Genetic Testing and Kidney Biopsies: Complementary to Another, not Substitutive

Introduction

Genetic testing is being more frequently used in the work-up of patients with kidney disease, including those with chronic kidney disease and cases detected at a young age.¹ This is in part because of the greater availability from commercial and academic laboratories lower cost that decreases barriers to testing. Typically, such testing also includes genetic counseling, which is beneficial for both the ordering provider and the patients. Patients developing kidney disease at a young age, with a family history of kidney disease, cystic or anatomic abnormalities, and with extrarenal manifestations (e.g., liver cysts, developmental delay, vision or hearing loss) will have a higher diagnostic yield of genetic testing; the 2024 Kidney Disease: Improving Global Outcomes guidelines include genetic testing as a tool for the work-up of chronic kidney disease.^1,2 Although a kidney biopsy can provide details regarding acute and/or chronic histopathologic features, genetic testing can provide insight into modes of inheritance from which the biopsy alone cannot divulge.

We describe a young patient who presented with hematuria and proteinuria, and underwent initial genetic testing that found evidence of a heterozygous likely pathogenic variant in COL4A3 associated with COL4A3-related Alport syndrome. A kidney biopsy subsequently performed revealed the absence of abnormality of the glomerular basement membrane (GBM) and the presence of a membranous nephropathy (MN). The case importantly illustrates the need for genetic testing to be interpreted together with kidney biopsy findings in order to provide the most appropriate and timely patient care, and leverage genetic counseling to assist nephrologists in the interpretation of results derived from such testing.

Case Presentation

A 20-year-old woman presented with development of generalized body aches, rash, and periorbital edema over 5 months. Past medical history was notable for dysmenorrhea, pagophagia, and Chlamydia infection, and there was no known family history of kidney disease. Initial laboratory testing showed 2+ proteinuria and 2+ hematuria, with otherwise normal renal function (further details are provided in Supplementary Table S1). Examination was notable for periorbital edema and an erythematous pruritic rash throughout the upper extremities and neck; notably absent were ocular or auditory deficits. Ultrasound examination showed only mild right-sided hydronephrosis. Based on the patient’s age and presentation, a large multigene, next generation sequencing panel was ordered and resulted with a heterozygous likely pathogenic variant in COL4A3 (c.190G>C [p.Gly64Arg]) associated with COL4A3-related Alport syndrome. A kidney biopsy was subsequently performed and revealed MN with extraglomerular deposits and positive glomerular staining for exostosin (EXT)-2 (EXT2), favored to be associated with systemic autoimmune disease; electron microscopy at this time was notably negative for evidence of GBM abnormalities (Figure 1). Shortly after, a skin biopsy showed evidence of interface dermatitis, and additional or repeated laboratory testing performed during a rheumatology consultation now showed positive autoimmune serologies (Supplementary Table S2), altogether fitting for systemic lupus erythematosus. With these additional data points, the diagnosed MN would more specifically represent membranous lupus nephritis, class V. Details of the patient’s subsequent clinical course and management are presented in Supplementary Table S3.

Figure 1.

Immunofluorescence microscopy (on a scale of 0 to 3+) revealed (a and b) global granular glomerular capillary loop and mesangial 3+ staining for IgG, kappa, and lambda, 2+ for C1q, and 1+ for C3; (a, arrows) in addition, there was evidence of extraglomerular deposits along vessel walls and very focally along tubular basement membranes. (c) Immunoperoxidase staining for exostosin-2 was positive within the glomeruli. Electron microscopy showed (d, arrows highlight select deposits) globally distributed subepithelial and (d, asterisk) mesangial, and (d, arrowhead) fewer subendothelial finely granular electron dense deposits; (d, encircled) few glomerular endothelial cells contained tubuloreticular inclusions. A total of 8 glomerular were evaluated using electron microscopy, none of which showed abnormal ultrastructural features of the glomerular basement membranes (i.e., no basketweave appearance, splitting, or fraying of the lamina densa, and the glomerular basement membranes were not thinned). Original magnifications for a and b at ×400; for c at ×600; and for d at ×7000.

Discussion

Genetic etiologies of kidney disease can be implicated in ≤70% of pediatric and 10% to 20% of adult patients with chronic kidney disease; thus, genetic testing in the appropriate scenario is a diagnostic tool in the work-up of certain patients.³ The 2024 Kidney Disease: Improving Global Outcomes guidelines include genetic testing as a tool in the evaluation of patients with chronic kidney disease, further providing nephrologists the impetus to order such testing.² Results from genetic testing can assist in establishing or confirming a diagnosis, identify modes of inheritance to assist in family planning, identify potential at-risk relatives, facilitate disease management, and others. However, the understanding of the given results by the end-using caring nephrologist is paramount to its appropriate application and integration into care. Complicating matters in this regard includes the nomenclature and nuances accompanying the results of examined variants such as “pathogenic”, “likely pathogenic”, and “variant of uncertain significance” as well as autosomal dominant versus autosomal recessive inheritance patterns such nuance is illustrated in this case.

The genetic variant identified in this patient is a missense change that converts a glycine to an arginine at position 64 in the COL4A3 gene. This amino acid change has not been previously reported in affected patients in the literature but has been observed at a frequency of < 0.01% (8/1,612,040 alleles) in the Broad gnomAD control dataset. This variant is located in the collagen triple helix repeat domain and disrupts the Gly-X-Y motif; other glycine amino acid changes in this region have been reported in individuals with Alport syndrome. Therefore, based on the available information, this variant was interpreted as likely pathogenic. Heterozygous pathogenic and likely pathogenic COL4A3 variants, genetically described as autosomal dominant Alport syndrome, are increasingly identified in a genotype-first approach. However, patient phenotypes often do not exhibit the classic ocular, auditory, and renal manifestations of “classic” Alport syndrome, typically described as being inherited as an autosomal recessive or X-linked recessive disease. Further, the classic structural changes observed on kidney biopsy, primarily GBM lamellation, are rare in heterozygotes, albeit cases may exhibit GBM thinning or more focal and subtle ultrastructural irregularities. As genotype-first testing becomes more prevalent in clinical practice, the interpretation of phenotypic manifestations relative to genetic results will continue to evolve and require expert assessment and interpretation.

The contemporary approach toward MN of determining related antigens has allowed nephrologists more targeted investigations into underlying associated etiologies. In cases of EXT1/EXT2-associated MN, patients are overwhelmingly (∼ 80%) found to have forms of systemic autoimmune disease, including systemic lupus erythematosus, mixed connective tissue disease and overlap syndromes, and Sjogren syndrome.⁴ In this case, although preliminary antinuclear antibody testing before the kidney biopsy was negative, the biopsy finding of EXT2-associated MN redirected further testing that would contribute to the patient soon after being diagnosed with systemic lupus erythematosus and subsequently started on an immunosuppressive regimen. Not only did the kidney biopsy provide direction for a thorough rheumatologic work-up, but as shown, was notable for the absence of GBM abnormalities. Although in the dichotomous “primary” versus “secondary” MN classification, this case would fall under the latter in part based on negative staining for phospholipase A2 receptor as well as extraglomerular immune complex deposits and would spurn on an autoimmune work-up among numerous other testing and lines of inquiry (infection, malignancy, etc.,), the finding of EXT2-positive staining provided a more narrowed and efficient subsequent patient evaluation. Of note, the subepithelial deposits were present but without related GBM remodeling (stage 1 MN features), thereby not confounding the examination of the GBM’s in this case.

The presented case highlights the importance of integrating genetic testing results with kidney biopsy findings (Table 1); if only the former were depended upon for decision making in the absence of a biopsy, the patient would have not received timely appropriate care. For this case in particular, based on the patient’s presentation, a kidney biopsy should have preceded genetic testing. Further illustrated is the critical role of integrating genetic counseling in the interpretation and contextualizing of genetic results, namely because it relates to the clinical impact, actionability, and practical meaning of any discovered genetic variants. As genetic testing becomes more widely used in nonacademic nephrology practice settings, the involvement and guidance of genetic counselors will be key in providing optimal clinical care. Lastly, the biopsy itself illustrates the importance of determining the related antigen in a case of MN, thereby allowing a narrowing of the differential of associated etiologies, and in the case of EXT1/EXT2-associated MN, systemic autoimmune disease.

Table 1.

Teaching points

Results from genetic testing should be used to complement histopathology.

The interpretation of genetic testing results may require discussion with and guidance from a geneticist.

Membranous glomerulopathy with extraglomerular deposits is often associated with systemic autoimmune disease.

Disclosure

All the authors declared no competing interests.

Patient Consent

The authors declare that they have obtained consent from the patient discussed in the report.

Supplementary Material

Supplementary File (PDF)

Table S1. Initial laboratory testing and results (before genetic testing and the kidney biopsy). Table S2. Additional or repeated laboratory testing performed during rheumatology consultation 2 weeks after the kidney biopsy. Table S3. Patient’s clinical course and management after the kidney biopsy.