How to treat multiple myeloma by CAR-T cell therapy?

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How to treat multiple myeloma by CAR-T cell therapy?

How to treat multiple myeloma by CAR-T cell therapy?  Multiple myeloma (MM), which accounts for 10% of blood cancers, is a plasma cell malignant tumor that originates in the bone marrow.

Over the years, more understanding of disease biology has led to parallel improvements in treatment methods, as evidenced by the emergence of new therapies, such as proteasome inhibitors (bortezomib, carfilzomib and ixazomib), immunomodulation Drugs (thalidomide, lenalidomide, and pomalidomide) and monoclonal antibodies (daratumumab and elotuzumab) approved by the U.S. Food and Drug Administration (FDA) [1,2,3,4].

Although these new therapies have led to better disease control, myeloma is still largely incurable, and high-risk patients cannot benefit from these therapies [4,5,6,7]. Drug resistance is inevitable, and disease recurrence is still a huge clinical challenge.

Chimeric antigen receptor (CAR) T cell therapy is one of the rapidly emerging and promising immunotherapy options, and it has shown unprecedented results in B cell malignancies [10-13]. It prolongs the patient’s survival and remission period, even for some patients who have failed standard treatment [14].

This review details the overall development of CAR T cell therapy and its role in the paradigm shift in the treatment of myeloma. The topics we discuss range from the current state of its clinical development to the latest technologies used in the manufacturing of myeloma CAR. Here, we also considered the future prospects and proposed ways to improve CART cell research, because we are working to completely change the CAR T scenario in myeloma.

1. Overview of CAR T cell therapy

T lymphocytes are bioengineered to express monoclonal antibodies (moAb) that recognize tumor-associated antigens (Figure 2). The binding of antibody-bound T cells to these homologous antigens will initiate a signal cascade in T cells, stimulating the release of pro-inflammatory cytokines (such as TNF-α, IFN-γ, IL2 and IL6), leading to cell lysis [20 ]. This unique CAR T cell characteristic can help alleviate the common limitations of T cell receptor (TCR) inducing immunity, including the loss of the major histocompatibility complex (MHC) of tumor cells and the low antigen binding affinity of T cells [21 ,22,23].

Figure 3 depicts the basic components of CAR. The single-chain variable fragment (ScFV) of the monoclonal antibody on the extracellular domain has an antigen recognition function and is connected to the intracellular domain through a hinge/transmembrane region, usually derived from CD8 or IgG4. The first generation of CAR T cells contained only the CD3ζ signal domain and lacked proliferation characteristics17. Current and routinely produced CAR T products contain one (second generation) or two (third generation) costimulatory domains (4-1BB, CD28, and/or OX-40) to promote effective CAR T cell signal transduction and lasting Sex and effectiveness [24,25,26,27]. The more complex fourth-generation CAR, T cell redirection for universal cytokine-mediated killing (TRUCK), consists of an additional transgene encoding a pro-inflammatory cytokine, which is released to mediate when induced by a signal molecule Lead to cytotoxicity [21,28,29].

CAR T cell manufacturing is a laborious process that takes 2-4 weeks to complete in a good manufacturing practice (GMP) environment (Figure 4). CAR T cell therapy is mainly autologous, and the manufacturing process starts with the collection of peripheral blood mononuclear cells (PBMC) from the patient through leukocyte removal. Before genetic modification with the CAR of interest, the T cell population is enriched. Gene transfer is carried out by viral transduction (γ-retrovirus or lentivirus) or non-viral transfer technology (DNA transposon or mRNA transfection) [30]. Subsequent immunophenotyping analysis is performed to ensure that T cells successfully obtain substances with CAR and cytolytic activity. These CAR T cells will then be expanded in vitro in a bioreactor containing a growth factor-rich medium, and finally frozen for storage or immediately transported to the clinic for infusion.

2. CAR-T therapy for MM treatment

The most widely studied CAR target for myeloma is the B cell maturation antigen (BCMA), a member of the tumor necrosis factor receptor superfamily 17 (TNFRSF17). It is considered an ideal antigen target because it is preferentially expressed on plasma cells rather than hematopoietic stem cells [31]. The combination and stimulation of ligand, B cell activating factor (BAFF) and proliferation-inducing ligand (APRIL) with BCMA can promote the growth and proliferation of plasma cells in bone marrow. Although the expression of BCMA is heterogeneous [32, 33], it is universally present in all MM cells [31, 34] and its overexpression has important prognostic value [35, 36].

The first BCMA-guided CAR was developed less than ten years ago, showing preclinical evidence of functional targeting [31]. Subsequently, the first human phase I clinical trial was conducted to test the efficacy of BCMA targeting CAR T cells in relapsed/refractory multiple myeloma (RRMM) (NCT02215967). The overall response rate (ORR) was 81%, of which 63% showed very good partial response (VGPR) or complete response (CR) [37,38]. Therefore, extensive efforts have been made to develop new anti-BCMA CARs and fine-tune old CARs.

Table 2 records the records of anti-BCMA CAR T cell clinical trials and the latest updates from ASH 2020 [38-40]. In 2017, bb2121 (BluebirdBio) won the FDA’s Breakthrough Therapy Designation Award and the European Medicines Agency’s (EMA) Priority Medicine (PRIME) Qualification Award. This is the first major breakthrough in the development of myeloma CAR T cells . This is due to its superior efficacy and long-lasting response. The latest reported ORR was 73%, and PFS in RRMM patients improved significantly (median PFS 8.8 months) [41,42]. Bb2121 is hailed as the latest leader in FDA-approved MM treatment. Its successor, bb21217, uses the same CAR construct design, but is cultured in the presence of an isolated PI3K inhibitor bb007, and its ORR is 55%, which is related to the enrichment and high peak expansion of memory T cell phenotypes [43] . In addition, JNJ-4528 is a bi-epitope CAR T cell product containing two BCMA targeting domains, designed to enhance antigen binding affinity, and also showed an impressive 94.8% ORR in the CARTITUDE-1 test[ 44,45]. This study provides convincing evidence that a single low-dose infusion of CAR T product is sufficient to induce an early, deep, and long-lasting response. JNJ-4528 uses the exact structure of LCAR-B38M, which is being evaluated in another geographic cohort (China) in the LEGEND-2 trial. The study reported a promising ORR of 82.4%, with good safety in a subgroup of high-risk patients [46].

Some other notable BCMA-CAR T products are humanized and fully humanized CARs (JCARH125, MCARH171, and FCARH143), which are designed to reduce graft-versus-host disease (GVHD) and extend the persistence of T cells. The ORRs for these treatments were significant 64%, 100%, and 82%, respectively. According to reports, especially the efficacy of JCARH125 is related to T cell products with high CD3+ purity and rich early memory phenotype and versatility.
Table 2 describes more ongoing and recruited BMCA CAR T trials, with different products showing different levels of efficacy. A recent meta-analysis study of a total of 23 different BCMA-CAR T cell products has been used in a total of 640 patients, reporting an average ORR of 80.5%, a CR of 44.8%, and a median PFS49 of 12.2 months.

3. Limitations of BCMA-CAR-T therapy

The promising results of anti-BCMA CAR T cell therapy are not without its own set of challenges.

Treatment resistance

The underlying mechanism of treatment resistance remains largely unclear, but involves tumor heterogeneity and antigen escape [50]. The heterogeneous expression of BMCA at the intratumoral level can lead to preferential targeting of cells with high BCMA, while retaining those with low/zero BCMA expression, leading to the growth of the latter clones [33,34,47]. In fact, BCMA often loses expression when the disease relapses after the first CAR T infusion, which indicates that CAR T cells have selected BCMA-negative MM clones [38,51,52,53].

For BCMA antigen escape, one of the most detailed methods is the error physiology of BCMA antigen. BCMA can inadvertently transfer from tumors to T cells in a process called phagocytosis, causing T cell fat death [3,50] or it can flow into the blood circulation (now called serum BCMA (sBCMA)) and be secreted by γ- Enzyme mediation 47. Both can lead to inhibition of tumor cell recognition [38,54]. Although lower sBCMA levels are indeed associated with good ORR [36,38,55,56], it is not always associated with CAR T dose response, and its expression is still low at relapses with increased disease burden [52 ], indicating that the drug resistance mechanism can be extended in addition to the internal factors of the tumor.

Related to this, a recent study using longitudinal single-cell transcriptomics and extensive genomic analysis emphasized molecular BCMA aberrations. They found that the patient’s resistance to bb2121 was related to biallelic loss of BCMA. Most MM cells show chr16p heterozygous loss, including the BCMA locus, accompanied by nonsense mutations of other alleles, resulting in biallelic inactivation of BCMA [52].
This was confirmed by another study, which revealed that genome instability is related to biallelic deletion of the BCMA locus after CAR T cell infusion. Both alleles were intact before CAR T treatment. Therefore, this indicates that branching evolution occurred during the treatment and that MM cells acquired genomic abnormalities when selecting BMCA-negative cells.

Toxicity

After CAR T cells are activated, cytokine release syndrome (CRS) and neurotoxicity mediated by pro-inflammatory cytokines are another long-standing problem. CRS manifests as fever, nausea, and flu in mild cases, but may escalate to hypotension, cardiac arrest, and liver failure in severe cases. On the other hand, neurotoxicity may range from mild confusion and delirium to severe retardation, seizures, and white matter degeneration [20]. Although it is a plasma cell marker, BCMA is also co-expressed on normal B lymphocytes. Therefore, BCMA CAR T cell therapy may also introduce targeted/non-tumor effects. Common manifestations include B cell hypoplasia and neutropenia Symptoms and immunosuppression that lead to an increased risk of infection.

In the same meta-analysis study described above, the average incidence of CRS and neurotoxicity reported were 80.3% and 10.5%, respectively [49]. Although toxicity is a common event, the patient’s toxicity-related fatality rate is controllable. High levels of toxicity are more commonly observed in patients with heavier tumor burden or higher CAR T cell doses [38]. Off-the-shelf drugs are usually used to control toxicity, such as IL6R antagonists (tocilizumab) for the treatment of CRS and corticosteroids for the treatment of neurological symptoms [20].

4.  Non-BCMA CAR T cells

People always look forward to the emergence of the next breakthrough molecule. Various surface antigens have been explored in this pursuit. This section documents the progress of some of the more well-studied myeloma antigens (Table 3).

SLAMF7/CS1

The attractiveness of using signaling lymphocyte activator molecule F7 (SLAMF7) for CAR targeting stems from elotuzumab, which is the first humanized SLAMF7 monoclonal antibody approved by the FDA for use in RRMM [57,58]. Anti-SLAMF7 CAR T cells derived from elotuzumab showed preclinical evidence of rapid cytolysis in primary myeloma samples and elimination of extramedullary cells in xenografts [59]. SLAMF7-CAR T cells incorporate an on/off suicide gene to improve safety and tolerability, and were later developed for clinical research (NCT03958656 and NCT03710421).  A European team also designed a new SLAMF7 CAR T cell model using the non-viral gene transfer method of the Sleeping Beauty transposon system. Their proprietary product is currently being tested in the Phase I/II CARAMBA trial (https://www.caramba-cart.eu/patients/)[60].

Marking a milestone, the allogeneic anti-SLAMF7-CAR T cell (UCARTCS1) became the first “off-the-shelf” CART cell product (MELANI-01) in MM to receive FDA clinical trial approval. UCARTCS1 is manufactured using healthy allogeneic T cells and TALEN gene editing technology to eliminate endogenous TCR and SLAMF7 expression, aiming to reduce the risk of GVHD and T cell cannibalism [61].

Importantly, a new anti-SLAMF7/BCMA bispecific CAR T cell product is under pre-clinical development to increase tumor coverage and overcome antigen loss. Compared with T cells expressing a single CAR, these CAR T cells with single-chain antibodies contain two tandem ligand binding domains (one for SLAMF7 and the other for BCMA), showing enhanced activity. Combined use with PD1 checkpoint inhibitors can accelerate tumor clearance in the body [62].

CD38

The clinical success of anti-CD38 moAb (daratumumab and second-generation isatuximab) in MM[63-65] is the basis for the development of anti-CD38 CAR T cells. Preclinical evidence that anti-CD38 CAR T cell therapy is effective against myeloma [66,67] has led to the initiation of various clinical trials, one of which used anti-CD38 CAR T cells as a monotherapy for RRMM (NCT03464916), while others People explored its binding to different CAR T cells, including anti-BCMA, anti-CD19, anti-CD138, anti-CD56 and anti-NY-ESO-13. A bispecific anti-CD38/BCMA CAR T cell product (NCT03767751) is being evaluated.
However, it is worth noting that CD38 is usually not highly expressed on myeloma cells, and its expression may be down-regulated in advanced diseases [67,68]. Therefore, anti-CD38-CAR-T may develop resistance. Since CD38 is also expressed on activated T cells (thus increasing the risk of T cell cannibalism), NK cells and normal prostate, neuron and muscle cells, it may also appear in target/non-tumor toxicity.

CD19

Although CD19 is not usually present in mature plasma cells, a small number of myeloma cells with unique reproductive characteristics express low CD19, which is related to drug resistance and the characteristics of promoting recurrence [3,54], making it a reasonable therapeutic target . A proof-of-concept pilot study was conducted using Kymriah, an anti-CD19 CAR T cell. Of the 12 patients treated, 6 received VGPR, 2 received PR, and the other 2 experienced disease progression (NCT02135406) ​​[13]. Importantly, the anti-CD19/anti-BCMA cocktail CAR T product can produce an impressive 100% ORR (NCT03455972). Although the cells expanded more than 1000-fold at the peak data point, the patient experienced only mild (grade 1-2) CRS with no neurotoxicity.

TACI

Transmembrane activators and CAML interactors (TACI), such as BCMA, are members of the TNF superfamily. TACI and BCMA share the same activation ligands APRIL and BAFF, when they bind to their receptors, they can promote the growth and survival of MM. Based on this biological characteristic, ligand-based CAR T cells for APRIL have been developed to simultaneously target the TACI and BCMA signaling pathways [69,70]. In an in vivo model, APRIL-targeted CAR T cells eradicated BCMA+TACI- and BCMA-TACI+ tumors, but the monospecific anti-BCMA CAR T cells failed to inhibit the proliferation of BCMA- cells, indicating that the potential application of APRIL-targeting is related to BCMA loss CAR T cells in the following cases [69]. Recently, another CAR T cell product targeting APRIL with fine molecular design has been reported [70]. Compared with its monomeric counterpart, this APRIL-based trimer CAR T cell has enhanced binding to BCMA and TACI receptors and has a higher cytolytic activity. Despite these promising preclinical data, so far the only clinical trial targeting CAR T cells targeting APRIL was terminated due to insufficient efficacy (NCT03287804).

Only one reported CAR T cell product is under development, specifically targeting the TACI surface antigen. Bispecific TACI/BCMA CAR T cells have effective in vitro and in vivo cytotoxicity to MM cells [71].
In addition to myeloma cells, TACI has also been found on immunosuppressive regulatory T cells (Tregs). Therefore, targeting TACI can not only cause cytotoxicity through direct cytolysis, but also produce cytotoxicity through indirect manipulation of the unfavorable microenvironment imposed by Tregs [34,72], thus providing a good two-pronged approach. method.

CD138

CD138 is a highly expressed surface molecule on MM cells and is clinically used as a selection marker for plasma cells. The Chinese People’s Liberation Army Hospital tested a CAR T cell product targeting 4-1BB/CD3 CD138 as a monotherapy for MM. However, only moderate treatment response (NCT01886976) was reported. The data was obtained from a small sample size (n = 5), so the results may not be sufficient for a meaningful interpretation. Anti-CD138 CART cells have also been included in other CAR T cell mixtures (BCMA, CD38, CD19, SLAMF7) trials, but no substantial data can be obtained (NCT03778346 and NCT03196414).

Although CD138 is overexpressed in myeloma cells, it should be noted that CD138 is present in other normal tissues, such as epithelial cells, endothelial cells, and vascular smooth muscle cells, again emphasizing the possibility of targeting/non-tumor effects. In past clinical trials, clinical trials of anti-CD138 antibody-drug conjugates for MM have clearly seen this shortcoming, and some patients have reported severe mucosal and skin toxicity [13].

Others: 

In the in-depth study of CAR T cell therapy, a variety of other target antigens were also examined, some of which are worth mentioning are the G protein-coupled receptor C class 5 group D members (GPRC5D), CD56, Lewis Y, CD44v6, And Kappa light chain. These molecules are studied because of their surface expression and prognostic influence [66]. Most of the comprehensive data from these CAR T cell product trials cannot be used for evaluation, but for some products that have published trial data, there is not much excitement because patients report moderate to low responsiveness [54]. Various other molecules involved in plasma cell biology, such as CD229, integrin β7, CD70, and CD126 (IL6R) are under preclinical research [3,73].

5.  Technical progress of CAR T cell manufacturing in myeloma

With the continuous development of CAR T cell therapy, the multi-modal engineering of effector cells provides prospects for coping with increasingly complex disease environments. In order to solve the problem of antigen escape when the disease recurs, CARs targeting the BCMA bi-epitope have been explored.

LCAR-B38BM and JNJ-4528 are used to express two BCMA binding domains to increase their binding affinity to myeloma cells with low BCMA expression. However, the caveat to this approach is that it is still unclear whether a certain threshold of BCMA expression is required for optimal tumor recognition and cytotoxicity.

A study by the University of Pennsylvania (NCT02546167) showed no correlation between baseline BCMA expression and response rate [74]; on the contrary, the FCARH143 trial (NCT03338972) found that there was a difference in BCMA expression on tumor cells between long-term responders and patients who had relapsed before treatment. In many BCMA clinical trials conducted to date, whether baseline BCMA is or is not the inclusion criteria, if it is, the range includes only observable expression (~1%) up to >50% (Table 2). This expected heterogeneity between studies makes it difficult to assess potential factors that contribute to safety and effectiveness. A single clinical trial with a series of different baseline levels may be able to inform the antigen threshold required for the desired anti-myeloma effect, and whether a lower baseline BCMA will lead to faster drug resistance/relapse.

In terms of clonal selection to overcome single antigen loss, dual/multiple antigen targeting has been achieved by: (a) infusing patients with multiple CAR T cell products in cocktail therapy, or (b) having multiple CAR T cell products in one T cell Double ScFV for different antigens. Some known antigens co-expressed with BCMA CAR are SLAMF7, CD19 and CD38 (Table 2). Considering that these targets remain stably expressed during progression in a BCMA-negative environment, this strategy is relevant [51]. However, it is important to consider the challenges that follow. For example, in (b), it may be technically challenging to control the positive ratio of each CAR in T cells. The difficulty in defining the expression level of any of these CARs on a single T cell will result in a highly heterogeneous CAR T cell pool, which may affect its tumor selectivity and ultimate efficacy. In addition, like single-target CARs, dual-target/multi-target CARs can also cause selective pressure. Due to the enhanced immune response, the selective pressure may be stronger, causing both antigens to escape at the same time. In addition to technical complexity, another obvious challenge is the risk of high-grade CRS, accompanied by concurrent antigen targeting to enhance T cell activation [75].

In order to reduce the toxicity and targeting/non-tumor specificity caused by over-activated CAR T cells, some CARs have been modified to contain suppressor/suicide genes, which can be triggered to turn off T cell signaling when needed to avoid subsequent toxicity . Suicide genes in myeloma are currently being studied, such as truncated EGFR (NCT03070327 and NCT03093168), iCasp9 (NCT03958656 and NCT03125577) and HSV-TK (NCT04097301). Other safety strategies, such as bispecific T cell adapters and tandem CAR have not been explored in myeloma, so it is worth studying [20].

So far, the general CAR model consists of CD28 or 4-1BB as the costimulatory domain. Choosing between these two CAR molecules is crucial because it determines the speed of immune activation and the persistence of CAR T cells. CD28-containing cells have been shown to have higher potency, rapid expansion and the ability to cause a rapid increase in cytokines after antigen stimulation, while CARs containing 4-1BB show lower signal strength, gradual activation and similar memory stem cells The phenotype of [76]. Although the former is related to rapid and powerful cell lysis activity, its active signal dynamics is related to faster T cell depletion, thus resulting in a less durable response. Compared with their 4-1BB counterparts, they lack continuous persistence, which makes them possible to be a poor choice of CAR structure [13,66]. MM’s low preference for CD28-CAR is obvious (Table 2). Therefore, after the first human-derived BCMA-CART test (NCI) of CD28, subsequent CAR T products mainly carry 4-1BB.

The short-term clinical remission of MM is attributed to the low persistence of BCMA CAR-T cells. To avoid this, bb21217 derived from its precursor bb2121 was cultured in vitro in the presence of the PI3K inhibitor bb007 to enrich T cells with a memory-like phenotype. When bb21217 instead of bb2121 induced tumor clearance during the second tumor challenge in the same mouse, the benefits of this approach were demonstrated without the need to re-administer the product [78]. Compared with bb2121, Bbb21217 showed higher levels of CCR7 and CD27, indicating higher levels of memory-like T cells, and lower levels of CD57 (a marker of T cell failure). The latest update of bb21217 shows that the ORR ratio is 55%, and CAR T cells can be detected within 18 months [39,43].

In addition, the contribution of immunosuppressive factors in the tumor microenvironment to the lack of durability of CAR-T cells is also worth noting. Recently, a new all-human BCMA-CAR has been reported, which has a dominant negative domain for immunosuppressive TGF-β, exhibits strong proliferation and persistence, and is functionally immune to hostility rich in TGF-β The influence of the microenvironment [79].

Another manufacturing optimization that has also attempted to amplify the effectiveness of CAR T cells in myeloma is T cell composition. The higher CD4:CD8 T cell ratio used in the pre-transduction and expansion stages is associated with greater in vivo BCMA-targeted CAR T cell expansion and response [74], confirming CD19-targeted CAR T cell therapy in lymphoma The report and ALL[80,81]. JCARH125 is injected into RRMM patients (EVOLVE) at the specified CD4:CD8 ratio of 1:1. The latest clinical data report a strong expansion of these products at all dose levels, and the patient’s ORR (82%) is closely related to compound T cells [82].

The gene delivery method is another variable in the production of MM CAR T, and virus transduction is most routinely selected. Given the risk of randomly inserting mutations in this system, and it is often associated with laborious and expensive manufacturing procedures, other non-viral methods of introducing CAR into T cells have been introduced. Poseida Therapeutics developed the first non-viral CAR T cell product for myeloma using the piggyback transposon technology (P-BCMA-101). Compared with those delivered by viruses, this mRNA electroporation method shows >95% efficiency and more stable and higher CAR expression, plus the prolonged activity of T cell expansion with memory stem cell phenotype [74,83 ,84]. In addition, another non-viral system, Sleeping Beauty, was used to introduce SLAMF7-CAR into effector cells and showed good integration [60].

Another CAR T manufacturing consideration is the source of moAb. One of the limitations of other successful CD19-targeted CAR T cell therapy in lymphoma is that T cells can kill each other due to the host immune anti-mouse CAR response [80]. Therefore, it is a pragmatic method to integrate humanized ScFV with low immunogenicity into T cells. In fact, the fully humanized CAR T cells targeting BCMA developed by MSKCC (FCARH143 and JCARH125) have shown rapid expansion, eradicating a large number of tumor burdens and a longer-lasting patient response [66,78]. A novel, completely human, heavy-chain anti-BCMA binding domain (FHVH33) instead of the complete ScFV is designed to avoid recognition by host immune cells, can induce deep and persistent responses, and has controllable toxicity [ 53].

6.  Expert Comments:

The search for new targetable myeloma antigens is critical; however, those that have been reviewed so far do not seem to be able to realize the potential of BCMA. As a single agent, non-BCMA CAR T cells lack meaningful results. Improvements to these CARS, such as making them bispecific, usually require BCMA as the accompanying target antigen. As bb2121 is about to receive FDA approval, research to clarify the potential resistance mechanisms and evasion methods is essential.

Synthetic biology is essential for designing more effective CAR T cells that target BCMA. However, maintaining tumor-associated BCMA expression and preventing antigen loss are necessary for their effectiveness. In this case, pharmacological methods have been explored. Gamma-secretase inhibitor (GSI) treatment can increase the density of BCMA on the tumor surface, while reducing the concentration of sBCMA and increasing the anti-tumor activity of CAR T cells [85,86]. It is currently studying oral GSI and CAR T cells in a phase I trial (NCT03502577). Among the 10 evaluable patients, 30% and 50% achieved sCR/CR and VGPR, respectively [85,87].
Recently, there have been new attempts to take advantage of the success of anti-CD19 CAR T therapy.

A new CD19 fusion protein model was developed to reactivate the targeting of recurrent NHL and HER2-positive solid tumors with low CD19 expression [90]. In the case of the successful production of artificial CD19 expression on the surface of these CD19 low/negative tumor cells, effective CD19-specific CAR T cell-mediated cytotoxicity was observed. MM can also consider this innovative strategy (Figure 5). In this case, the decoy CD19 antigen can be designed on the surface of myeloma cells by constructing a CD19-BCMA (or any other specific MM antigen) antibody complex, and the BCMA arm is connected to the tumor antigen. Overexpression of synthetic CD19 on MM will allow us to target CAR T cells (Kymriah or Yescarta) with commercially available CD19 that has been proven to be highly specific and potent.

The current standard of care has proven that the combination of multiple drugs for different mechanisms is a more effective method of disease management. However, research on the combination of BCMA CAR T therapy and multiple anti-myeloma drugs is quite limited. Preclinical studies combining CAR T cells with lenalidomide found that lenalidomide can enhance the durability and activity of the former [91,92]. Another study reported that the inhibition of BCMA by the new BCMA-targeted ADC (MEDI2228) can act synergistically with bortezomib in drug-resistant MM cells [93]. Various signal inhibitors such as BRD4i and MEKi have been shown to enhance the function of CAR T cells by blocking their immunosuppressive effects, but these have never been studied in myeloma 94,95. In a drug combination study, people can investigate whether a drug is added to CAR T cell infusion or for maintenance after CAR T treatment, which may be useful for patients with a huge burden of MM.

To date, the impressive results of CAR T cell therapy trials in advanced and heavily pretreated MM patients have raised the question of whether CAR T cells should be used as early therapy. The lower disease burden of newly diagnosed or early disease may represent a higher chance of achieving tumor clearance. In these cases, CARs with CD28 rather than 4-1BB costimulatory domains may be a better choice because they trigger a faster immune response. The rapid activity of CAR T cells equipped with CD28 will help to produce therapeutic effects quickly, thereby better controlling the disease at an early stage and preventing progression. For high-risk patients (such as heavy blows or functional high-risk myeloma), even with modern treatment strategies, the disease usually progresses rapidly, which seems more critical. The KarMMa-4 trial (NCT04196491)96 has begun an attempt to study bb2121 in high-risk newly diagnosed patients.

In addition, myeloma has long been associated with defective T cells in number and function, especially in progressive, refractory diseases. Obtaining T cells from healthy donors is an attractive solution to the problem of insufficient and inferior CAR T cells. Some known allogeneic compounds in MM have been approved by the FDA, namely UCARTCS1 (SLAMF7), CTX120 (BCMA), ALLO-715 (BCMA), CYAD-211 (BCMA) and PBCAR269A (BCMA) [97, 98]. These products use different technical concepts, including non-gene editing methods to eliminate TCR (CYAD-211) and single-step platforms to knock out CAR while knocking out TCR (PBCAR269A) [99]. Another BCMA allogeneic compound under clinical evaluation (ALLO-605) incorporates additional chimeric features to enhance cytokine signaling [100].

In addition to solving T cell quality issues, the ready availability of these products also means that we can bypass the long and complex self-manufacturing process, thereby achieving faster first-line treatment. The continued availability of these allogeneic CAR T cells will help eradicate residual malignant cells without the need for multiple rounds of painstaking autologous T cell collection from patients. In short, a cell bank with universal CAR T cells for allogeneic cell transfer will provide greater flexibility in applications and deliver faster to patients with lower cost-intensive manufacturing.

Recently, CAR NK has gradually attracted attention in myeloma because of its better safety, versatility of killing mechanism, and lower risk of GVHD, thereby improving the feasibility of allogeneic manufacturing, which translates into cost-effectiveness. [101]. Several preclinical studies have demonstrated that the use of CAR NK cells has cytotoxic activity and MM growth inhibitory effects on various targets, including CS1, CD138, BCMA and NKG2D ligand 102. There is currently an ongoing BCMA-CAR NK Phase 1/2 study (NCT03940833) in the RRMM. In addition, using the stem cell reprogramming system, an innovative CAR NK model was generated from induced pluripotent stem cells (iPSC), which contains a recombinant IL-15 signaling complex. Good synergy with moAb drugs (daratumumab, elotuzumab and anti-CD19) has shown therapeutic efficacy 103. With the reports on the safety characteristics of CAR NK cells relative to CAR T cells, we envision that CAR NK therapy research in MM will gain more and more interest, which will allow the diversification of MM cell therapy product portfolios, despite its potential The disadvantage is also that the weighing is cautious.

In addition to bioengineering and various clinical tests, basic mechanism research is also important because it provides insights into the basics of the immune response. Understanding the basic biological characteristics of T cells will enable us to produce “fitter” CAR T cells with enhanced proliferation and longer durability. With the advancement of CRISPR-Cas9 technology, a genome-wide screening process can be performed to identify new genes. For example, when manipulated, the flexibility and capacity of CAR T cells can be enhanced. Epigenetic mechanisms such as chromatin accessibility, DNA methylation status, RNA landscape in T cells, immune analysis, and identification of immune checkpoint features are some of the key areas for further research. Recoding T cell behavior through gene knock-in/knock-out methods will help solve the problems of innate T cell defects and inefficient effector cell pools. In order to understand the disease recurrence in the highly heterogeneous MM cell bank, the emergence of single-cell sequencing technology may help us have a high-level understanding of the genetic and epigenetic temporal evolution of immune cells. Recent single-cell studies in patients with plasma cell leukemia have revealed different CAR T cell subpopulations with different expression, proliferation, and cytotoxicity, indicating stage-specific changes along the developmental trajectory [104]. Considering this and the heterogeneity of MM, it will be interesting to observe how different genomics and transcriptomics (such as different TC classes and RNA editing characteristics) determine the expression of surface targets that can be used for immune targeting.

In addition to basic biology, in vitro and scaling processes also play an important role in CAR T cell production [37]. The logistical factors involved in the lapse of time between apheresis and the return of manufactured CAR T cells to the patient are critical, as the current 2-4 week turnaround time makes CAR T cell therapy unsuitable for patients with rapidly progressing diseases. An emerging technology to solve this lengthy manufacturing problem is GraCell Bio’s proprietary FasT CAR platform, which can achieve product release within 22–36 hours, using accelerated protocols to activate and transduce T cells without further in vitro expansion . Using this platform, a dual BCMA-CD19 CAR T treatment was developed, and the first study in humans showed that the effective ORR105 was 93.8%. Such a discovery should be a precursor to further identifying key bottlenecks in the entire CAR T cell manufacturing pipeline (Figure 4) and optimizing steps to reduce costs and cumbersome processes.

Since the first BCMA-targeted CAR was developed in 2013, MM’s CAR T cell field has indeed made considerable progress. Personalize rather than use the patient’s own immune system to destroy tumor treatment. BCMA has been recognized by the “Holy Grail” in the field of myeloma CAR T cells, which is comparable to the success of CD19-targeted CAR in lymphoma and ALL. CAR T cells are considered a “living drug” because as long as there is a targeted tumor antigen, they will be induced to make the therapeutic effect sustainable. If its effect is long-lasting, we are likely to be giving patients lasting relief. In view of this, there is every reason to be hopeful and optimistic that CAR T cell therapy will one day make MM a chronic but highly controllable and curable disease.

(source:internet, reference only)

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