Dynamic genomic changes in methotrexate-resistant human cancer cell lines beyond DHFR amplification suggest potential new targets for preventing drug resistance.

BACKGROUND: Although DHFR gene amplification has long been known as a major mechanism for methotrexate (MTX) resistance in cancer, the early changes and detailed development of the resistance are not yet fully understood. METHODS: We performed genomic, transcriptional and proteomic analyses of human colon cancer cells with sequentially increasing levels of MTX-resistance. RESULTS: The genomic amplification evolved in three phases (pre-amplification, homogenously staining region (HSR) and extrachromosomal DNA (ecDNA)). We confirm that genomic amplification and increased expression of DHFR, with formation of HSRs and especially ecDNAs, is the major driver of resistance. However, DHFR did not play a detectable role in the early phase. In the late phase (ecDNA), increase in FAM151B protein level may also have an important role by decreasing sensitivity to MTX. In addition, although MSH3 and ZFYVE16 may be subject to different posttranscriptional regulations and therefore protein expressions are decreased in ecDNA stages compared to HSR stages, they still play important roles in MTX resistance. CONCLUSION: The study provides a detailed evolutionary trajectory of MTX-resistance and identifies new targets, especially ecDNAs, which could help to prevent drug resistance. It also presents a proof-of-principal approach which could be applied to other cancer drug resistance studies.

Incidence and prognosis of olfactory and gustatory dysfunctions related to SARS-CoV-2 Omicron strain infection in China: A national multicenter survey of 35,566 individuals.

Objective: This cross-sectional study aimed to determine the epidemiology of olfactory and gustatory dysfunctions related to COVID-19 in China. Methods: This study was conducted by 45 tertiary Grade-A hospitals in China. Online and offline questionnaire data were obtained from patients infected with COVID-19 between December 28, 2022, and February 21, 2023. The collected information included basic demographics, medical history, smoking and drinking history, vaccination history, changes in olfactory and gustatory functions before and after infection, and other postinfection symptoms, as well as the duration and improvement status of olfactory and gustatory disorders. Results: Complete questionnaires were obtained from 35,566 subjects. The overall incidence of olfactory and taste dysfunction was 67.75%. Being female or being a cigarette smoker increased the likelihood of developing olfactory and taste dysfunction. Having received four doses of the vaccine or having good oral health or being a alcohol drinker decreased the risk of such dysfunction. Before infection, the average olfactory and taste VAS scores were 8.41 and 8.51, respectively; after infection, they decreased to 3.69 and 4.29 and recovered to 5.83 and 6.55 by the time of the survey. The median duration of dysosmia and dysgeusia was 15 and 12 days, respectively, with 0.5% of patients having symptoms lasting for more than 28 days. The overall self-reported improvement rate was 59.16%. Recovery was higher in males, never smokers, those who received two or three vaccine doses, and those that had never experienced dental health issues, or chronic accompanying symptoms. Conclusions: The incidence of dysosmia and dysgeusia following infection with the SARS-CoV-2 virus is high in China. Incidence and prognosis are influenced by several factors, including sex, SARS-CoV-2 vaccination, history of head-facial trauma, nasal and oral health status, smoking and drinking history, and the persistence of accompanying symptoms.

[In vitro study of expression and effects of HSP70/HSP90 in nasopharyngeal carcinoma cells after thermotherapy].

OBJECTIVE: To investigate the changes and effects of HSP70/HSP90 of in nasopharyngeal carcinoma cells HNE1 after thermotherapy. METHODS: HNE1 cells were incubated at 42 degrees C for 2 h. The changes of mRNA and protein level of HSP70/HSP90 were detected by real-time PCR and western-blot at different intervals. HNE1 cells were pretreated with quercetin, geldanamycin or quercetin plus geldanamycin, respectively, before heat treatment. Flow cytometry assay was applied to determine the apoptosis of HNE1 cells before thermotherapy and at 2 h, 4 h, 6 h, 8 h, 12 h and 24 h after thermotherapy. RESULTS: HSP70/HSP90 expression was up regulated at 2 h, reached to its peak at 4 h, descended at 8 h and returned to the normal level at 24 h after thermotherapy. The inhibition of HSP70 through quercetin induced up-regulation and delayed descent of HSP90. Combined pretreatment with quercetin and geldanamycin could significantly induce HNE1 apoptosis when compared with pretreatment with quercetin or geldanamycin alone (P<0.05). CONCLUSION: HSP70/HSP90 expression in HNE1 up regulated promptly after thermotherapy. Inhibition of expression and activity of HSP70/HSP90 before thermotherapy can increase sensitivity of the tumor cell to heat treatment.

[Screening and identification of human nasopharyngeal carcinoma-associated antigens].

OBJECTIVE: The purpose of this study was to provide an informative marker for clinical immunopathological diagnosis of nasopharyngeal carcinoma (NPC) and to lay a foundation for furture study of immunotherapy. METHODS: The NPC cDNA expression library was screened by serological analysis of recombinant cDNA expression libraries (SEREX) technology using xenogenic immune sera. The positive nucleotide sequences of cDNA inserts were determined and analysed with DNASIS and BLAST software in EMBL and GenBank. RESULTS: Eighteen positive clones encoding antigen genes were obtained after immunoscreening. These antigen genes included known genes, such as MAGE gene, CREM gene, beta2-microglobulin, Ribosomal Protein, et al, and unknown genes or ESTs in GenBank. To characterize the expression profile of these genes, the antibodies in sera from 40 nasopharyngeal carcinoma patients and 40 healthy donors were assayed with three antigens to screen specific and relative serum markers for nasopharyngeal carcinoma. The results showed that the positive rates for These results indicate nasopharyngeal carcinoma patients were higher than those for healthy donors. CONCLUSION: that some of the antigens may be related to nasopharyngeal carcinoma and may be valuable tumor markers in the diagnosis of nasopharyngeal carcinoma.

Co-inhibition of HSP70/HSP90 synergistically sensitizes nasopharyngeal carcinoma cells to thermotherapy.

The upregulation of both HSP70 and HSP90 frequently compromises the effects of thermotherapy. The co-inhibition of HSP70/HSP90 may be preferable to enhance the effects of thermotherapy on nasopharyngeal carcinoma cells. The changes of HSP70 and HSP90 were detected after thermotherapy in human nasopharyngeal cancer cell HNE1. 17-Dimethylaminoethylamino-17-demethoxygeldanamycin (17-DMAG) and quercetin were used to inhibit the activity of HSP90 and HSP70. The enhanced effects were evaluated in vitro and in vivo. Both HSP70 and HSP90 were upregulated promptly in HNE1 after thermotherapy. Single inhibition of HSP70 resulted in overexpression and delayed descent of HSP90. The co-inhibition of HSP70/HSP90 with quercetin plus 17-DMAG significantly increased apoptosis in hyperthermia-treated HNE1 cells both in vitro and in vivo. The co-inhibition of HSP70/HSP90 synergistically sensitizes nasopharyngeal carcinoma cells to hyperthermia.

[Study on the rates of smoking among the residents of Beijing, from 1997 to 2004].

OBJECTIVE: This study is to find out the trend of smoking rates among Beijing residents since the implementation of "Rules of Smoking Cessation in Public in Beijing" and to compare the data collected from 1997 and 2001. METHODS: Residents of Beijing, aged above 15 in a family were sampled by stratified sampling, and 4 districts were selected as the study locales. RESULTS: The over smoking rate of Beijing residents decreased by 7.6 percent with male smoking rate decreased by 13.03% and the female smoking rate ascended by 0.14% during that period. CONCLUSION: The smoking rate among residents of Beijing was decreasing which partly giving the credit to the implementation of the "Rules of Smoking Cessation in Public in Beijing".

[Surveys on the implementation of "rules on stop smoking in the public places in Beijing" from 1997 to 2004].

OBJECTIVE: To find out the situation of the implementation on "rules on stop smoking in the public places in Beijing", a study was carried out to compare the results from 1997 and 2001. METHODS: Thousands of residents aged above 15 in a family were sampled by stratified sampling, and 4 districts were selected as study locales. RESULTS: (1) About 40% of the department or school leaders considered much about the work on banning of smoking with 70 % of the departments or schools having posters of signs on 'ban on smoking' in the meeting rooms. (2) Some of the departments or schools had inspectors looking after the issue. (3) 60% of the people had never or 10% had seen that the inspectors ever criticizing or punishing the smokers who violating the rules. (4) People still often saw some one would smoke in public (the rates were 19.53% , 37.89% , 6.35% respectively in 1997,2001,2004 ). (5) People received the information of these rules through TV news or newspaper. CONCLUSION: Although success had been achieved since the implementation of the Rules, it did not seem to have had substantial progress.