Multiple Myeloma (MM) is a malignant disease with clonal abnormal plasma cell proliferation.
It is the second most common malignant tumor in hematological system tumors. The incidence
rate is 2-3/100000, and it is still incurable. In recent years, with the application of new
drug based regimens such as proteasome inhibitors, immunomodulators, and anti CD38 monoclonal
antibodies, the remission rate of MM patients has been significantly improved and
chemotherapy related toxic side effects have been reduced, increasing the survival period of
multiple myeloma from 2-3 years to more than 5 years. Sequential autologous hematopoietic
stem cell transplantation after intensified induction therapy is currently the preferred
treatment strategy for patients suitable for hematopoietic stem cell transplantation. For
relapsed refractory MM, new generation oral proteasome inhibitors, monoclonal antibodies, and
specific cellular immunotherapy have shown good therapeutic effects. However, multiple
myeloma is still an incurable disease, and more effective treatment methods need to be
explored to improve the efficacy and prolong survival time of relapsed and refractory
multiple myeloma.
In recent years, CAR-T has achieved significant therapeutic effects in hematological
malignancies. Chimeric antigen receptor T cell (CAR-T) immunotherapy is a rapidly developing
new approach to tumor adoptive immunotherapy in recent years. Its main feature is to obtain T
cells that recognize tumor antigen-specific receptors through genetic engineering
modification, and endow them with targeting, killing, and persistence. CAR-T studies with MM
antigen specificity have also been carried out and achieved good results. CAR-T cell therapy
has become an effective new treatment for MM, with B-cell mature antigen (BCMA) becoming a
clear target for CAR-T cell therapy. BCMA CAR-T cells can recognize and kill MM cells derived
from MM patients, and exert anti-tumor effects in MM model mice through the perforin pathway.
The overall effective rate of anti mouse derived BCMA CAR-T cell therapy for
recurrent/refractory MM can reach 81%, with a CR rate of 55.5% for patients and a strict
complete remission (sCR) of 13.3%. However, as the application time of CART cell therapy
increases, recurrence remains an unavoidable issue, with approximately 40% of patients
receiving CR still experiencing recurrence. Meanwhile, studies have shown that multi target
CAR-T may prolong the remission period and reduce recurrence. Expanding the coverage of MM
cell targets and clearing poorly differentiated MM cells may further improve the efficacy of
CAR-T in treating relapsed refractory MM. Studies in mouse experimental models have confirmed
that the combination of anti-CS1 CAR-T and anti-BCMA CAR-T has better therapeutic effects on
myeloma cells than using anti-BCMA CAR-T alone. Clinical trials have confirmed the use of
mouse derived anti BCMA CAR-T cells combined with humanized anti CD19 CAR-T cell infusion for
the treatment of relapsed and refractory MM, with an ORR of 95.2% and a negative MRD
conversion rate of 81.0%. However, with the extension of follow-up time, patients gradually
experience recurrence. Therefore, although CAR-T therapy has shown positive clinical results
in the treatment of multiple myeloma, recurrence remains a difficult problem to overcome, and
more treatment methods are needed to address this issue.
Although CAR-T cells have strong tumor killing ability and specific targeted binding, the
problems in CAR-T therapy cannot be ignored. Firstly, CAR-T therapy has related toxic side
effects, mainly including:
- (1) Autoimmune toxicity mainly refers to the "on target/off tumor"
toxic reaction.
CAR cells attack tumor cells while also attacking normal tissues expressing
the same antigen, causing damage to normal tissues.
- (2) Cells Factor release syndrome (CRS),
which is a non antigen-specific toxicity caused by high levels of immune activation beyond
the physiological state, is also the most prominent toxic response of CAR-T cell
immunotherapy.
CRS is often accompanied by an increase in various inflammatory cytokines,
such as IL-2, IL-6, IL-10, TNF-a, and IFN- γ Wait. The clinical manifestations of CRS vary
greatly, with fever being the most common, muscle pain, nausea, unstable hypotension, and
hypoxia frequently occurring. The overall symptoms can range from mild flu like symptoms to
severe life-threatening symptoms such as respiratory distress, multiple organ dysfunction,
and even failure.
- (3) Neurotoxicity also appeared in some subjects, which is an unexpected
toxic side effect that mainly includes symptoms such as blurred consciousness, coma, aphasia,
motor disorders, and seizures.
Secondly, the preparation cycle of CAR-T cells is long and not
suitable for patients with rapid disease progression. At present, the preparation of CAR-T
cells in the study population is mostly based on autologous peripheral blood collection and
lentivirus transfection. T cells need to undergo a series of operations such as sorting,
activation, transfection, and amplification before being reintroduced to patients, which
generally takes 2-3 weeks. On the one hand, it cannot meet the urgent treatment needs of
patients with rapid disease progression, and on the other hand, long-term in vitro
cultivation leads to excessive differentiation of CAR-T cells in vivo, resulting in reduced
survival, proliferation, and killing ability in vivo, reducing therapeutic efficacy. Finally,
the efficacy of CAR-T may be affected by early chemotherapy. At present, the clinical
application of CAR-T is mostly prepared by autologous T cells, and patients with
relapsed/refractory AML have received multiple chemotherapy treatments, resulting in impaired
T cell function, which affects the efficacy of CAR-T and patient prognosis. Therefore, it is
necessary to seek new alternative treatments.
NK cells are important immune cells in the body and an important component of innate
immunity. Under physiological conditions, NK cell inhibitory receptors recognize MHC class I
molecules widely expressed on the surface of normal tissue cells, which inhibits NK cell
function and prevents them from killing their own normal tissue cells. In tumor tissue, due
to the downregulation of MHC class I molecule expression on the surface of tumor cells,
ligands that activate receptors such as NKp30, NKp44, NKp46 are upregulated, leading to NK
cell activation and ultimately killing tumor cells. CAR-NK cells express CAR molecules on the
surface of NK cells, recognize target antigens through CAR, and further activate NK cells to
kill tumor cells. The CAR structure of CAR-NK cells usually consists of three parts, namely
the extracellular antigen binding domain, transmembrane domain, and intracellular activation
domain, which is very similar to the CAR structure used in CAR-T cell therapy. According to
the different intracellular domains, CAR structures will also be compared and validated
accordingly.
Compared to CAR-T cell therapy, CAR-NK cells have their unique advantages in adoptive cell
therapy: 1) The probability of CAR-NK cells experiencing cytokine storms is low. The
pro-inflammatory cytokines secreted by CAR-T cells, such as IL-1 and IL-6, are the main
cytokines causing CRS. Active CAR-NK cells typically produce TNF- α There is a significant
difference between the types of cytokines produced by granulocyte macrophage
colony-stimulating factor (GM-CSF) and T lymphocytes. Clinical studies have shown that CAR-NK
treatment significantly reduces the probability of cytokine storms. 2) CAR-NK cells have
multiple killing mechanisms that can kill tumor cells with low or no expression of target
antigens, reducing the recurrence of target antigen negative tumors. CAR-T cells are
difficult to recognize target cells with low or no expression of target antigens, and these
cells cannot be cleared, leading to tumor recurrence. CAR-NK cells not only recognize tumor
surface antigens through single chain antibodies to kill tumor cells, but also activate
various receptor recognition ligands to kill tumor cells. Such as natural cytotoxic receptors
(NKp46, NKp44, and NKp30), NKG2D, and DNAM-1. In addition, NK cells pass through Fc γ RIII
(CD16) induces antibody dependent cytotoxicity, and multiple mechanisms combine to kill
heterogeneous tumor cells, thereby reducing the risk of tumor recurrence. 3) Allogeneic
transplantation of CAR-NK cells does not cause graft-versus-host disease (GvHD). When CAR-T
cells in the test population are generated by healthy donors who do not match human leukocyte
antigen (HLA), Non autogenous MHCs expressed on allogeneic CAR-T cells can induce immune
rejection and cause severe hematological toxicity. NK cells are innate immune cells that do
not rely on MHCs molecular recognition. Allogeneic CAR-NK cells do not induce GvHD
production. 4) NK cells come from multiple sources to meet the needs of patients whose cell
quality and quantity are affected by multiple chemotherapy treatments. NK cells can be
obtained from human umbilical cord blood, peripheral blood, induced pluripotent stem cells,
and NK-92 cell lines. For patients who have difficulty collecting sufficient cells due to
multiple chemotherapy treatments, multiple sources of NK cells can meet the needs of CAR-NK
treatment. 5) CAR-NK cells can provide timely "spot type" treatment. NK cells are not limited
by major histocompatibility complexes, therefore, allogeneic NK cells can be used for
modification and development into off the shelf CAR-NK cell therapy. For patients with rapid
tumor progression, timely treatment can be provided to avoid disease progression due to
waiting for cell preparation.
The NKG2D receptor is an activating receptor that expresses NK cell expression. It can
recognize the NKG2D ligand (NKG2DL) expressed in tumor cells and activate NK cell killing
activity through NKG2D-NKG2DL interactions. NKG2DL is located at different positions on
chromosome 6 and belongs to two gene families, including MICA/MICB and ULBPs. The NKG2D
ligand is expressed at different levels on the surface of tumor cells or virus infected
cells, while it is not expressed on the surface of normal cells, with over 70% of human tumor
cells showing upregulation of the NKG2D ligand. Our previous research has shown that the U266
myeloma cell line and over 62% of MM patients express different levels of NKG2D ligands, and
immune cells can mediate the killing of MM cells through the interaction between NKG2D
receptors and ligands. Targeted NKG2DL CAR-NK cell therapy for MM achieved good therapeutic
effects in preclinical experiments. At the same time, 5 recurrent MM received NKG2D-CAR-NK
cells without treatment-related adverse reactions, dose limiting toxicity, and no CRS
response observed. A preclinical study has shown that NKG2D-CAR-NK has a more cytotoxic
effect on MM cells than simply expanded and activated NK cells, providing a basis for the
treatment of MM with NKG2D-CRA-NK.
Therefore, choosing NKG2DL as the target for treating AML has three main advantages: on the
one hand, its specific expression on tumor cells can ensure the safety of CAR-NK drugs and
avoid serious off target toxicity; In addition, the diversity of NKG2D ligands can to some
extent avoid tumor recurrence caused by MM cell heterogeneity or single target deficiency,
and better maintain drug efficacy; Finally, NKG2DL as a therapeutic target for MM has
obtained positive clinical data. Therefore, the investigators plan to evaluate the efficacy
and safety of combined infusion of NKG2D-CAR-NK cells in the treatment of recurrent multiple
myeloma. The clinical data of this study can provide data support for new treatment plans for
relapsed refractory MM.
