- Email: [email protected]
The prevalence of HLA alleles in a lupus nephritis population
Accepted Manuscript The prevalence of HLA alleles in a lupus nephritis population
Maria Izabel de Holanda, Evandro Klumb, Alicia Imada, Livia A. Lima, Isabela Alcântara, Flavia Gregório, Luis Fernando Christiani, Clarice Oliveira Martins, Branca Engel Timoner, Juliana Motta, Roberto Pozzan, Luis Cristóvão Pôrto PII: DOI: Reference:
S0966-3274(17)30090-4 doi:10.1016/j.trim.2018.02.001 TRIM 1125
To appear in:
Transplant Immunology
Received date: Revised date: Accepted date:
29 June 2017 30 January 2018 1 February 2018
Please cite this article as: Maria Izabel de Holanda, Evandro Klumb, Alicia Imada, Livia A. Lima, Isabela Alcântara, Flavia Gregório, Luis Fernando Christiani, Clarice Oliveira Martins, Branca Engel Timoner, Juliana Motta, Roberto Pozzan, Luis Cristóvão Pôrto , The prevalence of HLA alleles in a lupus nephritis population. The address for the corresponding author was captured as affiliation for all authors. Please check if appropriate. Trim(2018), doi:10.1016/j.trim.2018.02.001
This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
ACCEPTED MANUSCRIPT THE PREVALENCE OF HLA ALLELES IN A LUPUS NEPHRITIS POPULATION
Maria Izabel de Holanda 1, Evandro Klumb2, Alicia Imada 1, Livia A Lima 1, Isabela Alcântara 1, Flavia Gregório1, Luis Fernando Christiani 1, Clarice Oliveira Martins 2, Branca Engel Timoner3, Juliana Motta 3, Roberto Pozzan4, Luis Cristóvão Pôrto3
1 - Nephrology Service, Hospital Federal de Bonsucesso
RI
2 – Rheumatology Service, Rio de Janeiro State University
PT
Affiliations
SC
3 - Histocompatibility and Cryopreservation Laboratory, Rio de Janeiro State University 4 – Cardiology Service, Rio de Janeiro State University
NU
Corresponding Author Prof Luís Cristóvão Pôrto
Pav. Jose Roberto Feresin Moares Av. Marechal Rondon 381
MA
Laboratório de Histocompatibilidade e Criopreservação
ED
São Francisco Xavier 20950-003
Brazil
EP T
Rio de Janeiro
Tel +552123342426
AC C
[email protected]
Author Contributions
MIH – designed the study, followed up with patients, collected samples, analyzed the results and wrote the manuscript. AI, LAL, IA FG and LFC - followed up and retrieved historical records of transplanted patients. EK and COM followed up with lupus nontransplanted patients. BET and JM - performed histocompatibility tests. RP – performed statistical test analyses and interpreted results. LCP - designed the study, analyzed results and revised the manuscript. Conflict of Interest The authors declare no conflicts of interest.
ACCEPTED MANUSCRIPT Abstract Background: Systemic lupus erythematosus (SLE) is a severe autoimmune disease that involves multiple organ systems. Lupus nephritis (LN) is a complication of SLE and is associated with poor survival and high morbidity. Many genomic studies have been performed worldwide, and several histocompatibility leukocyte antigen (HLA) loci are
PT
linked to lupus susceptibility.
RI
Objective: The present study evaluated the association of HLA alleles in a lupus patient
SC
population, LN group and control group. The second objective evaluated whether HLA allele match or mismatch influenced kidney graft survival in a kidney transplanted
NU
lupus population.
Methods: This study was a retrospective study of 2 major groups: general lupus
MA
patients (GSLE - n=108) and a control group (GControl - n=216). Both groups were also divided into subgroups.
ED
Results: The control group was divided into two subgroups: a healthy control group
EP T
(HeCTRL) and transplant control group (TxCTRL). The GSLE group was composed of transplanted lupus patients (TxSLE) and non-transplanted lupus patients (nTxSLE).
AC C
Comparison of the demographics between groups did not reveal differences between ethnicity and gender. A difference in the prevalence of three alleles, B*08, DRB1*08 and DRB1*15, was observed. These alleles were more prevalent in the lupus subgroups compared to the control groups. Five-year survival was not different between patients carrying the allele DRB1*15 in either group (overall p=0.075; TxSLE p=0.419; TxCTRL=0.309). The presence of the match with this allele in the receptor was evaluated and did not demonstrate any difference in graft survival in both groups (p=0.146) or when analyzed separately in each group (TxCTRL p=0.739; TxSLE= 0.297).
ACCEPTED MANUSCRIPT Conclusion: This study demonstrated that the presence of HLA-DRB1*15 was a strong factor that predisposed patients to the development of SLE and LN, but did not
AC C
EP T
ED
MA
NU
SC
RI
PT
influence kidney graft survival.
ACCEPTED MANUSCRIPT BACKGROUND Systemic lupus erythematosus (SLE) is a severe autoimmune disease that involves multiple organ systems. SLE involves the production of autoreactive T cells and autoantibodies against nuclear, cytoplasmic and cell-surface antigens. The more
PT
common clinical features of SLE include skin and joint diseases, hematological abnormalities, renal disease and neuropsychiatric complications (Ghodke-Puranik and
RI
Niewold 2015).
SC
Lupus nephritis (LN) is one complication of SLE, and it is associated with poor survival
NU
and high morbidity, particularly in patients who develop end-stage renal disease (ESRD) (Hanly et al. 2015). The cause of LN is complex, and environmental and genetic
MA
factors are involved (Alarcon-Segovia et al. 2005). Monozygotic twins exhibit an approximately 10-fold higher risk for SLE than dizygotic twins (Block et al. 1975), and
ED
first-degree relatives of patients with SLE exhibit a 20-fold increased risk of developing
EP T
SLE compared to the healthy population (Alarcon-Segovia et al. 2005). The major histocompatibility complex (MHC) is located on the short arm of
AC C
chromosome 6, and its relationship to autoimmune diseases was studied extensively (Graham et al. 2007; Wahren-Herlenius and Dorner 2013). Approximately 421 genes are related to the MHC; 60% of these genes are expressed, and 22% are related to immunoregulatory functions (Fernando et al. 2008). Many worldwide genomic studies were performed, and several histocompatibility leukocyte antigen (HLA) loci were linked to the lupus susceptibility (Chung et al. 2014; Grumet et al. 1971; Morris et al. 2014). A recent meta-analysis, Niu et al (2015) suggested that distinct HLA Class II genetic variants conferred a predisposition and resistance to SLE and LN. HLA-DR4,
ACCEPTED MANUSCRIPT DR5, DR11 and DR14 were identified as protective against SLE, and HLA-DR3, DR9 and DR15 may be risk factors for SLE. The DR4 and DR11 alleles may be protective for LN, and DR3 and DR15 were risk factors (Niu et al. 2015). HLA class I and II genes were consistently associated with SLE susceptibility, especially
PT
B*18 and DRB1*15 (Cortes et al. 2004; Lin et al. 2016; Marchini et al. 2003; Segurado et al. 1991; Shimane et al. 2013; Suggs et al. 2011; Tian et al. 2000). This susceptibility
RI
may also be related to disease severity.
SC
The present study evaluated the association of HLA alleles in a lupus patient
NU
population, LN group and control group. The study also evaluated whether matched or mismatched HLA alleles influenced kidney graft survival in a kidney-transplanted lupus
AC C
EP T
ED
MA
population.
ACCEPTED MANUSCRIPT MATERIALS AND METHODS Study Population This study was a retrospective study of 2 major groups: general lupus patients (GSLE n=108) and a control group (GControl - n=216). Both groups included patients and
PT
controls with specific criteria (see below). Lupus Group
RI
The GSLE was composed of kidney-transplanted lupus patients (TxSLE) from the
SC
Bonsucesso Federal Hospital, Rio de Janeiro and non-transplanted lupus patients from
NU
the Rheumatology Department of Rio de Janeiro State University (nTxSLE), Rio de Janeiro, Brazil. Patients in both groups were matched by gender and ethnicity (white
MA
and non-white). All patients met the American College of Rheumatology classification criteria for the diagnosis of SLE (Hahn et al. 2012).
ED
Patients in the TxSLE group (n=44) received a kidney transplant during 1980-2016.
EP T
These patients had HLA typings before transplant as part of the pre-transplant routine. LN patients (LNSLE) from both health centers were subgrouped into patients who
AC C
presented with biopsy-proven LN class III/IV or V from the nTxSLE (n=32), considering that proliferative nephritis has a worse prognosis and 30% in 10 years will need dialysis (Mok 2016; Mok et al. 2006), and TxSLE groups (n=44, Figure 1). Control Group
The control groups were formed on a 2:1 proportion and matched by gender and ethnicity (white and non-white: including black and mestizos). This group was composed of 128 healthy people (HeCTRL) from the control group of a previous rheumatic arthritis study (Usnayo et al. 2011) and patients who received a kidney
ACCEPTED MANUSCRIPT transplant for other diseases in the Bonsucesso Federal Hospital (TxCTRL - n=88) during the same period. The control group of two different samples created uniformity of this group with the GSLE group. The control group (Gcontrol) was 216 individuals (Figure 1).
PT
HLA HLA class II-DR typing was performed using serology or polymerase chain reaction
RI
amplification with sequence-specific primers. HLA from the kidney-transplanted
SC
patients were collected from medical charts, and the antigens defined in the serology
NU
were converted to the most frequent possible allele according to the frequency of haplotypes and alleles in the Rio de Janeiro population (www.imunogenetica.org).
MA
All patients from the Rheumatology Department were invited to participate in the study, and participants signed a written informed consent form. Kidney-transplanted
ED
patients did not need to sign a consent form because consent was part of their pre-
EP T
transplant routine, and all data were collected after transplantation. This study was performed in accordance with standards of our hospital’s Ethics
AC C
Committees (142/09). Statistical Analysis:
The genetic distribution of HLA was performed using the Arlequin program. We compared all four groups initially, then performed two by two group comparisons: GSLE versus the GControl, followed by the LNSLE versus nLNSLE groups, and TxSLE versus the TxCTRL groups. We also analyzed the prevalence of HLA alleles in the LNspecific population compared to the nLNSLE and Gcontrol groups.
ACCEPTED MANUSCRIPT Categorical data are described using numbers and percentages. The frequencies of gender, ethnicity, and age were performed via direct counting. Haplotypes HLAA~B~DRB1 were generated using the expectation–maximization with Arlequin 3.5 (Excoffier and Lischer 2010).Data were analyzed using and EPI-INFO version 3.5.3 (CDC, Atlanta,
GA, USA) and SPSS version 22.0. Comparisons of demographic data and HLA allele
PT
frequencies between groups were performed using the chi-square and Fisher’s exact
RI
test. P<0.05 was considered statistically significant. Significant results in the HLA allele
SC
frequencies were multiplied by the total number of alleles found per locus (Locus A – 20, Locus B – 39 and Locus DRB1 – 13) to obtain a corrected value of P, termed Pc
NU
(Bonferroni correction). The T-test compared the mean ages between groups and
MA
different outcomes.
Survival analysis was performed using the Kaplan-Meier method. P values <0,05 were
ED
considered significant.
Demographics
EP T
RESULTS
AC C
The control group was divided into two groups: the health control group (HeCTRL) and the transplant control group (TxCTRL). The HeCTRL was composed of 111 women and 17 men (50 whites and 78 non white), and the mean age was 43.2 years (range 28-59 years). The TxCTRL group had 88 patients: 76 women and 43 whites, and the mean age was 39.5 years (range 11-71 years). The prevalence of underlying diseases in the TxCTRL group was 46 hypertension patients: 12 glomerulosclerosis, 11 polycystic kidney disease, 3 diabetes, 2 eclampsias and 14 presented with urological affections.
ACCEPTED MANUSCRIPT There was a total of 2,326 patients who received kidney transplants from 1980 to 2016 in the Bonsucesso Federal Hospital transplant center, and 56 patients were identified with LN. Twelve patients did not present a complete loci A-B-DRB1HLA typing or were lacking in information. The TxSLE group was comprised of 38 female patients, 24
PT
patients were white, and the mean age was 33 years (range 12-56 years). The nTxSLE group was comprised of 54 female patients, 23 patients were white, the mean age was
RI
37.5 years (range 20-59 years), and nephritis was present in 22 patients. Patients from
SC
these two group who did not present nephritis were sub-grouped in the nLNSLE group (n=44, Table 1). Comparison of demographics between groups revealed no differences
NU
in ethnicity or gender. The groups were homogeneous.
MA
Thirteen patients in nTxSLE exhibited serositis, nine patients had neurological involvement, 37 patients exhibited arthritis, 29 patients exhibited dermatological
ED
lesions, and 32 patients exhibited LN.
EP T
Transplant Patients Demographics and Characteristics The TxSLE group consisted of 44 patients of whom 29 were transplant recipients with live donors, 15 from deceased donors, 20 did not receive induction therapy, 18
AC C
induced with simulect® and five with thymoglobulin® and one with OKT3. Regarding previous transfusions, nine patients did not receive transfusions prior to transplantation, eight received one, six received two, nine received three, and twelve had several blood transfusions. The TxCTRL group, consisting of 88 patients, on induction therapy, forty were not transfused, 42 did simulect® and 6 did thymoglobulin®. Forty-nine patients in the TxCTRL group had previous transfusions, 13 had one blood transfusion, 12 had 2 transfusions, 6 had 3 transfusions, and 17 had multiple transfusions. Immunosuppression was based on a calcineurin and
ACCEPTED MANUSCRIPT corticosteroid inhibitor in both groups, but in the TxSLE group there were 11 patients using azathioprine associated with this scheme and in the TxCTRL 20 group, comparing the groups there was no statistical difference (p = 0.347). Regarding the type of renal replacement therapy (RRT) prior to transplantation, no
PT
difference was also observed between the groups. In the TxCTRL group, 3 patients underwent preemptive transplantation, 74 underwent hemodialysis (HD), 5 under
RI
peritoneal dialysis (PD) and 6 underwent both therapies. In the TxSLE 37 group they
SC
did HD before, one patient had PD and 6 did the two therapies. The mean time to RRT
NU
before transplantation was 50 months (approximately 4 years and 2 months, range 0172 months).
MA
Patients with acute rejection (RA) were those who presented renal biopsy confirming the diagnosis according to the Banff classification. Patients who had increased
ED
creatinine and / or decreased diuresis, and / or other signs of RA who underwent
clinical RA.
EP T
methylprednisolone pulse and presented response after therapy were classified with
AC C
Twenty-five (18.9%) patients had episodes of acute rejection (RA), 18 (20.4%) patients in the TxCTRL group and 7 (15.9%) patients in the TxSLE group. Only 10 patients underwent renal biopsy, five patients had acute humoral rejection, 1 in the TxSLE group and the remainder in the TxCTRL group, there was no difference between the groups (p = 0.353). The time of diagnosis of AR episodes ranged from 0-1752 days from the date of transplantation, and 16 patients had episodes within the first month after transplantation.
ACCEPTED MANUSCRIPT Twenty patients had viral infections, 12 patients in the TxCTRL group, two herpes virus infections and the remaining cytomegalovirus (CMV) in the TxSLE group were 8 patients with viral infections, one for herpes and 7 for CMV. There was no difference between groups (p = 0.763). Nine patients had these infections in the first year of
PT
transplantation. Thirty-one patients had bacterial infections, 53 of them were infectious, some of them had more than 3 episodes. The most frequent infection was
RI
the urinary tract infection. Patients in the TxCTRL group had more infections compared
SC
to the TxSLE group, 26 patients versus 5 TxSLE patients (p = 0.048).
NU
Locus frequency
Comparisons of pairs of population samples demonstrated that GLSE was genetically
MA
distant from Gcontrol (FST - 0.00409, p <0.05) and nTxSLE and TxSLE were genetically separate from HeControl (FST -0.00463 and 0.00425, respectively). Comparison of the
ED
allele genetic frequency between GSLE and Gcontrol revealed no differences between these groups in the A locus (p=0.914) or B locus (p=0.072). However, frequency in the
EP T
DRB1 locus was significantly different (p=0.0003). Comparison of the four subgroups also revealed differences in the DRB1 locus (p=0.0009), but the A and B loci were not
AC C
statistically different (p=0.852, p=0.113, respectively; Table 1S). Locus B was generally in pairwise linkage disequilibrium with loci A and DRB1, except in TxCTRL, in which locus B exhibited linkage disequilibrium with locus A. Hardy-Weinberg equilibrium was only absent for locus A in the TxCTRL subgroup (observed heterogeneity 0.78409, p=0. 0.034). Seventy-two possible HLA-A~B~DRB1 haplotypes were determined from the EM algorithm for the TxLSE group, and 96 of these haplotypes were determined for the
ACCEPTED MANUSCRIPT nTxSLE group. Twelve haplotypes were observed in at least two patients in the TxSLE group and in 23 patients in the nTxSLE. Sixteen of these nTxSLE haplotypes were not observed in the other subgroups, and 8 were not observed in the TxSLE group (Table 2S).
PT
Allele Frequencies The first analysis compared the GSLE (n=128) and the Control group (n=216). This
RI
analysis revealed eight alleles: HLA-B*08, -B*15, -B*27, -B*44, -DRB1*08, -DRB1*11, -
SC
DRB1*13 and -DRB1*15. The most prevalent alleles in the GSLE were -B*08, -DRB1*08
NU
and -DRB1*15. The other alleles were protective for lupus, primarily HLA B*27. No patients in the GSLE group presented these alleles. HLA DRB1*15 was the only allele
MA
that remained statistically significant after Bonferroni correction (Table 2). The four described subgroups were compared (Table 3). A difference in the prevalence
ED
of three alleles, HLA-B*08, -DRB1*08 and -DRB1*15, was demonstrated. These alleles
EP T
were more prevalent in the lupus subgroups than the control groups. No allele remained significant after Bonferroni correction. However, the HLA DRB1*15 was near
AC C
significant (p-value=0.0533).
Comparison between the LNSLE and nLNSLE group revealed a higher prevalence in HLA-A*02 (48.6% x 25.0%). The Alleles A*03, A*33, A*74 and DRB1*04 were more prevalent in the lupus without nephritis group (nLNSLE – Table 3S). However, no difference between groups was found after Bonferroni correction. The HLA DRB1*15 was present in 39.47% of patients in the LNSLE group and 46.88% in the nLNSLE group (p=0.307). We also examined whether a specific allele was associated with SLE without nephritis. The alleles with higher prevalence in the comparison of the nLNSLE with the
ACCEPTED MANUSCRIPT HeControl groups were HLA-A*02, -DRB1*11, -DRB1*13 in the control group and HLA B*07, -B*08, -B*15, -DRB1*08, and -DRB1*15 in the lupus group (Table 4S). We compared the TxCTRL group (n=88) with the TxSLE group (n=44). The results revealed a higher prevalence of alleles HLA-A*30, -B*18, -B*49, -DRB1*07 and -
PT
DRB1*15 in the TxSLE group. However, no alleles were statistically significant after Bonferroni correction (Table 4). The importance of the prevalence of HLA DRB1*15 in
RI
SLE was demonstrated in the literature. The association and link of some alleles, such
SC
as A*25~B*18~DRB1*15, is also well described in the literature. We evaluated this association in our population and only found this haplotype in the nLNSLE group (Table
NU
2S).
MA
The comparison between both control groups revealed no significant differences in allele prevalence. The prevalence of HLA-DRB1*15 was also analyzed by comparing
ED
ethnicity, and there were no differences between whites (27%) and non-whites (36%) (p=0.343).
EP T
Kidney Graft Survival and HLA DRB1*15 The overall 5-year kidney graft survival rate with death censored was 73.5% These
AC C
rates were 90.9% and 64.8% in the TxSLE and TxCTRL groups, respectively (p=0.05). We did not match for age in the control groups, and the TxCTRL group was older. Therefore, we used a t-test to evaluate whether age was a factor for worst outcome. No differences were found in age means and the outcome, and the mean age for patients with graft loss in five years was 37.9 years (sd 12.6) versus 37.2 years (sd13.4) (p=0.316) for patients who did not lose the graft. There was no difference between groups in donor type or deceased or live donors (p=0.350). Considering the possibility of the underlying disease being an influence factor in graft loss, we divided the control
ACCEPTED MANUSCRIPT group into hypertensive patients and with other diseases and compared them with the patients in the lupus group. We then compared 3 groups: Lupus, SAH and other diseases. The hypertensive patients had a cured survival rate of 54.2%, compared to 77.5% for other diseases and 90.2% for lupus (p = 0.105).
PT
When we evaluated the 5-year survival curve in relation to ethnicity, we observed a higher survival rate in whites (76.2%) vs. non-whites (55.1%; p = 0.008), who were
RI
censored for death, survival rate for whites was 85.7% and non-whites 62.3% (p =
SC
0.001). In the lupus group, the result found was 75% survival for non-whites and 90% for whites, although it was not significant (p = 0.144). However, when we censored for
NU
death, survival for whites was 100% and for non-whites, 83.3% (p = 0.040). There is a
MA
trend of worse survival for non-whites. The same happened in the control group, 65.1% survival for whites versus 44.4% for nonwhites (p = 0.047).
ED
Five-year survival was not different between patients carrying the DRB1*15 allele overall (p=0.075) or in TxSLE (p=0.419) and TxCTRL (p=0.304) groups or when both
EP T
groups were compared (p=0.059; Figure2). The presence of the match with this allele in the receptor was evaluated, but no difference in graft survival comparison of both
AC C
groups was demonstrated (p=0.075, Figure 3) or when this allele was analyzed separately in each group and overall (TxCTRL p=0.738; TxSLE p=0.297; overall p=0.275). The correlation between ethnicity, HLA-DRB1*15 and graft survival in the groups was also not significantly different (p=0.104). DISCUSSION The present study demonstrated the prevalence of HLA alleles in a Brazilian lupus population. We compared the prevalence to a healthy population and a control group
ACCEPTED MANUSCRIPT of kidney-transplanted patients with other diseases. Most published studies limited comparisons to a healthy population (Lin et al. 2016; Liphaus et al. 2007). The alleles HLA-B*08, -DRB1*08 and -DRB1*15 exhibited a higher prevalence in our lupus population. One of the limitations of our study was the low number of patients
PT
in the TxSLE group. HLA allele distribution in a population is linked to ethnicity, which presents variability in the determination of color-race in the Brazilian population(IBGE
RI
2008). Our groups were classified into white and non-white. This simplification may
SC
cause a bias in specific alleles that are over represented in Sub-Saharan Africans
NU
(blacks), such as HLA-A*30. HLA-A*30 is found in Brazilian black individuals on the Bone Marrow Donor Registry with a genetic frequency of 0.0804 compared to the
MA
white individuals (frequency of 0.0426) (www.imunogenetica.org). The allelic frequency in TxSLE patients also confirmed the important contribution of non-white
ED
ethnicity as a specific risk for lupus in this group. HLA-DRB1*08 frequency in the subgroups revealed an ethnic influence, and this allele was also more frequent in
EP T
Brazilian white individuals. The odds ratio in association with LN increased more than
AC C
double (3.68 x 8.00).
Our results are consistent with Liphaus et al. (Liphaus et al. 2002), who described a higher prevalence of HLA DR15 that was influenced by gender (female) and ethnicity (black individuals). This difference was not observed in our samples because we paired controls by gender and ethnicity. Araujo et al.(Araujo et al. 1997) also evaluated a Brazilian lupus population with C2 deficiency and demonstrated that patients with HLA-B*18 were more prevalent (55.6%) with a relative risk of 9.2 compared to the healthy population. HLA-B*18 was
ACCEPTED MANUSCRIPT also very prevalent in the families of patients with the same phenotypes. Our study demonstrated that the associations between HLA-DRB1*15 and HLA-B*18 were linked with -A*25 in the nTxSLE group (Table 2S). These results suggest that the alleles may be a risk factor for other manifestations of lupus because this haplotype HLA-
PT
A*25~B*18~DRB1*15 (Araujo et al. 1997) was not observed in the LNSLE group. HLAB*18~DRB1*15 was associated with other alleles of Locus A in the HeControl group,
RI
twice with A*24 and once with A*26 in the TxSLE group.
SC
Few studies in the literature evaluated the prevalence of HLA and lupus nephritis, and
NU
our study compared 2 nephritis population (with or without transplantation) to two different controls (transplant/others disease control and a healthy population). A
MA
genome-wide association study of LN demonstrated an association between LN and antigens HLA-DR2 and HLA-DR3. However, other genes outside the MHC were more
ED
evident than this correlation in this same study (Chung et al. 2014).
EP T
An LN sample evaluated the existence of a higher prevalence of specific alleles in patients with compromised renal function. However, it was not possible to
AC C
demonstrate a specific allele more prevalent in LN population. To our knowledge, this study is the first study to evaluate the prevalence of HLA in LN in a Brazilian population.
A recent meta-analysis demonstrated an association of lupus nephritis with DR3 and DR15 antigens, and DR14 and DR11 were protective genotypes for lupus nephritis (Niu et al. 2015). A Latin American meta-analysis also demonstrated a higher susceptibility for SLE with antigens DR2-DR3 and allele HLA-DRB1*15:01 (Castano-Rodriguez et al. 2008).
ACCEPTED MANUSCRIPT A Moroccan study in a lupus nephritis population demonstrated the prevalence of the HLA-DR15 antigen was also higher compared to the normal population, and the odds ratio for this population was 2.8 (Bhallil et al. 2017). Our positive result of HLA-DRB1*15 was also observed in Italian (Marchini et al. 2003),
PT
European (Morris et al. 2012), Brazilian (Fernandes et al. 1995), Japanese (Shimane et al. 2013) and Saudi populations (Wadi et al. 2014). Cortes et al. (Cortes et al. 2004) also
RI
demonstrated a higher prevalence of HLA-DQB1*04:02, -DQA1*01:02 and HLA
SC
DRB1*15 in a Mexican population, which was related to alleles from the ancestral
NU
haplotype HLA DQB1*06:02~DQA1*01:02~DRB1*15, which are found together as a result of linkage disequilibrium.
MA
The mechanism by which HLA-DR alleles determine a risk to develop LN is not completely defined. The association between HLA-DR2 and DR3 and renal involvement
ED
was demonstrated in European populations (Ceccarelli et al. 2015). Some authors demonstrated an association between antibodies against Ro to DR2 and antibodies
EP T
anti-Sm/RNP related to DR3 (Arnett et al. 1989; Graham et al. 2007; Olsen et al. 1993).
AC C
A recent study of HLA associations in diseases suggested that three amino acid positions (11, 13, and 26) on the epitope-binding groove of HLA-DR molecules were critical in conferring risk for SLE (Kim et al. 2014). It is presumed that these amino acid positions establish a pathogenic structure at the HLA-DRB1 epitope-binding groove in LN patients. The real mechanism by which MHC predisposes a person to autoimmune diseases is not certain. One hypothesis suggests a breakdown in immunological tolerance to selfantigens via aberrant class II presentation of own or foreign peptides to autoreactive T
ACCEPTED MANUSCRIPT lymphocytes. The specific MHC class II alleles determine the targeting of particular autoantigens, which results in a disease-specific association (Fernando et al. 2008). The susceptibility of interaction of child’s HLA molecules and maternal risk was also examined. Cruz et al. demonstrated that having children who are DRB1*04:01 allele-
PT
positive was associated with a two-fold increase in the risk of SLE in mothers who carried at least one DRB1 risk allele (*03:01, *15:01 or *08:01) (Cruz et al. 2016).
RI
Some alleles demonstrated protection for SLE. Our analysis revealed that HLA-B*15, -
SC
B*44, -B*27, -DRB1*11, -DRB1*13 alleles were protective. Furukawa et al. (Furukawa
NU
et al. 2014) demonstrated the protective effect of DRB1*13 against systemic autoimmune diseases. The HLA-DRB1*13:01 allele is protective in the European
MA
population (Arnett et al. 2010), and -DRB1*13:02 is protective in the Asian population (Furukawa et al. 2016; He et al. 2014; Lu et al. 1997). Several hypotheses for this
ED
protection were postulated. One hypothesis involves the dominant effect of DRB1*13
EP T
molecules. Future therapy at these targets may become curative treatment for these diseases (Furukawa et al. 2014; Furukawa et al. 2017). A Latin American meta-analysis
AC C
demonstrated protection related to DR5 and allele DRB1*11:01 (Castano-Rodriguez et al. 2008).
Kidney transplant is a safe therapy for lupus populations. Morbidity and mortality are lower in women with ESRD after kidney transplantation compared to women who remain supported with dialysis (Ward 2000). Some studies also demonstrated that African-Americans exhibit lower graft survival compared to Caucasian or HispanicAmericans (Contreras et al. 2014; Gonzalez-Suarez and Contreras 2017).
ACCEPTED MANUSCRIPT The kidney-transplanted patients with lupus in our study exhibited a better 5-year survival rate compared to the transplanted control population. Other studies that evaluated lupus transplant patients with transplant recipients did not observe differences in renal survival at 1 year or 5 years between the two groups, regardless of
PT
donor type (Bunnapradist et al. 2006; Moroni et al. 2005). However, Bunnapradist et al.(Bunnapradist et al. 2006) demonstrated a higher rate of AR in lupus patients with
RI
live donors compared to control.
SC
Regarding ethnicity, all groups analyzed in the study had a worse renal survival in non-
NU
white patients. However, we can not say that this is a trend only of SLE patients. This is the first study to evaluate the influence of the presence of an HLA allele that
MA
confers increased susceptibility to a disease and to correlate it with graft survival. We evaluated both the presence of this allele and its correlation with graft survival, as well
ED
as its presence and / or absence in the compatibility with the receptor. There were no
EP T
differences in the evaluation of graft survival with any of these factors. However, no disease activity, correlation with acute rejection and presence of specific donor
AC C
antibody against HLA antibodies specific to any susceptibility allele were assessed and correlated.
To our knowledge, this study is the first evaluation of match and mismatch using the more prevalent alleles (HLA DRB1*15) in kidney-transplanted SLE patients. These correlations were not relevant for graft survival, but we did not correlate the match and mismatch with lupus disease activity after transplant. This correlation may be investigated in future analyses. This result demonstrated that susceptibility before kidney transplant likely did not influence 5-year graft survival. More studies are
ACCEPTED MANUSCRIPT necessary to evaluate disease severity and activity with HLA molecules. Genetics and molecular therapies targeting HLA may be an option for the treatment of SLE. CONCLUSIONS The presence of HLA-DRB1*15 was a strong predisposition factor for lupus and lupus
PT
nephritis. No specific allele was identified solely for lupus nephritis. The presence of the allele HLA DRB1*15 and the match of this allele with the receptor did not influence
RI
5-year graft survival in lupus patients. This study demonstrated that HLA DRB1*13 and
AC C
EP T
ED
MA
NU
SC
HLA DRB1*11 were a protective factor for LN in SLE patients.
ACCEPTED MANUSCRIPT REFERENCES
AC C
EP T
ED
MA
NU
SC
RI
PT
Alarcon-Segovia D, Alarcon-Riquelme ME, Cardiel MH, Caeiro F, Massardo L, Villa AR, PonsEstel BA, Grupo Latinoamericano de Estudio del Lupus E (2005) Familial aggregation of systemic lupus erythematosus, rheumatoid arthritis, and other autoimmune diseases in 1,177 lupus patients from the GLADEL cohort. Arthritis Rheum 52:1138-47 Araujo MN, Silva NP, Andrade LE, Sato EI, Gerbase-DeLima M, Leser PG (1997) C2 deficiency in blood donors and lupus patients: prevalence, clinical characteristics and HLA associations in the Brazilian population. Lupus 6:462-6 Arnett FC, Gourh P, Shete S, Ahn CW, Honey RE, Agarwal SK, Tan FK, McNearney T, Fischbach M, Fritzler MJ, Mayes MD, Reveille JD (2010) Major histocompatibility complex (MHC) class II alleles, haplotypes and epitopes which confer susceptibility or protection in systemic sclerosis: analyses in 1300 Caucasian, African-American and Hispanic cases and 1000 controls. Ann Rheum Dis 69:822-7 Arnett FC, Hamilton RG, Reveille JD, Bias WB, Harley JB, Reichlin M (1989) Genetic studies of Ro (SS-A) and La (SS-B) autoantibodies in families with systemic lupus erythematosus and primary Sjogren's syndrome. Arthritis Rheum 32:413-9 Bhallil O, Ibrahimi A, Ouadghiri S, Ouzeddoun N, Benseffaj N, Bayahia R, Essakalli M (2017) HLA Class II with Lupus Nephritis in Moroccan Patients. Immunol Invest 46:1-9 Block SR, Winfield JB, Lockshin MD, D'Angelo WA, Christian CL (1975) Studies of twins with systemic lupus erythematosus. A review of the literature and presentation of 12 additional sets. Am J Med 59:533-52 Bunnapradist S, Chung P, Peng A, Hong A, Chung P, Lee B, Fukami S, Takemoto SK, Singh AK (2006) Outcomes of renal transplantation for recipients with lupus nephritis: analysis of the Organ Procurement and Transplantation Network database. Transplantation 82:612-8 Castano-Rodriguez N, Diaz-Gallo LM, Pineda-Tamayo R, Rojas-Villarraga A, Anaya JM (2008) Meta-analysis of HLA-DRB1 and HLA-DQB1 polymorphisms in Latin American patients with systemic lupus erythematosus. Autoimmun Rev 7:322-30 Ceccarelli F, Perricone C, Borgiani P, Ciccacci C, Rufini S, Cipriano E, Alessandri C, Spinelli FR, Sili Scavalli A, Novelli G, Valesini G, Conti F (2015) Genetic Factors in Systemic Lupus Erythematosus: Contribution to Disease Phenotype. J Immunol Res 2015:745647 Chung SA, Brown EE, Williams AH, Ramos PS, Berthier CC, Bhangale T, Alarcon-Riquelme ME, Behrens TW, Criswell LA, Graham DC, Demirci FY, Edberg JC, Gaffney PM, Harley JB, Jacob CO, Kamboh MI, Kelly JA, Manzi S, Moser-Sivils KL, Russell LP, Petri M, Tsao BP, Vyse TJ, Zidovetzki R, Kretzler M, Kimberly RP, Freedman BI, Graham RR, Langefeld CD, International Consortium for Systemic Lupus Erythematosus G (2014) Lupus nephritis susceptibility loci in women with systemic lupus erythematosus. J Am Soc Nephrol 25:2859-70 Contreras G, Pagan J, Chokshi R, Virmani S, Diego JM, Byers P, Isakova T, Munoz Mendoza J, Nayer A, Contreras JR, Panama G, Lenz O, Carpintero M, Muchayi T, Roth D (2014) Comparison of mortality of ESRD patients with lupus by initial dialysis modality. Clin J Am Soc Nephrol 9:1949-56 Cortes LM, Baltazar LM, Lopez-Cardona MG, Olivares N, Ramos C, Salazar M, Sandoval L, Lorenz MG, Chakraborty R, Paterson AD, Rivas F (2004) HLA class II haplotypes in Mexican systemic lupus erythematosus patients. Hum Immunol 65:1469-76 Cruz GI, Shao X, Quach H, Ho KA, Sterba K, Noble JA, Patsopoulos NA, Busch MP, Triulzi DJ, Wong WS, Solomon BD, Niederhuber JE, Criswell LA, Barcellos LF (2016) A Child's HLA DRB1 genotype increases maternal risk of systemic lupus erythematosus. J Autoimmun 74:201-207 Excoffier L, Lischer HE (2010) Arlequin suite ver 3.5: a new series of programs to perform population genetics analyses under Linux and Windows. Mol Ecol Resour 10:564-7
ACCEPTED MANUSCRIPT
AC C
EP T
ED
MA
NU
SC
RI
PT
Fernandes SRM, Persoli LB, Marques SBD, al. e (1995) HLA antigens and susceptibility to systemic lupus erythematosus in Brazilian patients. Lupus 4 Fernando MM, Stevens CR, Walsh EC, De Jager PL, Goyette P, Plenge RM, Vyse TJ, Rioux JD (2008) Defining the role of the MHC in autoimmunity: a review and pooled analysis. PLoS Genet 4:e1000024 Furukawa H, Kawasaki A, Oka S, Ito I, Shimada K, Sugii S, Hashimoto A, Komiya A, Fukui N, Kondo Y, Ito S, Hayashi T, Matsumoto I, Kusaoi M, Amano H, Nagai T, Hirohata S, Setoguchi K, Kono H, Okamoto A, Chiba N, Suematsu E, Katayama M, Migita K, Suda A, Ohno S, Hashimoto H, Takasaki Y, Sumida T, Nagaoka S, Tsuchiya N, Tohma S (2014) Human leukocyte antigens and systemic lupus erythematosus: a protective role for the HLA-DR6 alleles DRB1*13:02 and *14:03. PLoS One 9:e87792 Furukawa H, Oka S, Kawasaki A, Shimada K, Sugii S, Matsushita T, Hashimoto A, Komiya A, Fukui N, Kobayashi K, Osada A, Ihata A, Kondo Y, Nagai T, Setoguchi K, Okamoto A, Okamoto A, Chiba N, Suematsu E, Kono H, Katayama M, Hirohata S, Sumida T, Migita K, Hasegawa M, Fujimoto M, Sato S, Nagaoka S, Takehara K, Tohma S, Tsuchiya N (2016) Human Leukocyte Antigen and Systemic Sclerosis in Japanese: The Sign of the Four Independent Protective Alleles, DRB1*13:02, DRB1*14:06, DQB1*03:01, and DPB1*02:01. PLoS One 11:e0154255 Furukawa H, Oka S, Tsuchiya N, Shimada K, Hashimoto A, Tohma S, Kawasaki A (2017) The role of common protective alleles HLA-DRB1*13 among systemic autoimmune diseases. Genes Immun 18:1-7 Ghodke-Puranik Y, Niewold TB (2015) Immunogenetics of systemic lupus erythematosus: A comprehensive review. J Autoimmun 64:125-36 Gonzalez-Suarez ML, Contreras G (2017) Lower kidney allograft survival in African-Americans compared to Hispanic-Americans with lupus. Lupus:961203317699287 Graham RR, Ortmann W, Rodine P, Espe K, Langefeld C, Lange E, Williams A, Beck S, Kyogoku C, Moser K, Gaffney P, Gregersen PK, Criswell LA, Harley JB, Behrens TW (2007) Specific combinations of HLA-DR2 and DR3 class II haplotypes contribute graded risk for disease susceptibility and autoantibodies in human SLE. Eur J Hum Genet 15:823-30 Grumet FC, Coukell A, Bodmer JG, Bodmer WF, McDevitt HO (1971) Histocompatibility (HL-A) antigens associated with systemic lupus erythematosus. A possible genetic predisposition to disease. N Engl J Med 285:193-6 Hahn BH, McMahon MA, Wilkinson A, Wallace WD, Daikh DI, Fitzgerald JD, Karpouzas GA, Merrill JT, Wallace DJ, Yazdany J, Ramsey-Goldman R, Singh K, Khalighi M, Choi SI, Gogia M, Kafaja S, Kamgar M, Lau C, Martin WJ, Parikh S, Peng J, Rastogi A, Chen W, Grossman JM, American College of R (2012) American College of Rheumatology guidelines for screening, treatment, and management of lupus nephritis. Arthritis Care Res (Hoboken) 64:797-808 Hanly JG, Su L, Urowitz MB, Romero-Diaz J, Gordon C, Bae SC, Bernatsky S, Clarke AE, Wallace DJ, Merrill JT, Isenberg DA, Rahman A, Ginzler EM, Petri M, Bruce IN, Dooley MA, Fortin P, Gladman DD, Sanchez-Guerrero J, Steinsson K, Ramsey-Goldman R, Khamashta MA, Aranow C, Alarcon GS, Fessler BJ, Manzi S, Nived O, Sturfelt GK, Zoma AA, van Vollenhoven RF, Ramos-Casals M, Ruiz-Irastorza G, Lim SS, Kalunian KC, Inanc M, Kamen DL, Peschken CA, Jacobsen S, Askanase A, Theriault C, Thompson K, Farewell V (2015) Mood Disorders in Systemic Lupus Erythematosus: Results From an International Inception Cohort Study. Arthritis Rheumatol 67:1837-47 He D, Wang J, Yi L, Guo X, Guo S, Guo G, Tu W, Wu W, Yang L, Xiao R, Li Y, Chu H, Lai S, Jin L, Zou H, Reveille JD, Assassi S, Mayes MD, Zhou X (2014) Association of the HLA -DRB1 with scleroderma in Chinese population. PLoS One 9:e106939 IBGE (2008) Características étnico-raciais da população : um estudo das categorias de classificação de cor ou raça, Rio de Janeiro
ACCEPTED MANUSCRIPT
AC C
EP T
ED
MA
NU
SC
RI
PT
Kim K, Bang SY, Lee HS, Okada Y, Han B, Saw WY, Teo YY, Bae SC (2014) The HLA-DRbeta1 amino acid positions 11-13-26 explain the majority of SLE-MHC associations. Nat Commun 5:5902 Lin H, Sui W, Tan Q, Chen J, Zhang Y, Ou M, Xue W, Li F, Cao C, Sun Y, Dai Y (2016) Integrated analyses of a major histocompatibility complex, methylation and transcribed ultraconserved regions in systemic lupus erythematosus. Int J Mol Med 37:139-48 Liphaus BL, Goldberg AC, Kiss MH, Silva CA (2002) Analysis of human leukocyte antigens class II-DR in Brazilian children and adolescents with systemic lupus erythematosus. Rev Hosp Clin Fac Med Sao Paulo 57:277-82 Liphaus BL, Kiss MH, Goldberg AC (2007) HLA-DRB1 alleles in juvenile-onset systemic lupus erythematosus: renal histologic class correlations. Braz J Med Biol Res 40:591-7 Lu LY, Ding WZ, Fici D, Deulofeut R, Cheng HH, Cheu CC, Sung PK, Schur PH, Fraser PA (1997) Molecular analysis of major histocompatibility complex allelic associations with systemic lupus erythematosus in Taiwan. Arthritis Rheum 40:1138-45 Marchini M, Antonioli R, Lleo A, Barili M, Caronni M, Origgi L, Vanoli M, Scorza R (2003) HLA class II antigens associated with lupus nephritis in Italian SLE patients. Hum Immunol 64:462-8 Mok CC (2016) Towards new avenues in the management of lupus glomerulonephritis. Nat Rev Rheumatol 12:221-34 Mok CC, Ying KY, Ng WL, Lee KW, To CH, Lau CS, Wong RW, Au TC (2006) Long-term outcome of diffuse proliferative lupus glomerulonephritis treated with cyclophosphamide. Am J Med 119:355 e25-33 Moroni G, Tantardini F, Gallelli B, Quaglini S, Banfi G, Poli F, Montagnino G, Meroni P, Messa P, Ponticelli C (2005) The long-term prognosis of renal transplantation in patients with lupus nephritis. Am J Kidney Dis 45:903-11 Morris DL, Fernando MM, Taylor KE, Chung SA, Nititham J, Alarcon-Riquelme ME, Barcellos LF, Behrens TW, Cotsapas C, Gaffney PM, Graham RR, Pons-Estel BA, Gregersen PK, Harley JB, Hauser SL, Hom G, Langefeld CD, Noble JA, Rioux JD, Seldin MF, Systemic Lupus Erythematosus Genetics C, Vyse TJ, Criswell LA (2014) MHC associations with clinical and autoantibody manifestations in European SLE. Genes Immun 15:210-7 Morris DL, Taylor KE, Fernando MM, Nititham J, Alarcon-Riquelme ME, Barcellos LF, Behrens TW, Cotsapas C, Gaffney PM, Graham RR, Pons-Estel BA, Gregersen PK, Harley JB, Hauser SL, Hom G, International MHC, Autoimmunity Genetics N, Langefeld CD, Noble JA, Rioux JD, Seldin MF, Systemic Lupus Erythematosus Genetics C, Criswell LA, Vyse TJ (2012) Unraveling multiple MHC gene associations with systemic lupus erythematosus: model choice indicates a role for HLA alleles and non-HLA genes in Europeans. Am J Hum Genet 91:778-93 Niu Z, Zhang P, Tong Y (2015) Value of HLA-DR genotype in systemic lupus erythematosus and lupus nephritis: a meta-analysis. Int J Rheum Dis 18:17-28 Olsen ML, Arnett FC, Reveille JD (1993) Contrasting molecular patterns of MHC class II alleles associated with the anti-Sm and anti-RNP precipitin autoantibodies in systemic lupus erythematosus. Arthritis Rheum 36:94-104 Segurado OG, Iglesias-Casarrubios P, Martinez-Laso J, Corell A, Martin-Villa JM, Arnaiz-Villena A (1991) Autoimmunogenic HLA-DRB1*0301 allele (DR3) may be distinguished at the DRB1 non-coding regions of HLA-B8,DR3,Dw24 and B18,DR3,Dw25 haplotypes. Mol Immunol 28:189-92 Shimane K, Kochi Y, Suzuki A, Okada Y, Ishii T, Horita T, Saito K, Okamoto A, Nishimoto N, Myouzen K, Kubo M, Hirakata M, Sumida T, Takasaki Y, Yamada R, Nakamura Y, Kamatani N, Yamamoto K (2013) An association analysis of HLA-DRB1 with systemic lupus erythematosus and rheumatoid arthritis in a Japanese population: effects of *09:01 allele on disease phenotypes. Rheumatology (Oxford) 52:1172-82
ACCEPTED MANUSCRIPT
AC C
EP T
ED
MA
NU
SC
RI
PT
Suggs MJ, Majithia V, Lewis RE, Cruse JM (2011) HLA DRB1*1503 allelic haplotype predominance and associated immunodysregulation in systemic lupus erythematosus. Exp Mol Pathol 91:548-62 Tian W, Li LX, Guo SS (2000) [Correlative study on HLA-DR2 allelic polymorphism and systemic lupus erythematosus in the Han nationality in Hunan province]. Hunan Yi Ke Da Xue Xue Bao 25:15-7 Usnayo MJ, Andrade LE, Alarcon RT, Oliveira JC, Silva GM, Bendet I, Burlingame R, Porto LC, Pinheiro Gda R (2011) Study of the frequency of HLA-DRB1 alleles in Brazilian patients with rheumatoid arthritis. Rev Bras Reumatol 51:474-83 Wadi W, Elhefny NE, Mahgoub EH, Almogren A, Hamam KD, Al-Hamed HA, Gasim GI (2014) Relation between HLA typing and clinical presentations in Systemic Lupus Erythematosus patients in Al-Qassim region, Saudi Arabia. Int J Health Sci (Qassim) 8:159-65 Wahren-Herlenius M, Dorner T (2013) Immunopathogenic mechanisms of systemic autoimmune disease. Lancet 382:819-31 Ward MM (2000) Access to renal transplantation among patients with end-stage renal disease due to lupus nephritis. Am J Kidney Dis 35:915-22
ACCEPTED MANUSCRIPT Table 1- Age (mean), Gender (index Female/Male) and Ethnicity (white/non-white) of the transplanted SLE (TxSLE), non-transplanted SLE (nTxSLE), Kidney-transplanted for other diseases (TxCTRL) and healthy control (HeControl) subgroups. N (total patients)
Age Gender White/Nona b (range) (F/M) whitec 33.0 TxSLE 44 6.33 20/24 (12-56) Lupus (GSLE) 37.8 NTxSLE 64 5.40 23/41 (20-59) 39.5 TxCTRL 88 6.33 43/45 Control (11-71) (Gcontrol) 43.2 HeControl 128 6.52 50/78 (20-59) HeControl, nTxSLE and TxCTRL were paired to TxSLE for ethnicity and gender. a: X2 = 43.504 p = 0.0; b: X2 = 0.207, p = 0.976; c: X2 = 3.336, p = 0.342.
EP T
ED
MA
NU
SC
RI
PT
Subgroup
AC C
Group
ACCEPTED MANUSCRIPT
AC C
EP T
ED
MA
NU
SC
RI
PT
Table 2. Prevalence of Alleles in the Control Group (Gcontrol) X General Lupus group (GSLE) Allele GControl GSLE OR p-value Pc-value N=216 N=108 B*08 17 (17.9%) 20 (18.5%) 2.66 0.0048 0.1379 B*15 51 (23.6%) 15 (13.9%) 0.52 0.0265 0.7697 B*27 10 (4.7%) 0 (0.0%) 0.00 0.0161 0.4683 B*44 42 (19.4%) 12 (16.7%) 0.51 0.0384 1.1133 DRB1*08 20 (9.3%) 22 (20.4%) 2.50 0.0050 0.0653 DRB1*11 49 (22.7%) 13 (12%) 0.46 0.0140 0.1824 DRB1*13 65 (30%) 20 (18,5%) 0.52 0.0166 0.2159 DRB1*15 50 (23.2%) 45 (41.7%) 2.37 0.0005 0.0067 Total number of alleles found per locus (Locus A – 20, Locus B – 39 and Locus DRB1 – 13) were used to calculate Pc (Bonferroni correction).
ACCEPTED MANUSCRIPT
AC C
EP T
ED
MA
NU
SC
RI
PT
Table 3. Allele Prevalence in Health Control (HeControl), kidney transplanted for other diseases (TxCTRL), non-transplanted SLE (nTxSLE) and transplanted SLE (TxSLE) groups. Allele HeControl TxControl nTxSLE TxSLE p-value Pc-value N=128 N=88 N=64 N=44 B*08 8 9 13 7 0.0246 0.7134 (6.2%) (10.2%) (20.4%) (15.9%) DRB1*08 10 10 14 8 0.0324 0.4212 (7.8%) (11.4%) (21.9%) (18.2%) DRB1*15 43 12 24 21 0.0041 0.0533 (19.9%) (11.1%) (37,5%) (47,7%) Total number of alleles found per locus (Locus A: 20, Locus B: 39 and Locus DRB1: 13) were used calculating Pc (Bonferroni correction).
ACCEPTED MANUSCRIPT
AC C
EP T
ED
MA
NU
SC
RI
PT
Table 4. Prevalence of alleles in comparison of transplanted lupus patients (TxSLE) with transplanted control patients (TxCTRL) Allele TxSLE TxCTRL Odds p-value Pc –value N=44 N=88 Rate # A*30 14 (31.8%) 13 (14.7%) 2.69 0.021 0.426 B*18 7 (15.9%) 3 (3.4%) 5.36 0.016 0.459 B*49 14 (9.09%) 1 (1.14%) 8.7 0.042 1.221 DRB1*07 12 (27.2%) 12 (13.6%) 2.37 0.049 0.637 DRB1*15 21 (47.7%) 21 (23.8%) 2.91 0.005 0.065 #- HLA-A*30 was more frequent in non-white TxSLE patients (p= 0.048 – OR= 2.75).
ACCEPTED MANUSCRIPT Figure 1 – Lupus Patient and Control Groups
nTxSLE Lupus Nephritis without Transplant N=32
TxCTRL Ki deny Tra ns planted pa ti ents with others di s eases N=88
SC NU MA ED EP T AC C
N=128
RI
TxSLE kidney transplanted patients with Lupus Nephritis N=44
HeCTRL Hea lth people
PT
nTxSLE Lupus patients without nephritis N=32
ACCEPTED MANUSCRIPT
AC C
EP T
ED
MA
NU
SC
RI
PT
Figure 2. Cumulative survival in 5 years – HLA DRB1*15 comparison of both groups
ACCEPTED MANUSCRIPT
AC C
EP T
ED
MA
NU
SC
RI
PT
Figure 3. Cumulative survival in 5 years - HLA DRB1*15 Match-comparison of both groups
ACCEPTED MANUSCRIPT THE PREVALENCE OF HLA ALLELES IN A LUPUS NEPHRITIS POPULATION Highlights
Lupus nephritis (LN) is a complication of Systemic lupus erythematosus (SLE) and is associated with poor survival and high morbidity. Many genomic studies have been performed worldwide, and several histocompatibility leukocyte
PT
antigen (HLA) loci are linked to lupus susceptibility. This study was a retrospective study of 2 major groups: general lupus patients
SC
A difference in the prevalence of three alleles, B*08, DRB1*08 and DRB1*15,
NU
was observed. These alleles were more prevalent in the lupus subgroups compared to the control groups. The presence of HLA-DRB1*15 was a strong
MA
factor that predisposed patients to the development of SLE and LN, but did not
EP T
ED
influence kidney graft survival.
AC C
RI
(n=108) and a control group (n=216).
Maria Izabel de Holanda, Evandro Klumb, Alicia Imada, Livia A. Lima, Isabela Alcântara, Flavia Gregório, Luis Fernando Christiani, Clarice Oliveira Martins, Branca Engel Timoner, Juliana Motta, Roberto Pozzan, Luis Cristóvão Pôrto PII: DOI: Reference:
S0966-3274(17)30090-4 doi:10.1016/j.trim.2018.02.001 TRIM 1125
To appear in:
Transplant Immunology
Received date: Revised date: Accepted date:
29 June 2017 30 January 2018 1 February 2018
Please cite this article as: Maria Izabel de Holanda, Evandro Klumb, Alicia Imada, Livia A. Lima, Isabela Alcântara, Flavia Gregório, Luis Fernando Christiani, Clarice Oliveira Martins, Branca Engel Timoner, Juliana Motta, Roberto Pozzan, Luis Cristóvão Pôrto , The prevalence of HLA alleles in a lupus nephritis population. The address for the corresponding author was captured as affiliation for all authors. Please check if appropriate. Trim(2018), doi:10.1016/j.trim.2018.02.001
This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
ACCEPTED MANUSCRIPT THE PREVALENCE OF HLA ALLELES IN A LUPUS NEPHRITIS POPULATION
Maria Izabel de Holanda 1, Evandro Klumb2, Alicia Imada 1, Livia A Lima 1, Isabela Alcântara 1, Flavia Gregório1, Luis Fernando Christiani 1, Clarice Oliveira Martins 2, Branca Engel Timoner3, Juliana Motta 3, Roberto Pozzan4, Luis Cristóvão Pôrto3
1 - Nephrology Service, Hospital Federal de Bonsucesso
RI
2 – Rheumatology Service, Rio de Janeiro State University
PT
Affiliations
SC
3 - Histocompatibility and Cryopreservation Laboratory, Rio de Janeiro State University 4 – Cardiology Service, Rio de Janeiro State University
NU
Corresponding Author Prof Luís Cristóvão Pôrto
Pav. Jose Roberto Feresin Moares Av. Marechal Rondon 381
MA
Laboratório de Histocompatibilidade e Criopreservação
ED
São Francisco Xavier 20950-003
Brazil
EP T
Rio de Janeiro
Tel +552123342426
AC C
[email protected]
Author Contributions
MIH – designed the study, followed up with patients, collected samples, analyzed the results and wrote the manuscript. AI, LAL, IA FG and LFC - followed up and retrieved historical records of transplanted patients. EK and COM followed up with lupus nontransplanted patients. BET and JM - performed histocompatibility tests. RP – performed statistical test analyses and interpreted results. LCP - designed the study, analyzed results and revised the manuscript. Conflict of Interest The authors declare no conflicts of interest.
ACCEPTED MANUSCRIPT Abstract Background: Systemic lupus erythematosus (SLE) is a severe autoimmune disease that involves multiple organ systems. Lupus nephritis (LN) is a complication of SLE and is associated with poor survival and high morbidity. Many genomic studies have been performed worldwide, and several histocompatibility leukocyte antigen (HLA) loci are
PT
linked to lupus susceptibility.
RI
Objective: The present study evaluated the association of HLA alleles in a lupus patient
SC
population, LN group and control group. The second objective evaluated whether HLA allele match or mismatch influenced kidney graft survival in a kidney transplanted
NU
lupus population.
Methods: This study was a retrospective study of 2 major groups: general lupus
MA
patients (GSLE - n=108) and a control group (GControl - n=216). Both groups were also divided into subgroups.
ED
Results: The control group was divided into two subgroups: a healthy control group
EP T
(HeCTRL) and transplant control group (TxCTRL). The GSLE group was composed of transplanted lupus patients (TxSLE) and non-transplanted lupus patients (nTxSLE).
AC C
Comparison of the demographics between groups did not reveal differences between ethnicity and gender. A difference in the prevalence of three alleles, B*08, DRB1*08 and DRB1*15, was observed. These alleles were more prevalent in the lupus subgroups compared to the control groups. Five-year survival was not different between patients carrying the allele DRB1*15 in either group (overall p=0.075; TxSLE p=0.419; TxCTRL=0.309). The presence of the match with this allele in the receptor was evaluated and did not demonstrate any difference in graft survival in both groups (p=0.146) or when analyzed separately in each group (TxCTRL p=0.739; TxSLE= 0.297).
ACCEPTED MANUSCRIPT Conclusion: This study demonstrated that the presence of HLA-DRB1*15 was a strong factor that predisposed patients to the development of SLE and LN, but did not
AC C
EP T
ED
MA
NU
SC
RI
PT
influence kidney graft survival.
ACCEPTED MANUSCRIPT BACKGROUND Systemic lupus erythematosus (SLE) is a severe autoimmune disease that involves multiple organ systems. SLE involves the production of autoreactive T cells and autoantibodies against nuclear, cytoplasmic and cell-surface antigens. The more
PT
common clinical features of SLE include skin and joint diseases, hematological abnormalities, renal disease and neuropsychiatric complications (Ghodke-Puranik and
RI
Niewold 2015).
SC
Lupus nephritis (LN) is one complication of SLE, and it is associated with poor survival
NU
and high morbidity, particularly in patients who develop end-stage renal disease (ESRD) (Hanly et al. 2015). The cause of LN is complex, and environmental and genetic
MA
factors are involved (Alarcon-Segovia et al. 2005). Monozygotic twins exhibit an approximately 10-fold higher risk for SLE than dizygotic twins (Block et al. 1975), and
ED
first-degree relatives of patients with SLE exhibit a 20-fold increased risk of developing
EP T
SLE compared to the healthy population (Alarcon-Segovia et al. 2005). The major histocompatibility complex (MHC) is located on the short arm of
AC C
chromosome 6, and its relationship to autoimmune diseases was studied extensively (Graham et al. 2007; Wahren-Herlenius and Dorner 2013). Approximately 421 genes are related to the MHC; 60% of these genes are expressed, and 22% are related to immunoregulatory functions (Fernando et al. 2008). Many worldwide genomic studies were performed, and several histocompatibility leukocyte antigen (HLA) loci were linked to the lupus susceptibility (Chung et al. 2014; Grumet et al. 1971; Morris et al. 2014). A recent meta-analysis, Niu et al (2015) suggested that distinct HLA Class II genetic variants conferred a predisposition and resistance to SLE and LN. HLA-DR4,
ACCEPTED MANUSCRIPT DR5, DR11 and DR14 were identified as protective against SLE, and HLA-DR3, DR9 and DR15 may be risk factors for SLE. The DR4 and DR11 alleles may be protective for LN, and DR3 and DR15 were risk factors (Niu et al. 2015). HLA class I and II genes were consistently associated with SLE susceptibility, especially
PT
B*18 and DRB1*15 (Cortes et al. 2004; Lin et al. 2016; Marchini et al. 2003; Segurado et al. 1991; Shimane et al. 2013; Suggs et al. 2011; Tian et al. 2000). This susceptibility
RI
may also be related to disease severity.
SC
The present study evaluated the association of HLA alleles in a lupus patient
NU
population, LN group and control group. The study also evaluated whether matched or mismatched HLA alleles influenced kidney graft survival in a kidney-transplanted lupus
AC C
EP T
ED
MA
population.
ACCEPTED MANUSCRIPT MATERIALS AND METHODS Study Population This study was a retrospective study of 2 major groups: general lupus patients (GSLE n=108) and a control group (GControl - n=216). Both groups included patients and
PT
controls with specific criteria (see below). Lupus Group
RI
The GSLE was composed of kidney-transplanted lupus patients (TxSLE) from the
SC
Bonsucesso Federal Hospital, Rio de Janeiro and non-transplanted lupus patients from
NU
the Rheumatology Department of Rio de Janeiro State University (nTxSLE), Rio de Janeiro, Brazil. Patients in both groups were matched by gender and ethnicity (white
MA
and non-white). All patients met the American College of Rheumatology classification criteria for the diagnosis of SLE (Hahn et al. 2012).
ED
Patients in the TxSLE group (n=44) received a kidney transplant during 1980-2016.
EP T
These patients had HLA typings before transplant as part of the pre-transplant routine. LN patients (LNSLE) from both health centers were subgrouped into patients who
AC C
presented with biopsy-proven LN class III/IV or V from the nTxSLE (n=32), considering that proliferative nephritis has a worse prognosis and 30% in 10 years will need dialysis (Mok 2016; Mok et al. 2006), and TxSLE groups (n=44, Figure 1). Control Group
The control groups were formed on a 2:1 proportion and matched by gender and ethnicity (white and non-white: including black and mestizos). This group was composed of 128 healthy people (HeCTRL) from the control group of a previous rheumatic arthritis study (Usnayo et al. 2011) and patients who received a kidney
ACCEPTED MANUSCRIPT transplant for other diseases in the Bonsucesso Federal Hospital (TxCTRL - n=88) during the same period. The control group of two different samples created uniformity of this group with the GSLE group. The control group (Gcontrol) was 216 individuals (Figure 1).
PT
HLA HLA class II-DR typing was performed using serology or polymerase chain reaction
RI
amplification with sequence-specific primers. HLA from the kidney-transplanted
SC
patients were collected from medical charts, and the antigens defined in the serology
NU
were converted to the most frequent possible allele according to the frequency of haplotypes and alleles in the Rio de Janeiro population (www.imunogenetica.org).
MA
All patients from the Rheumatology Department were invited to participate in the study, and participants signed a written informed consent form. Kidney-transplanted
ED
patients did not need to sign a consent form because consent was part of their pre-
EP T
transplant routine, and all data were collected after transplantation. This study was performed in accordance with standards of our hospital’s Ethics
AC C
Committees (142/09). Statistical Analysis:
The genetic distribution of HLA was performed using the Arlequin program. We compared all four groups initially, then performed two by two group comparisons: GSLE versus the GControl, followed by the LNSLE versus nLNSLE groups, and TxSLE versus the TxCTRL groups. We also analyzed the prevalence of HLA alleles in the LNspecific population compared to the nLNSLE and Gcontrol groups.
ACCEPTED MANUSCRIPT Categorical data are described using numbers and percentages. The frequencies of gender, ethnicity, and age were performed via direct counting. Haplotypes HLAA~B~DRB1 were generated using the expectation–maximization with Arlequin 3.5 (Excoffier and Lischer 2010).Data were analyzed using and EPI-INFO version 3.5.3 (CDC, Atlanta,
GA, USA) and SPSS version 22.0. Comparisons of demographic data and HLA allele
PT
frequencies between groups were performed using the chi-square and Fisher’s exact
RI
test. P<0.05 was considered statistically significant. Significant results in the HLA allele
SC
frequencies were multiplied by the total number of alleles found per locus (Locus A – 20, Locus B – 39 and Locus DRB1 – 13) to obtain a corrected value of P, termed Pc
NU
(Bonferroni correction). The T-test compared the mean ages between groups and
MA
different outcomes.
Survival analysis was performed using the Kaplan-Meier method. P values <0,05 were
ED
considered significant.
Demographics
EP T
RESULTS
AC C
The control group was divided into two groups: the health control group (HeCTRL) and the transplant control group (TxCTRL). The HeCTRL was composed of 111 women and 17 men (50 whites and 78 non white), and the mean age was 43.2 years (range 28-59 years). The TxCTRL group had 88 patients: 76 women and 43 whites, and the mean age was 39.5 years (range 11-71 years). The prevalence of underlying diseases in the TxCTRL group was 46 hypertension patients: 12 glomerulosclerosis, 11 polycystic kidney disease, 3 diabetes, 2 eclampsias and 14 presented with urological affections.
ACCEPTED MANUSCRIPT There was a total of 2,326 patients who received kidney transplants from 1980 to 2016 in the Bonsucesso Federal Hospital transplant center, and 56 patients were identified with LN. Twelve patients did not present a complete loci A-B-DRB1HLA typing or were lacking in information. The TxSLE group was comprised of 38 female patients, 24
PT
patients were white, and the mean age was 33 years (range 12-56 years). The nTxSLE group was comprised of 54 female patients, 23 patients were white, the mean age was
RI
37.5 years (range 20-59 years), and nephritis was present in 22 patients. Patients from
SC
these two group who did not present nephritis were sub-grouped in the nLNSLE group (n=44, Table 1). Comparison of demographics between groups revealed no differences
NU
in ethnicity or gender. The groups were homogeneous.
MA
Thirteen patients in nTxSLE exhibited serositis, nine patients had neurological involvement, 37 patients exhibited arthritis, 29 patients exhibited dermatological
ED
lesions, and 32 patients exhibited LN.
EP T
Transplant Patients Demographics and Characteristics The TxSLE group consisted of 44 patients of whom 29 were transplant recipients with live donors, 15 from deceased donors, 20 did not receive induction therapy, 18
AC C
induced with simulect® and five with thymoglobulin® and one with OKT3. Regarding previous transfusions, nine patients did not receive transfusions prior to transplantation, eight received one, six received two, nine received three, and twelve had several blood transfusions. The TxCTRL group, consisting of 88 patients, on induction therapy, forty were not transfused, 42 did simulect® and 6 did thymoglobulin®. Forty-nine patients in the TxCTRL group had previous transfusions, 13 had one blood transfusion, 12 had 2 transfusions, 6 had 3 transfusions, and 17 had multiple transfusions. Immunosuppression was based on a calcineurin and
ACCEPTED MANUSCRIPT corticosteroid inhibitor in both groups, but in the TxSLE group there were 11 patients using azathioprine associated with this scheme and in the TxCTRL 20 group, comparing the groups there was no statistical difference (p = 0.347). Regarding the type of renal replacement therapy (RRT) prior to transplantation, no
PT
difference was also observed between the groups. In the TxCTRL group, 3 patients underwent preemptive transplantation, 74 underwent hemodialysis (HD), 5 under
RI
peritoneal dialysis (PD) and 6 underwent both therapies. In the TxSLE 37 group they
SC
did HD before, one patient had PD and 6 did the two therapies. The mean time to RRT
NU
before transplantation was 50 months (approximately 4 years and 2 months, range 0172 months).
MA
Patients with acute rejection (RA) were those who presented renal biopsy confirming the diagnosis according to the Banff classification. Patients who had increased
ED
creatinine and / or decreased diuresis, and / or other signs of RA who underwent
clinical RA.
EP T
methylprednisolone pulse and presented response after therapy were classified with
AC C
Twenty-five (18.9%) patients had episodes of acute rejection (RA), 18 (20.4%) patients in the TxCTRL group and 7 (15.9%) patients in the TxSLE group. Only 10 patients underwent renal biopsy, five patients had acute humoral rejection, 1 in the TxSLE group and the remainder in the TxCTRL group, there was no difference between the groups (p = 0.353). The time of diagnosis of AR episodes ranged from 0-1752 days from the date of transplantation, and 16 patients had episodes within the first month after transplantation.
ACCEPTED MANUSCRIPT Twenty patients had viral infections, 12 patients in the TxCTRL group, two herpes virus infections and the remaining cytomegalovirus (CMV) in the TxSLE group were 8 patients with viral infections, one for herpes and 7 for CMV. There was no difference between groups (p = 0.763). Nine patients had these infections in the first year of
PT
transplantation. Thirty-one patients had bacterial infections, 53 of them were infectious, some of them had more than 3 episodes. The most frequent infection was
RI
the urinary tract infection. Patients in the TxCTRL group had more infections compared
SC
to the TxSLE group, 26 patients versus 5 TxSLE patients (p = 0.048).
NU
Locus frequency
Comparisons of pairs of population samples demonstrated that GLSE was genetically
MA
distant from Gcontrol (FST - 0.00409, p <0.05) and nTxSLE and TxSLE were genetically separate from HeControl (FST -0.00463 and 0.00425, respectively). Comparison of the
ED
allele genetic frequency between GSLE and Gcontrol revealed no differences between these groups in the A locus (p=0.914) or B locus (p=0.072). However, frequency in the
EP T
DRB1 locus was significantly different (p=0.0003). Comparison of the four subgroups also revealed differences in the DRB1 locus (p=0.0009), but the A and B loci were not
AC C
statistically different (p=0.852, p=0.113, respectively; Table 1S). Locus B was generally in pairwise linkage disequilibrium with loci A and DRB1, except in TxCTRL, in which locus B exhibited linkage disequilibrium with locus A. Hardy-Weinberg equilibrium was only absent for locus A in the TxCTRL subgroup (observed heterogeneity 0.78409, p=0. 0.034). Seventy-two possible HLA-A~B~DRB1 haplotypes were determined from the EM algorithm for the TxLSE group, and 96 of these haplotypes were determined for the
ACCEPTED MANUSCRIPT nTxSLE group. Twelve haplotypes were observed in at least two patients in the TxSLE group and in 23 patients in the nTxSLE. Sixteen of these nTxSLE haplotypes were not observed in the other subgroups, and 8 were not observed in the TxSLE group (Table 2S).
PT
Allele Frequencies The first analysis compared the GSLE (n=128) and the Control group (n=216). This
RI
analysis revealed eight alleles: HLA-B*08, -B*15, -B*27, -B*44, -DRB1*08, -DRB1*11, -
SC
DRB1*13 and -DRB1*15. The most prevalent alleles in the GSLE were -B*08, -DRB1*08
NU
and -DRB1*15. The other alleles were protective for lupus, primarily HLA B*27. No patients in the GSLE group presented these alleles. HLA DRB1*15 was the only allele
MA
that remained statistically significant after Bonferroni correction (Table 2). The four described subgroups were compared (Table 3). A difference in the prevalence
ED
of three alleles, HLA-B*08, -DRB1*08 and -DRB1*15, was demonstrated. These alleles
EP T
were more prevalent in the lupus subgroups than the control groups. No allele remained significant after Bonferroni correction. However, the HLA DRB1*15 was near
AC C
significant (p-value=0.0533).
Comparison between the LNSLE and nLNSLE group revealed a higher prevalence in HLA-A*02 (48.6% x 25.0%). The Alleles A*03, A*33, A*74 and DRB1*04 were more prevalent in the lupus without nephritis group (nLNSLE – Table 3S). However, no difference between groups was found after Bonferroni correction. The HLA DRB1*15 was present in 39.47% of patients in the LNSLE group and 46.88% in the nLNSLE group (p=0.307). We also examined whether a specific allele was associated with SLE without nephritis. The alleles with higher prevalence in the comparison of the nLNSLE with the
ACCEPTED MANUSCRIPT HeControl groups were HLA-A*02, -DRB1*11, -DRB1*13 in the control group and HLA B*07, -B*08, -B*15, -DRB1*08, and -DRB1*15 in the lupus group (Table 4S). We compared the TxCTRL group (n=88) with the TxSLE group (n=44). The results revealed a higher prevalence of alleles HLA-A*30, -B*18, -B*49, -DRB1*07 and -
PT
DRB1*15 in the TxSLE group. However, no alleles were statistically significant after Bonferroni correction (Table 4). The importance of the prevalence of HLA DRB1*15 in
RI
SLE was demonstrated in the literature. The association and link of some alleles, such
SC
as A*25~B*18~DRB1*15, is also well described in the literature. We evaluated this association in our population and only found this haplotype in the nLNSLE group (Table
NU
2S).
MA
The comparison between both control groups revealed no significant differences in allele prevalence. The prevalence of HLA-DRB1*15 was also analyzed by comparing
ED
ethnicity, and there were no differences between whites (27%) and non-whites (36%) (p=0.343).
EP T
Kidney Graft Survival and HLA DRB1*15 The overall 5-year kidney graft survival rate with death censored was 73.5% These
AC C
rates were 90.9% and 64.8% in the TxSLE and TxCTRL groups, respectively (p=0.05). We did not match for age in the control groups, and the TxCTRL group was older. Therefore, we used a t-test to evaluate whether age was a factor for worst outcome. No differences were found in age means and the outcome, and the mean age for patients with graft loss in five years was 37.9 years (sd 12.6) versus 37.2 years (sd13.4) (p=0.316) for patients who did not lose the graft. There was no difference between groups in donor type or deceased or live donors (p=0.350). Considering the possibility of the underlying disease being an influence factor in graft loss, we divided the control
ACCEPTED MANUSCRIPT group into hypertensive patients and with other diseases and compared them with the patients in the lupus group. We then compared 3 groups: Lupus, SAH and other diseases. The hypertensive patients had a cured survival rate of 54.2%, compared to 77.5% for other diseases and 90.2% for lupus (p = 0.105).
PT
When we evaluated the 5-year survival curve in relation to ethnicity, we observed a higher survival rate in whites (76.2%) vs. non-whites (55.1%; p = 0.008), who were
RI
censored for death, survival rate for whites was 85.7% and non-whites 62.3% (p =
SC
0.001). In the lupus group, the result found was 75% survival for non-whites and 90% for whites, although it was not significant (p = 0.144). However, when we censored for
NU
death, survival for whites was 100% and for non-whites, 83.3% (p = 0.040). There is a
MA
trend of worse survival for non-whites. The same happened in the control group, 65.1% survival for whites versus 44.4% for nonwhites (p = 0.047).
ED
Five-year survival was not different between patients carrying the DRB1*15 allele overall (p=0.075) or in TxSLE (p=0.419) and TxCTRL (p=0.304) groups or when both
EP T
groups were compared (p=0.059; Figure2). The presence of the match with this allele in the receptor was evaluated, but no difference in graft survival comparison of both
AC C
groups was demonstrated (p=0.075, Figure 3) or when this allele was analyzed separately in each group and overall (TxCTRL p=0.738; TxSLE p=0.297; overall p=0.275). The correlation between ethnicity, HLA-DRB1*15 and graft survival in the groups was also not significantly different (p=0.104). DISCUSSION The present study demonstrated the prevalence of HLA alleles in a Brazilian lupus population. We compared the prevalence to a healthy population and a control group
ACCEPTED MANUSCRIPT of kidney-transplanted patients with other diseases. Most published studies limited comparisons to a healthy population (Lin et al. 2016; Liphaus et al. 2007). The alleles HLA-B*08, -DRB1*08 and -DRB1*15 exhibited a higher prevalence in our lupus population. One of the limitations of our study was the low number of patients
PT
in the TxSLE group. HLA allele distribution in a population is linked to ethnicity, which presents variability in the determination of color-race in the Brazilian population(IBGE
RI
2008). Our groups were classified into white and non-white. This simplification may
SC
cause a bias in specific alleles that are over represented in Sub-Saharan Africans
NU
(blacks), such as HLA-A*30. HLA-A*30 is found in Brazilian black individuals on the Bone Marrow Donor Registry with a genetic frequency of 0.0804 compared to the
MA
white individuals (frequency of 0.0426) (www.imunogenetica.org). The allelic frequency in TxSLE patients also confirmed the important contribution of non-white
ED
ethnicity as a specific risk for lupus in this group. HLA-DRB1*08 frequency in the subgroups revealed an ethnic influence, and this allele was also more frequent in
EP T
Brazilian white individuals. The odds ratio in association with LN increased more than
AC C
double (3.68 x 8.00).
Our results are consistent with Liphaus et al. (Liphaus et al. 2002), who described a higher prevalence of HLA DR15 that was influenced by gender (female) and ethnicity (black individuals). This difference was not observed in our samples because we paired controls by gender and ethnicity. Araujo et al.(Araujo et al. 1997) also evaluated a Brazilian lupus population with C2 deficiency and demonstrated that patients with HLA-B*18 were more prevalent (55.6%) with a relative risk of 9.2 compared to the healthy population. HLA-B*18 was
ACCEPTED MANUSCRIPT also very prevalent in the families of patients with the same phenotypes. Our study demonstrated that the associations between HLA-DRB1*15 and HLA-B*18 were linked with -A*25 in the nTxSLE group (Table 2S). These results suggest that the alleles may be a risk factor for other manifestations of lupus because this haplotype HLA-
PT
A*25~B*18~DRB1*15 (Araujo et al. 1997) was not observed in the LNSLE group. HLAB*18~DRB1*15 was associated with other alleles of Locus A in the HeControl group,
RI
twice with A*24 and once with A*26 in the TxSLE group.
SC
Few studies in the literature evaluated the prevalence of HLA and lupus nephritis, and
NU
our study compared 2 nephritis population (with or without transplantation) to two different controls (transplant/others disease control and a healthy population). A
MA
genome-wide association study of LN demonstrated an association between LN and antigens HLA-DR2 and HLA-DR3. However, other genes outside the MHC were more
ED
evident than this correlation in this same study (Chung et al. 2014).
EP T
An LN sample evaluated the existence of a higher prevalence of specific alleles in patients with compromised renal function. However, it was not possible to
AC C
demonstrate a specific allele more prevalent in LN population. To our knowledge, this study is the first study to evaluate the prevalence of HLA in LN in a Brazilian population.
A recent meta-analysis demonstrated an association of lupus nephritis with DR3 and DR15 antigens, and DR14 and DR11 were protective genotypes for lupus nephritis (Niu et al. 2015). A Latin American meta-analysis also demonstrated a higher susceptibility for SLE with antigens DR2-DR3 and allele HLA-DRB1*15:01 (Castano-Rodriguez et al. 2008).
ACCEPTED MANUSCRIPT A Moroccan study in a lupus nephritis population demonstrated the prevalence of the HLA-DR15 antigen was also higher compared to the normal population, and the odds ratio for this population was 2.8 (Bhallil et al. 2017). Our positive result of HLA-DRB1*15 was also observed in Italian (Marchini et al. 2003),
PT
European (Morris et al. 2012), Brazilian (Fernandes et al. 1995), Japanese (Shimane et al. 2013) and Saudi populations (Wadi et al. 2014). Cortes et al. (Cortes et al. 2004) also
RI
demonstrated a higher prevalence of HLA-DQB1*04:02, -DQA1*01:02 and HLA
SC
DRB1*15 in a Mexican population, which was related to alleles from the ancestral
NU
haplotype HLA DQB1*06:02~DQA1*01:02~DRB1*15, which are found together as a result of linkage disequilibrium.
MA
The mechanism by which HLA-DR alleles determine a risk to develop LN is not completely defined. The association between HLA-DR2 and DR3 and renal involvement
ED
was demonstrated in European populations (Ceccarelli et al. 2015). Some authors demonstrated an association between antibodies against Ro to DR2 and antibodies
EP T
anti-Sm/RNP related to DR3 (Arnett et al. 1989; Graham et al. 2007; Olsen et al. 1993).
AC C
A recent study of HLA associations in diseases suggested that three amino acid positions (11, 13, and 26) on the epitope-binding groove of HLA-DR molecules were critical in conferring risk for SLE (Kim et al. 2014). It is presumed that these amino acid positions establish a pathogenic structure at the HLA-DRB1 epitope-binding groove in LN patients. The real mechanism by which MHC predisposes a person to autoimmune diseases is not certain. One hypothesis suggests a breakdown in immunological tolerance to selfantigens via aberrant class II presentation of own or foreign peptides to autoreactive T
ACCEPTED MANUSCRIPT lymphocytes. The specific MHC class II alleles determine the targeting of particular autoantigens, which results in a disease-specific association (Fernando et al. 2008). The susceptibility of interaction of child’s HLA molecules and maternal risk was also examined. Cruz et al. demonstrated that having children who are DRB1*04:01 allele-
PT
positive was associated with a two-fold increase in the risk of SLE in mothers who carried at least one DRB1 risk allele (*03:01, *15:01 or *08:01) (Cruz et al. 2016).
RI
Some alleles demonstrated protection for SLE. Our analysis revealed that HLA-B*15, -
SC
B*44, -B*27, -DRB1*11, -DRB1*13 alleles were protective. Furukawa et al. (Furukawa
NU
et al. 2014) demonstrated the protective effect of DRB1*13 against systemic autoimmune diseases. The HLA-DRB1*13:01 allele is protective in the European
MA
population (Arnett et al. 2010), and -DRB1*13:02 is protective in the Asian population (Furukawa et al. 2016; He et al. 2014; Lu et al. 1997). Several hypotheses for this
ED
protection were postulated. One hypothesis involves the dominant effect of DRB1*13
EP T
molecules. Future therapy at these targets may become curative treatment for these diseases (Furukawa et al. 2014; Furukawa et al. 2017). A Latin American meta-analysis
AC C
demonstrated protection related to DR5 and allele DRB1*11:01 (Castano-Rodriguez et al. 2008).
Kidney transplant is a safe therapy for lupus populations. Morbidity and mortality are lower in women with ESRD after kidney transplantation compared to women who remain supported with dialysis (Ward 2000). Some studies also demonstrated that African-Americans exhibit lower graft survival compared to Caucasian or HispanicAmericans (Contreras et al. 2014; Gonzalez-Suarez and Contreras 2017).
ACCEPTED MANUSCRIPT The kidney-transplanted patients with lupus in our study exhibited a better 5-year survival rate compared to the transplanted control population. Other studies that evaluated lupus transplant patients with transplant recipients did not observe differences in renal survival at 1 year or 5 years between the two groups, regardless of
PT
donor type (Bunnapradist et al. 2006; Moroni et al. 2005). However, Bunnapradist et al.(Bunnapradist et al. 2006) demonstrated a higher rate of AR in lupus patients with
RI
live donors compared to control.
SC
Regarding ethnicity, all groups analyzed in the study had a worse renal survival in non-
NU
white patients. However, we can not say that this is a trend only of SLE patients. This is the first study to evaluate the influence of the presence of an HLA allele that
MA
confers increased susceptibility to a disease and to correlate it with graft survival. We evaluated both the presence of this allele and its correlation with graft survival, as well
ED
as its presence and / or absence in the compatibility with the receptor. There were no
EP T
differences in the evaluation of graft survival with any of these factors. However, no disease activity, correlation with acute rejection and presence of specific donor
AC C
antibody against HLA antibodies specific to any susceptibility allele were assessed and correlated.
To our knowledge, this study is the first evaluation of match and mismatch using the more prevalent alleles (HLA DRB1*15) in kidney-transplanted SLE patients. These correlations were not relevant for graft survival, but we did not correlate the match and mismatch with lupus disease activity after transplant. This correlation may be investigated in future analyses. This result demonstrated that susceptibility before kidney transplant likely did not influence 5-year graft survival. More studies are
ACCEPTED MANUSCRIPT necessary to evaluate disease severity and activity with HLA molecules. Genetics and molecular therapies targeting HLA may be an option for the treatment of SLE. CONCLUSIONS The presence of HLA-DRB1*15 was a strong predisposition factor for lupus and lupus
PT
nephritis. No specific allele was identified solely for lupus nephritis. The presence of the allele HLA DRB1*15 and the match of this allele with the receptor did not influence
RI
5-year graft survival in lupus patients. This study demonstrated that HLA DRB1*13 and
AC C
EP T
ED
MA
NU
SC
HLA DRB1*11 were a protective factor for LN in SLE patients.
ACCEPTED MANUSCRIPT REFERENCES
AC C
EP T
ED
MA
NU
SC
RI
PT
Alarcon-Segovia D, Alarcon-Riquelme ME, Cardiel MH, Caeiro F, Massardo L, Villa AR, PonsEstel BA, Grupo Latinoamericano de Estudio del Lupus E (2005) Familial aggregation of systemic lupus erythematosus, rheumatoid arthritis, and other autoimmune diseases in 1,177 lupus patients from the GLADEL cohort. Arthritis Rheum 52:1138-47 Araujo MN, Silva NP, Andrade LE, Sato EI, Gerbase-DeLima M, Leser PG (1997) C2 deficiency in blood donors and lupus patients: prevalence, clinical characteristics and HLA associations in the Brazilian population. Lupus 6:462-6 Arnett FC, Gourh P, Shete S, Ahn CW, Honey RE, Agarwal SK, Tan FK, McNearney T, Fischbach M, Fritzler MJ, Mayes MD, Reveille JD (2010) Major histocompatibility complex (MHC) class II alleles, haplotypes and epitopes which confer susceptibility or protection in systemic sclerosis: analyses in 1300 Caucasian, African-American and Hispanic cases and 1000 controls. Ann Rheum Dis 69:822-7 Arnett FC, Hamilton RG, Reveille JD, Bias WB, Harley JB, Reichlin M (1989) Genetic studies of Ro (SS-A) and La (SS-B) autoantibodies in families with systemic lupus erythematosus and primary Sjogren's syndrome. Arthritis Rheum 32:413-9 Bhallil O, Ibrahimi A, Ouadghiri S, Ouzeddoun N, Benseffaj N, Bayahia R, Essakalli M (2017) HLA Class II with Lupus Nephritis in Moroccan Patients. Immunol Invest 46:1-9 Block SR, Winfield JB, Lockshin MD, D'Angelo WA, Christian CL (1975) Studies of twins with systemic lupus erythematosus. A review of the literature and presentation of 12 additional sets. Am J Med 59:533-52 Bunnapradist S, Chung P, Peng A, Hong A, Chung P, Lee B, Fukami S, Takemoto SK, Singh AK (2006) Outcomes of renal transplantation for recipients with lupus nephritis: analysis of the Organ Procurement and Transplantation Network database. Transplantation 82:612-8 Castano-Rodriguez N, Diaz-Gallo LM, Pineda-Tamayo R, Rojas-Villarraga A, Anaya JM (2008) Meta-analysis of HLA-DRB1 and HLA-DQB1 polymorphisms in Latin American patients with systemic lupus erythematosus. Autoimmun Rev 7:322-30 Ceccarelli F, Perricone C, Borgiani P, Ciccacci C, Rufini S, Cipriano E, Alessandri C, Spinelli FR, Sili Scavalli A, Novelli G, Valesini G, Conti F (2015) Genetic Factors in Systemic Lupus Erythematosus: Contribution to Disease Phenotype. J Immunol Res 2015:745647 Chung SA, Brown EE, Williams AH, Ramos PS, Berthier CC, Bhangale T, Alarcon-Riquelme ME, Behrens TW, Criswell LA, Graham DC, Demirci FY, Edberg JC, Gaffney PM, Harley JB, Jacob CO, Kamboh MI, Kelly JA, Manzi S, Moser-Sivils KL, Russell LP, Petri M, Tsao BP, Vyse TJ, Zidovetzki R, Kretzler M, Kimberly RP, Freedman BI, Graham RR, Langefeld CD, International Consortium for Systemic Lupus Erythematosus G (2014) Lupus nephritis susceptibility loci in women with systemic lupus erythematosus. J Am Soc Nephrol 25:2859-70 Contreras G, Pagan J, Chokshi R, Virmani S, Diego JM, Byers P, Isakova T, Munoz Mendoza J, Nayer A, Contreras JR, Panama G, Lenz O, Carpintero M, Muchayi T, Roth D (2014) Comparison of mortality of ESRD patients with lupus by initial dialysis modality. Clin J Am Soc Nephrol 9:1949-56 Cortes LM, Baltazar LM, Lopez-Cardona MG, Olivares N, Ramos C, Salazar M, Sandoval L, Lorenz MG, Chakraborty R, Paterson AD, Rivas F (2004) HLA class II haplotypes in Mexican systemic lupus erythematosus patients. Hum Immunol 65:1469-76 Cruz GI, Shao X, Quach H, Ho KA, Sterba K, Noble JA, Patsopoulos NA, Busch MP, Triulzi DJ, Wong WS, Solomon BD, Niederhuber JE, Criswell LA, Barcellos LF (2016) A Child's HLA DRB1 genotype increases maternal risk of systemic lupus erythematosus. J Autoimmun 74:201-207 Excoffier L, Lischer HE (2010) Arlequin suite ver 3.5: a new series of programs to perform population genetics analyses under Linux and Windows. Mol Ecol Resour 10:564-7
ACCEPTED MANUSCRIPT
AC C
EP T
ED
MA
NU
SC
RI
PT
Fernandes SRM, Persoli LB, Marques SBD, al. e (1995) HLA antigens and susceptibility to systemic lupus erythematosus in Brazilian patients. Lupus 4 Fernando MM, Stevens CR, Walsh EC, De Jager PL, Goyette P, Plenge RM, Vyse TJ, Rioux JD (2008) Defining the role of the MHC in autoimmunity: a review and pooled analysis. PLoS Genet 4:e1000024 Furukawa H, Kawasaki A, Oka S, Ito I, Shimada K, Sugii S, Hashimoto A, Komiya A, Fukui N, Kondo Y, Ito S, Hayashi T, Matsumoto I, Kusaoi M, Amano H, Nagai T, Hirohata S, Setoguchi K, Kono H, Okamoto A, Chiba N, Suematsu E, Katayama M, Migita K, Suda A, Ohno S, Hashimoto H, Takasaki Y, Sumida T, Nagaoka S, Tsuchiya N, Tohma S (2014) Human leukocyte antigens and systemic lupus erythematosus: a protective role for the HLA-DR6 alleles DRB1*13:02 and *14:03. PLoS One 9:e87792 Furukawa H, Oka S, Kawasaki A, Shimada K, Sugii S, Matsushita T, Hashimoto A, Komiya A, Fukui N, Kobayashi K, Osada A, Ihata A, Kondo Y, Nagai T, Setoguchi K, Okamoto A, Okamoto A, Chiba N, Suematsu E, Kono H, Katayama M, Hirohata S, Sumida T, Migita K, Hasegawa M, Fujimoto M, Sato S, Nagaoka S, Takehara K, Tohma S, Tsuchiya N (2016) Human Leukocyte Antigen and Systemic Sclerosis in Japanese: The Sign of the Four Independent Protective Alleles, DRB1*13:02, DRB1*14:06, DQB1*03:01, and DPB1*02:01. PLoS One 11:e0154255 Furukawa H, Oka S, Tsuchiya N, Shimada K, Hashimoto A, Tohma S, Kawasaki A (2017) The role of common protective alleles HLA-DRB1*13 among systemic autoimmune diseases. Genes Immun 18:1-7 Ghodke-Puranik Y, Niewold TB (2015) Immunogenetics of systemic lupus erythematosus: A comprehensive review. J Autoimmun 64:125-36 Gonzalez-Suarez ML, Contreras G (2017) Lower kidney allograft survival in African-Americans compared to Hispanic-Americans with lupus. Lupus:961203317699287 Graham RR, Ortmann W, Rodine P, Espe K, Langefeld C, Lange E, Williams A, Beck S, Kyogoku C, Moser K, Gaffney P, Gregersen PK, Criswell LA, Harley JB, Behrens TW (2007) Specific combinations of HLA-DR2 and DR3 class II haplotypes contribute graded risk for disease susceptibility and autoantibodies in human SLE. Eur J Hum Genet 15:823-30 Grumet FC, Coukell A, Bodmer JG, Bodmer WF, McDevitt HO (1971) Histocompatibility (HL-A) antigens associated with systemic lupus erythematosus. A possible genetic predisposition to disease. N Engl J Med 285:193-6 Hahn BH, McMahon MA, Wilkinson A, Wallace WD, Daikh DI, Fitzgerald JD, Karpouzas GA, Merrill JT, Wallace DJ, Yazdany J, Ramsey-Goldman R, Singh K, Khalighi M, Choi SI, Gogia M, Kafaja S, Kamgar M, Lau C, Martin WJ, Parikh S, Peng J, Rastogi A, Chen W, Grossman JM, American College of R (2012) American College of Rheumatology guidelines for screening, treatment, and management of lupus nephritis. Arthritis Care Res (Hoboken) 64:797-808 Hanly JG, Su L, Urowitz MB, Romero-Diaz J, Gordon C, Bae SC, Bernatsky S, Clarke AE, Wallace DJ, Merrill JT, Isenberg DA, Rahman A, Ginzler EM, Petri M, Bruce IN, Dooley MA, Fortin P, Gladman DD, Sanchez-Guerrero J, Steinsson K, Ramsey-Goldman R, Khamashta MA, Aranow C, Alarcon GS, Fessler BJ, Manzi S, Nived O, Sturfelt GK, Zoma AA, van Vollenhoven RF, Ramos-Casals M, Ruiz-Irastorza G, Lim SS, Kalunian KC, Inanc M, Kamen DL, Peschken CA, Jacobsen S, Askanase A, Theriault C, Thompson K, Farewell V (2015) Mood Disorders in Systemic Lupus Erythematosus: Results From an International Inception Cohort Study. Arthritis Rheumatol 67:1837-47 He D, Wang J, Yi L, Guo X, Guo S, Guo G, Tu W, Wu W, Yang L, Xiao R, Li Y, Chu H, Lai S, Jin L, Zou H, Reveille JD, Assassi S, Mayes MD, Zhou X (2014) Association of the HLA -DRB1 with scleroderma in Chinese population. PLoS One 9:e106939 IBGE (2008) Características étnico-raciais da população : um estudo das categorias de classificação de cor ou raça, Rio de Janeiro
ACCEPTED MANUSCRIPT
AC C
EP T
ED
MA
NU
SC
RI
PT
Kim K, Bang SY, Lee HS, Okada Y, Han B, Saw WY, Teo YY, Bae SC (2014) The HLA-DRbeta1 amino acid positions 11-13-26 explain the majority of SLE-MHC associations. Nat Commun 5:5902 Lin H, Sui W, Tan Q, Chen J, Zhang Y, Ou M, Xue W, Li F, Cao C, Sun Y, Dai Y (2016) Integrated analyses of a major histocompatibility complex, methylation and transcribed ultraconserved regions in systemic lupus erythematosus. Int J Mol Med 37:139-48 Liphaus BL, Goldberg AC, Kiss MH, Silva CA (2002) Analysis of human leukocyte antigens class II-DR in Brazilian children and adolescents with systemic lupus erythematosus. Rev Hosp Clin Fac Med Sao Paulo 57:277-82 Liphaus BL, Kiss MH, Goldberg AC (2007) HLA-DRB1 alleles in juvenile-onset systemic lupus erythematosus: renal histologic class correlations. Braz J Med Biol Res 40:591-7 Lu LY, Ding WZ, Fici D, Deulofeut R, Cheng HH, Cheu CC, Sung PK, Schur PH, Fraser PA (1997) Molecular analysis of major histocompatibility complex allelic associations with systemic lupus erythematosus in Taiwan. Arthritis Rheum 40:1138-45 Marchini M, Antonioli R, Lleo A, Barili M, Caronni M, Origgi L, Vanoli M, Scorza R (2003) HLA class II antigens associated with lupus nephritis in Italian SLE patients. Hum Immunol 64:462-8 Mok CC (2016) Towards new avenues in the management of lupus glomerulonephritis. Nat Rev Rheumatol 12:221-34 Mok CC, Ying KY, Ng WL, Lee KW, To CH, Lau CS, Wong RW, Au TC (2006) Long-term outcome of diffuse proliferative lupus glomerulonephritis treated with cyclophosphamide. Am J Med 119:355 e25-33 Moroni G, Tantardini F, Gallelli B, Quaglini S, Banfi G, Poli F, Montagnino G, Meroni P, Messa P, Ponticelli C (2005) The long-term prognosis of renal transplantation in patients with lupus nephritis. Am J Kidney Dis 45:903-11 Morris DL, Fernando MM, Taylor KE, Chung SA, Nititham J, Alarcon-Riquelme ME, Barcellos LF, Behrens TW, Cotsapas C, Gaffney PM, Graham RR, Pons-Estel BA, Gregersen PK, Harley JB, Hauser SL, Hom G, Langefeld CD, Noble JA, Rioux JD, Seldin MF, Systemic Lupus Erythematosus Genetics C, Vyse TJ, Criswell LA (2014) MHC associations with clinical and autoantibody manifestations in European SLE. Genes Immun 15:210-7 Morris DL, Taylor KE, Fernando MM, Nititham J, Alarcon-Riquelme ME, Barcellos LF, Behrens TW, Cotsapas C, Gaffney PM, Graham RR, Pons-Estel BA, Gregersen PK, Harley JB, Hauser SL, Hom G, International MHC, Autoimmunity Genetics N, Langefeld CD, Noble JA, Rioux JD, Seldin MF, Systemic Lupus Erythematosus Genetics C, Criswell LA, Vyse TJ (2012) Unraveling multiple MHC gene associations with systemic lupus erythematosus: model choice indicates a role for HLA alleles and non-HLA genes in Europeans. Am J Hum Genet 91:778-93 Niu Z, Zhang P, Tong Y (2015) Value of HLA-DR genotype in systemic lupus erythematosus and lupus nephritis: a meta-analysis. Int J Rheum Dis 18:17-28 Olsen ML, Arnett FC, Reveille JD (1993) Contrasting molecular patterns of MHC class II alleles associated with the anti-Sm and anti-RNP precipitin autoantibodies in systemic lupus erythematosus. Arthritis Rheum 36:94-104 Segurado OG, Iglesias-Casarrubios P, Martinez-Laso J, Corell A, Martin-Villa JM, Arnaiz-Villena A (1991) Autoimmunogenic HLA-DRB1*0301 allele (DR3) may be distinguished at the DRB1 non-coding regions of HLA-B8,DR3,Dw24 and B18,DR3,Dw25 haplotypes. Mol Immunol 28:189-92 Shimane K, Kochi Y, Suzuki A, Okada Y, Ishii T, Horita T, Saito K, Okamoto A, Nishimoto N, Myouzen K, Kubo M, Hirakata M, Sumida T, Takasaki Y, Yamada R, Nakamura Y, Kamatani N, Yamamoto K (2013) An association analysis of HLA-DRB1 with systemic lupus erythematosus and rheumatoid arthritis in a Japanese population: effects of *09:01 allele on disease phenotypes. Rheumatology (Oxford) 52:1172-82
ACCEPTED MANUSCRIPT
AC C
EP T
ED
MA
NU
SC
RI
PT
Suggs MJ, Majithia V, Lewis RE, Cruse JM (2011) HLA DRB1*1503 allelic haplotype predominance and associated immunodysregulation in systemic lupus erythematosus. Exp Mol Pathol 91:548-62 Tian W, Li LX, Guo SS (2000) [Correlative study on HLA-DR2 allelic polymorphism and systemic lupus erythematosus in the Han nationality in Hunan province]. Hunan Yi Ke Da Xue Xue Bao 25:15-7 Usnayo MJ, Andrade LE, Alarcon RT, Oliveira JC, Silva GM, Bendet I, Burlingame R, Porto LC, Pinheiro Gda R (2011) Study of the frequency of HLA-DRB1 alleles in Brazilian patients with rheumatoid arthritis. Rev Bras Reumatol 51:474-83 Wadi W, Elhefny NE, Mahgoub EH, Almogren A, Hamam KD, Al-Hamed HA, Gasim GI (2014) Relation between HLA typing and clinical presentations in Systemic Lupus Erythematosus patients in Al-Qassim region, Saudi Arabia. Int J Health Sci (Qassim) 8:159-65 Wahren-Herlenius M, Dorner T (2013) Immunopathogenic mechanisms of systemic autoimmune disease. Lancet 382:819-31 Ward MM (2000) Access to renal transplantation among patients with end-stage renal disease due to lupus nephritis. Am J Kidney Dis 35:915-22
ACCEPTED MANUSCRIPT Table 1- Age (mean), Gender (index Female/Male) and Ethnicity (white/non-white) of the transplanted SLE (TxSLE), non-transplanted SLE (nTxSLE), Kidney-transplanted for other diseases (TxCTRL) and healthy control (HeControl) subgroups. N (total patients)
Age Gender White/Nona b (range) (F/M) whitec 33.0 TxSLE 44 6.33 20/24 (12-56) Lupus (GSLE) 37.8 NTxSLE 64 5.40 23/41 (20-59) 39.5 TxCTRL 88 6.33 43/45 Control (11-71) (Gcontrol) 43.2 HeControl 128 6.52 50/78 (20-59) HeControl, nTxSLE and TxCTRL were paired to TxSLE for ethnicity and gender. a: X2 = 43.504 p = 0.0; b: X2 = 0.207, p = 0.976; c: X2 = 3.336, p = 0.342.
EP T
ED
MA
NU
SC
RI
PT
Subgroup
AC C
Group
ACCEPTED MANUSCRIPT
AC C
EP T
ED
MA
NU
SC
RI
PT
Table 2. Prevalence of Alleles in the Control Group (Gcontrol) X General Lupus group (GSLE) Allele GControl GSLE OR p-value Pc-value N=216 N=108 B*08 17 (17.9%) 20 (18.5%) 2.66 0.0048 0.1379 B*15 51 (23.6%) 15 (13.9%) 0.52 0.0265 0.7697 B*27 10 (4.7%) 0 (0.0%) 0.00 0.0161 0.4683 B*44 42 (19.4%) 12 (16.7%) 0.51 0.0384 1.1133 DRB1*08 20 (9.3%) 22 (20.4%) 2.50 0.0050 0.0653 DRB1*11 49 (22.7%) 13 (12%) 0.46 0.0140 0.1824 DRB1*13 65 (30%) 20 (18,5%) 0.52 0.0166 0.2159 DRB1*15 50 (23.2%) 45 (41.7%) 2.37 0.0005 0.0067 Total number of alleles found per locus (Locus A – 20, Locus B – 39 and Locus DRB1 – 13) were used to calculate Pc (Bonferroni correction).
ACCEPTED MANUSCRIPT
AC C
EP T
ED
MA
NU
SC
RI
PT
Table 3. Allele Prevalence in Health Control (HeControl), kidney transplanted for other diseases (TxCTRL), non-transplanted SLE (nTxSLE) and transplanted SLE (TxSLE) groups. Allele HeControl TxControl nTxSLE TxSLE p-value Pc-value N=128 N=88 N=64 N=44 B*08 8 9 13 7 0.0246 0.7134 (6.2%) (10.2%) (20.4%) (15.9%) DRB1*08 10 10 14 8 0.0324 0.4212 (7.8%) (11.4%) (21.9%) (18.2%) DRB1*15 43 12 24 21 0.0041 0.0533 (19.9%) (11.1%) (37,5%) (47,7%) Total number of alleles found per locus (Locus A: 20, Locus B: 39 and Locus DRB1: 13) were used calculating Pc (Bonferroni correction).
ACCEPTED MANUSCRIPT
AC C
EP T
ED
MA
NU
SC
RI
PT
Table 4. Prevalence of alleles in comparison of transplanted lupus patients (TxSLE) with transplanted control patients (TxCTRL) Allele TxSLE TxCTRL Odds p-value Pc –value N=44 N=88 Rate # A*30 14 (31.8%) 13 (14.7%) 2.69 0.021 0.426 B*18 7 (15.9%) 3 (3.4%) 5.36 0.016 0.459 B*49 14 (9.09%) 1 (1.14%) 8.7 0.042 1.221 DRB1*07 12 (27.2%) 12 (13.6%) 2.37 0.049 0.637 DRB1*15 21 (47.7%) 21 (23.8%) 2.91 0.005 0.065 #- HLA-A*30 was more frequent in non-white TxSLE patients (p= 0.048 – OR= 2.75).
ACCEPTED MANUSCRIPT Figure 1 – Lupus Patient and Control Groups
nTxSLE Lupus Nephritis without Transplant N=32
TxCTRL Ki deny Tra ns planted pa ti ents with others di s eases N=88
SC NU MA ED EP T AC C
N=128
RI
TxSLE kidney transplanted patients with Lupus Nephritis N=44
HeCTRL Hea lth people
PT
nTxSLE Lupus patients without nephritis N=32
ACCEPTED MANUSCRIPT
AC C
EP T
ED
MA
NU
SC
RI
PT
Figure 2. Cumulative survival in 5 years – HLA DRB1*15 comparison of both groups
ACCEPTED MANUSCRIPT
AC C
EP T
ED
MA
NU
SC
RI
PT
Figure 3. Cumulative survival in 5 years - HLA DRB1*15 Match-comparison of both groups
ACCEPTED MANUSCRIPT THE PREVALENCE OF HLA ALLELES IN A LUPUS NEPHRITIS POPULATION Highlights
Lupus nephritis (LN) is a complication of Systemic lupus erythematosus (SLE) and is associated with poor survival and high morbidity. Many genomic studies have been performed worldwide, and several histocompatibility leukocyte
PT
antigen (HLA) loci are linked to lupus susceptibility. This study was a retrospective study of 2 major groups: general lupus patients
SC
A difference in the prevalence of three alleles, B*08, DRB1*08 and DRB1*15,
NU
was observed. These alleles were more prevalent in the lupus subgroups compared to the control groups. The presence of HLA-DRB1*15 was a strong
MA
factor that predisposed patients to the development of SLE and LN, but did not
EP T
ED
influence kidney graft survival.
AC C
RI
(n=108) and a control group (n=216).