Fig. 1 Mutation burden and spectrum in unrelated donors. (A) Number and types of somatic mutations detected in donors. Of the donor cohort, 44% (11 of 25) harbored at least one somatic mutation. (B) Mutation spectrum of detected mutations in donors. (C) Types of nucleotide changes. (D) Age of donors with and without detected SNV(s). Boxes show the 25th and 75th percentiles, as well as median (P = 0.03, two-sided Wilcoxon rank-sum test). (E) Clonal dynamics of engrafted mutations in recipients after HSCT.
Fig. 2 Mutation burden and spectrum of donor clonal mutations engrafted in the recipients. (A) Mutation spectrum in recipients at different time points after HCST. “Engrafted” mutations are known to be from the donor, and “New” mutations were not observed before HSCT in either donor or recipient specimens. Genes with ≥1 engrafted and ≥ 1 new mutations are shown. (B) Mutation spectrum of new mutations detected in recipients after HSCT. Genes with ≥2 new mutations were represented. (C) Violin plots showing mutation burden at different time points after HSCT. The P value was calculated using one-sided Wilcoxon rank-sum test. (D to F) Representative plots showing ddPCR experiment results. The blue dots in the panels indicate positive mutant droplets, green dots indicate positive wild-type (WT) droplets, and gray dots indicate empty droplets. (D) ddPCR results demonstrating the presence of a de novo, persistent mutation in DNAH2 in donor-derived cells engrafted in the recipient (PID_0589). The mutation was not detected in the donor before HSCT. (E) ddPCR results demonstrating the presence of a de novo, transient mutation in STAG2 in donor-derived cells engrafted in the recipient (PID_0450). The mutation was not detected in the donor before HSCT, and it reduced in VAF by 1 year after HSCT. (F) ddPCR results demonstrating the presence of an engrafted donor-derived mutation in CREBBP that was detected at an extremely low VAF in the donor (PID_0450). This mutation increased in VAF at D100 and 1 year after HSCT in the recipient.
- Table 1 Somatic mutations detected in donors that were predicted to be pathogenic according to CADD.
Six mutations were found to be associated with various malignancies, and three were specifically associated with hematologic malignancies (*).
Gene Type Amino acid change CADD COSMIC Engrafted COL12A1 Missense p.I530L 22.1 COSM271996 Yes CREBBP Missense p.T1242I 25.5 – Yes DNMT3A Nonsense p.W288X 40 COSM1130818 Yes DNMT3A Missense p.R174S 26.5 – Yes DNMT3A Missense p.G398R 30 COSM256035* Yes DNMT3A Missense p.I158M 23.6 – Yes DNMT3A Missense p.Q222P 26.1 – Yes DNMT3A Missense p.H669P 23.3 – Yes FAT1 Missense p.D1554N 25.8 COSM1429043 Yes SRCAP Indel T:TGCTTCGCC 29 – Yes STAG2 Missense p.Y188D 26.3 – Yes TET2 Splicing c.3954 + 1G > A 34 COSM87141* Yes TET2 Missense p.Y1345C 32 – Yes TP53 Missense p.R150W 25.7 COSM99925* Yes USP34 Missense p.H1874R 22.5 – Yes WT1 Missense p.R74W 28.7 – Yes - Table 2 Demographic information of recipients and the corresponding matched donors in relation to engraftment of donor-derived mutations. CR, complete remission; MAC, myeloablative conditioning.
Characteristic Category No donor mutation
(n = 14)Mutation engrafted
(n = 11)P Test performed Donor age Median (range) 24 (21 to 39) 36 (20 to 58) 0.03 Wilcoxon rank-sum test Donor gender Male 10 (71.4%) 8 (72.7%) 0.99 Fisher’s exact test Female 4 (28.6%) 3 (27.3%) Recipient age Median (range) 51 (27 to 65) 55 (19 to 69) 0.66 Wilcoxon rank-sum test Recipient gender Male 13 (92.9%) 7 (63.6%) 0.13 Fisher’s exact test Female 1 (7.1%) 4 (36.4%) Primary disease AML/MDS 7 (50%) 9 (81.8%) 0.21 Fisher’s exact test Non-AML 7 (50%) 2 (18.2%) Disease status prior to
transplantCR 7 (50%) 5 (45.4%) 0.99 Fisher’s exact test Non-CR 7 (50%) 5 (45.4%) Unknown 0 (0%) 1 (9.1%) Conditioning MAC 8 (57.1%) 7 (63.6%) 0.99 Fisher’s exact test Non-MAC 6 (42.9%) 4 (36.4%) HLA mismatch No mismatch 13 (92.9%) 9 (81.8%) 0.56 Fisher’s exact test Mismatch 1 (7.1%) 2 (18.2%)
Supplementary Materials
stm.sciencemag.org/cgi/content/full/12/526/eaax6249/DC1
Fig. S1. Engrafted donor mutations in recipients.
Fig. S2. Clonal expansion of mutations reaching the threshold for CHIP (≥0.02 VAF) in three patients after HSCT.
Fig. S3. Types of somatic substitutions in recipients after HSCT.
Fig. S4. The sequencing depth of each ECS library at all time points.
Fig. S5. New mutations detected in recipients after HSCT.
Fig. S6. ECS calls validated by ddPCR.
Fig. S7. Number of detected mutations in genes according to gene length.
Fig. S8. Leukemia-free survival of recipients with or without persistent engraftment of donor-derived mutations.
Fig. S9. Cumulative incidence of chronic GvHD in recipients with or without persistent engraftment of donor-derived mutations.
Data file S1. Detected somatic mutations in donors.
Data file S2. Detected somatic mutations in recipients after HSCT after removing recipient’s own hematopoietic clones.
Data file S3. Shared variants in pre-HSCT and post-HSCT recipient samples due to incomplete clearance of recipient’s hematopoietic clones after HSCT.
Data file S4. Analysis of recipient clinical outcomes in relation to engraftment of donor-derived mutations.
Data file S5. Recurrently mutated genes in adult and pediatric AML.
Data file S6. ddPCR probe sequences.
Additional Files
The PDF file includes:
- Fig. S1. Engrafted donor mutations in recipients.
- Fig. S2. Clonal expansion of mutations reaching the threshold for CHIP (≥0.02 VAF) in three patients after HSCT.
- Fig. S3. Types of somatic substitutions in recipients after HSCT.
- Fig. S4. The sequencing depth of each ECS library at all time points.
- Fig. S5. New mutations detected in recipients after HSCT.
- Fig. S6. ECS calls validated by ddPCR.
- Fig. S7. Number of detected mutations in genes according to gene length.
- Fig. S8. Leukemia-free survival of recipients with or without persistent engraftment of donor-derived mutations.
- Fig. S9. Cumulative incidence of chronic GvHD in recipients with or without persistent engraftment of donor-derived mutations.
- Legends for data files S1 to S6
Other Supplementary Material for this manuscript includes the following:
- Data file S1 (Microsoft Excel format). Detected somatic mutations in donors.
- Data file S2 (Microsoft Excel format). Detected somatic mutations in recipients after HSCT after removing recipient’s own hematopoietic clones.
- Data file S3 (Microsoft Excel format). Shared variants in pre-HSCT and post-HSCT recipient samples due to incomplete clearance of recipient’s hematopoietic clones after HSCT.
- Data file S4 (Microsoft Excel format). Analysis of recipient clinical outcomes in relation to engraftment of donor-derived mutations.
- Data file S5 (Microsoft Excel format). Recurrently mutated genes in adult and pediatric AML.
- Data file S6 (Microsoft Excel format). ddPCR probe sequences.
