Acute HUS: fast-track genetics

Timely genetic analysis in hemolytic uremic syndrome (HUS) has remained elusive; however, in this issue of Blood, Yousfi et al¹ describe the utility of nanopore sequencing to deliver results in hours to days.

Hemolytic uremic syndrome (HUS) was first described in 1955 by Gasser and colleagues, characterized by renal failure, hemolytic anemia, and thrombocytopenia.² However, differentiation from other thrombotic microangiopathies, in particular thrombotic thrombocytopenia purpura, was not achieved until the late 1990s, with the identification of ADAMTS13 (a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13), but also appreciation that HUS is caused by dysregulation of the alternative complement pathway. The abnormality in HUS was factor H, initially characterized in 1998.³ Ultimately, this led to the development of complement inhibitor therapy, licensed since 2012, which has revolutionized the outcome of patients with complement-mediated HUS (CM-HUS).

There are over 500 variants identified in complement genes, including those causing loss or gain of function of complement regulators (complement factors H and I and membrane cofactor protein) or effecting complement 3 convertase (C3 and complement factor B). The impact of variants is diverse with respect to time of presentation and influence on renal outcome. There is the additional impact of other genetic modifiers and environmental elements, known as the “second-hit” hypothesis.⁴ 

CM-HUS is a clinical diagnosis that can be confirmed by complement genetic analysis. However, genetic analysis may take weeks (or months). Yousfi and colleagues present the results of nanopore sequencing in 3 retrospective and 18 prospective HUS cases, with results available in days, compared with standard exome sequencing, which takes weeks (or months). The authors were originally using whole-exome sequencing for single-nucleotide variants and multiple ligation-dependent probe amplification for copy variant numbers, with a turnaround for results of up to 8 weeks⁵ (see figure). With the goal of improving turnaround times, they investigated Oxford Nanopore technology, which provided additional advantages over standard sequencing platforms. Specifically, long-read sequencing detects complex structural variants that may be more difficult to identify with short-read current standard methodologies. There is comparable detection of single-nucleotide variants but greater detection of hybrid genes and intronic variants when compared with the current methodology. The significant benefit of nanopore technology is the speed of results, within hours to days, associated with a lower cost due to fewer consumables and a lower computing expense. Compared with standard whole-genome sequencing, there was a 50% reduction in price with nanopore technology. The authors validated their results on 2 nanopore platforms, confirming a higher sensitivity and specificity of variant detection.

Yousfi et al also describe challenging clinical scenarios, including cobalamin C disease, identifying a homozygous metabolism of the cobalamin-associated C gene. Cobalamin C disease is an inborn metabolic disorder, typically presenting in childhood; the identification within the faster genetic panel could have improved outcome of patients by the administration of vitamin B12 replacement therapy. A further example includes complement factor H, the most severe form of complement factor dysregulation, which allowed genetic confirmation within 3 days and prompt initiation of complement inhibitor therapy. Finally, there is Shiga toxin–producing Escherichia coli HUS, for which complement inhibitor therapy is not licensed, but genetic analysis may identify an underlying complement genetic variant that may alter the therapeutic plan in favor of a complement inhibitor treatment.

The speedy availability of genetic results may expedite the rationale for complement inhibitor therapy and may avert the need for kidney biopsy in acute kidney injury, especially with associated thrombocytopenia and hypertension, usually in critically ill patients with acute worsening of their condition. It is conceivable that the genomic “pick-up” rate will increase. How will those cases with a negative genetic profile be managed? Currently, 60% of CM-HUS cases have a detectable complement-associated genetic variant identified.⁶ Although there have been national variations,⁷ there remains a cohort of patients with no identified genetic abnormality but a clinical picture consistent with a clinical diagnosis of CM-HUS and a very good response to complement inhibitor therapy. Genotype-phenotype associations have suggested these cases have a very low relapse rate,⁸ and therefore clinical management still requires consideration of a negative gene profile.

Nanopore technology uses electrical signals rather than conventional light signal platforms.⁹ Its utility is scalable from “point-of-care” to high-throughput analysis.

Yousfi et al have identified significant additional benefits with this technology from a small cohort of patients. Further validation in a larger study population may not only lead to a time-critical diagnostic solution but may also identify further genetic variants as precipitants of this acute thrombotic microangiopathy.

Conflict-of-interest disclosure: M.S. has received speakers’ fees from Takeda, Sanofi, Alexion, and Octapharma and research grants from Takeda and Alexion.