Figure 5: PT-Cy enabled the generation of myeloma-driven CD8⁺ T cell exhaustion.

B6 MM-bearing recipients were transplanted with 5 × 10⁶ BM with 0.5 × 10⁶ CD8⁺ and 0.5 × 10⁶ CD4⁺ T cells from C3H.SW donors. (A) MM-bearing recipients of T cell replete grafts were untreated (alloBMT) or administered 50 mg/kg cyclophosphamide on D+3 and D+4 (PT-Cy) and monitored for myeloma burden using M-band (log G/A; n = 24 from 4 experiments; Mann-Whitney U test) and survival (n = 17 from 3 experiments; Log-rank test). (B) MM-bearing recipients of T cell deplete BM grafts (TCD) were treated as above and monitored for M-band (n = 5 from 1 experiment; Student’s t-test). (C-I) MM-bearing and MM-naive recipients were treated as above and BM was harvested at 7 weeks post-transplant for immunophenotyping using flow cytometry (n = 8 – 14 from 2 experiments). (C) Representative contour plots. (D) CD8⁺, conventional CD4⁺ (FoxP3⁻), and regulatory CD4⁺ T cell enumeration per femur. (E) Frequency of naïve T (T_N; CD44⁻CD62L⁻), central memory T (T_CM; CD44⁺CD62L⁺), effector memory T (T_EM; CD44⁺CD62L⁻), and effector T (T_EFF; CD44⁻CD62L⁻), (F) DNAM-1 and TIGIT expressing cells, and (G) TOX⁺, CD101⁺ and TIM-3⁺ cells within CD8⁺ T cells. (H) Frequency of DNAM-1 and TIGIT expressing cells and (I) TOX⁺, CD101⁺ and TIM-3⁺ cells within conventional CD4⁺ T cells. Data represent mean ± SEM. One-way ANOVA with Tukey’s multiple comparisons test or Kruskal-Wallis test with Dunn’s multiple comparisons test. * p < 0.05 ** p < 0.01 *** p <0.001