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B-cell maturation antigen-targeted CAR T cell as a salvage
therapy for progressive multiple myeloma after anti-G
protein-coupled receptor, class C group 5 member D CAR
T-cell therapy
by Xiaotian Zhang, Fan Xia, Kunming Qi, Jieyun Xia, Dian Zhou, Qian Sun, Huanxin Zhang,
Xiangmin Wang, Wei Sang, Depeng Li, Zhiling Yan, Alex H Chang, Jiang Cao and Bin Pan
Received: May 21, 2026.
Accepted: September 17, 2026.
Citation: Xiaotian Zhang, Fan Xia, Kunming Qi, Jieyun Xia, Dian Zhou, Qian Sun, Huanxin Zhang,
Xiangmin Wang, Wei Sang, Depeng Li, Zhiling Yan, Alex H Chang, Jiang Cao and Bin Pan. B-cell maturation
antigen-targeted CAR T cell as a salvage therapy for progressive multiple myeloma after anti-G protein-coupled
receptor, class C group 5 member D CAR T-cell therapy.
Haematologica. 2026 Sept 24. doi: 10.3324/haematol.2026.301282 [Epub ahead of print]
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1
B-cell maturation antigen -targeted CAR T cell as a salvage
therapy for progressive multiple myeloma after anti-G protein-
coupled receptor, class C group 5 member D CAR T-cell therapy
Xiaotian Zhang1,2*, Fan Xia1,2*, Kunming Qi1,2*, Jieyun Xia1,2, Dian Zhou1,2, Qian
Sun1,2, Huanxin Zhang1,2, Xiangmin Wang1,2, Wei Sang1,2, Depeng Li1,2, Zhiling
Yan1,2, Alex H Chang3,4#, Jiang Cao1,2#, Bin Pan1,2#
1Department of Hematology, The Affiliated Hospital of Xuzhou Medical University,
Xuzhou, Jiangsu, China
2Blood Diseases Institute, Xuzhou Medical University, Xuzhou, Jiangsu, China
3Shanghai YaKe Biotechnology, Shanghai, China
4Engineering Research Center of Gene Technology, Ministry of Education, Institute of
Genetics, School of Life Sciences, Fudan University, Shanghai, China
*XZ, FX, and KQ contributed equally to this work.
#Correspondence:
Alex H Chang : Shanghai YaKe Biotechnology, 1600 Guo Quan Bei Lu, Shanghai
200438, China, e-mail: changah@yakebiotech.com;
Jiang Cao: The Affiliated Hospital of Xuzhou Medical University, 99 West Huaihai
Road, Xuzhou 221002, China, e-mail: zimu05067@163.com;
Bin Pan : The Affiliated Hospital of Xuzhou Medical University, 99 West Huaihai
Road, Xuzhou 221002, China, e-mail: panbinsam@163.com.
Running heads
BCMA CAR T-cells for RRMM after GPRC5D CAR T-cells
Data availability
Original data are available with panbinsam@163.com upon request.
Competing interests
AHC is a founding member of Shanghai YaKe Biotechnology, a
biotechnology company focusing on research and development of tumour cellular
immunotherapy. All other authors declare no competing interests.
Funding
This work is supported by funding from the National Natural Science Foundation
of China (82370218, 82570268, 82341203, 82570272), Jiangsu Science Fund
for Distinguished Young Scholars (BK20250055), Advanced Program of The
Affiliated Hospital of Xuzhou Medical University (PYJH2025101), General Project
of Jiangsu
2
3
Commission of Health (M2022103) and Jiangsu Province High -Level Hospital
Construction Project (GSPJS202504).
Authors' contributions
BP, ZY , JC, and AHC designed the study. XZ, KQ, JX, DZ, QS, HZ, and XW
collected research data. XZ and FX did the statistical analysis. XZ drafted the
manuscript. BP, ZY , JC, and AHC performed critical revision of the manuscript for
important content. All authors contributed to the manuscript and interpretation of the
data and approved the final version of the manuscript.
Acknowledgements
We sincerely appreciate all the participants in this study. We also thank all physicians,
nurses, and other staff members who coordinated the clinical study.
Clinical trial registration information
Clinical trials were registered at www.chictr.org.cn as # ChiCTR1900026219, #
ChiCTR2000033567 and www.clinicaltrials.gov as # NCT05509530.
4
Abstract
Repeat infusion of chimeric antigen receptor (CAR) T -cell is effective for patients
with refractory or relapsed multiple myeloma (RRMM) after failing a prior CAR T-
cell therapy. It is unknown whether B-cell maturation antigen (BCMA) CAR T-cell is
effective for patients with RRMM after anti-G protein-coupled receptor, class C group
5 member D (GPRC5D) CAR -T cell therapy. In this study, 20 subjects with RRMM
after a prior anti -GPRC5D CAR T-cell therapy received BCMA-targeted CAR T-cell
therapy as a salvage treatment . During a median follow-up of 13.4 months (IQR 3.6 -
21.2), 13 subjects showed responses including 9 with stringent complete response
(sCR) and 4 with partial response. The median duration of response was 10.2 months
(95% CI, 2.6 - not reached ). The median progression-free survival (PFS) were 9.2
months (95% CI, 3.0 - 13.6) in all subjects and 20.8 months (95% CI, 5.3 - not
reached) in subjects with sCR. There is no statistic difference in overall response rate
(ORR) and median PFS between this study and our previous study, which enrolled 37
subjects with RRMM receiving anti-GPRC5D CAR T-cells after failing anti-BCMA
CAR T-cell therapies (ORR: 65% vs 84%, P = 0.18; PFS: 9.2 vs 4.5 months, P = 0.93).
The most common a dverse event s were grade ≥3 haematological toxicities (n=18).
Fourteen subjects (78%) had cytokine release syndrome (grade ≥3, n=1) . In
conclusion, anti-BCMA CAR T-cell therapy was effective for RRMM after failing an
anti-GPRC5D CAR T-cell therapy, and the safety profile was acceptable.
5
Introduction
B-cell maturation antigen (BCMA) is the most widely explored and used target for
immunotherapies in multiple myeloma (MM), while G protein -coupled receptor, class
C group 5 member D (GPRC5D) is emerging as another potential target. We and
others previously reported that both anti-BCMA chimeric antigen receptor (CAR) T-
cell and anti-GPRC5D CAR T-cell therapies showed high response rates in relapsed
or refractory MM (RRMM) including overall response rates (ORR) of 70%-100% and
complete response rates (CRR) of 30%-80%.1-6 However, a considerable part of
patients experienced disease progression, including refractory and relapse after
response.4,5,7,8 There is no standard -of-care therapy for these patients after failing
CAR T-cell therapies. Repeat infusion of CAR T-cell might be an option.
Different therapies were used to treat patients with RRMM after BCMA-targeted
CAR T -cell therapy ,9 and immunotherapies induced responses in these patients.
Repeat infusion of CAR T -cells showed efficacy in patients with RRMM who had
failed a prior CAR T-cell therapy. In KarMMa trial 28 subjects with RRMM received
a second infusion of idecabtagene vicleucel. Six responded to the second infusion of
CAR T-cell including 1 with very good partial response and 5 with partial response
(PR).10 In a multicenter study 7 subjects with RRMM failing anti-BCMA CAR T-cell
therapies received anti-GPRC5D CAR T-cell, 6 responded to anti-GPRC5D CAR T-
cell and the median progression-free survival ( PFS) was 18.2 months (95%
confidence interval [CI], 14 .4 - not estimatable ).11 These findings suggested that a
second infusion of CAR T -cells was feasible and changing the target of CAR T -cell
6
seemed to be more effective.
It is important to evaluate whether anti -BCMA T-cell and anti-GPRC5D CAR T-
cell could be salvage therapies for each other, and which treatment sequence would be
better. In our previous study, 37 subjects with RRMM who failed a prior anti-BCMA
CAR T -cell therapy received anti-GPRC5D CAR T -cell. Thirty -one subjects
responded to anti-GPRC5D CAR T-cell, including 13 with complete response (CR). 12
The number of patients treated by anti-GPRC5D CAR-T cell was limited, and those
who received a second infusion of CAR T -cell were even fewer. Thus, few studies
have systematically evaluated the efficacy of repeat infusion of CAR T-cell in patients
who had failed a prior anti-GPRC5D CAR -T cell therapy. In this study, we
retrospectively investigated the efficacy and safety of anti-BCMA CAR T-cell therapy
for 20 subjects with RRMM after anti-GPRC5D CAR-T cell therapy, and compared
the efficacy with that in the cohort of receiving anti-GPRC5D CAR T-cell after failing
anti-BCMA CAR T-cell therapies.
Methods
Study design and subjects
From March 2022 to October 2025, 24 subjects with RRMM involved in 3 clinical
trials (ChiCTR1900026219, ChiCTR2000033567, and NCT05509530) conducted in
our institution were screened for this study . All the subjects had RRMM after a prior
anti-GPRC5D CAR T -cell therapy and underwent apheresis for the productions of
BCMA-targeted CAR T -cells. Inclusion and exclusion criteria was described
7
previously.13-15 Four subjects did not receive BCMA-targeted CAR T -cells due to
CAR T-cell production failure, withdrawal of consent, and death . Twenty subjects
who received BCMA-targeted CAR T-cells were included in the analysis (Figure 1).
This study was approved by the Ethics Committee of the Affiliated Hospital of
Xuzhou Medical University (XYFY-2018-KL003-01, XYFY2020 -KL062-01,
XYFY2022-KL118-01), and informed consent was obtained from each participant, in
compliance with the Declaration of Helsinki.
CAR-T cell manufacturing
The production s of anti-BCMA CAR T-cells, anti-BCMA/CD19 CAR T -cells, and
anti-BCMA/GPRC5D CAR T -cells were performed as previously described .13-15
Briefly, peripheral blood mononuclear cells were obtained by a blood cell separator,
and T lymphocytes were sorted using anti -CD3 magnetic beads. T lymphocytes were
transduced with lentiviruses encoding the second -generation CAR structural
fragments (scFv/ 4-1BB/CD3ζ) and amplified in vitro for 5 -10 days prior to
examination and infusion .13-15 The bispecific BCMA/CD19 -CAR scFv and t he
bispecific BCMA/GPRC5D scFv constructs were described previously.14,15
Clinical procedures
Subjects could receive bridging antimyeloma therapies, which should be ended at
least 14 days before lymphodepletion. Expressions of BCMA on myeloma cells were
confirmed by flow cytometry for all subjects before lymphodepletion (Table S1,
Figure S1 ), which comprised of fludarabine (30 mg/m 2/day) for 3 days and
cyclophosphamide (750 mg/m2/day) for 1 day followed by 2 days of interval before
8
CAR T-cell infusion. Subjects received a median dose of intravenous CAR T -cell at
1.6 × 10⁶ cells per kg (IQR 1.3-2.0).
Response and toxicity assessment
After CAR T-cell infusion, responses were assessed since day 14 according to the
International Myeloma Working Group criteria.16 The assessment indicators included
measurable residual disease (MRD) in the bone marrow, serum paraprotein ,
immunoglobulin concentration, immunofixation electrophoresis, free immunoglobulin
light chains, and quantitation of 24-hour urine protein. MRD negativity was defined
as the absence of phenotypically aberrant clonal plasma cells, which was assessed by
multi-colour flow cytometry according to the EuroFlow protocol (minimum
sensitivity, 1 in 10⁵) .17 The assessment of extramedullary disease included imaging
techniques and physical examination. Cytokine release syndrome (CRS) and immune
effector cell associated neurotoxicity syndrome were graded according to the
American Society for Transplantation and Cellular Therapy criteria.18 Haematological
toxicities, other neurologic toxicities and adverse events (AEs) were graded according
to the National Cance r Institute Common Terminology Criteria for Adverse Events
version 5.0.
Outcomes
The primary end point was ORR referring to best overall response which included
stringent complete response (sCR), CR, very good partial response and PR as assessed
according to the International Myeloma Working Group criteria.16 Secondary end
points were PFS, duration of response (DOR) and safety (including CRS, neurologic
9
toxicities and other AEs).
Statistical analysis
Descriptive statistics include medians with inter-quartile range (IQR) for continuous
variables and counts and percentages for categorical variables. Clopper-Pearson 95%
CIs and Fisher’s exact test were used for categorical variables. Continuous variables
were tested by Mann -Whitney U test for two groups. DOR and PFS of subjects were
determined using Kaplan -Meier and compared using the log -rank test . All analyses
were performed with Statistical Package for the Social Sciences 26 or GraphPad
Prism 10. P values < 0.05 (two-tailed) were considered significant.
Results
Baseline characteristics
A total of 20 subjects with RRMM receiving a second CAR T-cell infusion were
included in the study. Clinical characteristics of subjects are presented in Table 1 and
Table S1. Median age was 61 years (IQR 58 - 64) and 12 of 20 (60%) subjects were
male. The median time at the second CAR T-cell infusion since diagnosis was 69.3
months ( IQR 31.7 - 85.2). Fourteen subjects ( 70%) had Second Revision of the
International Staging System III/IV disease ,19 5 subjects ( 25%) had extramedullary
disease, and 16 subjects (80%) had high -risk cytogenetic abnormalities. The 20
subjects had received a median of 6 lines of previous therapy (IQR 4 - 8), 17 (85%)
were refractory to at least one proteasome inhibitor and 18 (90%) were refractory to at
least one immunomodulatory drug. Six subjects (30%) had received auto-transplants
10
and 10 (50%) had triple-class refractory diseases. The median DOR was 10.3 months
(95% CI, 4.7 - 18.2) in 12 evaluable subjects who achieved PR or better in the prior
anti-GPRC5D CAR T-cell therapies. The median time from anti-GPRC5D CAR T-cell
infusion to anti-BCMA CAR T-cell infusion was 14.4 months (IQR 4.8 - 22.3).
Efficiency
During a median follow-up of 13.4 months (IQR 3.6 - 21.2), 13 of 20 subjects (65%)
achieved responses, including 9 (45%) with sCR and 4 with (20%) PR (Figure 2A, B).
The median time to initial response was 26 days (IQR 14 - 57), and the median time
to best response was 31 days (IQR 18 - 57). Bone marrow MRD was evaluated in 17
subjects. In the 4 subjects who were bone marrow MRD-negative before anti-BCMA
CAR T-cell infusion, 2 achieved sCR and 1 achieved PR after CAR T-cell infusion. In
the 13 subjects who were bone marrow MRD-positive before anti-BCMA CAR T-cell
infusion, bone marrow MRD turned negative after CAR T-cell infusion in 11 subjects
including 7 with sCR and 2 with PR (Figure 2C).
In the 13 subjects with responses, the median DOR was 10.2 months (95% CI, 2.6 -
not reached) (Figure 3A), and 7 subjects experienced disease progression during the
follow-up (Figure 2A). The median PFSs were 9.2 months (95% CI, 3.0 - 13.6) in all
subjects with responses and 20.8 months (95% CI, 5.3 - not reached) in the 9 subjects
with sCR (Figure 3B). Although there was no statistically significant difference in
ORR and CRR between this study and our recent study (ORR: P = 0.18, CRR: P =
0.57),12 the ORR in this study seems lower than that in the cohort of our recent study,
which reported the ORR was 84% (n=31) in 37 subjects with RRMM receiving anti -
11
GPRC5D CAR T-cells after failing a prior anti-BCMA CAR T-cell therapy.12 We also
compared the PFS with that in our recent study, which showed the m edian PFS was
4.5 months (95% CI, 2.9 - 6.2) in all subjects and was not reached in subjects with CR
or better during a median follow-up of 12.6 months,12 and no statistical difference was
observed (all subjects: 9.2 vs. 4.5 months, P = 0.93; CR or better: 20.8 vs. not reached,
P = 0.20).12
Following disease progression after anti-GPRC5D CAR T-cell therapy, 11 subjects
received anti -myeloma therapies before anti -BCMA CAR T-cell treatment, with 5
subjects receiving 2 lines of treatment. Eight subjects received anti-CD38 antibody-
based therapies, and 1 of 3 responders achieved CR. One of 3 subjects receiving
aponermin-based therapies achieved an objective response . None responded to triplet
or quadruplet combination of approved agents (Table S2). Among the 9 subjects who
achieved sCR in the salvage anti-BCMA CAR-T cell therapy, 8 had response s in the
prior anti-GPRC5D CAR-T cell therapy including 6 with sCR and 2 with PR . Among
the 7 subjects refractory to the prior anti-GPRC5D CAR-T cell therapy, 4 responded
(sCR, n=1; PR, n= 2; minimal response, n=1) to the salvage anti-BCMA CAR-T cell
therapy (Table S2).
Safety
The AEs within the first 30 days are evaluated in 18 subjects ( Table 2 ). Before
lymphodepletion, 13 (72%) subjects had cytopenia to different extent, including 11
(61%) with neutropenia, 8 (44%) with lymphopenia , 9 (50%) with anaemia, and 9
(50%) with thrombocytopenia. After infusion of anti-BCMA CAR-T cells, all the 18
12
subjects had grade ≥3 haematological AEs, including lymphopenia ( 18 [100%]),
leukopenia (n= 15 [83%]), and neutropenia (n= 15 [83%]). The median duration of
grade ≥3 lymphopenia was 11 days (IQR 5 - 15), leukopenia 10 days (IQR 3 - 12),
and neutropenia 9 days (IQR 3 - 19). Eight subjects (44%) had grade ≥3
thrombocytopenia and 5 (28%) were with grade ≥3 anaemia.
Within the first 30 days a fter anti-BCMA CAR T-cell infusion, 14 (78%) subjects
had CRS including 1 with grade 3 CRS. The median time to onset of CRS was 7 days
(IQR 5 - 9), and the median duration was 4 days ( IQR 3 - 7). Nine subjects (64%)
received glucocorticoids, and 1 received tocilizumab. Levels of IL-6, ferritin, and C -
reactive protein were measured twice weekly after infusion of anti -BCMA CAR T-
cells. The median time of reaching peak levels of IL-6, ferritin, and C-reactive protein
were 9 days (IQR 8 - 11), 11 days (IQR 9 - 15), and 10 days (IQR 7 - 11) after anti-
BCMA CAR T-cell infusion, respectively. In the subjects with CRS, the peak levels of
IL-6, ferritin, and C-reactive protein were significantly higher than baselines (IL-6: P
< 0.05, ferritin: P = 0.05, C-reactive protein: P < 0.0001) (Figure S2 ). Neurotoxic
event was observed in 1 subject (5%) who showed brief self-limited headache but
without disturbance of consciousness, paraphasia or dysgraphia. Immune effector cell
associated neurotoxicity syndrome was not observed in all the 18 subjects.
All non-hematologic toxicity effects resolved quickly with supportive care without
severe organ damage. Grade ≥3 non-hematological toxic events occurred in 4 subjects
including 1 with grade 3 increase of alanine aminotransferase level, 1 with grade 3
increase of aspartate aminotransferase level, 1 with decrease of fibrinogen level, and 1
13
with infection . The other non-hematological toxic events included grade 1 or 2
malignant nausea (n=8 [4 4%]), hypophosphataemia (n=1 2 [67%]), hypocalcaemia
(n=7 [39%]), hypoalbuminemia (n= 7 [39%]), and prolonged activated partial
thromboplastin time (n = 10 [56%]).
Lymphocyte subset data were available in 13 subjects. B-cell percentage in
peripheral blood lymphocytes decreased during the first 30 days after CAR T -cell
infusion (Figure S3). In 17 evaluable subjects, 15 showed decreased levels of two or
three uninvolved immunoglobulins, and 1 showed decreased level of one uninvolved
immunoglobulin (IgA) within 60 days after CAR T-cell infusion (Figure S4).
Discussion
There is no standard-of-care therapy for patients with RRMM after failing CAR T-cell
therapies. In this study, 20 subjects with RRMM received salvage anti-BCMA CAR T-
cell therapies after failing a prior anti -GPRC5D CAR T-cell therapy. Thirteen (65%)
subjects showed responses to anti -BCMA CAR T-cell therapies, and the median PFS
was 20.8 months in the 9 (45%) subjects who achieved sCR. The efficacy in this study
is comparable to that in the cohort of our recent study, in which 37 subjects with
RRMM received anti-GPRC5D CAR T-cells after failing a prior anti -BCMA CAR T-
cell therapy.12 Our studies systematically documented that the two targets (BCMA and
GPRC5D) could be used alternatively when repeat infusion of CAR T -cell is
considered for treating RRMM . The same is true for other BCMA or GPRC5D -
directed therapies. Joiner and colleagues reported BCMA-directed therap ies,
14
including bi-specific antibodies and CAR T -cell, were effective in subjects with
RRMM after failing GPRC5D-directed therapies.20 In subjects with RRMM receiving
GPRC5D bi-specific antibodies as a salvage treatment after anti-BCMA CAR T-cell
therapies, the ORR and CRR were 70%-80% and 40%, and the 12-month DOR was
50%-60%.21,22 Repeat infusion of CAR T -cell also showed effectiveness in other
hematological malignancies. In a phase -1/-2 trial 44 subjects with refractory B -cell
malignancies (acute lymphoblastic leukemia, n=14; chronic lymphocytic leukemia,
n=9; non-Hodgkin lymphoma, n=21) failing a first infusion of anti-CD19 CAR T-cells
received a second infusion of anti -CD19 CAR T-cells. 39% responded to the second
infusion, and 2-year PFS and OS were 23% (95% CI, 9-59) and 36% (95% CI, 19-71)
in subjects who achieved PR or better at day 28 after the second infusion. 23 In a
phase-1 trial anti -CD22 CAR T -cell therapy was used to treat 3 7 subjects with
advanced LBCL who relapsed after anti-CD19 CAR T -cell therapies. The ORR and
CRR were 68% and 53% , and the median DOR was 27.8 months (95% CI, 5.1 -not
evaluable).24
Although statistical significance was not reached ( P = 0.18), t he ORR in the anti-
BCMA after anti-GPRC5D cohort of this study seems lower than that in the anti -
GPRC5D after anti -BCMA cohort of our previous report.12 We deduce the lower
response rate might result from the disease per se , because the percent of subjects
with high-risk cytogenetic profile in this study was higher than that in the previous
study (80% vs. 43%).12 BCMA is more extensively expressed on myeloma cells as
compared with GPRC5D.2,12,25 Anti-BCMA CAR T-cells used in front might eliminate
15
myeloma sub -clones more extensively, comparing with anti -GPRC5D CAR T -cells
used in front . Others reported that anti -GPRC5D CAR T-cells potentially limited the
expansion of anti -BCMA CAR-T cells when they were used concurren tly.26 There
was a 14.4 -month interval between infusion of the two types of CAR T -cells in this
study, and both CAR T-cells could expand well in vivo when used as salvage therapies
in our studies (data not shown).12 Thus, the two types of CAR T-cells might not have a
direct impact on each other herein. These findings suggested the treatment sequence
of anti-GPRC5D CAR T-cells after anti-BCMA CAR T-cells might be a better choice
for RRMM. Nonetheless, the PFS in this study was comparable to that in our previous
report,12 which indicated the anti-BCMA CAR T-cells were effective even been used
as a salvage therapy. At the cutoff date of follow -up, 8 of 9 subjects with sCR in this
study are alive.
The efficacy of repeat infusion of CAR T -cell could be affected by factors such as
antigen escape and T -cell fitness. Loss of BCMA or GPRC5D antigen was observed
among patients with MM who relapsed after anti-BCMA CAR T-cell or anti-GPRC5D
CAR T -cell therapies .27-30 We found one patient with deletion mutations of both
TNFRSF17 (encoding BCMA) and GPRC5D genes after sequential anti-BCMA CAR
T-cell and anti -GPRC5D CAR T -cell therapies. 30 These findings suggest that
myeloma cells heterogeneity and genomic instability can facilitate clonal outgrowth
of antigen -negative myeloma cells under selective pressure of CAR T -cells. In
addition, modification of proteins also causes antigen-loss such as gamma-secretase
mediated shedding of BCMA from membrane of myeloma cells. 31,32 We recently
16
reported that plasma cell tumors were GPRC5D-negative in 8 of 10 subjects with MM
relapsing after anti -GPRC5D CAR T -cell therapies, 30 and it is reasonable to change
the target when repeat infusion of CAR T -cell is considered for these patients. Tumor
cells were BCMA-positive in all subjects before receiving anti-BCMA CAR T-cells in
this study. These results could interpret why anti-BCMA CAR T-cells remain effective
after failing anti-GPRC5D CAR T -cell therapies. Both the anti-GPRC5D after anti -
BCMA sequence and anti-BCMA after anti-GPRC5D sequence showed effectiveness
in our studies, which indicated that neither anti -GPRC5D CAR -T cells nor anti -
BCMA CAR T-cells significantly decreased the expression of the other target ant igen.
However, the underlying mechanism is unknown.
T-cell function and persistence are crucial factors associated with CAR T -cell
efficacy.33 Our previous studies showed both anti -GPRC5D CAR -T cells and anti -
BCMA CAR T-cells expanded well in vivo in most patients who had not been treated
by CAR T -cells, and the CAR T -cells persisted as long as 1 year in a small part of
patients.2,13-15 When be used after a prior CAR T -cell therapy, the expansion and
persistence of anti -GPRC5D CAR -T cells and anti -BCMA CAR T-cells mig ht be
affected by more complex factors such as a higher tumor burden and a more
immunosuppressive tumor microenvironment which mediated resistance to CAR T -
cells.34-36
CAR T -cells often lead to significant toxic reactions because of activation o f
immune system. It is important to re -evaluate the safety when repeat of CAR T -cell
therapy is conducted. In this study and our previous study ,12 the grade ≥3 CRS was
17
comparable (6% vs. 5%) and most of AEs were manageable, and none died from CAR
T-cell infusion. The most common grade ≥3 AEs were early h aematotoxicities, which
was potentially resulted from chemotherapy drugs and inflammation induced by CAR
T-cells. Other studies also showed the most common grade ≥3 AEs were
haematotoxicities in RRMM subjects receiving anti-BCMA CAR T-cell therapy, with
incidences of more than 85%.4,13,14,37 The incidences of CRS were 75% to 95%, and
the majority were grade 1 or 2 .4,13,14,37 The incidences of AEs were not increased in
subjects with B-cell malignancies or plasma cell tumors after the second infusion of
CAR T-cells.12,24,38 These studies, together with ours, indicated that a second use of
CAR T-cell therapy might not increase AEs.
Our study has limitations. The findings from this retrospective study with limited
sample size need to be confirmed in a larger clinical trial. The long-term efficacy and
safety of repeat infusion of CAR T-cell reported in this study is unknown and needs to
be validated in a prolonged follow-up duration . Nevertheless, this study has
implications for the options after failing a CAR T-cell therapy in patients with RRMM.
In conclusion, anti -BCMA CAR T -cell as a salvage therapy was effective for
patients with RRMM who had been treated by anti-GPRC5D CAR T -cells, and the
safety was acceptable.
18
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24
Table 1: Baseline characteristics
Characteristic Value
Age
Median age, years 61 (58-64)
Gender
Female 8 (40%)
Male 12 (60%)
ECOG performance status score
0 7 (35%)
1 7 (35%)
2 6 (30%)
Median time since diagnosis, months 69.3 (31.7-85.2)
Median time of interval between two infusions of
CAR T-cells, months
14.4 (4.8-22.3)
Monoclonal globulin
IgA 1 (5%)
IgD 2 (10%)
IgG 14 (70%)
Κ light chain 1(5%)
λ light chain 2 (10%)
R2-ISS
I 2 (10%)
II 4 (20%)
III 9 (45%)
IV 5 (25%)
Extramedullary disease* 5 (25%)
High-risk cytogenetic profile† 16 (80%)
Previous therapy lines 6 (4-8)
Previous autotransplant 6 (30%)
Triple-class refractory disease‡ 10 (50%)
Infused cells
Isolated BCMA 11 (55%)
Bispecific BCMA and CD19 7 (35%)
Bispecific BCMA and GPRC5D 2(10%)
Median dose (×106/kg) 1.6(1.3-2.0)
Data are median (IQR) or n (%). Percentages might be not total 100 because of rounding.
BCMA, B-cell maturation antigen; CAR , chimeric antigen receptor; ECOG , Eastern Cooperative
Oncology Group; GPRC5D, G protein -coupled receptor, class C group 5 member D; R2 -ISS,
Second Revision of the International Staging System.
*Extramedullary diseases included tissue masses in extraosseous locations and bone -related
plasmacytomas. †High-risk cytogenetic prof ile was defined as the presence of del(17p), t(4;14),
t(14;16), and amp(1q). ‡Triple -class refractory disease was defined as disease refractory to an
immunomodulatory agent, a proteasome inhibitor, and an anti-CD38 monoclonal antibody.
25
Table 2: Adverse events within the first 30 days after anti-BCMA
CAR T-cell infusion
All grades Grade 3-4
Haematological disorders
Leukopenia 18(100%) 15(83%)
Neutropenia 18(100%) 15(83%)
Lymphopenia 18(100%) 18(100%)
Anaemia 15(83%) 5(28%)
Thrombocytopenia 15(83%) 8(44%)
Gastrointestinal disorders
Nausea 8(44%) 0
Diarrhoea 3(17%) 0
Abdominal discomfort 1(6%) 0
Metabolism and nutrition disorders
Hypophosphataemia 12(67%) 0
Hypocalcaemia 7(39%) 0
Hyponatraemia 5(28%) 0
Hypokalaemia 5(28%) 0
Hypoalbuminemia 7(39%) 0
Hypoglycemia 1(6%) 0
Respiratory and cardiac disorders
Cough 3(17%) 0
Productive cough 2(11%) 0
Nasal discharge 3(17%) 0
Others
Prolonged activated partial thromboplastin time 10(56%) 0
Decreased fibrinogen 5(28%) 1(6%)
Elevated alanine aminotransferase level 3(17%) 1(6%)
Elevated aspartate aminotransferase level 3(17%) 1(6%)
Elevated cardiac troponin T level 4(22%) 0
Increased lactate dehydrogenase 4(22%) 0
Heart failure 2(11%) 0
Headache 1(6%) 0
Muscular soreness 1(6%) 0
Facial edema 1(6%) 0
Fatigue 2(11%) 0
Infection 5(28%) 1(6%)
Cytokine release syndrome 14(78%) 1(6%)
Immune effector cell–associated neurotoxicity
syndrome
0 0
Data are n (%).
26
Figure Legends
Figure 1. Trial profile.
BCMA, B-cell maturation antigen; CAR, chimeric antigen receptor.
Figure 2. Overall responses.
(A) Swimmer plot of clinical responses to anti -BCMA CAR T-cells. (B) The number
of overall response, minimal response, stable disease, and progressive disease in all
20 enrol led subjects. (C) The number of overall response, minimal response, and
stable disease in all 17 subjects receiving evaluation of MRD after infusion . BCMA,
B-cell maturation antigen; CAR, chimeric antigen receptor ; MRD, minimal residual
disease.
Figure 3. Kaplan-Meier analysis of the DOR and PFS.
(A) Kaplan -Meier curve of DOR in subjects with partial response or better. (B)
Kaplan-Meier curve of PFS in patients treated with anti -BCMA CAR T-cells. DOR,
defined as the time from the date of first ≥ PR evidence to disease progression or
death from any cause. PFS, defined as the time from CAR T -cell infusion to disease
progression or death from any cause. BCMA, B -cell maturation antigen; CAR,
chimeric antigen receptor; DOR, duration of response; GPRC5 D, G protein -coupled
receptor, class C group 5 member D; PFS, progression-free survival.
Supplementary Data
Table S1. Baseline characteristics
Characteristic Patient 1 Patient 2 Patient 3 Patient 4 Patient 5 Patient 6 Patient 7 Patient 8 Patient 9 Patient 10
Age, years 64 58 63 46 58 68 60 59 62 60
Gender Male Male Female Male Female Male Female Female Male Male
ECOG performance status 1 0 2 0 0 1 2 1 2 0
Time since diagnosis, months 8·9 9·5 42·1 86·3 45·1 80·2 71·1 29·5 75·0 100·9
Time of interval between two infusions of CAR T -
cells, months
3.4 2.7 5.0 6.6 19.5 16.9 4.8 22.8 10.3 27.5
Monoclonal globulin IgG IgG IgG IgG IgG λ light chain IgG IgG IgD IgG
R2-ISS disease stage II II III I IV III III III IV I
Extramedullary disease* None None Liver, uterus None None None soft tissue None None None
High-risk cytogenetic profile† None None amp(1q) amp(1q) amp(1q),
del(17p) amp(1q) NE amp(1q) amp(1q) amp(1q)
BCMA antigen expression BCMA+ BCMA+ BCMA+ BCMA+ BCMA+ BCMA+ BCMA+ BCMA+ BCMA+ BCMA+
Previous therapy lines 3 3 5 4 4 8 9 3 8 6
Previous autotransplant None None None None None Yes None None Yes None
Previous proteasome inhibitors
Bortezomib Exposed Yes Yes Yes Yes Yes Yes Yes Yes Yes Yes
Refractory‡ Yes Yes Yes None Yes Yes Yes Yes Yes Yes
Ixazomib Exposed None None None None None Yes Yes Yes Yes Yes
Refractory None None None None None NE Yes NE Yes None
Carfilzomib Exposed None None None None None None None None Yes None
Refractory None None None None None None None None Yes None
Previous immunomodulatory drugs
Thalidomide Exposed None None None Yes Yes Yes Yes None None Yes
Refractory None None None None None None Yes None None Yes
Lenalidomide Exposed Yes Yes Yes Yes Yes Yes Yes Yes Yes None
Refractory Yes Yes Yes None Yes Yes Yes None Yes None
Pomalidomide Exposed None None None None None None Yes None Yes None
Refractory None None None None None None Yes None Yes None
Previous selinexor
Exposed None None None None None None Yes None Yes None
Refractory None None None None None None Yes None Yes None
Previous anti-CD38 monoclonal antibodies
Exposed Yes None None Yes None Yes Yes None Yes None
Refractory None None None None None None Yes None Yes None
Triple-class refractory disease§ None None None None None None Yes None Yes None
Infused CAR T-cells anti-BCMA/
CD19
anti-BCMA/
CD19
anti-BCMA/
CD19
anti-BCMA/
CD19 anti-BCMA anti-BCMA anti-BCMA/
CD19
anti-BCMA/
CD19 anti-BCMA anti-BCMA
Average dose (× 106/kg) 1.3 1.7 1.5 0.9 1.5 1.5 1.6 1.7 1.1 0.9
(continued in next column)
Supplementary
Table S1. Baseline characteristics (continued)
Characteristic Patient 11 Patient 12 Patient 13 Patient 14 Patient 15 Patient 16 Patient 17 Patient 18 Patient 19 Patient 20
Age, years 64 62 60 68 60 57 63 62 49 67
Gender Male Female Male Male Male Male Female Female Male Female
ECOG performance status 2 2 0 0 1 1 0 2 1 1
Time since diagnosis, months 31·4 18·0 86·4 74·3 81·9 32·6 124·7 88·5 48·4 67·5
Time of interval between two infusions of CAR T -
cells, months 18.0 3.2 9.6 12.6 16.1 3.9 30.0 20.8 32.8 27.5
Monoclonal globulin IgA IgD λ light chain IgG Κ light chain IgG IgG IgG IgG IgG
R2-ISS disease stage III III IV III II IV II IV III III
Extramedullary disease None None None None Pleura/thoracic
cavity soft tissue None Thoracic cavity None None
High-risk cytogenetic profile amp(1q) amp(1q) amp(1q),
del(17p) NE del(17p) amp(1q),
del(17p) amp(1q) amp(1q),
del(17p) amp(1q) t(4:14)
BCMA antigen expression BCMA+ BCMA+ BCMA+ BCMA+ BCMA+ BCMA+ BCMA+ BCMA+ BCMA+ BCMA
Previous therapy lines 7 4 7 5 11 9 6 12 6 8
Previous autotransplant Yes None None None None Yes None Yes None Yes
Previous proteasome inhibitors
Bortezomib Exposed Yes Yes Yes Yes Yes Yes Yes Yes Yes Yes
Refractory Yes None None None Yes Yes Yes Yes Yes Yes
Ixazomib Exposed None None Yes None Yes None None Yes None None
Refractory None None Yes None Yes None None Yes None None
Carfilzomib Exposed Yes None None None Yes Yes Yes Yes Yes Yes
Refractory Yes None None None Yes Yes None Yes Yes Yes
Previous immunomodulatory drugs
Thalidomide Exposed None None None None None None None Yes None Yes
Refractory None None None None None None None Yes None None
Lenalidomide Exposed Yes Yes Yes Yes Yes Yes Yes Yes None Yes
Refractory Yes Yes Yes Yes Yes Yes None Yes None Yes
Pomalidomide Exposed Yes None Yes Yes Yes Yes Yes Yes Yes Yes
Refractory Yes None Yes Yes Yes Yes Yes Yes Yes Yes
Previous selinexor
Exposed None None None None None Yes None Yes Yes Yes
Refractory None None None None None Yes None Yes Yes None
Previous anti-CD38 monoclonal antibodies
Exposed Yes None Yes None Yes Yes Yes Yes Yes Yes
Refractory Yes None Yes None Yes Yes Yes Yes Yes Yes
Triple-class refractory disease Yes None Yes None Yes Yes Yes Yes Yes Yes
Infused CAR T-cells anti-BCMA anti-BCMA anti-BCMA anti-BCMA anti-BCMA/
CD19 anti-BCMA anti-BCMA anti-BCMA/
GPRC5D
anti-BCMA/
GPRC5D anti-BCMA
Average dose (× 106/kg) 2.6 1.9 1.3 1.6 2 1.4 1.4 2 2 2
BCMA, B-cell maturation antigen ; CAR, Chimeric Antigen Receptor ; ECOG, Eastern Cooperative Oncology Group ; GPRC5D, G protein-coupled receptor class C group 5
member D; NE, not estimable; R2-ISS, Second Revision of the International Staging System.
*Extramedullary disease was defined as tissue masses in extraosseous locations. †High -risk cytogenetic profile was defined as the presence of del(17p), t(4;14), t(14;16), and
amp(1q). ‡Refractory was defined as disease progression on or within 60 days after the last dose of the most recent drug giv en in each drug class. § Triple-class refractory
disease was defined as disease refractory to an immunomodulatory agent, a proteasome inhibitor, and an anti -CD38 monoclonal antibody.
Table S2. Efficacy of anti-GPRC5D T-cells, antimyeloma therapies and anti-BCMA CAR T-cells
Patients Best response to anti-GPRC5D CAR
T-cells
Antimyeloma therapies/best response before anti-BCMA CAR
T-cells
Best response to anti-BCMA CAR T-
cells
patient 1 PD Dd/PR MR
patient 2 PR None/None sCR
patient 3 PD None/None SD
patient 4 PD D-Vd/PR SD
patient 5 sCR None/None sCR
patient 6 sCR Dd/PD sCR
patient 7 SD D-MRd combined with radiotherapy/SD PR
patient 8 sCR None/None MR
patient 9 sCR SKCd/PD sCR
patient 10 PR None/None sCR
patient 11 sCR D-Kd/PD sCR
patient 12 PD None/None sCR
patient 13 sCR None/None sCR
patient 14 sCR None/None sCR
patient 15 PD D-Kd/PD, Apo-Vd/PR PR
patient 16 SD SKCd/PD, Apo-Kd/PD PD
patient 17 sCR None/None PR
patient 18 VGPR MPT/PD, Apo-Vd/PD PD
patient 19 PR D-KPd/CR, D-SKd/PD PD
patient 20 CR KPd/SD, D-Kd/PD PR
Apo-Kd, Aponermin-carfilzomib-dexamethasone; Apo-Vd, Aponermin-bortezomib-dexamethasone;
BCMA, B-cell maturation antigen; CAR, chimeric antigen receptor; Dd, daratumumab-dexamethasone;
D-Kd, daratumumab-carfilzomib-dexamethasone; D-KPd, daratumumab-carfilzomib-pomalidomide-
dexamethasone; D-MRd, daratumumab-melphalan-lenalidomide-dexamethasone; D-SKd,
daratumumab-selinexor-carfilzomib-dexamethasone; D-Vd, daratumumab-bortezomib-dexamethasone;
GPRC5D, G protein -coupled receptor class C group 5 member D ; KPd, carfilzomib-pomalidomide-
dexamethasone; MPT, melphalan-prednisone-thalidomide; MR, Minimal response ; PD, progressive
disease; PR, partial response; sCR, stringent complete response ; SD, stable disease; SKCd, selinexor-
carfilzomib-cyclophosphamide-dexamethasone.
Supplementary Figure Legends
Figure S1. BCMA expression on bone marrow plasma cells by flow cytometry.
Bone marrow was collected before lymphodepletion. The expression of BCMA on myeloma cells was
detected according to EuroFlow antibody panels for plasma cell disorders. These are the representative
detection results of BCMA expression on bone marrow plasma cells from three patients. BCMA, B-cell
maturation antigen.
Multiple myeloma Flow-MRD: antibody panel.
PacO APC-CY7 APC PerCP-CY5.5 PE-CY7 FITC PE
CD45 CD38 CD138 CD19 BCMA CyIgκ CyIgλ
Procedure
Flow cytometric analysis was performed on fresh specimens and isotype control antibody was used in
each sample for normalization. Plasma cells were gated according to the expression of CD45 and CD38.
Phenotype of plasma cells was defined as low level of CD45 and intermediate -to-high level of CD38,
which was abbreviated as CD45low CD38 int/hi thereafter. Plasma cells were further analyzed for the
expression BCMA. No less than 1.0× 105 cells were acquired in analysis for each sample.
Figure S2. Comparison between baseline and peak levels of IL -6, ferritin, and CRP after CAR T-
cell infusion. Median ± interquartile range values are shown. (Peak value of serum (A) IL -6, (B)
Ferritin and (C) CRP in patients with CRS. * p<0.05, *** p<0.001, **** p<0.0001. CAR , chimeric
antigen receptor; CRP, C-reactive protein; CRS, cytokine release syndrome; IL, interleukin.
Figure S3 Percent of peripheral blood B lymphocytes after CAR T -cell infusion. Each line
represents a single patient. CAR, chimeric antigen receptor.
Figure S4 Uninvolved immunoglobulin concentration change after CAR T-cell infusion. Changes
in immunoglobulin (A) IgA, (B) IgM, (C) IgG concentration from baseline. Each line represents a
single patient. CAR, chimeric antigen receptor.
Figure S1
Figure S2
A
Baseline Peak
0
200
400
600
800
1000
IL-6(pg/ml)
✱✱✱
B
Baseline Peak
0
2000
4000
6000
8000
Ferritin(ng/ml)
✱
C
Baseline Peak
0
50
100
150
CRP(mg/L)
✱✱✱✱
Figure S3
0 10 20 30
0
2
4
6
8
Time since infusion (days)
The percentage of B-cell in total
lymphocytes of peripheral blood(%) Patient1
Patient2
Patient4
Patient6
Patient9
Patient11
Patient12
Patient13
Patient14
Patient5
Patient15
Patient16
Patient18
Figure S4
A
0 20 40 60 80
0.0
0.5
1.0
1.5
Time since infusion (days)
Immunoglobulin (IgA g/L)
Patient1
Patient2
Patient4
Patient5
Patient6
Patient7
Patient8
Patient9
Patient10
Patient12
Patient13
Patient14
Patient3
Patient15
Patient16
Patient17
B
0 20 40 60 80
0.0
0.5
1.0
1.5
2.0
Time since infusion (days)
Immunoglobulin (IgM g/L)
Patient1
Patient2
Patient4
Patient5
Patient6
Patient7
Patient8
Patient9
Patient10
Patient11
Patient12
Patient13
Patient14
Patient15
Patient16
Patient17
C
0 20 40 60 80
0
2
4
6
8
10
Time since infusion (days)
Immunoglobulin (IgG g/L)
Patient6
Patient9
Patient11
Patient12
Patient13
Patient15