Anti-CD38 targeted radiation is effective and enhances CAR-T cell recruitment and expansion in resistant extramedullary multiple myeloma Free

Multiple myeloma (MM) cell proliferation is typically confined to the bone marrow (BM) niche (intramedullary disease, IMD). However, a subset of patients develop extramedullary disease (EMD), where MM cells proliferate outside the BM. EMD currently poses a significant challenge to MM treatment because of its aggressive nature and resistance to current therapies, including the anti-CD38 antibody daratumumab (Dara) and CAR-T cells. Recent studies, including our own, have demonstrated that, at relapse, most Dara-treated patients (~80%) maintain targetable CD38 expression on their IMD MM clones, and recent data have also shown that patients carrying EMD lesions exhibit lower CD38 levels and reduced immune cell infiltration, potentially contributing to immune therapy resistance, compared to effects in IMD (Jelinek et al. Leukemia 2022).

Our in vivo data also reveal that, when MM cells form EMD tumors in mice, Luc+ CAR-T cells directed toward CS-1 lack anti-MM activity and are poorly recruited to the site of the tumor, compared to levels observed in mice with IMD tumor engraftment. Specifically, the effect size of CS1 CAR-T cell therapy, as measured by Cohen's d, was substantially greater in IMD mice (d=4.50, 95%CI: 2.25–6.74) than in EMD mice (d=0.12, 95%CI: -0.81–1.04), and the area under the curve for CAR-T cell recruitment and expansion as measured by bioluminescence imaging (BLI) for a period of 2 weeks was 9.9 (95% CI: 8.1–11.6) x10¹⁴ photons in IMD mice vs 4.2 (95% CI: 2.4–6.0) x10¹² photons in EMD mice.

Furthermore, in humanized NSG mice with IMD, Dara therapy initially induces a deep response, but relapses occur, with CD38+ MM cells forming diffuse EMD lesions. Notably, these relapsed mice show no MM engraftment in the BM and exhibit limited immune infiltration in EMD tumors, supporting the role of tumor localization in therapeutic resistance.

To investigate if CD38 remains a viable targetable in EMD, independently of immune system engagement, we conjugated Dara to the alpha-emitting radionuclide Actinium-225 (²²⁵Ac-Dara) as a targeted radionuclide therapy (TRT) toward CD38-expressing cancer cells. Treatment with a single 200 nCi (~5 µg) dose of ²²⁵Ac-Dara significantly extended median survival in mice with EMD (46 days, 95%CI: 38–47; n=10) compared to levels in untreated controls (28 days, 95%CI:25–32; n=10; p=0.001), indicating the presence of targetable CD38 receptors on cancer cells.

In EMD-bearing mice, the therapeutic efficacy of CD38-TRT could be further improved by a second (2xTRT) and a third (3xTRT) infusion at an interval of 1 half-life of ²²⁵Ac (~10 days). While increased radiation dosing in IMD mice led to toxicity that outweighed therapeutic benefits, EMD-bearing mice showed improved survival with repeated dosing. Median survival increased from 43 days (95%CI: 43–46) with a single dose to 56 days with two doses (95%CI: 50–60; p=0.006) and 60 days with three doses (95%CI: 56–60; p<0.001), without evidence of increased toxicity. Immunohistochemical analysis of EMD tumors revealed significant changes in tumor architecture, including reduced vascularization and increase in tumor necrosis, in CD38-TRT–treated mice compared to controls (p=0.03).

These findings led us to investigate if CD38-TRT could facilitate CAR-T cell infiltration and expansion within EMD tumors. We randomized mice with comparable EMD masses into 3 groups: untreated (n=9), treated with 3x10⁶ Luc+ CS1-CAR T cells as a monotherapy (n=9), or 200 nCi CD38-TRT followed by these CAR T cells (n=11).

Mice pre-treated with CD38-TRT exhibited significantly greater recruitment and expansion of CS1-CAR-T cells at both early (5–7 days) and late (8–13 days) timepoints compared to levels in mice receiving CAR-T cells alone (p<0.001). After one week, the area under the CAR-T BLI curve for the TRT–pretreated mice and those that were not TRT–pretreated were 2.1 (95%CI: 1.0–3.2) x10¹² vs 4.5 (95%CI: 3.2–5.8) x10¹¹photons, respectively.By the end of the second week, these values were 6.1 (95%CI: 1.5-10.7) x10¹³ and 3.8 (95%CI: 2.0–5.6) x10¹² photons, respectively. Moreover, the TRT–pretreated mice had significantly higher tumor necrosis when compared to the other treatment groups.

In sum, we report for the first time that CD38-TRT not only provides therapeutic benefit in immune-resistant EMD but also enhances CAR-T cell infiltration and expansion within these tumors. These findings suggest that CD38-TRT may improve the efficacy of T cell–based therapies in EMD.