HCFC1-Related Disorders via the HCFC1 Gene

  • Summary and Pricing
  • Clinical Features and Genetics
  • Citations
  • Methods
  • Ordering/Specimens
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NextGen Sequencing

Test Code Test Copy GenesPriceCPT Code Copy CPT Codes
3475 HCFC1$640.00 81479 Add to Order
Pricing Comments

Our most cost-effective testing approach is NextGen sequencing with Sanger sequencing supplemented as needed to ensure sufficient coverage and to confirm NextGen calls that are pathogenic, likely pathogenic or of uncertain significance. If, however, full gene Sanger sequencing only is desired (for purposes of insurance billing or STAT turnaround time for example), please see link below for Test Code, pricing, and turnaround time information.

For Sanger Sequencing click here.
Targeted Testing

For ordering sequencing of targeted known variants, please proceed to our Targeted Variants landing page.

Turnaround Time

The great majority of tests are completed within 20 days.

Clinical Sensitivity

Clinical sensitivity of this test should be high in patients with HCFC1 deficiency. For example, Yu and colleagues (2013) reported pathogenic variants in 13 out of 17 male patients who were suspected of or cellularly diagnosed with cblc type methylmalonic aciduria and homocystinuria but did not harbor pathogenic variants in the MMACHC gene. The clinical sensitivity of HCFC1 testing in patients suspected of X-linked intellectual disability (XLID) is more difficult to predict, given that ~0.15-0.3% of the population is predicted to be affected with this disorder (Koufaris et al. 2016), and HCFC1 variants would be expected explain only a subset of those cases. Analytical sensitivity of this test is expected to be high as all variants reported to date should be detectable via sequencing.

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Clinical Features

Defects in the HCFC1 gene have been reported to be causative for both X-linked intellectual disability type 3 (XLID 3) and combined methylmalonic aciduria and homocystinuria, cblX type (Huang et al. 2012; Yu et al. 2013; Jolly et al. 2015).

In the reported XLID 3 patients, most were classified as having mild-to-moderate intellectual disability, though a few have been reported with severe intellectual disability. Other commonly reported features include behavioral problems such as aggression or hyperactivity, anxiety, and autistic features. Some of the patients have been reported to present with microcephaly and facial dysmorphism (Huang et al. 2012; Jolly et al. 2015; Koufaris et al. 2016).

Patients diagnosed with the cblX disorder have mainly presented during early infancy with severe developmental delays, intractable epilepsy, and failure to thrive. Other clinical features that have been observed include hypotonia, brain abnormalities, microcephaly, ketoacidosis, hyperammonemia, choreoathetosis, and multiple congenital anomalies that may include genital abnormalities. Biochemically, the cblX patients also have signs of combined methylmalonic aciduria and homocystinuria, including increased methylmalonic acid in the serum and/or urine, increased total homocysteine levels, and decreased methionine (Yu et al. 2013; Gérard et al. 2015; Jolly et al. 2015; Niranjan et al. 2015). The cblX form of HCFC1 deficiency tends to be more severe than the XLID type 3 form (Jolly et al. 2015).

Combined methylmalonic aciduria and homocystinuria can also be caused by defects in the MMACHC, MMADHC, ABCD4 and LMBRD1 genes (Carrillo-Carrasco et al. 2013). Though cellular complementation studies can typically pinpoint the specific form of cobalamin deficiency in a patient, the genetic cause of a number of cblC type methylmalonic aciduria and homocystinuria cases (MMACHC deficiency) has not been found (Yu et al. 2013). Recent studies have shown that pathogenic variants in the HCFC1 gene can explain a large percentage of these cases (Yu et al. 2013).


XLID type 3 and methylmalonic aciduria and homocystinuria cblX type are caused by pathogenic variants in HCFC1, which is located on the X chromosome. These disorders are inherited in an X-linked recessive fashion (Huang et al. 2012; Yu et al. 2013). Thus far, all affected individuals that have been reported have been male. Variants may be inherited or arise de novo (Huang et al. 2012; Yu et al. 2013). To date, more than 10 pathogenic variants have been reported in the HCFC1 gene. With the exception of one regulatory variant found in a large XLID type 3 family, most variants have been missense (Human Gene Mutation Database). The most commonly reported variant is c.218C>T (p.Ala73Val), which was reported in nine cblX patients (Yu et al. 2013). There does appear to be a genotype-phenotype correlation, with loss-of-function variants being associated with the more severe cblX presentation and partial loss-of-function variants being associated with XLID type 3 (Jolly et al. 2015). The severity also seems to be dependent upon location within the protein. Thus far, nearly all variants that have been linked with the cblX presentation have been located in the Kelch domain region of the protein or within the nuclear localization signal sequence (Yu et al. 2013; Gérard et al. 2015; Jolly et al. 2015; Niranjan et al. 2015). Variants associated with XLID have been found in other locations within the HCFC1 protein (Huang et al. 2012; Jolly et al. 2015; Koufaris et al. 2016).

The HCFC1 gene encodes a transcriptional co-regulator protein that has been shown to play a role in regulating the expression of the MMACHC gene, in addition to many other genes involved in cellular proliferation and other critical functions (Jolly et al. 2015). Both MMACHC and HCFC1 defects lead to neurological impairment, though HCFC1 deficiency tends to be more severe. It is believed that this is because the HCFC1 protein is involved in the regulation of many other genes in addition to MMACHC (Jolly et al. 2015).

Testing Strategy

For this NextGen test, the full coding regions plus ~10 bp of non-coding DNA flanking each exon are sequenced for the gene listed below. Sequencing is accomplished by capturing specific regions with an optimized solution-based hybridization kit, followed by massively parallel sequencing of the captured DNA fragments. Additional Sanger sequencing is performed for any regions not captured or with an insufficient number of sequence reads. All pathogenic, likely pathogenic, or variants of uncertain significance are confirmed by Sanger sequencing.

Indications for Test

Patients suspected of cblC type methylmalonic aciduria and homocystinuria who have not been found to harbor pathogenic variants in the MMACHC, MMADHC, ABCD4 or LMBRD1 genes are good candidates for this test, as are those with clinical features suggestive of HCFC1 deficiency. Testing is also indicated for family members of patients with known HCFC1 sequence variants. We will also sequence the HCFC1 gene to determine carrier status.


Official Gene Symbol OMIM ID
HCFC1 300019
Inheritance Abbreviation
Autosomal Dominant AD
Autosomal Recessive AR
X-Linked XL
Mitochondrial MT

Related Test

Autism Spectrum Disorders and Intellectual Disability (ASD-ID) Comprehensive Sequencing Panel with CNV Detection


Genetic Counselors
  • Carrillo-Carrasco N. et al. 2013. Disorders of Intracellular Cobalamin Metabolism. In: Pagon RA, Adam MP, Ardinger HH, Bird TD, Dolan CR, Fong C-T, Smith RJ, and Stephens K, editors. GeneReviews(®), Seattle (WA): University of Washington, Seattle. PubMed ID: 20301503
  • Gérard M. et al. 2015. European Journal of Medical Genetics. 58: 148-53. PubMed ID: 25595573
  • Huang L. et al. 2012. American Journal of Human Genetics. 91: 694-702. PubMed ID: 23000143
  • Human Gene Mutation Database (Bio-base).
  • Jolly L.A. et al. 2015. Human Molecular Genetics. 24: 3335-47. PubMed ID: 25740848
  • Koufaris C. et al. 2016. Biomedical Reports. 4: 215-218. PubMed ID: 26893841
  • Niranjan T.S. et al. 2015. Plos One. 10: e0116454. PubMed ID: 25679214
  • Yu H.C. et al. 2013. American Journal of Human Genetics. 93: 506-14. PubMed ID: 24011988
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NextGen Sequencing using PG-Select Capture Probes

Test Procedure

We use a combination of Next Generation Sequencing (NGS) and Sanger sequencing technologies to cover the full coding regions of the listed genes plus ~10 bases of non-coding DNA flanking each exon.  As required, genomic DNA is extracted from the patient specimen.  For NGS, patient DNA corresponding to these regions is captured using an optimized set of DNA hybridization probes.  Captured DNA is sequenced using Illumina’s Reversible Dye Terminator (RDT) platform (Illumina, San Diego, CA, USA).  Regions with insufficient coverage by NGS are often covered by Sanger sequencing.

For Sanger sequencing, Polymerase Chain Reaction (PCR) is used to amplify targeted regions.  After purification of the PCR products, cycle sequencing is carried out using the ABI Big Dye Terminator v.3.0 kit.  PCR products are resolved by electrophoresis on an ABI 3730xl capillary sequencer.  In nearly all cases, cycle sequencing is performed separately in both the forward and reverse directions.

Patient DNA sequence is aligned to the genomic reference sequence for the indicated gene region(s). All differences from the reference sequences (sequence variants) are assigned to one of five interpretation categories, listed below, per ACMG Guidelines (Richards et al. 2015).

(1) Pathogenic Variants
(2) Likely Pathogenic Variants
(3) Variants of Uncertain Significance
(4) Likely Benign Variants
(5) Benign Variants

Human Genome Variation Society (HGVS) recommendations are used to describe sequence variants (  Rare variants and undocumented variants are nearly always classified as likely benign if there is no indication that they alter protein sequence or disrupt splicing.

Analytical Validity

As of March 2016, 6.36 Mb of sequence (83 genes, 1557 exons) generated in our lab was compared between Sanger and NextGen methodologies. We detected no differences between the two methods. The comparison involved 6400 total sequence variants (differences from the reference sequences). Of these, 6144 were nucleotide substitutions and 256 were insertions or deletions. About 65% of the variants were heterozygous and 35% homozygous. The insertions and deletions ranged in length from 1 to over 100 nucleotides.

In silico validation of insertions and deletions in 20 replicates of 5 genes was also performed. The validation included insertions and deletions of lengths between 1 and 100 nucleotides. Insertions tested in silico: 2200 between 1 and 5 nucleotides, 625 between 6 and 10 nucleotides, 29 between 11 and 20 nucleotides, 25 between 21 and 49 nucleotides, and 23 at or greater than 50 nucleotides, with the largest at 98 nucleotides. All insertions were detected. Deletions tested in silico: 1813 between 1 and 5 nucleotides, 97 between 6 and 10 nucleotides, 32 between 11 and 20 nucleotides, 20 between 21 and 49 nucleotides, and 39 at or greater than 50 nucleotides, with the largest at 96 nucleotides. All deletions less than 50 nucleotides in length were detected, 13 greater than 50 nucleotides in length were missed. Our standard NextGen sequence variant calling algorithms are generally not capable of detecting insertions (duplications) or heterozygous deletions greater than 100 nucleotides. Large homozygous deletions appear to be detectable.   

Analytical Limitations

Interpretation of the test results is limited by the information that is currently available.  Better interpretation should be possible in the future as more data and knowledge about human genetics and this specific disorder are accumulated.

When Sanger sequencing does not reveal any difference from the reference sequence, or when a sequence variant is homozygous, we cannot be certain that we were able to detect both patient alleles.  Occasionally, a patient may carry an allele which does not amplify, due to a large deletion or insertion.   In these cases, the report will contain no information about the second allele.  Our Sanger and NGS Sequencing tests are generally not capable of detecting Copy Number Variants (CNVs).

We sequence all coding exons for each given transcript, plus ~10 bp of flanking non-coding DNA for each exon.  Test reports contain no information about other portions of the gene, such as regulatory domains, deep intronic regions or any currently uncharacterized alternative exons.

In most cases, we are unable to determine the phase of sequence variants.  In particular, when we find two likely causative mutations for recessive disorders, we cannot be certain that the mutations are on different alleles.

Our ability to detect minor sequence variants due to somatic mosaicism is limited.  Sequence variants that are present in less than 50% of the patient’s nucleated cells may not be detected.

Runs of mononucleotide repeats (eg (A)n or (T)n) with n >8 in the reference sequence are generally not analyzed because of strand slippage during PCR.

Unless otherwise indicated, DNA sequence data is obtained from a specific cell-type (usually leukocytes from whole blood).   Test reports contain no information about the DNA sequence in other cell-types.

We cannot be certain that the reference sequences are correct.

Rare, low probability interpretations of sequencing results, such as for example the occurrence of de novo mutations in recessive disorders, are generally not included in the reports.

We have confidence in our ability to track a specimen once it has been received by PreventionGenetics.  However, we take no responsibility for any specimen labeling errors that occur before the sample arrives at PreventionGenetics.

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Ordering Options

myPrevent - Online Ordering
  • The test can be added to your online orders in the Summary and Pricing section.
  • Once the test has been added log in to myPrevent to fill out an online requisition form.
  • A completed requisition form must accompany all specimens.
  • Billing information along with specimen and shipping instructions are within the requisition form.
  • All testing must be ordered by a qualified healthcare provider.


(Delivery accepted Monday - Saturday)

  • Collect 3 ml -5 ml (5 ml preferred) of whole blood in EDTA (purple top tube) or ACD (yellow top tube). For Test #500-DNA Banking only, collect 10 ml -20 ml of whole blood.
  • For small babies, we require a minimum of 1 ml of blood.
  • Only one blood tube is required for multiple tests.
  • Ship blood tubes at room temperature in an insulated container. Do not freeze blood.
  • During hot weather, include a frozen ice pack in the shipping container. Place a paper towel or other thin material between the ice pack and the blood tube.
  • In cold weather, include an unfrozen ice pack in the shipping container as insulation.
  • At room temperature, blood specimen is stable for up to 48 hours.
  • If refrigerated, blood specimen is stable for up to one week.
  • Label the tube with the patient name, date of birth and/or ID number.


(Delivery accepted Monday - Saturday)

  • Send in screw cap tube at least 5 µg -10 µg of purified DNA at a concentration of at least 20 µg/ml for NGS and Sanger tests and at least 5 µg of purified DNA at a concentration of at least 100 µg/ml for gene-centric aCGH, MLPA, and CMA tests, minimum 2 µg for limited specimens.
  • For requests requiring more than one test, send an additional 5 µg DNA per test ordered when possible.
  • DNA may be shipped at room temperature.
  • Label the tube with the composition of the solute, DNA concentration as well as the patient’s name, date of birth, and/or ID number.
  • We only accept genomic DNA for testing. We do NOT accept products of whole genome amplification reactions or other amplification reactions.


(Delivery preferred Monday - Thursday)

  • PreventionGenetics should be notified in advance of arrival of a cell culture.
  • Culture and send at least two T25 flasks of confluent cells.
  • Some panels may require additional flasks (dependent on size of genes, amount of Sanger sequencing required, etc.). Multiple test requests may also require additional flasks. Please contact us for details.
  • Send specimens in insulated, shatterproof container overnight.
  • Cell cultures may be shipped at room temperature or refrigerated.
  • Label the flasks with the patient name, date of birth, and/or ID number.
  • We strongly recommend maintaining a local back-up culture. We do not culture cells.
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