Using IGV and gnomAD to assess a putative autoimmune disease variant

Use IGV, ClinGen, and gnomAD to assess clinically relevant sequence variation near a self-reactive TCR target.
ClinGen
IGV
gnomAD
Author

Alice MacQueen

Published

June 18, 2024

There is a cool biotech under development to identify the antigens for specific T cell receptors, by exposing the T cell to a library of (thousands to hundreds of thousands) cells expressing a wide variety of antigens. If the T cell recognizes an antigen, a specific reporter fluorescence molecule will be activated, and the library can be sorted for reporter activation and the sequence of the antigen determined using next generation sequencing (NGS) (Kula et al. 2019).

Graphical Abstract for Kula et al., 2019 (Kula et al. 2019).

This technology is exciting because it can be used to screen for self-reactive TCRs, which could be important for developing therapeutics for specific autoimmune diseases and for developing therapeutics for cancer. For example, the authors identify four human proteins recognized by a self-reactive TCR specific for a HLA-A1-restricted epitope of the gene MAGE-A3 (Kula et al. 2019; Karanikas et al. 2003): MAGE-A3, MAGE-A6, PLD5, and FAT2.

Now, let’s check one of these genes in IGV, ClinGen, and gnomAD. Here are a few interesting questions to explore:

  1. What sequence variation is present at these genes?

  2. Is this variation is implicated in skin cancer, or any other disease? (aka, is there additional evidence that this gene is involved in cancers or autoimmune conditions?)

  3. Is common genetic variation present in the identified epitopes? (Variation can change TCR reactivity.)

IGV

I pulled up the PLD5 gene in the web version of IGV - the epitope from the paper is at amino acids 179 to 187. To get a first overview of the variation within PLD5 and known disease implications, I searched for PLD5 in IGV, and pulled up the common SNPs and GWAS annotation tracks. There are five GWAS associations within PLD5:

GWAS associations within PLD5

There are a couple of associations for N-glycosylation of IgG; one association is with coronary artery calcification among African Americans, and one is for a drug response to cytidine analogues. None of these seem particularly relevant to melanoma.

The fifth association is with squamous cell lung carcinoma - while not melanoma, this association with a familial disposition towards cancer could be of interest.

ClinGen

I next searched ClinGen for these genes. However, none of them were in ClinGen’s databases of curated clinically relevant disease variants.

PLD5 is not in ClinGen

So next I decided to check gnomAD, particularly as by doing so I could also check ClinVar.

gnomAD

I wanted to look at SNPs in the exon of PLD5 which had the epitope recognized by the self-reactive TCR. As the epitope is at amino acids 179 to 187, I looked for sites with impacts on these amino acids (aka, HGVS Consequence matching 179 - 187). I downloaded the full gnomAD variant dataset for this gene to make searching it slightly easier.

PLD5 info from NCBI
Assembly Location
GRCh38.p14 (GCF_000001405.40) NC_000001.11 (242082986..242530546, complement) 
pld5 |> 
  arrange(desc(Position)) |> 
1  # filter(grepl("[1][78][0-79]", `HGVS Consequence`))
  filter(between(Position, 242265383, 242265409)) |> 
  select(`gnomAD ID`, Chromosome, Position, Reference, Alternate, `HGVS Consequence`, `VEP Annotation`, `ClinVar Clinical Significance`, `Allele Frequency`, everything()) |> 
  kable()
1
Original search string to find amino acid changes from 179-187; it pulled up more than just amino acid changes, so I ultimately subset by Position.
gnomAD ID Chromosome Position Reference Alternate HGVS Consequence VEP Annotation ClinVar Clinical Significance Allele Frequency rsIDs Source Filters - exomes Filters - genomes Transcript Protein Consequence Transcript Consequence ClinVar Variation ID Flags Allele Count Allele Number Homozygote Count Hemizygote Count Filters - joint GroupMax FAF group GroupMax FAF frequency cadd revel_max spliceai_ds_max pangolin_largest_ds phylop sift_max polyphen_max Allele Count African/African American Allele Number African/African American Homozygote Count African/African American Hemizygote Count African/African American Allele Count Admixed American Allele Number Admixed American Homozygote Count Admixed American Hemizygote Count Admixed American Allele Count Ashkenazi Jewish Allele Number Ashkenazi Jewish Homozygote Count Ashkenazi Jewish Hemizygote Count Ashkenazi Jewish Allele Count East Asian Allele Number East Asian Homozygote Count East Asian Hemizygote Count East Asian Allele Count European (Finnish) Allele Number European (Finnish) Homozygote Count European (Finnish) Hemizygote Count European (Finnish) Allele Count Middle Eastern Allele Number Middle Eastern Homozygote Count Middle Eastern Hemizygote Count Middle Eastern Allele Count European (non-Finnish) Allele Number European (non-Finnish) Homozygote Count European (non-Finnish) Hemizygote Count European (non-Finnish) Allele Count Amish Allele Number Amish Homozygote Count Amish Hemizygote Count Amish Allele Count South Asian Allele Number South Asian Homozygote Count South Asian Hemizygote Count South Asian Allele Count Remaining Allele Number Remaining Homozygote Count Remaining Hemizygote Count Remaining
1-242265407-T-C 1 242265407 T C p.Gln179Gln synonymous_variant NA 6.0e-07 NA gnomAD Exomes PASS NA ENST00000536534.7 p.Gln179Gln c.537A>G NA NA 1 1612620 0 0 PASS NA NA 8.26 NA 0.00 0.04 1.080 NA NA 0 75000 0 0 0 59848 0 0 0 29568 0 0 0 44812 0 0 0 63994 0 0 0 6046 0 0 1 1179350 0 0 0 912 0 0 0 90658 0 0 0 62432 0 0
1-242265405-T-G 1 242265405 T G p.Asn180Thr missense_variant NA 6.0e-07 NA gnomAD Exomes PASS NA ENST00000536534.7 p.Asn180Thr c.539A>C NA NA 1 1612690 0 0 PASS NA NA 22.60 0.051 0.00 0.01 3.120 0.02 0.031 0 74996 0 0 0 59850 0 0 0 29574 0 0 0 44810 0 0 0 63980 0 0 0 6046 0 0 1 1179376 0 0 0 912 0 0 0 90706 0 0 0 62440 0 0
1-242265404-A-G 1 242265404 A G p.Asn180Asn synonymous_variant NA 1.2e-06 rs760794546 gnomAD Exomes PASS NA ENST00000536534.7 p.Asn180Asn c.540T>C NA NA 2 1612710 0 0 PASS NA NA 8.78 NA 0.02 0.03 -0.561 NA NA 0 75014 0 0 0 59834 0 0 0 29574 0 0 1 44812 0 0 0 63980 0 0 0 6046 0 0 0 1179396 0 0 0 912 0 0 0 90704 0 0 1 62438 0 0
1-242265402-A-G 1 242265402 A G p.Ile181Thr missense_variant NA 6.0e-07 NA gnomAD Exomes PASS NA ENST00000536534.7 p.Ile181Thr c.542T>C NA NA 1 1612808 0 0 PASS NA NA 23.80 0.374 0.01 0.02 6.330 0.00 0.395 0 75008 0 0 0 59858 0 0 0 29574 0 0 0 44822 0 0 0 64000 0 0 0 6046 0 0 1 1179444 0 0 0 912 0 0 0 90704 0 0 0 62440 0 0
1-242265401-A-G 1 242265401 A G p.Ile181Ile synonymous_variant NA 6.0e-07 NA gnomAD Exomes PASS NA ENST00000536534.7 p.Ile181Ile c.543T>C NA NA 1 1612790 0 0 PASS NA NA 8.81 NA 0.01 0.02 -0.114 NA NA 0 75002 0 0 0 59858 0 0 0 29566 0 0 0 44820 0 0 0 63994 0 0 0 6048 0 0 1 1179436 0 0 0 912 0 0 0 90718 0 0 0 62436 0 0
1-242265400-C-T 1 242265400 C T p.Glu182Lys missense_variant NA 1.2e-06 rs1244994670 gnomAD Exomes PASS NA ENST00000536534.7 p.Glu182Lys c.544G>A NA NA 2 1612500 0 0 PASS nfe 3e-07 21.60 0.171 0.00 -0.01 8.900 0.88 0.015 0 74958 0 0 0 59830 0 0 0 29570 0 0 0 44814 0 0 0 63962 0 0 0 6046 0 0 2 1179340 0 0 0 912 0 0 0 90640 0 0 0 62428 0 0
1-242265392-C-T 1 242265392 C T p.Lys184Lys synonymous_variant NA 6.0e-07 NA gnomAD Exomes PASS NA ENST00000536534.7 p.Lys184Lys c.552G>A NA NA 1 1612778 0 0 PASS NA NA 10.10 NA 0.02 0.04 5.040 NA NA 0 74970 0 0 0 59876 0 0 0 29556 0 0 0 44820 0 0 0 63978 0 0 0 6052 0 0 0 1179464 0 0 0 912 0 0 1 90720 0 0 0 62430 0 0
1-242265389-T-C 1 242265389 T C p.Leu185Leu synonymous_variant NA 6.0e-07 NA gnomAD Exomes PASS NA ENST00000536534.7 p.Leu185Leu c.555A>G NA NA 1 1612956 0 0 PASS NA NA 14.20 NA 0.71 0.51 1.030 NA NA 0 75012 0 0 0 59876 0 0 0 29578 0 0 0 44826 0 0 0 64004 0 0 0 6052 0 0 1 1179528 0 0 0 912 0 0 0 90722 0 0 0 62446 0 0
1-242265387-A-G 1 242265387 A G p.Val186Ala missense_variant Uncertain significance 1.9e-06 rs775543164 gnomAD Exomes,gnomAD Genomes PASS PASS ENST00000536534.7 p.Val186Ala c.557T>C 3214673 NA 3 1612882 0 0 PASS nfe 7e-07 21.50 0.172 0.14 0.18 6.330 0.72 0.012 0 74886 0 0 0 59858 0 0 0 29576 0 0 0 44834 0 0 0 64012 0 0 0 6074 0 0 3 1179574 0 0 0 912 0 0 0 90732 0 0 0 62424 0 0
1-242265386-C-T 1 242265386 C T p.Val186Val synonymous_variant NA 6.0e-07 rs1364495756 gnomAD Genomes NA PASS ENST00000536534.7 p.Val186Val c.558G>A NA NA 1 1612794 0 0 PASS NA NA 11.10 NA 0.27 0.18 3.590 NA NA 1 74878 0 0 0 59854 0 0 0 29574 0 0 0 44836 0 0 0 63984 0 0 0 6072 0 0 0 1179550 0 0 0 912 0 0 0 90716 0 0 0 62418 0 0
1-242265383-A-T 1 242265383 A T p.Ser187Arg missense_variant NA 1.9e-06 rs772623416 gnomAD Exomes PASS NA ENST00000536534.7 p.Ser187Arg c.561T>A NA NA 3 1612900 0 0 PASS nfe 3e-07 24.00 0.348 0.07 0.14 0.424 0.02 0.927 0 74978 0 0 0 59850 0 0 0 29578 0 0 0 44822 0 0 0 63998 0 0 0 6050 0 0 2 1179572 0 0 0 912 0 0 1 90690 0 0 0 62450 0 0

So gnomAD has 11 SNPs in the epitope region of PLD5. None of them are particularly common (they all have allele frequencies of 1.860024e-06 or lower). Only one is in ClinVar as having Clinical Significance, but it is a missense variant of ‘Uncertain signficiance’. All but three of these rare alleles appear predominantly in the non-Finnish European population, with two in South Asian, one in East Asian, and one in an African American population.

Conclusions

The gnomAD and ClinVar data suggest that the epitope of PLD5 recognized by a self-reactive TCR is robustly present in humans without any common variation in this region. This suggests that if this TCR is present, it will likely be able to recognize this self-antigen, which improves its possible utility as a TCR targeting a tumor antigen.

References

Karanikas, Vaios, Christophe Lurquin, Didier Colau, Nicolas van Baren, Charles De Smet, Bernard Lethe', Thierry Connerotte, et al. 2003. “Monoclonal Anti-MAGE-3 CTL Responses in Melanoma Patients Displaying Tumor Regression After Vaccination with a Recombinant Canarypox Virus.” The Journal of Immunology 171 (9): 4898–4904. https://doi.org/10.4049/jimmunol.171.9.4898.
Kula, Tomasz, Mohammad H. Dezfulian, Charlotte I. Wang, Nouran S. Abdelfattah, Zachary C. Hartman, Kai W. Wucherpfennig, Herbert Kim Lyerly, and Stephen J. Elledge. 2019. “T-Scan: A Genome-Wide Method for the Systematic Discovery of T Cell Epitopes.” Cell 178 (4): 1016–1028.e13. https://doi.org/10.1016/j.cell.2019.07.009.

Citation

BibTeX citation:
@online{macqueen2024,
  author = {MacQueen, Alice},
  title = {Using {IGV} and {gnomAD} to Assess a Putative Autoimmune
    Disease Variant},
  date = {2024-06-18},
  url = {https://alice-macqueen.github.io/posts/2024-06-03-olga/},
  langid = {en}
}
For attribution, please cite this work as:
MacQueen, Alice. 2024. “Using IGV and gnomAD to Assess a Putative Autoimmune Disease Variant.” June 18, 2024. https://alice-macqueen.github.io/posts/2024-06-03-olga/.