Polymorphisms in TYMS for Prediction of Capecitabine-Induced Hand-Foot Syndrome in Chinese Patients with Colorectal Cancer / Journal of the Korean Cancer Association, 대한암학회지

Purpose@#Capecitabine is an extensively used oral prodrug of 5-fluorouracil in treatment of colon cancer and is known to cause hand-foot syndrome (HFS). As the target enzyme for capecitabine, thymidylate synthase (TYMS) plays a key role for 5-fluorouracil metabolism and has been associated with some side effects caused by capecitabine. The aim of our study is to identify the possible genetic predictors of capecitabine-induced HFS (CAP-HFS) in Chinese colorectal cancer patients. @*Materials and Methods@#Whole exons of TYMS were sequenced for 288 extreme phenotype HFS patients, including 144 severe or early-onset (first 2 cycles) moderate HFS extreme cases and 144 extreme controls with no reported HFS. The associations between polymorphisms and CAP-HFS were analyzed using logistic regression under an additive model. @*Results@#We identified a novel risk mutation (c.1A>G, chr18:657743), was associated with severe HFS in an extreme case who was affected during the first cycle of treatment. Moreover, we identified three new variants, rs3786362, rs699517, rs2790, and two previously reported variants, 5’VNTR 2R/3R and 3′-untranslated region 6-bp ins-del, which were significantly associated with CAP-HFS (p < 0.05). In silico analysis revealed that the effect of these polymorphisms in the TYMS region on the development of HFS might not be restricted solely to the regulation of TYMS expression, but also the TYMS catalytic activity through the indirect effect on ENOSF1 expression. @*Conclusion@#This study identified new polymorphisms in TYMS gene significantly associated with CAP-HFS, which may serve as useful genetic predictors for CAP-HFS and help to elucidate the underlying mechanism of HFS.

Polymorphisms in TYMS for Prediction of Capecitabine-Induced Hand-Foot Syndrome in Chinese Patients with Colorectal Cancer / Journal of the Korean Cancer Association, 대한암학회지

Purpose@#Capecitabine is an extensively used oral prodrug of 5-fluorouracil in treatment of colon cancer and is known to cause hand-foot syndrome (HFS). As the target enzyme for capecitabine, thymidylate synthase (TYMS) plays a key role for 5-fluorouracil metabolism and has been associated with some side effects caused by capecitabine. The aim of our study is to identify the possible genetic predictors of capecitabine-induced HFS (CAP-HFS) in Chinese colorectal cancer patients. @*Materials and Methods@#Whole exons of TYMS were sequenced for 288 extreme phenotype HFS patients, including 144 severe or early-onset (first 2 cycles) moderate HFS extreme cases and 144 extreme controls with no reported HFS. The associations between polymorphisms and CAP-HFS were analyzed using logistic regression under an additive model. @*Results@#We identified a novel risk mutation (c.1A>G, chr18:657743), was associated with severe HFS in an extreme case who was affected during the first cycle of treatment. Moreover, we identified three new variants, rs3786362, rs699517, rs2790, and two previously reported variants, 5’VNTR 2R/3R and 3′-untranslated region 6-bp ins-del, which were significantly associated with CAP-HFS (p < 0.05). In silico analysis revealed that the effect of these polymorphisms in the TYMS region on the development of HFS might not be restricted solely to the regulation of TYMS expression, but also the TYMS catalytic activity through the indirect effect on ENOSF1 expression. @*Conclusion@#This study identified new polymorphisms in TYMS gene significantly associated with CAP-HFS, which may serve as useful genetic predictors for CAP-HFS and help to elucidate the underlying mechanism of HFS.

Status of AIDS discrimination among freshmen of a university in Hefei City / 上海预防医学

[Objective] To investigate the knowledge and discrimination of AIDS among freshmen in a university of Hefei City, so as to provide evidence for AIDS education and improve discrimination status.[Methods] A total of 317 students from a university of Hefei City were randomly selected using cluster sampling method and a questionnaire survey was conducted on them about AIDS knowledge and discrimination. [Results]The awareness rate of AIDS/HIV knowledge was 67.2％ in students. The total scores of AIDS discrimination knowledge attitude, emotional attitude and AIDS discrimination attitude were higher in the students without knowledge of AIDS than those with knowledge of AIDS, and the differences were statistically significant (P<0.05). In the discrimination behavior tendency, the score of the students in the knowledge group was higher than that in the no knowledge group, and the difference was statistically significant (P < 0.05). [Conclusion] The awareness of AIDS/HIV knowledge among freshmen in a university of Hefei City is good, but discrimination is widespread, and the degree of AIDS related knowledge directly affects the discrimination degree of AIDS discrimination. In future, we should strengthen the AIDS knowledge system education and pay more attention to the anti-discrimination propaganda intervention, especially the comprehensive intervention against behavior tendency.

Research development of Carpesii Fructus / 中国药理学与毒理学杂志

Carpesii Fructus is the fruit of Carpesium abrotanoides L. and is recorded in the Chinese Pharmacopoeia (2015 Edition). Carpesium abrotanoides broadly distribute in China. Traditionally, Carpesii Fructus was used as a parasiticide,especially for ascariasis,pinworms and tapeworm disease. In ancient times,the Carpesium plants were used as traditional Chinese,Korean and Japanese herbal medicines for the treatment of several diseases.Carpesii Fructus was first recorded in the book"Newly Revised Canon of Materia Medica"in the Tang Dynasty of China.The original plant is Compositae Arte-misia santonica (Seriphidium cinum) from middle east Persian. At present, Carpesium abrotanoides issometimes confused with the Lappula family in species classification. In the Song Dynasty of China,"KaiYang Materia Medica"recorded that the best Carpesii Fructus was from Persian.The main compo-nents of Carpesii Fructusare terpenes,phenolic compounds,flavonoids and coumarins. Including telekin, 3-epi-isotelekin, 11β-13-dihydro-1-epi-inuviscolide, carabrone, carabrol, terpene lactone, gerilin, carpesia, valeric acid, oleic acid, linolenic acid, thirty-one alkane, sterol, etc. The chemical components isolated from whole plants of carpesia are more than 143. In clinical practice, Carpesii Fructus is mainly used as antiparasitic drugs and usually combined with other drugs since the poor efficacy as single drug. Its toxic reaction is closely related to the dose of the drug.Carpesia,asa main component of Carpesii Fructus, might lead to adverse reactions also. At present, Major issuesof Carpesii Fructusare the lack of phar-macological research,as well as lack of in-depth study on the material basis.Therefore,further studies are needed on the drug development and clinical usuage.

The regulation of FFAR1 on insulin secretion and the development of related drugs / 药学学报

Free fatty acid receptor 1 (FFAR1), also known as G protein-coupled receptor 40 (GPR40), is a receptor for diverse free fatty acids. This review aims at summarizing effects and mechanisms of FFAR1 on insulin secretion and related blood glucose and lipids metabolism. FFAR1 is involved in the occurrence and development of type 2 diabetes, but its specific mechanism has not been clarified. FFAR1 is expressed in the wide variety of issues, especially β-cells in the pancreatic islets, as well as α-cells in islets, central nervous tissue, subcutaneous fat, skeletal muscle, gastrointestinal tract, etc. FFAR1 can act on islet β-cells to promote the secretion of insulin, promote α-cells on glucagon secretion, and regulate the secretion of endocrine cells in the gastrointestinal tract to balance the level of glucose and lipids. Existing research found that FFAR1 agonists have significant advantages. They promote insulin release, reduce weight and protect pancreatic β-cells, and have no risk of hypoglycemia. To in-depth understand the role of FFAR1 as a drug target in the treatment of diabetes, further pharmacological studies are still needed in order to obtain safer and more effective drugs against type 2 diabetes.

A single nucleotide polymorphism in the Epstein-Barr virus genome is strongly associated with a high risk of nasopharyngeal carcinoma / 癌症

<p><b>BACKGROUND</b>Epstein-Barr virus (EBV) commonly infects the general population and has been associated with nasopharyngeal carcinoma (NPC), which has a high incidence in certain regions. This study aimed to address how EBV variations contribute to the risk of NPC.</p><p><b>METHODS</b>Using logistic regression analysis and based on the sequence variations at EBV-encoded RPMS1, a multi-stage association study was conducted to identify EBV variations associated with NPC risk. A protein degradation assay was performed to characterize the functional relevance of the RPMS1 variations.</p><p><b>RESULTS</b>Based on EBV-encoded RPMS1 variations, a single nucleotide polymorphism (SNP) in the EBV genome (locus 155391: G>A, named G155391A) was associated with NPC in 157 cases and 319 healthy controls from an NPC endemic region in South China [P < 0.001, odds ratio (OR) = 4.47, 95% confidence interval (CI) 2.71-7.37]. The results were further validated in three independent cohorts from the NPC endemic region (P < 0.001, OR = 5.20, 95% CI 3.18-8.50 in 168 cases vs. 241 controls, and P < 0.001, OR = 5.27, 95% CI 4.06-6.85 in 726 cases vs. 880 controls) and a non-endemic region (P < 0.001, OR = 7.52, 95% CI 3.69-15.32 in 58 cases vs. 612 controls). The combined analysis in 1109 cases and 2052 controls revealed that the SNP G155391A was strongly associated with NPC (P(combined) < 0.001, OR = 5.27, 95% CI 4.31-6.44). Moreover, the frequency of the SNP G155391A was associated with NPC incidence but was not associated with the incidences of other EBV-related malignancies. Furthermore, the protein degradation assay showed that this SNP decreased the degradation of the oncogenic RPMS1 protein.</p><p><b>CONCLUSIONS</b>Our study identified an EBV variation specifically and significantly associated with a high risk of NPC. These findings provide insights into the pathogenesis of NPC and strategies for prevention.</p>

Plasma microRNA profiling in nasopharyngeal carcinoma patients reveals miR-548q and miR-483-5p as potential biomarkers / 癌症

MicroRNAs (miRNAs), which play a role in tumorigenesis, may also serve as diagnostic or prognostic biomarkers. However, studies on human miRNA profiles in plasma from nasopharyngeal carcinoma (NPC) patients are in their infancy. Here, we used microarrays to perform systematic profiling of human miRNAs in plasma from NPC patients. We subsequently used real-time quantitative polymerase chain reaction (Q-PCR) to validate miRNAs with aberrant expression that could serve as potential biomarkers. By comparing the plasma miRNA profiles of 31 NPC patients and 19 controls, 39 of 887 human miRNAs were found to be aberrantly expressed. Considering the fold change and P value, miR-548q and miR-483-5p were validated in 132 samples from 82 NPC patients and 50 controls. Moreover, high expression of miR-548q and miR-483-5p was further found in 3 NPC cell lines and clinical biopsy tissues from 54 NPC patients and 22 controls. Our results revealed that miR-548q and miR-483-5p are potential biomarkers of NPC. Combining the receiver operating characteristic (ROC) analyses of these 2 miRNAs, an area under the ROC curve (AUC) of 0.737 with 67.1% sensitivity and 68.0% specificity were obtained, showing the preliminary diagnostic value of plasma miRNAs. Moreover, most NPC patients with a poor outcome exhibited high expression (> median) of miR-548q (70.6%) and miR-483-5p (64.7%) in tissue samples, indicating their prognostic value. The high expression levels of miR-548q and miR-483-5p in plasma, cell lines, and clinical tissues of NPC patients indicate that their roles in NPC should be explored in the future.

Interaction of body mass index and a polymorphism in gene of catalytic subunit of glutamate cysteine ligase on breast cancer risk among Chinese women / 中华流行病学杂志

Objective To investigate the interaction of body mass index (BMI) and a single nucleotide polymorphism (SNP,rs17883901) in catalytic subunit of glutamate-cysteine ligase (GCLC) on breast cancer risk.Methods A total of 839 women with incident breast cancer and 863 age-matched controls without cancer were recruited at the same period in three affiliated hospitals of Sun Yat-sen University in Guangzhou from October 2008 to June 2010.GCLC rs17883901 was genotyped by MALDI-TOF-MS.Binary unconditional logistic regression was applied to calculate odds ratios and 95％ confidence intervals.Results The difference of present BMI and BMI at age 20 was not statistically significant between cases and controls,either as the genotypes of GCLC.No association was found between BMI at present and premenopausal or postmenopausal breast cancer risk.But we found that women who had a BMI at age 20 of 18.5 to 22.9 had a marginally decreased risk of premenopausal breast cancer [OR and 95％CI:0.69 (0.48,1.00)].Among women with CT/TT genotypes,whose present BMI was greater than 25 had a increased risk [OR and 95％CI:1.91 (1.09,3.36)] of breast cancer and a decreased risk [OR and 95％CI:0.56(0.31,0.99)] with a BMI at age 20 of 18.5 to 22.9.There was a interaction between GCLC gene (rs17883901)and BMI at present in breast cancer risk (P=0.043),which was not found between rs17883901 and BMI at age 20.Conclusion Our findings indicate BMI at age 20 may be a protective factor of premenopausal breast cancer,while no association appears between GCLC (rs17883901) and breast cancer.Obesity at present may significantly increase the risk of breast cancer among women with CT/TT genotypes of GCLC (rs17883901).

Centrifugation: an important pre-analytic procedure that influences plasma microRNA quantification during blood processing / 癌症

Circulating microRNAs are robustly present in plasma or serum and have become a research focus as biomarkers for tumor diagnosis and prognosis. Centrifugation is a necessary procedure for obtaining high-quality blood supernatant. Herein, we investigated one-step and two-step centrifugations, two centrifugal methods routinely used in microRNA study, to explore their effects on plasma microRNA quantification. The microRNAs obtained from one-step and two-step centrifugations were quantified by microarray and TaqMan-based real-time quantitative polymerase chain reaction (Q-PCR). Dynamic light scattering was performed to explore the difference underlying the two centrifugal methods. The results from the microarray containing 1,347 microRNAs showed that the signal detection rate was greatly decreased in the plasma sample prepared by two-step centrifugation. More importantly, the microRNAs missing in this plasma sample could be recovered and detected in the precipitate generated from the second centrifugation. Consistent with the results from microarray, a marked decrease of three representative microRNAs in two-step centrifugal plasma was validated by Q-PCR. According to the size distribution of all nanoparticles in plasma, there were fewer nanoparticles with size >1,000 nm in two-step centrifugal plasma. Our experiments directly demonstrated that different centrifugation methods produced distinct quantities of plasma microRNAs. Thus, exosomes or protein complexes containing microRNAs may be involved in large nanoparticle formation and may be precipitated after two-step centrifugation. Our results remind us that sample processing methods should be first considered in conducting research.

GWAS signals across the HLA regions: revealing a clue for common etiology underlying infectious tumors and other immunity diseases / 癌症

Increasing evidence suggests that multiple genes in the human leukocyte antigen(HLA) regions play an important role in development of cancers and immunity disorders. However, the biological mechanisms of the HLA associations are not well understood. We recently conducted a survey of all genome-wide association studies (GWAS) with significant findings in the HLA regions and concluded that diseases such as cancer and immune disorders are more likely to be associated with genetic variants located in the HLA regions than other diseases. This finding is suggestive for testing a hypothesis of a common etiology of infectious tumors and other immunity diseases.

Comparative study on risk factors and family history of familial and sporadic nasopharyngeal carcinoma patients / 中华预防医学杂志

<p><b>OBJECTIVE</b>To explore the difference between familial and sporadic nasopharyngeal carcinoma patients on risk factors and family history and provide evidence on genetic counseling and screening strategy for relatives of nasopharyngeal carcinoma patients in Guangdong province.</p><p><b>METHODS</b>The Cantonese nasopharyngeal carcinoma patients diagnosed in Cancer Center, Sun Yat-sen University from October, 2005 to October, 2007 were recruited as subjects. 1877 patients were collected, including 181 familial nasopharyngeal carcinoma patients and 1696 sporadic nasopharyngeal carcinoma patients. The demographic characteristics, clinical characteristics, risk factors and family history between two groups were compared. Moreover, the distribution of nasopharyngeal carcinoma patients in first-degree relatives and the time interval between proband and the affected first-degree relatives in familial nasopharyngeal carcinoma patients was analyzed.</p><p><b>RESULTS</b>All 9.64% of 1877 nasopharyngeal carcinoma patients had affected relatives in first-degree relatives, among them, 58.49% (124/212) were siblings and 41.51% (88/212) were parents. The mean time interval between siblings and proband were (7.40 +/- 5.41) years while the mean time interval between parents and proband were (15.55 +/- 10.61) years when nasopharyngeal carcinoma occurred, and the difference was statistically significant (t = -5.78, P < 0.01). More than 80% patients of the two group were at advanced stage when they were diagnosed. There were no difference (P values were all > 0.05) both in adulthood and childhood in salted fish (OR = 1.01; 95% CI: 0.59 - 1.75 vs OR = 1.31; 95% CI: 0.92 - 1.86), preserved vegetables (OR = 0.93; 95% CI: 0.58 - 1.49 vs OR = 1.12; 95% CI: 0.80 - 1.57), fermented pastes (OR = 0.37; 95% CI: 0.14 - 1.01 vs OR = 1.61; 95% CI: 0.99 - 2.48), fresh fruits (OR = 0.87; 95% CI: 0.60 - 1.26 vs OR = 0.65; 95% CI: 0.20 - 2.12) and cured meat (OR = 1.26; 95% CI: 0.87 - 1.83 vs OR = 1.28; 95% CI: 0.71 - 2.30) diet. No significant difference (P > 0.05) was obtained on smoking (OR = 0.99; 95% CI: 0.68 - 1.45) and incidence of other cancers in first-degree relatives (OR = 0.85; 95% CI: 0.56 - 1.28) in the two groups.</p><p><b>CONCLUSION</b>Familial nasopharyngeal carcinoma was 9.64% in the observed subjects. In the familial nasopharyngeal carcinoma, the time interval at diagnosis was shorter between proband and siblings as compared with parents. Most of the patients were at advanced stage. So, we recommend the first-degree relatives of nasopharyngeal carcinoma patients, especially siblings, should be screened regularly according to the specific conditions.</p>

Association of nasopharyngeal carcinoma risk with cytochrome P450 CYP1A1 gene polymorphisms / 中华预防医学杂志

<p><b>OBJECTIVE</b>To investigate the association between CYP1A1 gene polymorphisms and susceptibility of nasopharyngeal carcinoma in Cantonese nuclear families through family-based association study.</p><p><b>METHODS</b>A total of 457 Cantonese nuclear families,consisting of 2134 members, were recruited as subjects. Each family included two parents and at least one offspring with nasopharyngeal carcinoma. Two single nucleotide polymorphisms (SNP) in CYP1A1 named m1 (rs4646903) and m2 (rs1048943), were genotyped by PCR-RFLP assay and verified by directly sequencing. The genotype data were analyzed with family-based association test (FBAT) software to check the linkage and association between the two genetic markers and susceptibility of nasopharyngeal carcinoma.</p><p><b>RESULTS</b>FBAT analysis showed that the minor allele frequencies (MAF) of the two SNP were 0.442 (C) and 0.339 (G) respectively. For m1 polymorphism in CYP1A1 gene was not significantly associated with nasopharyngeal carcinoma in our study population whether stratified with VCA-IgA or not (without stratification: chi2 = 2.399, P = 0.301; with stratification: low-titer group (VCA-IgA <1:80), MAF = 0.457 (C), chi2 = 1.221, P = 0.543; high-titer group (VCA-IgA > or = 1:80), MAF = 0.427 (C), chi2 =2.832, P = 0.243) . For m2 polymorphism, when VCA-IgA <1:80, the G allele showed decreased transmission under additive and dominant model (MAF = 0.347 (G); Zadditive = -2.120, Padditive = 0.034; Zdominant = - 2.303, Pdominant = 0.021) and a boundary P value was got with global statistic (chi2 = 5.394, P = 0.067) . Haplotype TG (0.057), constructed by m1 and m2, might decrease nasopharyngeal carcinoma risk (Z= -2.002, P=0.045). A boundary P value was also got with global statistic (chi2 =7.067, P=0.070).</p><p><b>CONCLUSION</b>There was no statistical significance between m1 polymorphism and susceptibility of nasopharyngeal carcinoma in Cantonese nuclear families. And this study showed that m2 polymorphism might associated with the decrease of nasopharyngeal carcinoma in Cantonese nuclear families.</p>

Observation on therapeutic effect of digital acupoint pressure for treatment of the nerve root type of cervical spondylosis / 中国针灸

<p><b>OBJECTIVE</b>To observe the therapeutic effect of digital acupoint pressure for treatment of the nerve root type of cervical spondylosis.</p><p><b>METHODS</b>Four hundred cases were randomly divided into a digital acupoint pressure group (DAP group) and a medicine group, 200 cases in each group. Acupoints of Quepen (ST 12), Jianjing (GB 21) and Tianzong (ST 11) etc. were used for digital pressing in the DAP group; and Chinese herb medicine of Gentongping was routinely taken in the medicine group. After three treatment courses, the symptoms of pain and numbness, the signs of pressure measurement by compression of head, brachial plexus drawer test and arm myodynamia, as well as the total cumulative scores of daily living capability, were compared.</p><p><b>RESULTS</b>After treatment, the total cumulative scores of numbness, pressure measurement by compression of head, brachial plexus drawer test, arm myodynamia and daily living capability in both groups were obviously better than those of before treatment (all P<0.01); but there was a significant difference on the total cumulative score of the symptoms and signs between the two groups. The cured rate of 78.0% and total effective rate of 99.0% in the DAP group were better than those of 61.0% and 87.0% in the medicine group, respectively (both P<0.01).</p><p><b>CONCLUSION</b>Digital acupoint pressure plays an active role in improving the symptoms and signs on patients with nerve root type of cervical spondylosis, which is better than Chinese herb medicine of Gentongping.</p>

Family-based association study of XRCC1 gene polymorphisms in nasopharyngeal carcinoma / 中华预防医学杂志

<p><b>OBJECTIVE</b>To test the association between XRCC1 gene polymorphisms and susceptibility of nasopharyngeal carcinoma in Cantonese nuclear families through a family-based association study.</p><p><b>METHODS</b>A total of 2134 study subjects from 457 Cantonese nuclear families were recruited in the study. Each family had two parents and at least one offspring with nasopharyngeal carcinoma. Genotyping for three single nucleotide polymorphisms in XRCC1 gene, including rs1799782 (C > T), rs25489 (G > A) and rs25487 (G > A), were performed with PCR-RFLP assay. The genotype data were analyzed with family-based association test (FBAT) software to check linkage and association between the three genetic markers and susceptibility of nasopharyngeal carcinoma.</p><p><b>RESULTS</b>FBAT analysis showed XRCC1 gene genotypes and haplotypes were not significantly associated with nasopharyngeal carcinoma in our study population (rs1799782: chi(2) = 1.006, P = 0.605; rs25489: chi(2) = 0.470, P = 0.790; rs25487: chi(2) = 2.563, P = 0.278; haplotype: chi(2) = 3.004, P = 0.557, global statistic). For rs25487, the G allele (major allele) showed increased transmission under dominant model (Z = 1.985, P = 0.047). Whereas the C allele (minor allele) exhibited reduced transmission under recessive model (Z = -1.985, P = 0.047). However, no increased/reduced transmission was observed under additive model and with global statistic.</p><p><b>CONCLUSION</b>There is no evidence of an association between polymorphisms in XRCC1 gene and susceptibility of nasopharyngeal carcinoma in Cantonese nuclear families is observed in this study.</p>

Study on genetic polymorphisms of CYP2F1 gene in Guangdong population of China / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To investigate the genetic polymorphism of CYP2F1 gene, a member of CYP450 gene family in the healthy population and the patients with nasopharyngeal carcinoma (NPC) of Guangdong province, and furthermore analyze the relationship between CYP2F1 genetic polymorphism and the risk of developing NPC.</p><p><b>METHODS</b>By direct gene sequencing, all of 10 exons of CYP2F1 gene were detected in 40 peripheral blood specimens of patients with primary NPC. For the genetic polymorphism with high allelic frequency, mismatch PCR-RFLP technique was developed to identify the different frequency between 368 NPC cases and 344 cancer-free controls.</p><p><b>RESULTS</b>There were totally 35 SNPs identified in all of 10 exons and exon-intron junctions of CYP2F1 gene from 40 NPC patients, which included 10 missense mutations and 1 frame shift mutation. The most important mutation was C insertion located in 15-16 bp, which caused the frame shift. The allelic frequency of C insertion was 25%. However, there was no significant difference found between 368 NPC cases and 344 controls in allelic frequency of 15-16 bp C insertion mutation (P>0.05).</p><p><b>CONCLUSION</b>A lot of genetic polymorphism of CYP2F1 gene is found in Guangdong population of China. However, no single genetic polymorphism associated with the individual susceptibility to NPC can be identified. The cooperated operations with multiple genetic polymorphisms of one or more genes may be critical factors contributing to the development and progression of NPC.</p>