This is a preprint, a preliminary version of a manuscript that has not completed peer review at a journal. Research Square does not conduct peer review prior to posting preprints. The posting of a preprint on this server should not be interpreted as an endorsement of its validity or suitability for dissemination as established information or for guiding clinical practice.
Research article
Chang-ming Huang
Fujian Medical University Union Hospital
Corresponding Author
ORCiD: https://orcid.org/0000-0002-0019-885X
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Background: We previously demonstrated that CDK5RAP3 acts as a tumour suppressor in gastric cancer through negative regulation of the Wnt/β-catenin signalling pathway, but its function in chemotherapeutic responsiveness of gastric cancer has not been investigated. In this study, we aimed to examine the clinical significance of CDK5RAP3 to predict chemotherapeutic responsiveness in gastric cancer.
Methods: A collection of 188 pairs of tumour tissue microarray specimens from Fujian Medical University were employed for the discovery set, and 310 tumour tissue samples of gastric cancer patients were employed for the internal validation set. Eight-five tumour tissue samples from Qinghai University Hospital were used as the external validation set 1. Transcriptomic and clinical data of 299 gastric cancer patients from TCGA were used as the external validation set 2. CDK5RAP3 expression, microsatellite instability (MSI) status, and tumour-infiltrating lymphocytes (TIL) were examined with immunohistochemistry. Clinical outcomes of patients were compared with Kaplan-Meier curves and the Cox model.
Results: In a multi-centre evaluation, increased CDK5RAP3 indication of better prognosis depends mainly on MSI-L/MSS status or TILhigh. High CDK5RAP3 expression predicts sensitive therapeutic responsiveness to postoperative adjuvant chemotherapy in gastric cancer. In a stratification analysis based on CDK5RAP3 combined with TIL or MSI status, patients with CKD5RAP3low TILlow showed no significant difference in prognosis after receiving chemotherapy, whereas patients with CKD5RAP3low TILhigh, CKD5RAP3high TILlow, and CKD5RAP3high TILhigh had better responsiveness to chemotherapy. In addition, patients with CKD5RAP3high MSI-L/MSS status benefitted the most from adjuvant chemotherapy among all patients evaluated.
Conclusions: CKD5RAP3 can be used as an effective marker to evaluate individualized chemotherapy regimens in gastric cancer patients dependent on their TIL and MSI status.
Keywords
CDK5RAP3 , tumour , Wnt/β-catenin , chemotherapeutic responsiveness, microsatellite instability (MSI) status
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This is a list of supplementary files associated with this preprint. Click to download.
- SupplementaryFigure1.pdf
Reduced CDK5RAP3 expression is observed in gastric tumour tissues, and Wnt/β-catenin signalling is upregulated in CKD5RAP3low tumours. (A) The expression of CKD5RAP3 in gastric cancer tissue and adjacent tissue from the discovery set (n = 188). (B) The transcriptomic data of CKD5RAP3 in gastric cancer tissue and normal gastric tissue from the discovery set (n = 299). (C) Gene set enrichment analysis (GSEA) revealed an enrichment of Wnt/β catenin signalling involved in CKD5RAP3low tumours. Data are presented as the mean ± SEM; *, P <0.05; **, P <0.01; ***, P <0.001, two-tailed Student’s t-test were performed. NES, non-enrichment score.
- SupplementaryFigure2.pdf
Scoring criteria for CDK5RAP3 in gastric tissue. A, Tissue microarray specimens (TMAs) staining results. B, immunohistochemical (IHC) staining results. Magnification: X4 and X40. Scale bar = 400 µm.
- SupplementaryFigure3.pdf
Immunohistochemical scoring criteria for MSI under X4 magnification. Scale bar = 400 µm.
- SupplementaryFigure4.pdf
(A-B) Immunohistochemical scoring criteria for CD3CT, CD3IM, CD8CD, and CD8IM. (C) Immunoscore definition for TIL. Scale bar = 400 µm.
- SupplementaryFigure5.pdf
Heat map presenting data on CDK5RAP3, CD3, CD8, and TIL in patients from three centres.
- SupplementaryFigure6.pdf
TILhigh indicates better responsiveness to adjuvant chemotherapy in gastric cancer patients. (A, B) Overall survival analysis of adjuvant chemotherapy (ACT) in patients from the discovery set (n = 188) after TIL stratification is presented with Kaplan-Meier curves. (C, D) Overall survival analysis of ACT in patients from the internal validation set (n = 310) after TIL stratification is presented with Kaplan-Meier curves. (E, F) Overall survival analysis of ACT in patients from the external validation set 1 (n = 85) after TIL stratification is presented using Kaplan-Meier curves. (G) The proportional hazards models were established for the chemotherapy factor in the discovery set (n = 188), internal validation set (n = 310), and external validation set 1 (n = 85). P-values for all survival analyses were calculated using the log-rank test; CI indicates confidence interval; HR, hazard ratio.
- SupplementaryFigure7.pdf
Combined effect of CDK5RAP3 and TIL to observe the chemotherapy response in gastric cancer patients from external validation set 1 (n = 85). (A-D) Kaplan-Meier curves show the overall survival analysis of adjuvant chemotherapy (ACT) in patients after stratification analysis based on CDK5RAP3 and TIL. (E) Sector graph showing the number of patients in the four subgroups. (F) The proportional hazards model was established for the chemotherapy factor in external validation set 1 (n = 85). P-values for all survival analyses were calculated using the log-rank test; CI indicates confidence interval; HR, hazard ratio.
- SupplementaryFigure8.pdf
Patients with MSI-L/MSS status showed a better prognosis after chemotherapy. (A, B) Overall survival analysis of adjuvant chemotherapy (ACT) in patients from the discovery set (n = 188) after MSI status stratification is presented with Kaplan-Meier curves. (C, D) Overall survival analysis of ACT in patients from the internal validation set (n = 310) after MSI status stratification is presented with Kaplan-Meier curves. (E, F) Overall survival analysis of ACT in patients from external validation set 1 (n = 85) after MSI status stratification is presented with Kaplan-Meier curves. (G, H) Overall survival analysis of ACT in patients from external validation set 2 (n = 299) after MSI status stratification is presented with Kaplan-Meier curves. (I, J) The proportional hazards models were established for the chemotherapy factor in the discovery set (n = 188), internal validation set (n = 310), external validation set 1 (n = 85), and external validation Set 2 (n = 299). P-values for all survival analyses were calculated using the log-rank test; CI indicates confidence interval; HR, hazard ratio.
- SupplementaryFigure9.pdf
Patients in three subgroups (CKD5RAP3low MSI-H status, CKD5RAP3low MSI-L/MSS status, and CKD5RAP3high MSI-H status) showed no additional significant benefit from chemotherapy. (A-C) Kaplan-Meier curves show the overall survival analysis of adjuvant chemotherapy (ACT) in patients from the discovery set (n = 188) after stratification analysis based on CDK5RAP3 and MSI status. (D-F) Kaplan-Meier curves show the overall survival analysis of ACT in patients from the internal validation set (n = 310) after stratification analysis based on CDK5RAP3 and MSI status. (G-I) Kaplan-Meier curves show the overall survival analysis of ACT in patients from external validation set 1 (n = 85) after stratification analysis based on CDK5RAP3 and MSI status. (J-L) Kaplan Meier curves show the overall survival analysis of ACT in patients from external validation set 2 (n = 299) after stratification analysis based on CDK5RAP3 and MSI status. P-values for all survival analyses were calculated using the log-rank test.
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This is a preprint, a preliminary version of a manuscript that has not completed peer review at a journal. Research Square does not conduct peer review prior to posting preprints. The posting of a preprint on this server should not be interpreted as an endorsement of its validity or suitability for dissemination as established information or for guiding clinical practice.
Research article
Chang-ming Huang
Fujian Medical University Union Hospital
Corresponding Author
ORCiD: https://orcid.org/0000-0002-0019-885X
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
History
CURRENT STATUS: Posted
Version 1
Posted 21 Dec, 2020
No community comments so far
Metrics
Comments: 0
PDF Downloads: ...
HTML Views: ...
Subject Areas
Cancer Biology
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Background: We previously demonstrated that CDK5RAP3 acts as a tumour suppressor in gastric cancer through negative regulation of the Wnt/β-catenin signalling pathway, but its function in chemotherapeutic responsiveness of gastric cancer has not been investigated. In this study, we aimed to examine the clinical significance of CDK5RAP3 to predict chemotherapeutic responsiveness in gastric cancer.
Methods: A collection of 188 pairs of tumour tissue microarray specimens from Fujian Medical University were employed for the discovery set, and 310 tumour tissue samples of gastric cancer patients were employed for the internal validation set. Eight-five tumour tissue samples from Qinghai University Hospital were used as the external validation set 1. Transcriptomic and clinical data of 299 gastric cancer patients from TCGA were used as the external validation set 2. CDK5RAP3 expression, microsatellite instability (MSI) status, and tumour-infiltrating lymphocytes (TIL) were examined with immunohistochemistry. Clinical outcomes of patients were compared with Kaplan-Meier curves and the Cox model.
Results: In a multi-centre evaluation, increased CDK5RAP3 indication of better prognosis depends mainly on MSI-L/MSS status or TILhigh. High CDK5RAP3 expression predicts sensitive therapeutic responsiveness to postoperative adjuvant chemotherapy in gastric cancer. In a stratification analysis based on CDK5RAP3 combined with TIL or MSI status, patients with CKD5RAP3low TILlow showed no significant difference in prognosis after receiving chemotherapy, whereas patients with CKD5RAP3low TILhigh, CKD5RAP3high TILlow, and CKD5RAP3high TILhigh had better responsiveness to chemotherapy. In addition, patients with CKD5RAP3high MSI-L/MSS status benefitted the most from adjuvant chemotherapy among all patients evaluated.
Conclusions: CKD5RAP3 can be used as an effective marker to evaluate individualized chemotherapy regimens in gastric cancer patients dependent on their TIL and MSI status.
Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
This is a list of supplementary files associated with this preprint. Click to download.
- SupplementaryFigure1.pdf
Reduced CDK5RAP3 expression is observed in gastric tumour tissues, and Wnt/β-catenin signalling is upregulated in CKD5RAP3low tumours. (A) The expression of CKD5RAP3 in gastric cancer tissue and adjacent tissue from the discovery set (n = 188). (B) The transcriptomic data of CKD5RAP3 in gastric cancer tissue and normal gastric tissue from the discovery set (n = 299). (C) Gene set enrichment analysis (GSEA) revealed an enrichment of Wnt/β catenin signalling involved in CKD5RAP3low tumours. Data are presented as the mean ± SEM; *, P <0.05; **, P <0.01; ***, P <0.001, two-tailed Student’s t-test were performed. NES, non-enrichment score.
- SupplementaryFigure2.pdf
Scoring criteria for CDK5RAP3 in gastric tissue. A, Tissue microarray specimens (TMAs) staining results. B, immunohistochemical (IHC) staining results. Magnification: X4 and X40. Scale bar = 400 µm.
- SupplementaryFigure3.pdf
Immunohistochemical scoring criteria for MSI under X4 magnification. Scale bar = 400 µm.
- SupplementaryFigure4.pdf
(A-B) Immunohistochemical scoring criteria for CD3CT, CD3IM, CD8CD, and CD8IM. (C) Immunoscore definition for TIL. Scale bar = 400 µm.
- SupplementaryFigure5.pdf
Heat map presenting data on CDK5RAP3, CD3, CD8, and TIL in patients from three centres.
- SupplementaryFigure6.pdf
TILhigh indicates better responsiveness to adjuvant chemotherapy in gastric cancer patients. (A, B) Overall survival analysis of adjuvant chemotherapy (ACT) in patients from the discovery set (n = 188) after TIL stratification is presented with Kaplan-Meier curves. (C, D) Overall survival analysis of ACT in patients from the internal validation set (n = 310) after TIL stratification is presented with Kaplan-Meier curves. (E, F) Overall survival analysis of ACT in patients from the external validation set 1 (n = 85) after TIL stratification is presented using Kaplan-Meier curves. (G) The proportional hazards models were established for the chemotherapy factor in the discovery set (n = 188), internal validation set (n = 310), and external validation set 1 (n = 85). P-values for all survival analyses were calculated using the log-rank test; CI indicates confidence interval; HR, hazard ratio.
- SupplementaryFigure7.pdf
Combined effect of CDK5RAP3 and TIL to observe the chemotherapy response in gastric cancer patients from external validation set 1 (n = 85). (A-D) Kaplan-Meier curves show the overall survival analysis of adjuvant chemotherapy (ACT) in patients after stratification analysis based on CDK5RAP3 and TIL. (E) Sector graph showing the number of patients in the four subgroups. (F) The proportional hazards model was established for the chemotherapy factor in external validation set 1 (n = 85). P-values for all survival analyses were calculated using the log-rank test; CI indicates confidence interval; HR, hazard ratio.
- SupplementaryFigure8.pdf
Patients with MSI-L/MSS status showed a better prognosis after chemotherapy. (A, B) Overall survival analysis of adjuvant chemotherapy (ACT) in patients from the discovery set (n = 188) after MSI status stratification is presented with Kaplan-Meier curves. (C, D) Overall survival analysis of ACT in patients from the internal validation set (n = 310) after MSI status stratification is presented with Kaplan-Meier curves. (E, F) Overall survival analysis of ACT in patients from external validation set 1 (n = 85) after MSI status stratification is presented with Kaplan-Meier curves. (G, H) Overall survival analysis of ACT in patients from external validation set 2 (n = 299) after MSI status stratification is presented with Kaplan-Meier curves. (I, J) The proportional hazards models were established for the chemotherapy factor in the discovery set (n = 188), internal validation set (n = 310), external validation set 1 (n = 85), and external validation Set 2 (n = 299). P-values for all survival analyses were calculated using the log-rank test; CI indicates confidence interval; HR, hazard ratio.
- SupplementaryFigure9.pdf
Patients in three subgroups (CKD5RAP3low MSI-H status, CKD5RAP3low MSI-L/MSS status, and CKD5RAP3high MSI-H status) showed no additional significant benefit from chemotherapy. (A-C) Kaplan-Meier curves show the overall survival analysis of adjuvant chemotherapy (ACT) in patients from the discovery set (n = 188) after stratification analysis based on CDK5RAP3 and MSI status. (D-F) Kaplan-Meier curves show the overall survival analysis of ACT in patients from the internal validation set (n = 310) after stratification analysis based on CDK5RAP3 and MSI status. (G-I) Kaplan-Meier curves show the overall survival analysis of ACT in patients from external validation set 1 (n = 85) after stratification analysis based on CDK5RAP3 and MSI status. (J-L) Kaplan Meier curves show the overall survival analysis of ACT in patients from external validation set 2 (n = 299) after stratification analysis based on CDK5RAP3 and MSI status. P-values for all survival analyses were calculated using the log-rank test.
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