Establishment of papillary thyroid cancer organoid lines from clinical specimens.

Papillary thyroid cancer (PTC) is a common malignancy of the endocrine system, and its morbidity and mortality are increasing year by year. Traditional two-dimensional culture of cell lines lacks tissue structure and is difficult to reflect the heterogeneity of tumors. The construction of mouse models is inefficient and time-consuming, which is difficult to be applied to individualized treatment on a large scale. Clinically relevant models that recapitulate the biology of their corresponding parental tumors are urgently needed. Based on clinical specimens of PTC, we have successfully established patient-derived organoids by exploring and optimizing the organoid culture system. These organoids have been cultured stably for more than 5 passages and successfully cryopreserved and retried. Histopathological and genome analysis revealed a high consistency of the histological architectures as well as mutational landscapes between the matched tumors and organoids. Here, we present a fully detailed method to derive PTC organoids from clinical specimens. Using this approach, we have developed PTC organoid lines from thyroid cancer samples with a success rate of 77.6% (38/49) until now.

Papillary thyroid cancer organoids harboring BRAFV600E mutation reveal potentially beneficial effects of BRAF inhibitor-based combination therapies.

BACKGROUNDS: Papillary thyroid cancer (PTC), which is often driven by acquired somatic mutations in BRAF genes, is the most common pathologic type of thyroid cancer. PTC has an excellent prognosis after treatment with conventional therapies such as surgical resection, thyroid hormone therapy and adjuvant radioactive iodine therapy. Unfortunately, about 20% of patients develop regional recurrence or distant metastasis, making targeted therapeutics an important treatment option. Current in vitro PTC models are limited in representing the cellular and mutational characteristics of parental tumors. A clinically relevant tool that predicts the efficacy of therapy for individuals is urgently needed. METHODS: Surgically removed PTC tissue samples were dissociated, plated into Matrigel, and cultured to generate organoids. PTC organoids were subsequently subjected to histological analysis, DNA sequencing, and drug sensitivity assays, respectively. RESULTS: We established 9 patient-derived PTC organoid models, 5 of which harbor BRAFV600E mutation. These organoids have been cultured stably for more than 3 months and closely recapitulated the histological architectures as well as mutational landscapes of the respective primary tumors. Drug sensitivity assays of PTC organoid cultures demonstrated the intra- and inter-patient specific drug responses. BRAFV600E inhibitors, vemurafenib and dabrafenib monotherapy was mildly effective in treating BRAFV600E-mutant PTC organoids. Nevertheless, BRAF inhibitors in combination with MEK inhibitors, RTK inhibitors, or chemotherapeutic agents demonstrated improved efficacy compared to BRAF inhibition alone. CONCLUSIONS: These data indicate that patient-derived PTC organoids may be a powerful research tool to investigate tumor biology and drug responsiveness, thus being useful to validate or discover targeted drug combinations.

Vitamin C sensitizes BRAFV600E thyroid cancer to PLX4032 via inhibiting the feedback activation of MAPK/ERK signal by PLX4032.

BACKGROUND: BRAFV600E mutation is the most common mutation in thyroid cancer. It strongly activates MAPK/ERK pathway and indicates an invasive subtype of thyroid cancer. PLX4032 is a selective oral inhibitor of the BRAFV600 kinase although with limited effect in treating this panel of thyroid cancer, due to the feedback activation of MAPK/ERK as well as PI3K/AKT pathways. It was investigated that Vitamin C plays a positive role in inhibiting these pathways in thyroid cancer. However, whether Vitamin C could enhance the antitumor effect of PLX4032 remains largely unclear. METHODS: The antitumor efficacy of combination therapy with PLX4032 and Vitamin C on BRAFMT thyroid cancer cell was assessed by the MTT assay, EdU assay and colony formation, Chou-Talalay way was employed to analyze the synergistic effect. Flow cytometry were employed to assess cells' apoptosis and cell cycle arrest in response to combination therapy. Xenograft models were used to test its in vivo antitumor activity. Western blot and IHC were applied to investigate the mechanism underlying synergistic effect. RESULTS: PLX4032 or Vitamin C monotherapy was mildly effective in treating BRAFMT thyroid cancer cell and xenografts model. The combination therapy significantly inhibited cancer cell proliferation and tumor growth in nude mice, and induced cell apoptosis and cell cycle arrest compared to either monotherapy. PLX4032 monotherapy induced feedback activation of MAPK/ERK as well as PI3K/AKT pathway; while combination therapy significantly relieved this feedback. CONCLUSION: Vitamin C promotes the antitumor effect of PLX4032 in BRAFMT thyroid cancer cell and xenografts model via relieving the feedback activation of MAPK/ERK as well as PI3K/AKT pathway. PLX4032/Vitamin C combination may be a potential therapeutic approach to treat BRAFMT thyroid cancer.

The transcutaneous electromyography recording method for intraoperative neuromonitoring of recurrent laryngeal nerve during minimally invasive parathyroidectomy.

Intraoperative neuromonitoring (IONM) facilitates recurrent laryngeal nerve (RLN) protection in thyroid and parathyroid surgeries. This study aimed to investigate a novel transcutaneous electromyography (EMG) recording method for IONM of the RLN during minimally invasive parathyroidectomy (MIP). Twenty patients with primary hyperparathyroidism undergoing MIP were enrolled. Two paired needle electrodes were percutaneously inserted into the bilateral laminas of thyroid cartilage for monitoring the vagus nerve and RLN. A standardized IONM procedure (V1-R1-R2-V2 signals) was strictly followed, and the RLN was routinely located and mapped. Pre- and postoperative laryngofiberoscopy was performed to confirm vocal cord function. The proposed technique was successfully used in all patients, and typical EMG signals were effectively detected. No significant change in EMG signals before and after tumor resection was noted, and a normal vocal cord movement was ensured in all patients with postoperative laryngofiberoscopy. IONM helped localize the position of the RLN and facilitated the safe resection of the parathyroid tumor during MIP. The novel transcutaneous EMG recording method proposed in this study was feasible, convenient, reliable, and inexpensive.

Modified arytenoid muscle electrode recording method for neuromonitoring during thyroidectomy.

BACKGROUND: Intraoperative neuromonitoring (IONM) is an important application for protecting recurrent laryngeal nerve (RLN) during thyroid surgery. The method for recording arytenoid muscle electromyography (EMG) signals is reported to be feasible and reliable. However, the parameters of EMG signals are not provided. This study aimed to analyze the clinical characteristics of EMG signal parameters by modifying the insertion direction of needle electrodes. METHODS: A total of 92 patients who were scheduled to undergo thyroidectomy were recruited. Two paired needle electrodes were inserted in bilateral angle points between rectus cricothyroid muscle and inferior margin of thyroid cartilage (TC) intraoperatively, and then the information from the EMG signals was recorded according to four-step method (V1-R1-R2-V2). Pre-and post-operative laryngo-fiberoscopy was performed to confirm the vocal cord function. RESULTS: A total of 122 RLNs were successfully recorded during thyroidectomy, with the mean EMG amplitude and latency were 1,857±1,718/2,347±2,323 µV and 3.89±1.12/2.26±0.05 ms for V1/R1 signals before resection, and 1,924±1,705/2,450±2,345 µV and 3.87±1.17/2.27±0.08 ms for R2/V2 signals after resection. There were no significant changes before and after resection, and a normal vocal cord movement was observed postoperatively. The amplitude of left nerves was higher than that of the right ones. Furthermore, the latency of the right vagus was shorter than the left ones, but there was no difference in the amplitude and latency between age, sex and pathological types. CONCLUSIONS: Modified arytenoid muscle EMG recording method was considered to be safe, feasible and reliable. The latency of right vagus EMG signals were shorter than the left ones, and the amplitude of EMG signals might be related to different sides.

[Evaluation of radionuclide therapy for the residue after surgery in papillary thyroid carcinoma].

OBJECTIVE: To assess the efficacy of radioactive iodine (RAI) for the treatment of residual papillary thyroid cancer (PTC) after surgery. METHODS: A total of 20 patients diagnosed with PTC and underwent 2-6 courses of RAI therapy for residual PTC after surgery in other hospitals were included our study. Of these, 13 were in stage I, 3 in stage III and 4 in stage IV. All the cases were operated again due to the presence of suspicious residual tumors indicated by CT. Excision of thyroid tumor residue was performed in 5 cases and neck dissection in 15 cases (20 sides). The suspicious thyroid or neck residual tumors were examined pathologically after surgery. Response Evaluation Criteria in Solid Tumors (RECIST) was used to evaluate the efficacy of surgery treatment on residual tumor. T-test was used to identify variables associated to RAI and to calculate the propensity score to receive RAI after surgery. RESULTS: The patients aged 22-58 years, with a median age of 40 years. The mean times of surgeries received before RAI was 1.5 and the mean dose of applied RAI was 318 mCi (210-660 mCi). No significant difference in tumor size between pre-RAI and post-RAI was found (t = 1.177, P > 0.05). With postoperative pathological examination, the suspicious thyroid or neck residual tumors were confirmed as PTC or the cervical lymph metastasis of PTC. CONCLUSIONS: For the residue or metastasis of PTC after operation, reoperation should be a priority, while RAI therapy has no obvious therapeutic effect and it should be limited to selected cases such as those with distant metastasis or unsuitable for operation but with iodine uptake function, or taken as an adjuvant treatment after radical resection of cervical lesions.

Survival Outcome of Squamous Cell Carcinoma Arising from Sinonasal Inverted Papilloma.

BACKGROUND: Sinonasal inverted papilloma (IP) is a rare benign tumor of the nasal cavities and paranasal sinuses. It is destructive or bone-remodeling, tends to recur after surgical resection, and has a significant malignant potential. The present study aimed to perform a retrospective analysis of patients with squamous cell carcinoma (SCC) arising from IP, including characteristics, survival outcome, and predictors of associated malignancy. METHODS: The medical records of 213 patients diagnosed with IP from January 1970 to January 2014 were retrospectively reviewed. Eighty-seven patients were diagnosed with SCC/IP; their clinical characteristics, treatments, and survival outcomes were analyzed. RESULTS: Of the 87 patients with SCC/IP, the 5- and 10-year overall survival outcomes were 39.6% and 31.8%, respectively. Twenty-nine of these patients received surgery and 58 received combined surgery and radiation. Of the patients with stages III-IV, the 5-year survival rate was 30.7% for those treated with surgery only and 39.9% for those given the combination treatment (P = 0.849). Factors associated with significantly poor prognosis were advanced-stage, metachronous tumors, or with cranial base and orbit invasion. Age, synchronous or metachronous tumors, and pathological stage were independent risk factors for mortality, shown by multivariate analysis. CONCLUSION: Patients with SCC/IP had low overall survival outcomes. Advanced age, stage, and metachronous tumors are the main factors affecting prognosis. Treatment planning should consider high-risk factors to improve survival outcome.

[Treatment and prognosis of the carcinoma arising from nasal and sinonasal inverted papilloma: report of 62 cases and systematic review].

OBJECTIVE: To analyze the factors affecting prognosis of the carcinoma arising from nasal and sinonasal inverted papilloma. METHODS: The clinicopathological data of sixty-two patients treated in our hospital from January 1974 to February 2012 were retrospectively analyzed. Of the 62 cases, 10 were at stage I or II, 24 at stage III, and 28 at stage IV. Twenty-six patients were treated with surgery alone, and 36 with surgery combined with radiation therapy. Kaplan-Meier method and log-rank test were used for the survival analysis. RESULTS: The overall 5-years survival rate was 39.6%. The 5-years survival rate was 67.5% in the stage I or II patients,51.3% in the stage III patients, and 18.3% in the stage IV patients (P<0.05). The 5-years survival rate was 65.7% in patients who had no surgical history, and 29.9% in patients who had surgery (P<0.05). The 5-year survival rate was 17.6% in the group who relapsed after the treatment, and 49.6% in the non-relapsed patients (P<0.05). The 5-year survival rate was 23.4% in the patients who had involvement of cranial base and (or) orbit, and 47.6% in the patients whose cranial base and (or) orbit were clean (P<0.05). Of the patients at the same stage (III-IV), the 5-year survival rate of patients treated with surgery alone was 32.4%, and those treated with combination therapy was 36.2%(P=0.89). The univariate analysis showed that clinical stage, surgical history before malignization, involvement of the cranial base and (or) orbit organs, and post-operative relapse are significantly correlated to prognosis of the patients (P<0.05 for all). Multivariate analysis showed that age, clinical stage, and previous history of surgery were independent factors affecting the prognosis of the patients. Distant metastasis was the major cause of death, mostly lung metastases. CONCLUSIONS: Age, clinical stage and surgical history are the main factors affecting the prognosis of the patients. The history of recurrence and involvement of cranial base or orbit also play an important role for the prognosis. Distant metastasis is the main cause of death in the patients with carcinoma arising from nasal and sinonasal inverted papilloma.

[Analysis of the effectiveness of radiotherapy in postoperative residual tumors in patients with thyroid papillary carcinoma].

OBJECTIVE: The aim of this study was to assess the impact of radiotherapy on patients with postoperative residual or recurrent papillary thyroid cancer (PTC). METHODS: We retrospectively reviewed the medical records of 34 patients with PTC, who underwent surgery and radiotherapy in other hospitals, and treated at the Department of Head and Neck Surgery at Cancer Institute & Hospital CAMS from January 2011 to January 2014. Among the 34 cases, 22 were in stage I, 5 in stage II and 7 in stage IVa. The 34 patients received 1.5 times of surgery before radiotherapy in average. All the cases received radiotherapy (mean, 56 Gy; range, 50-70 Gy). The patients were re-operated in our hospital, and the specimens were examined by pathology. The pre- and post-radiotherapy images (CT and B-ultrasound) were compared, and the changes of tumor volume were examined. The objective effect of treatment on the tumor residual focus was evaluated using RECIST, and analyzed by t-test (SPSS 17.0). RESULTS: All the re-resected lesions after radiotherapy were proved by pathology to be papillary thyroid cancer (PTC) or metastatic PTC in cervical lymph nodes. Among the 34 patients, 22 cases showed mild or moderate cell degeneration and the other 12 cases showed no obvious degeneration. The largest tumor diameter was 27.18 mm before radiotherapy and 27.76 mm after radiotherapy, with a non-significant difference between them (t=-1.618, P>0.05). Among the 34 patients, only 3 patients received reoperation, all other 31 cases had complete resection, and no severe complications were observed except recurrent laryngeal nerve injury in one case. CONCLUSIONS: Radiotherapy has few therapeutic benefit to PTC patients after surgery with residual tumor or local recurrence. It should be used in the PTC patients, in which the tumor invasion involves important organ tissues and is difficult for a single operation to achieve safe resection margin, or in patients who can't bear a surgery because of severe coronary heart disease or others.