Hereditary Cystic Kidney Diseases Panel

Hereditary cystic kidney diseases manifest in children and adults with variable expression of renal cysts as well as unique extra-renal manifestations in each disease. These diseases can be recognized in terms of underlying etiology (ciliopathies or phakomatoses) or morphologic appearance (size, location, and complexity) (Dillman et al. 2017. PubMed ID: 28493804; Bergmann. 2017. PubMed ID: 29479522). In addition to radiological imaging, genetic testing of a panel of relevant genes simultaneously is able to help ensure a conclusive diagnosis.

The kidney ciliopathic disorders primarily include autosomal dominant polycystic kidney disease (ADPKD), autosomal recessive polycystic kidney disease (ARPKD), nephronophthi­sis and medullary cystic kidney disease (MCKD). Manifestations in skeletal (Jeune syndrome), central nervous system (CNS) (Meckel-Gruber syndrome and Joubert syndrome) and other internal organs (ADPKD and ARPKD) are well documented. The cysts in renal ciliopathies can be macroscopic (ADPKD) or microscopic (ARPKD, nephronophthi­sis and MCKD).

The phakomatoses comprise a heterogeneous group of hereditary neurocutaneous multi­system disorders. Tissues of ectodermal origin such as CNS, eyes, and skin are commonly affected. Abdominal manifestations are also common in this group of disorders such as renal cystic lesions in patients with tuberous sclerosis complex (TSC) and von Hippel–Lindau syndrome. The cysts in the phakomatoses are generally macroscopic and scattered throughout the kidney(s).

In addition, HNF1B-nephropathy represents a common phenocopy of ARPKD and ADPKD (Bergmann. 2017. PubMed ID: 29479522). Beyond renal cysts, its wide clinical spectrum encompasses diabetes, genital tract malformations, elevated liver enzymes, hyperuricemia and electrolyte disturbances.

Hereditary cystic kidney diseases can be categorized into two primary groups in regards of underlying etiology: ciliopathies and the phakomatoses (Dillman et al. 2017. PubMed ID: 28493804).

Ciliopathies represent a group of disorders caused by genetic defects in genes encoding proteins that are involved in formation or function of the primary cilia. The primary cilia are sensory organelles with critical roles in cellular signaling pathways including proliferation and differentiation, cellular motility, and cellular polarity. The kidneys are one of the major affected organs in ciliopathies. Hereditary cystic kidney diseases within this category primarily include ADPKD, ARPKD, nephronophthi­sis and medullary cystic kidney disease (MCKD).

PKD1 and PKD2 are the two major causative genes for ADPKD (Rossetti et al. 2007. PubMed ID: 17582161; Audrézet et al. 2012. PubMed ID: 22508176). Accounting for a small fraction of ADPKD-spectrum cases, defects in the GANAB, DNAJB11, and IFT140 genes result in an atypical (mild) form of ADPKD (Porath et al. 2016. PubMed ID: 27259053; Cornec-Le Gall et al. 2018. PubMed ID: 29706351; Senum et al. 2022. PubMed ID: 34890546).

PKHD1 is the primary causative gene for ARPKD (Bergmann. 2017. PubMed ID: 29479522; Ward et al. 2002. PubMed ID: 11919560). Accounting for a small fraction of genetically positive cases, DZIP1L was newly identified as the second causative gene for ARPKD (Lu et al. 2017. PubMed ID: 28530676; Hartung and Guay-Woodford. 2017. PubMed ID: 28736432).

Nephronophthi­sis is a group of genetically heterogeneous disorders inherited in an autosomal recessive manner (Hildebrandt et al. 2009. PubMed ID: 19118152). To date, defects in at least 20 genes have been reported to cause nephronophthisis (Srivastava et al. 2017. PubMed ID: 29379777).

The causative genes for autosomal dominant medullary cystic kidney disease (MCKD), also known as autosomal dominant tubulointerstitial kidney disease (ADTKD), include HNF1B, UMOD, MUC1, REN and SEC61A1 (Cornec-Le Gall et al. 2019. PubMed ID: 30819518; Hart et al. 2002. PubMed ID: 12471200; Kirby et al. 2013. PubMed ID: 23396133). Of note, the HNF1B-related renal disorders, termed HNF1B-nephropathy, have a wide clinical phenotypic spectrum and variable age of onset from in utero to adulthood, including congenital anomalies of the kidney and urinary tract (CAKUT), tubular transport abnormalities, chronic tubulointerstitial and cystic renal disease (Izzi et al. 2020. PubMed ID: 33305128). Around 30–50% of pathogenic HNF1B variants arise de novo. A large deletion (~1.4Mb) including the entire HNF1B gene can be found in ~50% of patients with HNF1B-nephropathy. HNF1B encodes hepatocyte nuclear factor-1-beta, which is the master regulator of a number of polycystic kidney disease genes.

The phakomatoses in this panel include two autosomal dominant disorders: tuberous sclerosis complex (TSC) (with the contiguous gene syndrome caused by a contiguous deletion at PKD1 and TSC2) and von Hippel–Lindau syndrome. TSC is caused by defects in the TSC1 and TSC2 genes while Von Hippel-Lindau disease is due to defects in the VHL gene (Northrup et al. 2018. PubMed ID: 20301399; Nordstrom-O'Brien et al. 2010. PubMed ID: 20151405). These genes encode tumor suppressors.

This panel also includes some other genes that have been associated with renal cystic lesions including COL4A1, CRB2, LRP5, JAG1, NOTCH2, OFD1, PAX2, SEC61A1 and DICER1.

To our knowledge, genetic testing sensitivity of each gene included in this panel in a large clinically heterogeneous cohort of patients with hereditary cystic kidney diseases has not been reported in the literature. The clinical sensitivities listed as below are based on individual well-defined disease entities.

In two large cohort studies of autosomal dominant polycystic kidney disease (ADPKD), the overall pathogenic variants detection rate of PKD1 and PKD2 is about 89%, in which defects in PKD1 and PKD2 explain approximately 85% and 15% of genetically positive ADPKD cases, respectively (Rossetti et al. 2007. PubMed ID: 17582161; Audrézet et al. 2012. PubMed ID: 22508176). Large deletions and duplications in PKD1 and PKD2 are relatively rare (<4%) in ADPKD patients (Bataille et al. 2011. PubMed ID: 22008521; Audrézet et al. 2012. PubMed ID: 22508176). After PKD1 and PKD2, IFT140 LoF variants likely represent the third most common cause of cystic kidney disease, accounting for >1% of ADPKD-spectrum-affected individuals (Senum et al. 2022. PubMed ID: 34890546). Defects in GANAB and DNAJB11 account for another ~0.3% and ~ 1% of total ADPKD (Porath et al. 2016. PubMed ID: 27259053; Cornec-Le Gall et al. 2018. PubMed ID: 29706351).

Since we primarily use Next Generation Sequencing (NGS) to test the PKD1 gene (see Testing Strategy section), gene conversions can be missed. However, our internal data suggested gene conversions are rare (<0.5%) in PKD1. These events have been found by long-range PCR based Sanger sequencing, but not by NGS only. Therefore, to increase detection rate (but by a very limited amount) of PKD1 pathogenic variants, Sanger sequencing for exons 1 to 33 (homologous regions) of PKD1 may be ordered.

Homozygous or compound heterozygous pathogenic variants in PKHD1 can be found in ~80% of ARPKD patients regardless of disease severity. Approximately 95% of affected individuals were found to have at least one pathogenic variant in PKHD1 (Bergmann. 2017. PubMed ID: 29479522). Defects in the DZIP1L gene were found in only two of 743 (~0.3%) unrelated individuals with suspected ARPKD or sporadic PKD (Lu et al. 2017. PubMed ID: 28530676).

HNF1B pathogenic variants were found via Sanger sequencing in up to 7% of patients/fetuses with renal hypodysplasia in three large cohort studies (Weber et al. 2006. PubMed ID: 16971658; Thomas et al. 2011. PubMed ID: 21380624; Madariaga et al. 2013. PubMed ID: 23539225). A large deletion (~1.4Mb) including the entire HNF1B gene can be found in ~50% of patients with HNF1B-nephropathy (Bergmann. 2017. PubMed ID: 29479522). The current NGS panel test can detect this large deletion.

Sensitivity of genetic testing for nephronophthisis is approximately 30% overall (Hildebrandt et al. 2009. PubMed ID: 19118152). Approximately 20% of individuals with nephronophthisis have a homozygous deletion encompassing the NPHP1 gene. The current NGS panel test can detect the ~279kb deletion in the NPHP1 gene in both heterozygous and homozygous states.

Pathogenic variants can be identified in approximately 85% of individuals with tuberous sclerosis complex (TSC); 15% of individuals with TSC will not have a pathogenic variant identified (Northrup et al. 2018. PubMed ID: 20301399).

Genetic testing for VHL achieves a molecular diagnosis in 90-100% of patients with VHL disease (Nordstrom-O'Brien et al. 2010. PubMed ID: 20151405).

This test is performed using Next-Gen sequencing with additional Sanger sequencing as necessary.

DNA analysis of the PKD1 gene is complicated and challenging due to the presence of several PKD1 pseudogenes. There is high sequence similarity of exons 1 to 33 between PKD1 and its pseudogenes (Audrézet et al. 2012. PubMed ID: 22508176). We have validated Next Generation Sequencing (NGS) to reliably sequence these exons.

For the PKD1 gene, including exons 1 to 33 (homologous regions), we primarily use Next Generation Sequencing (NGS) (~96%) complimented with Sanger sequencing for low-coverage regions (~4%). For any pathogenic, likely pathogenic, and uncertain variants found in exons 1 to 33 (homologous regions) via NGS, we use long-range PCR based Sanger sequencing to confirm them. Therefore, this test provides full coverage of all coding exons of the PKD1 gene plus 10 bases of flanking noncoding DNA in all available transcripts along with other non-coding regions in which pathogenic variants have been identified at PreventionGenetics or reported elsewhere. We define full coverage as >20X NGS reads or Sanger sequencing. PGnome panels typically provide slightly increased coverage over the PGxome equivalent. PGnome sequencing panels have the added benefit of additional analysis and reporting of deep intronic regions (where applicable).

Due to homologous sequence, gene conversion events in the PKD1 gene have been reported in the literature and found at PreventionGenetics. Our internal data suggested gene conversions are rare (<0.5%) in PKD1. These events have been found by long-range PCR based Sanger sequencing, but not by NGS only. Therefore, Sanger sequencing for exons 1 to 33 (homologous regions) of PKD1 may also be ordered.

To date, the only documented pathogenic variant in MUC1 causing medullary cystic kidney disease is the insertion of a single cytosine in one copy of the repeat unit comprising the extremely long (∼1.5-5 kb), GC-rich (>80%) coding variable-number tandem repeat (VNTR) sequence (Kirby et al. 2013. PubMed ID: 23396133). Our current sequencing methodology has not been validated to detect this variant.

Regarding copy number variants (CNVs) analysis, because of the paucity of CNVs and the complicated nature of sequence in PKD1, CNV analysis for this gene can be performed via the multiplex ligation-dependent amplification (MLPA) assay with limited increased sensitivity (compared to Next-Gen sequencing CNV analysis), and can be ordered separately (Test #2058).

This panel typically provides 98.8% coverage of all coding exons of the genes, PKD1 and PKD2 are covered 100%, plus 10 bases of flanking noncoding DNA in all available transcripts along with other non-coding regions in which pathogenic variants have been identified at PreventionGenetics or reported elsewhere. We define coverage as ≥20X NGS reads or Sanger sequencing. PGnome panels typically provide slightly increased coverage over the PGxome equivalent. PGnome sequencing panels have the added benefit of additional analysis and reporting of deep intronic regions (where applicable).

Dependent on the sequencing backbone selected for this testing, discounted reflex testing to any other similar backbone-based test is available (i.e., PGxome panel to whole PGxome; PGnome panel to whole PGnome).