Influence of recipient KRAS gene rs712 polymorphisms on the overall survival rate of Hepatocellular carcinoma after hepatic transplantation

doi:10.21203/rs.3.rs-4717757/v1

Download PDF

Research Article

Influence of recipient KRAS gene rs712 polymorphisms on the overall survival rate of Hepatocellular carcinoma after hepatic transplantation

https://doi.org/10.21203/rs.3.rs-4717757/v1

This work is licensed under a CC BY 4.0 License

You are reading this latest preprint version

Background

Hepatocellular carcinoma (HCC) recurrence appears commonly after liver transplantation (LT), and it severely affected the long-term survival of patients. Previous studies have proved that Rap1A is involved in hepatocarcinogenesis and metastasis, and demonstrated the significant association between KRAS rs712 polymorphism and HCC. However, the relationship between KRAS rs712 polymorphism and HCC recurrence after LT remained unclear.

Methods

A total of 93 HCC patients who underwent LT from March 2008 to Dec 2015 was analyzed. The genotypes of both donors and recipients had been confirmed as KRAS rs712. The independent risk factors that associated with HCC recurrence were investigated with univariate and multivariate logistic regression analysis. The recurrence-free (RFS) and overall survival (OS) were calculated with Cox regression analysis. The KRAS rs712 genotype frequencies were determined using the Χ² test and the minor allele frequencies (MAFs) of KRAS rs712 genotypes were calculated by Hardy-Weinberg equilibrium.

Results

We found that the recipient KRAS rs712 polymorphism was significantly associated with HCC recurrence after LT. Moreover, the Milan criteria, microvascular invasion and recipient KRAS rs712 genotype were proved to be independent risk factors for HCC recurrence after LT. Patients with donor TG/TT genotypes had a significantly higher RFS and OS than TT genotype. The TNM stage, Milan criteria, microvascular invasion, Milan criteria, treatment and recipient KRAS rs712 genotype were independent factors for the RFS of LT patients.

Conclusions

Recipient KRAS rs712 polymorphism is associated with HCC recurrence after liver transplantation and plays as a promising bio-predictor of overall survival rate of HCC risks after hepatic transplantation.

hepatocellular carcinoma

KRAS polymorphism

recurrence

predictive marker

Hepatocellular carcinoma (HCC) can be known as the top 2 causes of cancer-related deaths, accounting for approximately 750,000 deaths per year globally. However, the therapeutic option is limited^[1]. Besides, about 780,000 cases are newly diagnosed each year while about 80% of patients are in Asia^[2]. The best treatment option for HCC patients is liver transplantation, but the selection of the best candidates is important to decrease the recurrence rates and prolong survival ^[3]. The Milan criteria are the most common criteria to determine HCC patients following the hepatic transplantation, for instance, microvascular invasion, tumor grades, AFP, tumor diameter, and vascular invasion^[3]. However, patients meeting the Milan criteria still have a 15–20% chance of tumor recurrence after hepatic transplantation^[4]. Some proposals have been used as an expansion of the criteria, leading to good outcomes and high tumor recurrence risks. The mutation of several key genes may affect the progression and occurrence of HCC^[5–7].

RAS accounts for about 30% of all cancers and is known as the most common mutation oncogene in human cancers^[8]. It is also seen as a therapy and vaccine target in cancer^{[9, 10]}. In the isoform of RAS (N-RAS, H-RAS, and K-RAS), KRAS is the most frequent mutation isoform. It has been acknowledged that the mutation of KRAS may lead to tumorigenesis^[11]. KRAS mutation may also cause poor prognosis, however, efficient treatment schemes have not been discovered to specifically cure the cancers expressing mutant forms of the KRAS oncoprotein^[12].

KRAS rs712 polymorphisms are proved to have correlations with gastric cancer, oral squamous cell carcinoma, colorectal cancer, and non-small-cell lung cancer^[13–17]. Previous studies have also revealed that KRAS rs712 TT genotype frequencies in HBV-positive HCC patients are higher than HBV-negative HCC patients ^[18]. However, the recurrent HCC after hepatic transplantation in the context of KRAS rs712 mutation has never been investigated yet.

Patients

From the liver transplantation surgery department, Shanghai General Hospital, 93 Chinese patients with HCC undergoing hepatic transplantation from May 2008 to June 2015 were chosen as research subjects. Patients were aged from 32 years old to 66 years old, with a median age of 48 years old. There were 82 male cases and 11 female cases. Patients were followed from 2.5 months to 113 months, with an average value of 55.5 months. Hepatic transplantation criteria for HCC patients were the absences of extrahepatic malignancies, extrahepatic metastasis of HCC, and macroscopic tumor thrombosis. Patients were cured with conventional immunosuppression plans, including triple drug combinations of prednisone, mycophenolate (MMF), tacrolimus (FK506), or cyclosporin (CsA). Hepatitis B immune globulin (HBIG) and Lamivudine were applicable to avoid the recurrence of hepatitis B for positive HBV DNA patients. Meanwhile, some patients might be treated with antineoplastic prophylaxis of targeted therapies and chemotherapy. All patients were required to inspect the tumor metastasis or recurrence through ultrasonography, AFP test, emission computed tomography, and chest X-ray every 3 months for the first two years, and then once every 6 months.

Ethical statements

Organ donation and transplantation had been approved by the Institutional Review Board, Shanghai General Hospital, confirmed to the instructions specified by the Ethics Committee of the Hospital, Declaration of Helsinki and Declaration of Istanbul. All of the patients had written the informed consent. All organs were donated voluntarily.

Data Acquisition

Clinicopathological data had been recorded, for instance, gender, hepatitis B status, age, cirrhosis, serum AFP level before hepatic transplantation, tumor size, TNM stage, microvascular invasion, multinodular status, Milan criteria, treatment schemes, and histologic grades (well, moderately and poorly differentiated). All data were acquired 24 hours before transplantation.

DNA Isolation & Polymorphism Genotyping

Hepatic transplantation patients were assigned to the HCC recurrence group and nonrecurrence control group. Prior to genotyping, the acquired samples were stored at -80^oC. The AllPrep DNA/RNA Mini kit (Qiagen, Venlos, the Netherlands) was used to isolate DNA from the recipient’s and donor’s liver tissues or EDTA-anticoagulated whole blood. PCR reaction system (10 ul) included 3.0 mM Mg2+, 1xGC buffer, 0.3 mM dNTP, 1 ul PCR primer pair, 1 ul templates DNA and 1 U HotStarTaq polymerase. The amplification primers were R: 5'-GAATCATCATCAGGAAGCCCAT-3' and F: 5'-ATGACAGTGGAAGTTTTTTTTTCCTC-3'. The PCR amplification conditions and the mass extension reactions were also prepared. More than 10% of the samples were sequenced twice to evaluate the genotyping reliability. Single nucleotide polymorphism (SNP) genotyping was implemented through Sequenom Mass ARRAY SNP genotyping platform (Sequenom, San Diego, CA) [19].

Statistical analysis

All of the analysis data were processed by R (version 4.0.3) and R Studio (version 1.4) software. Quantitative variables were expressed as the medians (±SD) and compared through Wilcoxon signed-rank test or Student’s t-test. Categorical variables were denoted as the percentage values and compared by Fisher’s exact test or Pearson’s Χ² test. The Χ² test was used to account for allele and genotype frequencies. Hardy-Weinberg equilibrium was carried out through package“GWASExactHW” in R (version 4.0.3). The Logistic regression analysis was used to study the influence factors of recurrent HCC. Kaplan-Meier analysis was used to calculate the overall survival (OS) and recurrence-free survival (RFS). Statistical significance was expressed as P < 0.05.

Patient characteristics

93 HCC patients were divided into the nonrecurrence group (n=49, 52.7%) and the recurrence group (n=44, 47.3%), according to the criteria for disease states. In the recurrence group, the follow-up lasts from 2.5 to 97 months, with a median duration of 30.4 months; and in the nonrecurrence group, it varies from 30–113 months, with the median of 78 months, with 32 deaths recorded (34.4%) (Table 1). The Student's t-test revealed a significant difference between the two groups (P<0.001), indicating a tight relationship between OS with recurrence and follow-up time ? Additionally, Milan criteria, tumor size, TNM stage, number of nodules, microvascular invasion, and antineoplastic prophylaxis after LT were all significantly correlated with recurrence (P<0.050)，but not sex, age, tumor differentiation, hepatitis B, Child-Pugh grade, cirrhosis, and pre-LT serum AFP level (Table 1).

KRAS genotype distribution and the relationship with HCC recurrence

Minor allele frequencies (MAFs) of KRAS rs712 genotypes were more than 0.01 in both recipients and donors, as determined by Hardy-Weinberg equilibrium. When donors and recipients with KRAS rs712 polymorphisms were compared, the latter showed very wide distributions in the nonrecurrence and recurrence groups, while the former did not, which can be seen in Table 2. It is critical to note that patients with the KRAS rs712 TG/GG genotype had a significantly greater incidence of recurrence after LT than those with the TT genotype (79.5% vs20.5%, P<0.001). However, the donor’s KRAS rs712 genotype had no effect on the recurrence rate of HCC after LT.

Risk factors for HCC recurrence after LT

Due to prior research indicating that the histologic grade and AFP level also correlate with the recurrence rate, the study included these two predictive variables in the logistic regression analysis. We conducted the univariate and multivariate logistic regression analysis, shown in Table 3. The Milan criteria, tumor size, TNM stage, microvascular invasion, number of nodules, and recipient KRAS rs712 genotypes are all significant factors of HCC recurrence after hepatic transplantation. To prevent collinearity, the number of nodules and tumor size were omitted, and the remaining variables did not exhibit collinearity (tolerance>0.1). TNM stage (OR=12.43, 95% CI 4.58~39.02; P<0.001), tumor size (OR=6.01, 95% CI 2.44~15.86; P<0.001), microvascular invasion (OR=17.12, 95% CI 5.19~81.75; P<0.001), number of nodules (OR=4.43, 95% CI 1.68~12.94; P=0.025), Milan criteria (OR=10.42, 95% CI 3.87~32.41; P<0.001), Treatment (OR=4.86, 95% CI 1.37~23.98; P=0.018) and recipient KRAS rs712 genotypes (OR=0.21, 95% CI 0.04~0.73; P=0.018) were related with HCC recurrence (Table 3). Additionally, multivariate analysis revealed that Milan criteria (OR=0.60, 95% CI 0.36–0.99; P=0.045) was an independent risk factor of HCC recurrence after liver transplantation.

Association between recipients rs712 polymorphisms and survival rates

According to univariate Cox regression analysis, recipients’ KRAS rs712 genotype polymorphisms, but not donors’, contributed significantly to the different recurrence in both RFS and OS. The RFS and OS of recipients bearing the donor’s TT, TG, and GG alleles were subsequently examined using Kaplan-Meier survival and the log-rank test to ascertain the impact of the rs712 polymorphism on prognosis in various recipients. Distinctions in the RFS and OS of recipients with the TT, TG, and GG genotypes were detected (P<0.001 and P<0.001, respectively; Fig. 1A and 1B). However, donor rs712 genotypes did not demonstrate an impact on OS and RFS (Supplementary Figure 1A and 1B). Simultaneously, the matching rate between donor and recipient genotype polymorphisms has no effect on the patients’ survival rate (Supplementary Figure 2A and 2B).

Risk factors for the survival rate with no recurrence and overall survival rate by Cox regression analysis

We performed multivariate Cox regression analysis on more significant factors, including histologic grade (poorly differentiated), pre-LT serum AFP level (>400ng/ml), tumor size (5cm), TNM stage (stages 3-4), Milan criteria (beyond), number of nodules (3), microvascular invasion, and donor rs712 TG/TT genotype. Tumor size and the number of nodules were excluded to prevent collinearity. There was no evidence of collinearity in the other variables (tolerance>0.1). The pre-LT serum AFP level (HR=2.400, 95% CI 1.092~5.278; P=0.020), as well as the TNM stage (HR=4.241, 95% CI 1.252~14.36; P=0.010), were deemed to be independent risk factors for RFS (Table 4). Additionally, TNM stage (HR=3.5249, 95% CI 2.221~5.594; P<0.001), tumor size (HR=2.602, 95% CI 1.645~4.114; P<0.001), microvascular invasion (HR=3.386, 95% CI 2.11~5.43; P<0.001), the Milan criteria (HR=2.654, 95% CI 1.663~4.236; P<0.001), and recipients rs712 TG/TT genotype (HR=0.397, 95% CI 0.218~0.725; P=0.002) can be seen as independent risk.

Oncogenic KRAS mutations are one of the most common types of mutations in human cancer, but they are also the most difficult to target [20]. According to a previous study, the KRAS rs712 TT genotype was substantially more prevalent in HBV-positive HCC patients than in HBV-negative HCC patients[18]. However, this research found that the non-recurrence group of HCC patients with liver transportation had a higher rate of recipient KRAS rs712 mutation. Additionally, whether or not recipients and donors are matched has little effect on the recurrence of HCC. How is it possible for a single gene to have a reversible role in the recurrence of single cancer?

The conclusion has two hypotheses: (1) Alterations in the environment’s metabolism may eventually alter the progression and occurrence of cancer [21]. KRAS-mutant cells show a glycolytic switch that is associated with increased glucose-derived metabolite channeling into the tricarboxylic acid cycle and glutathione biosynthesis, leading to improved glutathione-mediated detoxification. Carcinogenesis is primarily driven by oncogenic KRAS-downstream signaling that result in persistent proliferation [20, 22]. Mutant KRAS copy gain results in unique metabolic dependencies, which can be used to target these aggressive mutant KRAS tumors selectively [23]. The metabolic changes after LT may disrupt the sensitive metabolic environment in the liver. (2) On the other side, the recurrence of liver cancer depends upon the constitution of the liver environment and the balance between the immune system and cancer cells. The liver-resident innate immune cell may sustain a balance in the liver after LT, between pro- and anti-inflammation[24]. Thanks to the recognition of peptide epitopes presented on MHC molecules, the cytotoxic CD8+ T cell eliminates tumor cells in tumor tissue[25]. Some antigen-specific T cells with special T cell receptors (TCR) can combine with the MHC molecules on the cell surface to stimulate an anti-tumor response[26, 27]. After cancer cells are stimulated by the immune system, cancer patients may generate specific T cells that identify peptides produced from mutant KRAS variations, allowing them to develop immunological responsiveness to mutated KRAS variants [28-30]. T cells with mutant KRAS-reactive TCRs may substantially inhibit tumor development and prolong survival in patients with such malignancies [31]. T-cell responses to mutant KRAS have been shown to exist for up to 15 years [28, 29]. The process of liver transplantation will not affect the existence of T cells with KRAS-reactive TCRS. Moreover, the immune response after LT can enhance the T-cell clonal expansion[32]. These mutated KRAS-reactive TCRs staying in the peripheral blood after LT can be applied to inhibit the recurrence tendency of liver cancer induced by such mutation and improve the overall survival rate. The detailed mechanism needs further research.

The research weaknesses were evident in three areas. The first was the small sample size of patients, 91.4% of whom had HBV-induced liver disease. To receive more accurate findings, it is necessary to increase the patient sample size and include more HCC samples produced by other causes excluding HBV. Second, although we found that the variant genotype had an effect on HCC recurrence in patients after LT surgery, further investigation of the mechanism and function of KRAS rs712 in HCC recurrence is necessary. Third, the recurrence rate in our study is a high incidence compared to most of published series. The higher recurrence rate was due to samples from single center, if the amounts of samples increase, the recurrence rate may decrease. In addition, the center mainly deals with critical cases, which might be an explanation for the high recurrence rate.

In conclusion, this is the first study to examine the relationship between KRAS gene rs712 polymorphisms and post-LT HCC patients. Our findings established a positive correlation between recipient KRAS rs712 and HCC recurrence and overall survival (OS) in LT patients. Additionally, variables such as the TNM stage, Milan criteria, number of nodules, tumor size, microvascular invasion, and anticancer prophylaxis after liver transplantation can be seen as clinical prognostic indicators for liver transplantation.

HCC

Hepatocellular Carcinoma

RFS

Recurrent Free Survival

Overall Survival

liver transplantation

AFP

alpha-fetoprotein

HWE

Hardy-Weinberg equilibrium

Funding

This work was supported by the National Natural Science Foundation of China (NSFC-82300160).

Author Contribution

YL and YZ designed and conceived this project. TC and RZ performed experiments and generated data. XL, LL, ZN and HQ analyzed and interpreted data. YL made suggestions for this project. TC wrote the draft. YL and YZ revised and finalized the manuscript. All authors contributed to and approved the manuscript.

Acknowledgements

We thank for Dr. Yannan Zhao from the Department of Medical Oncology, Fudan University Shanghai Cancer Center, Shanghai Medical College, Fudan university, for technical assistance.

Disclosure Statement

The authors have no conflict of interest.

Comprehensive and Integrative Genomic Characterization of Hepatocellular Carcinoma. Cell, 2017. 169(7).
Desdín-Micó, G., G. Soto-Heredero, and M. Mittelbrunn, Mitochondrial activity in T cells. Mitochondrion, 2018. 41: p. 51–57.
Sapisochin, G. and J. Bruix, Liver transplantation for hepatocellular carcinoma: outcomes and novel surgical approaches. Nature reviews. Gastroenterology & hepatology, 2017. 14(4): p. 203–217.
Ma, E., et al., Development of a predictive nomogram for early recurrence of hepatocellular carcinoma in patients undergoing liver transplantation. Annals of translational medicine, 2021. 9(6): p. 468.
Zhang, R., et al., Donor polymorphisms of Rap1A rs494453 contribute to a higher risk of hepatocellular carcinoma recurrence following liver transplantation. Journal of Cancer, 2020. 11(10): p. 3082–3088.
Mohamed, A.A., et al., Clinical significance of SNP (rs2596542) in histocompatibility complex class I-related gene A promoter region among hepatitis C virus related hepatocellular carcinoma cases. Journal of advanced research, 2017. 8(4): p. 343–349.
Chen, D., et al., Donor interleukin 6 gene polymorphisms predict the recurrence of hepatocellular carcinoma after liver transplantation. International journal of clinical oncology, 2016. 21(6): p. 1111–1119.
Uprety, D. and A.A. Adjei, KRAS: From undruggable to a druggable Cancer Target. Cancer treatment reviews, 2020. 89: p. 102070.
Tran, E., et al., T-Cell Transfer Therapy Targeting Mutant KRAS in Cancer. The New England journal of medicine, 2016. 375(23): p. 2255–2262.
Zhang, Y., et al., Cancer vaccines: Targeting KRAS-driven cancers. Expert review of vaccines, 2020. 19(2): p. 163–173.
Waters, A.M. and C.J. Der, KRAS: The Critical Driver and Therapeutic Target for Pancreatic Cancer. Cold Spring Harbor perspectives in medicine, 2018. 8(9).
Haigis, K.M., KRAS Alleles: The Devil Is in the Detail. Trends in cancer, 2017. 3(10): p. 686–697.
Zhao, W.-H., et al., Association of rs712 polymorphism in Kras gene 3'-luntranslated region and cancer risk: a meta-analysis. Journal of B.U.ON.: official journal of the Balkan Union of Oncology, 2015. 20(1): p. 309–316.
Patricia Gallegos-Arreola, M., et al., Protective effect of rs712 polymorphism in a let-7 microRNA-binding of KRAS gene in breast cancer of a Mexican population. Journal of B.U.ON.: official journal of the Balkan Union of Oncology, 2020. 25(1): p. 176–181.
Gallegos-Arreola, M.P., et al., Association of rs712 polymorphism in a let-7 microRNA-binding site of gene with colorectal cancer in a Mexican population. Iranian journal of basic medical sciences, 2019. 22(3): p. 324–327.
Uras, I.Z., H.P. Moll, and E. Casanova, Targeting Mutant Non-Small-Cell Lung Cancer: Past, Present and Future. International journal of molecular sciences, 2020. 21(12).
Romero, R., et al., Keap1 loss promotes Kras-driven lung cancer and results in dependence on glutaminolysis. Nature medicine, 2017. 23(11): p. 1362–1368.
Xiong, D., et al., An let-7 KRAS rs712 polymorphism increases hepatocellular carcinoma risk. Genetics and molecular research: GMR, 2015. 14(4): p. 14050–14055.
Gabriel, S., L. Ziaugra, and D. Tabbaa, SNP genotyping using the Sequenom MassARRAY iPLEX platform. Current protocols in human genetics, 2009. Chapter 2: p. Unit 2.12.
Hamarsheh, S.a., et al., Immune modulatory effects of oncogenic KRAS in cancer. Nature communications, 2020. 11(1): p. 5439.
Vander Heiden, M.G. and R.J. DeBerardinis, Understanding the Intersections between Metabolism and Cancer Biology. Cell, 2017. 168(4): p. 657–669.
Raho, S., et al., KRAS-regulated glutamine metabolism requires UCP2-mediated aspartate transport to support pancreatic cancer growth. Nature metabolism, 2020. 2(12): p. 1373–1381.
Kerr, E.M., et al., Mutant Kras copy number defines metabolic reprogramming and therapeutic susceptibilities. Nature, 2016. 531(7592): p. 110–113.
Huang, H., et al., Innate Immune Cells in Immune Tolerance After Liver Transplantation. Frontiers in immunology, 2018. 9: p. 2401.
Sim, M.J.W., et al., High-affinity oligoclonal TCRs define effective adoptive T cell therapy targeting mutant KRAS-G12D. Proceedings of the National Academy of Sciences of the United States of America, 2020. 117(23): p. 12826–12835.
He, Q., et al., Targeting cancers through TCR-peptide/MHC interactions. Journal of hematology & oncology, 2019. 12(1): p. 139.
Todo, S., et al., A pilot study of operational tolerance with a regulatory T-cell-based cell therapy in living donor liver transplantation. Hepatology (Baltimore, Md.), 2016. 64(2): p. 632–643.
Linard, B., et al., A ras-mutated peptide targeted by CTL infiltrating a human melanoma lesion. Journal of immunology (Baltimore, Md.: 1950), 2002. 168(9): p. 4802–4808.
Kubuschok, B., et al., Naturally occurring T-cell response against mutated p21 ras oncoprotein in pancreatic cancer. Clinical cancer research: an official journal of the American Association for Cancer Research, 2006. 12(4): p. 1365–1372.
Gjertsen, M.K., et al., HLA-A3 restricted mutant ras specific cytotoxic T-lymphocytes induced by vaccination with T-helper epitopes. Journal of molecular medicine (Berlin, Germany), 2003. 81(1): p. 43–50.
Wang, Q.J., et al., Identification of T-cell Receptors Targeting KRAS-Mutated Human Tumors. Cancer immunology research, 2016. 4(3): p. 204–214.
Dhanasekaran, R., Management of Immunosuppression in Liver Transplantation. Clinics in liver disease, 2017. 21(2): p. 337–353.

Table 1: Clinical characteristics of patients with and without HCC recurrence
Parameter	Recurrence (n=44)	Nonrecurrence (n=49)	P value
Recipient age	44 (33-64)	49 (32-66)	0.207
Recipient sex
Male	39(88.6%)	43(87.8%)
Female	5(11.4%)	6(12.2%)	0.897
Follow up time(month)	30.4 (2.5-97)	78.0 (30-113)
Hepatitis B
Yes	39(88.6%)	46(93.9%)
No	5(11.4%)	3(0.061%)	0.374
Cirrhosis
Yes	33(75%)	43(87.8%)
No	11(25%)	6(12.2%)	0.114
Child-Pugh grading
A	25(56.8%)	32(65.3%)
B+C	19(43.2%)	17(34.7%)	0.407
Histologic grade
Well+moderately	39(88.6%)	45(91.8%)
Poorly	5(11.4%)	4(8.2%)	0.607
Milan criteria
Within	6(13.6%)	31(63.3%)
Beyond	38(86.4%)	18(36.7%)	<0.001
TNM stage
I~II	6(13.6%)	33(67.3%)
III~IV	38(86.4%)	16(32.7%)	<0.001
Tumor size(cm)
<5	15(35.7%)	39(79.6%)
≥5	27(64.3%)	10(20.4%)	<0.001
Number of nodules
<3	25(56.8%)	40(81.6%)
≥3	19(43.2%)	9(18.4%)	<0.050
Microvascular invasion
Yes	24(54.5%)	3(6.1%)
No	20(45.5%)	46(93.9%)	<0.001
Pre-LT serum AFP level
≤400(ng/ml)	35(79.5%)	32(65.3%)
>400(ng/ml)	9(20.5%)	17(34.7%)	0.129
Antineoplastic prophylaxis after LT
Yes	11(25%)	3(6.1%)
No	33(75%)	46(93.9%)	<0.050

Table2: Donor and Recipient KRAS genotype distribution and the association with HCC recurrence
	Genotype distribution, n (%)		P value	HWE value
	Recurrence(n=44)	Nonrecurrence(n=49)	P value	HWE value
Recipient rs712
GG	11(25%)	3(6.1%)		0.180
TG	24(54.5%)	15(30.6%)	0.248
TT	9(20.5%)	31(63.3%)	<0.001
Dominant model
GG	11(25%)	3(6.1%)
TG/TT	33(75%)	46(93.9%)	<0.050
Recessive model
TT	9(20.5%)	31(63.3%)
TG/GG	35(79.5%)	18(36.7%)	<0.001
Additive model
TG	24(54.5%)	15(30.6%)
GG/TT	20(45.5%)	34(69.4%)	<0.050
Donor rs712
GG	25(56.8%)	27(55.1%)		0.084
TG	15(34.1%)	17(34.7%)	0.915
TT	4(9.1%)	5(10.2%)	0.840
Dominant model
GG	25(56.8%)	27(55.1%)
TG/TT	19(43.2%)	22(44.9%)	0.868
Recessive model
TT	4(9.1%)	5(10.2%)
TG/GG	40(90.9%)	44(89.8%)	0.856
Additive model
TG	15(34.1%)	17(34.7%)
GG/TT	29(65.9%)	32(65.3%)	0.951

Table3: Univariate and Multivariate logistic regression analysis of risk factors associated with HCC recurrence after LT
Variables	Univariate analysis		Multivariate analysis
	Odds ratio(95%CI)	P value	Odds ratio(95%CI)	P value
Recipient age(0 ≤50,1>50)	0.58(0.23~1.42)	0.270
Recipient sex (0 female,1 male)	1.08(0.29~ 4.16)	1.000
Hepatitis B(0 No, 1 Yes)	0.52(0.10~2.36)	0.470
Cirrhosis(0 No,1 Yes)	0.43(0.13~1.26)	0.180
Child-Pugh grade (0 Grade A, 1 Grade B+C)	1.42(0.61~3.34)	0.520
Histologic grade (0 well+moderately, 1 poorly)	1.43(0.34~6.40)	0.730
Pre-LT serum AFP level (0 ≤400ng/ml,1>400ng/ml)	0.59(0.22~1.52)	0.340
TNM stage(0 stageI-II, 1 stageIII-IV)	12.43(4.58~39.02)	< 0.001	0.58(0.33~1.02)	0.058
Tumor size(cm) (0<5, 1 ≥5)	6.01(2.44~15.86)	< 0.001
Number of nodules (0<3, 1 ≥3)	4.43(1.68~12.94)	0.003
Microvascular invasion (0 No, 1 Yes)	17.12(5.19~81.75)	< 0.001	0.61(0.33~1.10)	0.102
Milan criteria (0 within, 1 beyond)	10.42(3.87~32.41)	< 0.001	0.60(0.36~0.99)	0.047
Treatment(0 No, 1 Yes)	4.86(1.37~23.98)	0.018	1.13(0.73~1.74)	0.584
Recpicant KRAS rs712 genotype(0 GG,1 TG/TT)	0.21(0.04~0.73)	0.018	1.59(0.84~3.02)	0.158
Donar KRAS rs712 genotype (0 GG,1 TG/TT)	1(0.41~2.14)	1.000

Table4: Cox regression analysis of the risk factors for recurrence-free survival time
Variables	Univariate analysis		Multivariate analysis
	Hazard ratio(95% CI)	P value	Hazard ratio(95% CI)	P value
Pre-LT serum AFP level (0 ≤400ng/ml,1>400ng/ml)	2.40(1.09~5.28)	0.020	0.80(0.46~1.40)	0.442
TNM stage(0 stageI-II, 1 stageIII-IV)	4.24(1.25~14.36)	0.010	3.99(2.41~6.61)	6.90E-08
Tumor size(cm) (0<5, 1 ≥5)	2.19(1.14~4.20)	0.020
Number of nodules (0<3, 1 ≥3)	1.32(0.71~2.44)	0.400
Microvascular invasion(0 No, 1 Yes)	1.46(0.77~2.74)	0.200
Milan criteria(0 within, 1 beyond)	2.23(0.77~6.49)	0.100
Treatment(0 No, 1 Yes)	0.92(0.46~1.84)	0.800
Recpicant KRAS rs712 genotype(0 GG,1 TG/TT)	0.62(0.30~1.27)	0.200
Donar KRAS rs712 genotype(0 GG,1 TG/TT)	1.06(0.70~1.61)	0.800
Matched KRAS rs712 genotype(0 Unpaired, 1 paired)	1.24(0.82~1.88)	0.300

Table5: Cox regression analysis of the risk factors for overall survival time
Variables	Univariate analysis		Multivariate analysis
	Hazard ratio (95% CI)	P value	Odds ratio(95%CI)	P value
Pre-LT serum AFP level (0 ≤400ng/ml,1>400ng/ml)	0.60(0.35~1.02)	0.060
TNM stage(0 stageI-II, 1 stageIII-IV)	3.52(2.22~5.59)	< 0.001	2.16(1.40~4.08)	0.021
Tumor size(cm) (0<5, 1 ≥5)	2.60(1.65~4.11)	< 0.001
Number of nodules (0<3, 1 ≥3)	2.09(1.31~3.33)	0.002
Microvascular invasion(0 No, 1 Yes)	3.39(2.11~5.43)	< 0.001	2.23(1.19~4.20)	0.008
Milan criteria(0 within, 1 beyond)	2.65(1.66~4.24)	< 0.001	1.21(0.62~2.34)	0.726
Treatment(0 No, 1 Yes)	1.30(0.73~2.34)	0.400
Recpicant KRAS rs712 genotype(0 GG,1 TG/TT)	0.40(0.212~0.73)	0.002	0.32(0.15~0.66)
Donar KRAS rs712 genotype(0 GG,1 TG/TT)	0.95(0.63~1.44)	0.800
Matched KRAS rs712 genotype(0 Unpaired, 1 paired)	1.10(0.73~1.68)	0.600

No competing interests reported.

Download PDF

Reviewers invited by journal
15 Jul, 2024
Editor assigned by journal
13 Jul, 2024
Submission checks completed at journal
13 Jul, 2024
First submitted to journal
10 Jul, 2024

You are reading this latest preprint version

Influence of recipient KRAS gene rs712 polymorphisms on the overall survival rate of Hepatocellular carcinoma after hepatic transplantation

Status:

Version 1

Abstract

Background

Methods

Results

Conclusions

Figures

Introduction

Materials and Methods

Results

Discussion

Abbreviations

Declarations

Funding

Author Contribution

Acknowledgements

Disclosure Statement

References

Tables

Additional Declarations

Supplementary Files

Status:

Version 1