tissues (PCHNTs). Real-time methylation specific-PCR was used to investigate XAF1 promoter methylation in the same panel of GC tissues, their PCHNTs and sera. RESULTS: We confirmed frequent XAF1 down-regulation in both mRNA and protein levels in GC tissues as compared to normal controls and PCHNTs. XAF1 hypermethylation was evidenced in 83.2% (168/202) of GC tissues and 27.2% (55/202) of PCHNTs, while no methylation was detected in the 88 normal controls. The methylation level in GC tissues was significantly higher than that in PCHNTs (p<0.05). The hypermethylation of XAF1 significantly correlated with the down-regulation of XAF1 in GC tissues in both mRNA and protein levels (p<0.001 each). Moreover, we detected high frequency of XAF1 methylation (69.8%, 141 out of 202) in the sera DNAs from the same patients, while the sera DNAs from 88 non-tumor controls were negative for XAF1 methylation. The XAF1 methylation in both GC tissues and in the sera could be a good biomarker for diagnosis of GC (AUC = 0.85 for tissue and AUC = 0.91 for sera) and significantly correlated with poorer prognosis (p<0.001). In addition, after-surgery negative-to-positive transition of XAF1 methylation in sera strongly associated with tumor recurrence. CONCLUSIONS: 1) Dysfunction of XAF1 is frequent and is regulated through XAF1 promoter hypermethylation; 2) Detection of circulating methylated XAF1 DNAs in the serum may be a useful biomarker in diagnosis, evaluating patient's outcome (prognosis and recurrence) for GC patients.

[Methylation status of CDH1 gene in preoperative abdominal lavage of patients with gastric cancer and its clinical significance].

OBJECTIVE: To explore the association between the progression of gastric cancer and the aberrant methylation of CDH1 gene in preoperative abdominal lavage fluid. METHODS: Real-time methylation-specific polymerase chain reaction(qMSP) was used to investigate the methylation status of the CDH1 gene promoter 5'-CpG islands from preoperative abdominal lavage fluid in 92 patients with gastric cancer. The associations between methylation of CDH1 genes and clinicopathologic features and prognosis were investigated. RESULTS: Among the 92 patients with gastric cancer, aberrant methylation of CDH1 gene was detected in 45(48.9%) patients, including total aberrant methylation in 12(13.0%) cases and partly aberrant methylation in 33(35.9%) cases. Significant associations were found between CDH1 methylation status and tumor size, growth pattern, differentiation, lymphovascular invasion, infiltration depth, lymph node metastasis, distant metastasis, and clinical staging(all P<0.05). However, there were no significant associations between CDH1 methylation status with gender, age, tumor location, or Helicobacter pylori infection(all P>0.05). The median progression-free survival was 20 months for CDH1 methylation group and 38 months for non-methylated group, and the difference was statistically significant(P<0.01). Cox model analysis revealed that CDH1 methylation status in preoperative peritoneal lavage fluid was an independent factor associated with postoperative survival in patients with gastric cancer(P=0.000, RR=332.88, 95%CI:21.71-5105.07). CONCLUSIONS: The aberrant methylation of 5'-CpG of CDH1 gene promoter is common in gastric cancer. The examination of CDH1 methylation status of abdominal lavage should be considered in the progression of gastric cancer.

Stepwise cumulation of RUNX3 methylation mediated by Helicobacter pylori infection contributes to gastric carcinoma progression.

BACKGROUND: Helicobacter pylori has been recognized as a definite carcinogen for gastric cancer (GC); however, the pathogenesis of H. pylori infection remains unclear. Runt-related transcription factor 3 (RUNX3) is a candidate tumor suppressor gene whose deficiency is causally related to GC. However, in H. pylori infection-associated GC, the role of RUNX3 has not been studied. METHODS: The authors used real-time methylation-specific polymerase chain reaction analysis to determine methylation status of the RUNX3 promoter in a spectrum of gastric lesions, including 220 samples of chronic atrophic gastritis, 196 samples of intestinal metaplasia, 134 samples of gastric adenoma, 102 samples of dysplasia, and 202 samples of GC with paired noncancerous mucosa tissues and corresponding blood specimens. The association of abnormal methylation with precancerous gastric lesions was evaluated along with the association between RUNX3 methylation and H. pylori infection, and the concordance of methylation levels was investigated between serum and tissues. RESULTS: The results indicated that increasing RUNX3 promoter methylation was correlated with distinct stages of GC progression. GC tissues had the highest methylation proportion (75.2%) compared with precancerous gastric lesions, including chronic atrophic gastritis (15.9%), intestinal metaplasia (36.7%), gastric adenoma (41.8%), and dysplasia (54.9%). H. pylori infection, a major risk factor for GC, contributed to the inactivation of RUNX3 in gastric epithelial cells through promoter hypermethylation. The levels of RUNX3 methylation in serum were in significant concordance with the methylation levels observed in GC tissues (P = .887). CONCLUSIONS: The current findings supported RUNX3 methylation as a risk factors for the carcinogenesis of chronic atrophic gastritis with H. pylori infection and indicated that circulating RUNX3 methylation is a valuable biomarker for the detection of early GC.

CDH1 methylation in preoperative peritoneal washes is an independent prognostic factor for gastric cancer.

BACKGROUND AND OBJECTIVES: To investigate the clinical value of CDH1 methylation in preoperative peritoneal washes (PPW) from gastric cancer patients. METHODS: CDH1 methylation was detected by real-time methylation specific-PCR in tumor tissues and corresponding PPW from 92 gastric cancer patients, gastric mucosa from 40 chronic gastritis patients and 48 normal persons. RESULTS: CDH1 methylation was found in 75 of 92 (81.5%) gastric cancer tissues, which significantly correlated with size, growth pattern, differentiation, lymphatic invasion, venous invasion, invasion depth, lymph node metastasis, distant metastasis, and TNM stage of tumor (all P < 0.05), but its relationship to age, gender, tumor site, and H. pylori infection was not found (all P > 0.05). The percentage of CDH1 methylation in PPW was 48.9%, of which the Aζ value of ROC curve was 0.8 compared to that in gastric cancer tissues. Kaplan-Meier analysis showed that there was a significant difference in disease-free survival (DFS) between the patients with or without methylated CDH1 in their PPW (χ(2) = 109.64, P < 0.000). Cox regression analysis revealed CDH1 methylation in PPW was an independent risk factor for gastric cancer patients, with a remarkable decrease in DFS after postoperative 30 months. CONCLUSIONS: Methylated CDH1 in PPW predicts poor prognosis for gastric cancer patients.

Heterogeneity of chemosensitivity in esophageal cancer using ATP-tumor chemosensitivity assay.

AIM: Current chemotherapy for esophageal cancer is conducted on the basis of empirical information from clinical trials, which fails to take into account the known heterogeneity of chemosensitivity between patients. This study was aimed to demonstrate the degree of heterogeneity of chemosensitivity in esophageal cancers. METHODS: A total of 42 esophageal cancer specimens were collected. The heterogeneity of chemosensitivity in esophageal cancer specimens was examined using an ex vivo ATP-tumor chemosensitivity assay (ATP-TCA). RESULTS: Thirty eight specimens produced evaluable results (90.5%). The most active single agent tested was nedaplatin, to which 28.9% of samples were sensitive. Combinations of chemotherapy agents exhibited much higher sensitivity: cisplatin + paclitaxel was sensitive in 16 of 38 (42.1%) of samples, while nedaplatin+paclitaxel was more effective, which was sensitive in 20 of 38 cases (52.6%). CONCLUSION: There was a marked heterogeneity of chemosensitivity in esophageal cancer. Chemosensitivity testing may provide a practical method for testing new regimens before clinical trials in esophageal cancer patients.

Hypermethylation-modulated downregulation of RASSF1A expression is associated with the progression of esophageal cancer.

BACKGROUND AND AIMS: Chromosome 3p21 is an important locus harboring critical tumor suppressor genes (TSGs) implicated in the pathogenesis of multiple tumors including esophageal carcinoma (EC). Aberrant promoter methylation is a fundamental mechanism of inactivation of TSGs in cancer. RASSF1A, a candidate tumor suppressor gene, recently cloned from the lung tumor locus at 3p21.3, is frequently inactivated by hypermethylation of its promoter region in a number of malignancies. We undertook this study to investigate the methylation status of RASSF1A and its significance in esophageal squamous cell carcinoma (ESCC). METHODS: Real-time RT-PCR and real-time methylation-specific PCR (real-time MSP) were used to detect RASSF1A expression and the methylation status of the RASSF1A promoter, respectively, in 124 primary ESCC tissues. RESULTS: Hypermethylation, partial methylation and unmethylation of the promoter region of RASSF1A were detected in 56 (45.2%), 23 (18.6%) and 45 (36.2%) of 124 ESCC samples, respectively. Unmethylation of the promoter region of RASSF1A was detected in 119 (96%) of the 124 corresponding noncancerous tissues. Five (4.0%) of 124 noncancerous tissues showed partial methylation. The presence of hypermethylation was statistically associated with loss of RASSF1A mRNA expression in primary ESCC (p <0.05). There were statistically significant correlations between the presence of hypermethylation and regional lymph node involvement (p=0.000), histological differentiation (p=0.009) and tumor stage (p=0.000). CONCLUSIONS: Our results suggest that RASSF1A may be one of the ESCC-related TSGs located at 3p21, and hypermethylation of the CpG island promoter of the RASSF1A is associated with the progression of ESCC.

Aberrant methylation of different DNA repair genes demonstrates distinct prognostic value for esophageal cancer.

BACKGROUND: DNA mismatch repair (MMR) deficiency results in a strong mutator phenotype and high-frequency microsatellite instability (MSI-H), which are the hallmarks of many tumors. AIM: The objective of this study is to investigate the promoter CpG island methylation status of mismatch repair genes human mutL homolog 1 (hMLH1), human mutS homolog 2 (hMSH2), and O(6)-methylguanine-DNA methyltransferase (MGMT) in esophageal squamous cell carcinoma (ESCC) and its roles in alkylating agents chemotherapy. METHODS: Real-time methylation-specific polymerase chain reaction (PCR) (real-time MSP) was employed to detect promoter CpG island methylation of the hMLH1, hMSH2, as well as MGMT genes in 235 surgical tumor tissue samples from ESCC patients and their corresponding normal tissue samples. RESULTS: Promoter CpG island methylation of hMLH1, hMSH2, and MGMT were detectable in 43.4, 28.9, and 40.4% of ESCC tumor DNA, respectively, and the loss rates of hMLH1, hMSH2, and MGMT protein expression were 48.6, 34.5, and 40.9% in tumor tissues, respectively. For the entire population of 235 ESCC patients who were enrolled in operating treatment combined with radiotherapy and chemotherapy with alkylating agents, there was a significant difference in the overall survival between patients with methylated MGMT promoter and those with an unmethylated MGMT promoter (P < 0.05). CONCLUSION: Promoter CpG island methylation may be a frequent event in ESCC carcinogenesis. Detection of the methylated sequences of hMLH1, hMSH2, and MGMT appears to be promising as a predictive factor in primary ESCC.

Hypermethylation-modulated down-regulation of CDH1 expression contributes to the progression of esophageal cancer.

CDH1, a cell adhesion molecule, which plays a key role in maintaining the epithelial phenotype, is regarded as an invasion-suppressor gene in light of accumulating evidence from in vitro experiments and clinical observations. In an attempt to clarify the mechanism responsible for inactivation of this gene in carcinomas, we investigated the methylation status of the CDH1 gene 5'-CpG islands and its regulatory mechanism in the progression of esophageal squamous cell carcinoma. Real-time methylation-specific polymerase chain reaction (qMSP) and treatment with the demethylating agent 5-aza-2'-deoxycytidine (5-Aza-CdR) were conducted to analyze the methylation status at the CDH1 promoter region in the human esophageal carcinoma cell lines, EC1 and EC9706. A total of 235 invasive esophageal squamous cell carcinomas (ESCC) at stages I-IV and their corresponding normal tissue samples, were included in an immunohistochemistry study and methylation analysis of CDH1. The results demonstrate that in EC1 and EC9706 cells, the CDH1 promoter is methylated and treatment with 5-Aza-CdR restored CDH1 expression. Enhanced CDH1 expression decreased cell migration, invasion ability and increased adhesion ability. Decreased CDH1 expression was detected in 59.6% of ESCC tissues, compared with their adjacent non-neoplastic epithelia, which had a close correlation with the primary tumor status, lymph node status, distant metatasis and clinicopathologic stage. Hypermethylation at the CDH1 promoter was detected in 97.9% of 140 cases of ESCC with low CDH1 expression. The methylation of CDH1 promoters (P=0.929) was closely correlated with the lack of expression of their corresponding proteins. The Cox regression model for survival analysis showed that increases in CDH1 methylation had a greater impact on the prognosis than tumor clinical stage. These findings suggest that CDH1 gene silencing by promoter hypermethylation and the resultant reduction of CDH1 expression may play an important role in the progression of ESCC. CDH1 methylation was a significant predictor of survival in ESCC patients after surgery.

[Sentinel lymph node detection and intra-operational diagnosis in patients with early stage cervical cancer].

OBJECTIVE: To study the effective method for rapid detection of sentinel lymph nodes in patients with early stage cervical cancer and clinical value thereof. METHODS: Thirty female patients with early stage cervical cancer, 14 at FIGO stage IB1 and 16 at stage IIA, underwent injection of 99mTc-labelled sulfur colloid 0.4 mci/0.4 ml at the positions of 3, 6, 9, and 12 o'clock of the cervix 5 h or 18 h before operation, and injection of 1 ml of methylene blue at the same cervical positions as mentioned above after the abdomen was opened. The blue-stained lymph nodes or the lymph nodes directed by the blue-stained lymph vessels were identified as SLNs: gamma-detector was used to position the hot nodes. The SLNs were resected and then radical hysterectomy and pelvic lymphadenectomy were performed. SLN frozen section and imprint cytology were conducted during the operation, the results were compared with that of HE staining RESULTS: SLN were detected successfully in 29 of the 30 patients with a detection rate of 96.7%. Routine HE staining indicated pelvic lymph node metastasis in 9 patients. The SLN was positive in 8 of the 29 patients, negative in 20 patients, and false negative in 1 patient. The sensitivity, accuracy, and false-negative rates of SLN detection were 88.9%, 96.6%, and 11.1% respectively. The sensitivity, specificity, accuracy, and positive and negative predictive rates were 92.3%, 100%, 98.9%, 100%, and 98.8% for frozen section, and 92.3%, 97.6%, 96.8%, 85.7%, and 98.8% for imprint cytology respectively. CONCLUSION: Frozen section and imprint cytology may be effective rapid methods to diagnose SLN metastasis during operation. SLN detection can predict pelvic lymph node metastasis in cervical cancer.