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Case Reports
. 2024 Jul 23;8(14):3754-3759.
doi: 10.1182/bloodadvances.2023011875.

A novel transmembrane CXCR4 variant that expands the WHIM genotype-phenotype paradigm

Katarina Zmajkovicova  1 , Sumit Pawar  2 , Svetlana O Sharapova  3 , Christoph B Geier  4   5 , Ivana Wiest  2 , Chi Nguyen  1 , Halenya Monticelli  1 , Sabine Maier-Munsa  2 , Kelly Chen  6 , John W Sleasman  7 , Svetlana Aleshkevich  3 , Ekaterina Polyakova  3 , Inga Sakovich  3 , Klaus Warnatz  4   5 , Bodo Grimbacher  4   5 , Michele Proietti  8   9 , Neal Sondheimer  10 , Boglarka Ujhazi  11 , Sumai Gordon  11 , Maryssa Ellison  11 , Melis Yilmaz  11   12 , Jolan E Walter  11   12   13 , Adriana Badarau  2 , Arthur G Taveras  6 , Jadee L Neff  14 , Jacob R Bledsoe  15 , Teresa K Tarrant  16   17
Affiliations
Case Reports

A novel transmembrane CXCR4 variant that expands the WHIM genotype-phenotype paradigm

Katarina Zmajkovicova et al. Blood Adv. .
No abstract available

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Conflict of interest statement

Conflict-of-interest disclosure: X4 Pharmaceuticals funded the development of this manuscript. T.K.T. is a principal investigator in the phase 3 trial of mavorixafor for WHIM syndrome and consultant to X4 Pharmaceuticals. N.S. is a consultant to X4 Pharmaceuticals. K.Z., C.N., H.M., K.C., and A.G.T. are employees, and K.Z., C.N., H.M., and A.G.T. hold shares of X4 Pharmaceuticals. A.B., S.P., S.M.-M., and I.W. hold shares of X4 Pharmaceuticals. J.E.W. has received funding for investigator initiated and sponsored clinical trials from X4 Pharmaceuticals; served as consultant to X4 Pharmaceuticals in the past; and is a principal investigator of investigator-initiated and sponsored studies funded by X4 Pharmaceuticals. J.L.N. serves as a speaker for EUSA Pharma; an adviser for EUSA Pharma and AbbVie; and a consultant for Guidepoint Global. The remaining authors declare no competing financial interests.

Figures

Figure 1.
Figure 1.
Impact of CXCR4D84H on receptor function and effects of mavorixafor treatment in P1. (A) PBMCs isolated from HDs (HD-SC and HD-IH) and P1 were stimulated with CXCL12 (vehicle, 1 nM, 10 nM, and 100 nM) for 45 minutes. CXCR4 receptor internalization was assessed by surface expression of CXCR4 by flow cytometry. Values are expressed as percent of remaining CXCR4 expression compared with vehicle-treated cells. PBMCs were subtyped using fluorescent mAbs specific for CD3 (T cells), CD19 (B cell), and CD3 and CD56 (NKT cells). Statistical analysis was not performed owing to low sample numbers. Mean ± standard deviation (SD) of 2 to 4 from 2 biological replicates. (B) K562 cells were transiently transfected with indicated CXCR4 constructs and stimulated with CXCL12 (vehicle, 1 nM, 10 nM, and 100 nM) for 45 minutes. CXCR4 receptor internalization was assessed by surface expression of CXCR4 via flow cytometry. Statistical significance determined by Mann-Whitney tests comparing variants with the WT at each respective concentration of ligand. Values are expressed as percent of remaining CXCR4 compared with vehicle-treated cells. Values represent mean ± standard error of mean (SEM) of 3 to 17. ∗P < .05; ∗∗P < .01; ∗∗∗P < .001. (C) Transwell chemotaxis assay: PBMCs isolated from HDs (HD-SC and HD-IH) and P1 migrated toward 2, 10, and 50 nM CXCL12 or medium only for 2.5 hours. The total number of migrated cells was normalized to the HD-SC with 10 nM CXCL12 in each cell type. PBMCs were subtyped using fluorescent mAbs specific for CD3 (T cells), CD19 (B cell), and CD3 and CD56 (NKT cells). Statistical analysis in PBMCs was not performed owing to low sample numbers. Mean ± SD of 2. (D) Transwell chemotaxis assay: K562 cells transiently transfected with indicated CXCR4 constructs migrated toward 0.4, 2, and 10 nM CXCL12 or medium only for 4 hours. The total number of migrated cells was normalized to WT with 2 nM CXCL12 in each assay. Statistical significance was determined by Mann-Whitney tests (0.4 nM and 10 nM) and 1-sample Wilcoxon signed-rank test (2 nM) comparing variants with the WT at each respective concentration of ligand. Mean ± SEM of 5 to 20. ∗P < .05; ∗∗P < .01; ∗∗∗P < .001. (E) PBMCs isolated from HDs (HD-SC and HD-IH) and P1 were stimulated with vehicle, 100 nM or 1 µM CXCL12 to measure Ca2+ mobilization. Statistical analysis was not performed owing to low sample numbers; n = 2 technical replicates; the experiment is representative of 2 independent experiments. (F) K562 cells with stable CXCR4 expression were stimulated with serial dilutions of CXCL12 to measure Ca2+ mobilization. Relative fluorescence units measured in WT-expressing cell line at 1 µM CXCL12 represented 100%. Mean ± SEM of 11 to 12. (G) K562 cells with stable CXCR4 expression were stimulated with forskolin ± 100 nM CXCL12 for 30 minutes. cAMP production was measured by enzyme-linked immunosorbent assay. Percent of inhibition of cAMP production by CXCL12 was calculated respective to forskolin-only treated cells. Statistical significance was determined by Mann-Whitney tests comparing variants with the WT. Values represent mean ± SEM of 9 to 12. ∗P < .05; ∗∗P < .01; ∗∗∗P < .001. (H) Time course of WBC counts, ANC, and ALC for P1 before receiving oral mavorixafor (day –1) and on the first day of mavorixafor treatment (day 1). ALC, absolute lymphocyte count; AMC, absolute monocyte count; ANC, absolute neutrophil count; CD, cluster of differentiation; HD-IH, healthy donor in-house; HD-SC, healthy donor shipping control; mAb, monoclonal antibody; NK, natural killer; NKT, natural killer T; ns, not significant; WBC, white blood cell; WT, wild type.
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References

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