- Case report
- Open Access
A case of refractory systemic lupus erythematosus with monocytosis exhibiting somatic KRAS mutation
Inflammation and Regeneration volume 42, Article number: 10 (2022)
Systemic lupus erythematosus (SLE), an autoimmune disorder that damages various organ systems, is caused by a combination of genetic and environmental factors. Although germline mutations of several genes are known to cause juvenile SLE, most of the susceptibility genetic variants of adult SLE are common variants of the population, somatic mutations that cause or exacerbate SLE have not been reported. We hereby report a refractory SLE case with monocytosis accompanying somatic KRAS mutation that have been shown to cause lupus-like symptoms.
A 60-year-old female patient who had been diagnosed with SLE was admitted to our hospital. Although prednisolone and tacrolimus treatments had kept her thrombocytopenia and anti-DNA Ab level at bay for more than 4 years, a diagnosis of transverse myelitis was made when she became acutely ill with pleocytosis. Elevated cells (predominately monocytes), protein, IgG, and IL-6 levels were also found in the cerebrospinal fluid (CSF) of the patient. Standard pulse treatments of methylprednisolone, high-dose of prednisolone, and intravenous cyclophosphamide in combination with plasma exchange could not alleviate the refractory neural and autoimmune manifestation. Monocytosis of peripheral blood was also noted. Flow cytometric analysis revealed elevated ratio of CD14+CD16+ atypical monocytes, which excluded the possibility of chronic myelomonocytic leukemia. Lupus-like symptoms with monocytosis reminded us of Ras-associated autoimmune leukoproliferative disorder, and Sanger sequencing of KRAS and NRAS genes from the patients’ peripheral blood mononuclear cells (PBMC), sorted CD3+ lymphocytes and CD14+ monocytes, and cerebrospinal fluid were performed. An activating KRAS somatic mutation was found in the patients’ DNA at the time of encephalomyelitis diagnosis.
Somatic mutations of some genes including KRAS may cause the refractoriness of SLE.
Systemic lupus erythematosus (SLE) is a chronic autoimmune disease that affects any parts of the body, characterized by the production of autoantibodies. Cumulative evidences have pointed to the conglomerate of genetic and environmental factors that constitute the pathogenesis of SLE.
Although SLE is a polygenic disorder, monogenic cases have been reported . Such monogenic SLE patients can be grouped into interferonopathy-like, self-tolerance breakdown-associated, complement deficiency-like, RASopathy-like, and others. RASopathies are rare neurodevelopmental syndromes that are caused by dominantly inherited mutations in several genes within the RAS/MAPK pathway (KRAS, NRAS, PTPN11, RAF, SHOC2, and SOS1). Meanwhile, Ras-associated autoimmune leukoproliferative disorder (RALD) is caused by somatic, gain-of-function NRAS or KRAS mutation within hematopoietic lineage. It was initially reported in a patient presenting autoimmune lymphoproliferative syndrome (ALPS)-like symptom with normal FAS-mediated apoptotic pathway . Both KRAS and NRAS activating mutations, produce characteristic and persistent monocytosis accompanied by high frequency of autoimmunity and antinuclear antibody production , symptoms reminiscent of SLE patients.
A 60-year-old female SLE patient was admitted to our hospital due to fever and paralysis of both of her lower legs. Four years before this admission, she had joint swelling on her fingers, wrists, right shoulder, knees, and non-palpable purpura on the forearms and thighs. Thrombocytopenia, elevated erythrocyte sedimentation rate, serum C-reactive protein (CRP), and total serum IgG; as well as reduced complement activity were reported. The patient presented antinuclear antibody (ANA) of 1:320 (homogeneous/speckled type), anti-dsDNA antibody (anti-DNA Ab) of 156 IU/ml, anti-SS-A Ab, circulating immune complex, and prolonged lupus anticoagulant. Based on the 2019 revised European Alliance of Associations for Rheumatology (EULAR) /American College of Rheumatology (ACR) classification criteria, she was diagnosed with SLE. Thrombocytopenia and anti-DNA Ab level were relieved and maintained by the treatment of prednisolone and tacrolimus for more than 4 years (Fig. 1). Four days before the admission, however, she became acutely ill with a fever of 38°. Subsequently, the patient suffered from sudden laxative total paralysis in both lower limbs. Her bladder and rectum showed dysfunction with high intensity signal on the cervical cord of MRI (Fig. 2). Pleocytosis, along with elevated protein, IgG and IL-6 levels in the cerebrospinal fluid (CSF) were detected. The patient presented mild leukocytosis with neutrophil predominance, moderate thrombocytopenia, elevated CRP and serum IgG amount, prolonged active prothrombin time, anti-DNA Ab, anti-ribosomal P antibody and marked reduction of complement activity (Table 1). The diagnosis of transverse myelitis due to the flare of SLE was made; then, standard pulse therapy of methylprednisolone was followed by high-dose of prednisolone, intravenous cyclophosphamide in combination with plasma exchange. In the following months, her refractory neurological and autoimmune symptoms continued to fluctuate, and failed to achieve remission with the appropriate treatments (Fig. 1). The apparent histologically normal monocytes (Fig. 3A) continued to cloud the clinical features of the patient whose monocytosis persisted. Flow cytometry analyses revealed an elevated ratio of CD14+CD16+ atypical monocytes, CD14+CD16+ atypical granulocytes as well as CD4+CD8+TCRαβ+ double positive T cells as compared to the healthy control (Fig. 3B). Increased atypical monocytes excluded chronic myelomonocytic leukemia ; and lupus-like symptoms with monocytosis together with these immune-phenotypes reminded us of previously-reported RALD cases, although these immunophenotypes can be seen in SLE cases. Therefore, we performed Sanger sequencing of KRAS and NRAS genes from the patients’ PBMC (Fig. 4A), magnetic-activated cell sorting (MACS)-sorted CD3+ lymphocytes and CD14+ monocytes, together with CSF (Fig. 4B). A point mutation in codon 12 of KRAS (c.G35A, p.G12D) was detected. Since the mutation was almost absent in the buccal mucosa sample of the patient (Fig. 4A), the mutation is likely to be somatic and not germline, as in RASopathies. A backtrack of the patient’s cell-free DNA obtained by the stocked serum samples showed that the mutation manifested at the time of encephalomyelitis diagnosis, and persisted for 5 years (Fig. 4C). This mutation seems to be involved in lupus flare of this case, as it was not detected when she was diagnosed of SLE (Fig. 4C), or in another SLE patient (Fig. 4A).
Discussion and conclusions
KRAS G12D mutation is a missense mutation that results in a constitutive activating function . In the immune system, the KRAS molecule regulates normal hematopoiesis . Constitutive active mutation of the RAS protein results in breakdowns of B cell self-tolerance and productions of autoantibody , potentially by disruption of the RAS/MAPK regulatory pathway.
Given the many overlapping symptoms between various autoimmune disorders, it is still difficult to have a clear cut in the diagnosis of various similar connective tissue autoimmune diseases. Since RALD patients are known to present symptoms mimicking SLE  and KRAS mutation can also give rise to monogenic pediatric SLE , one could argue whether this case was SLE or RALD. Although she did not show the typical SLE symptoms like marlar rash or glomerulonephritis, she satisfied the 2019 EULAR/ACR criteria with 22 points (one can be classified as SLE with ≥ 10 points). Therefore, there is no doubt that she had SLE at least at the time of the onset, when we could not detect the mutation using the cell-free DNA in the stocked serum. However, it is not clear whether KRAS mutation was responsible for the recurrence and refractoriness of SLE.
The somatic KRAS mutation detected could be explained by clonal hematopoiesis. Throughout the genome, about 20 somatic mutations are estimated to accumulate in human hematopoietic stem cells (HSCs) each year [10, 11], and about 0.1 mutations in protein-coding exons . By the age of 70, we may harbor from 350,000 to 1,400,000 coding mutations within the HSC pool . When one of these mutations was able to expand due to certain selective advantages, it is coined clonal hematopoiesis. The current accepted definition of clonal hematopoiesis is when a clone reaches a proportion of 4% of cells measured in the peripheral blood . Although most clonal hematopoiesis are linked to hematologic cancers, some mutations found their ways in circulating immune cells like monocytes, which could alter the immune responses of the patients . Unfortunately, we did not collect samples and conduct necessary experiments to check if 4% of her cells in the peripheral blood have this clone, while the patient was residing in our hospital. With that said, when the KRAS mutation was identified, the patient was in her 60s, approaching the age when an acquired mutation would have enough time to accumulate and then expand to reach the classical definition of ~ 4% of cells in the peripheral blood, or of a variant allele frequency of over 2% in the blood . Comparing to genes that are more commonly mutated in clonal hematopoiesis (DNMT3A, 48.3%), KRAS mutations contribute to about 1.3% of the current cases discovered .
Unlike the limited number of genes such as DNMT3A, which enhances self-renewal of stem and progenitor cells ; KRAS, a small GTPase could be causing some of the resistance in the treatments for our patient. Autoreactive immature B cells usually reside in the bone marrow, but Teodorovic et al. have shown that activation of Ras can change this pattern, altering the Ras-erk pathway, and resulting in secretion of autoantibodies. Although not the same mutation (N-RasD12), KRAS (G12D) is also an activating mutation, which could give rise to the production of autoantibodies. Using serial transplantation mouse studies might be able to help elucidate KRAS (G12D) function in the treatment resistance. Given the molecular complexity of different autoimmune diseases, we could only deduce that this is a case of refractory SLE with monocytosis, accompanying a somatic KRAS mutation. Since other genetic alterations can also trigger SLE-like and RALD-like conditions, we would need to conduct a more in-depth investigation of other potential somatic mutations such as PTPN11, RAF, SHOC2 and SOS1 to understand this case better. Even so, we believe this case provides an additional reference for understanding complicated SLE pathogenesis.
Availability of data and materials
No datasets were generated or analyzed during the current study.
Systemic lupus erythematosus
American College of Rheumatology
Kirsten Rat Sarcoma Proto-Oncogene
- GTPase; NRAS:
Neuroblastoma rat sarcoma proto-oncogene
- GTPase; MAPK:
Mitogen-activated protein kinase
Protein tyrosine phosphatase non-receptor type 11
Raf-1 proto-oncogene, serine/threonine kinase
Suppressor of clear homolog (C. Elegans)
Leucine-rich repeat scaffold protein 2
Son of Sevenless Homolog
Guanine nucleotide exchange factor 1
Ras-associated autoimmune leukoproliferative disorder
Autoimmune lymphoproliferative syndrome
Fas cell surface death receptor
- Anti-DNA Ab:
Magnetic-activated cell sorting
Peripheral blood mononuclear cells
Demirkaya E, Sahin S, Romano M, Zhou Q, Aksentijevich I. New horizons in the genetic etiology of systemic lupus erythematosus and lupus-like disease: monogenic lupus and beyond. J Clin Med. 2020;9(3).
Oliveira JB, Bidere N, Niemela JE, Zheng L, Sakai K, Nix CP, et al. NRAS mutation causes a human autoimmune lymphoproliferative syndrome. Proc Natl Acad Sci U S A. 2007;104(21):8953–8. https://doi.org/10.1073/pnas.0702975104.
Calvo KR, Price S, Braylan RC, Oliveira JB, Lenardo M, Fleisher TA, et al. JMML and RALD (Ras-associated autoimmune leukoproliferative disorder): common genetic etiology yet clinically distinct entities. Blood. 2015;125(18):2753–8. https://doi.org/10.1182/blood-2014-11-567917.
Tarfi S, Harrivel V, Dumezy F, Guy J, Roussel M, Mimoun A, et al. Multicenter validation of the flow measurement of classical monocyte fraction for chronic myelomonocytic leukemia diagnosis. Blood Cancer J. 2018;8(11):114. https://doi.org/10.1038/s41408-018-0146-8.
Seeburg PH, Colby WW, Capon DJ, Goeddel DV, Levinson AD. Biological properties of human c-Ha-ras1 genes mutated at codon 12. Nature. 1984;312(5989):71–5. https://doi.org/10.1038/312071a0.
Damnernsawad A, Kong G, Wen Z, Liu Y, Rajagopalan A, You X, et al. Kras is required for adult hematopoiesis. Stem cells. 2016;34(7):1859–71. https://doi.org/10.1002/stem.2355.
Teodorovic LS, Babolin C, Rowland SL, Greaves SA, Baldwin DP, Torres RM, et al. Activation of Ras overcomes B-cell tolerance to promote differentiation of autoreactive B cells and production of autoantibodies. Proc Natl Acad Sci U S A. 2014;111(27):E2797–806. https://doi.org/10.1073/pnas.1402159111.
Li G, Li Y, Liu H, Shi Y, Guan W, Zhang T, et al. Genetic heterogeneity of pediatric systemic lupus erythematosus with lymphoproliferation. Medicine. 2020;99(20):e20232. https://doi.org/10.1097/MD.0000000000020232.
Ragotte RJ, Dhanrajani A, Pleydell-Pearce J, Del Bel KL, Tarailo-Graovac M, van Karnebeek C, et al. The importance of considering monogenic causes of autoimmunity: A somatic mutation in KRAS causing pediatric Rosai-Dorfman syndrome and systemic lupus erythematosus. Clinical immunology. 2017;175:143–6. https://doi.org/10.1016/j.clim.2016.12.006.
Lee-Six H, Øbro NF, Shepherd MS, Grossmann S, Dawson K, Belmonte M, et al. Population dynamics of normal human blood inferred from somatic mutations. Nature. 2018;561(7724):473–8. https://doi.org/10.1038/s41586-018-0497-0.
Osorio FG, Rosendahl Huber A, Oka R, Verheul M, Patel SH, Hasaart K, et al. Somatic mutations reveal lineage relationships and age-related mutagenesis in human hematopoiesis. Cell Rep. 2018;25(9):2308–16.e4.
Welch JS, Ley TJ, Link DC, Miller CA, Larson DE, Koboldt DC, et al. The origin and evolution of mutations in acute myeloid leukemia. Cell. 2012;150(2):264–78. https://doi.org/10.1016/j.cell.2012.06.023.
Jaiswal S. Clonal hematopoiesis and nonhematologic disorders. Blood. 2020;136(14):1606–14. https://doi.org/10.1182/blood.2019000989.
Challen GA, Goodell MA. Clonal hematopoiesis: mechanisms driving dominance of stem cell clones. Blood. 2020;136(14):1590–8. https://doi.org/10.1182/blood.2020006510.
Jeong M, Park HJ, Celik H, Ostrander EL, Reyes JM, Guzman A, et al. Loss of Dnmt3a Immortalizes Hematopoietic Stem Cells In Vivo. Cell Rep. 2018;23(1):1–10. https://doi.org/10.1016/j.celrep.2018.03.025.
The present study was partly supported by the research grant from Foundation for Biomedical Research and Innovation at Kobe.
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This study was designed in accordance with the Helsinki Declaration, and was approved by Kyoto University Graduate School and Faculty of Medicine, Ethics Committee (approval number: G1006) and written informed consent was obtained from the patient.
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Law, SM., Akizuki, S., Morinobu, A. et al. A case of refractory systemic lupus erythematosus with monocytosis exhibiting somatic KRAS mutation. Inflamm Regener 42, 10 (2022). https://doi.org/10.1186/s41232-022-00195-w
- Systemic lupus erythematosus
- Somatic mutation