How to assess clinical response for multiple myeloma

In this interview, Cerba Research explains how its teams were able to assess clinical response in multiple myeloma (MM) patients who were receiving monoclonal antibody therapy.

Can you give our readers a brief overview of multiple myeloma (MM)?

in multiple myeloma (mm)a type of bone marrow cancer, malignant plasma cells secrete high levels of monoclonal immunoglobulin protein (M-protein) that can be detected by blood protein electrophoresis (SPEP) or immunofluorescence electrophoresis (IFE).

International Myeloma Working Group (IMWG) criteria state that serum samples should present negative results for M protein by SPEP/IFE to determine complete response (CR) or strict CR (sCR).

What role do monoclonal antibodies play, and why are they important?

Monoclonal antibodies have proven to be therapeutic effectiveness in a number of malignancies, but they can hamper how IFE data are interpreted. For example, Daratumumab is a CD38 IgG1κ monoclonal antibody (mAb) in clinical development for the treatment of MM.⁵ that have shown clinical responses that increase in severity over time. This entails assessment of CR/sCR by SPEP/IFE.

Approximately half of patients with MM produce IgG1κ M protein. In a subset of patients, daratumumab or the daratumumab-anti-idiotype complex may co-migrate with endogenous M protein. Steady-state concentrations of daratumumab (at a dose of 16 mg/kg weekly, biweekly, then monthly) are readily detectable in the majority of SPEP and IFE assays.

What are the main goals when evaluating clinical response in MM?

The primary goal is to validate and implement a daratumumab interference reflex test (DIRA) that differentiates the M protein from daratumumab, as assessed by IFE, to determine if additional testing for CR/sCR evaluation (ie, bone marrow examination) is warranted. To do this, we obtain human serum samples from MM patients (n = 51) from a commercial source or patients treated with daratumumab in clinical trials (n = 33).

How are clinical samples evaluated?

Serpa conducts a series of serum IFE assays using Maxikit Hydragel 9IF Kits (Sebia Electrophoresis, Norcross, GA) according to the manufacturer’s specifications.

Antisera against immunoglobulin gamma (IgG), alpha and mu heavy chains and free and bound kappa ( ) and lambda light chains are used to identify the monoclonal protein present in each sample.

We then incubate the serum samples of patients treated with daratumumab and baseline with or without a placebo antibody mAb (mouse anti-huMaxCD38; clone 5-3-9-4) at room temperature for 15 min before analysis with IFE using IgG and Igκ sera. .

To demonstrate that the anti-idiotype antibody binds to and alters daratumumab without having an effect on detection and migration of endogenous M protein, commercially available serum samples from MM patients (n = 51) were raised with daratumumab, anti-idiotype, or daratumumab + anti- idiotype (500 and 1000 μg/ml; ratio 1:1) IgG and Igκ), and then analyzed by IFE to assess changes in M ​​protein translocation.

What other key parts of the analysis must be identified in order to judge whether the evaluation is effective?

There are many factors that we must take into account. The lower limit of detection (LOD) is determined by evaluating daratumumab ± anti-idiotype on a clinically relevant dynamic scale to determine the lowest concentration detected by parameter ≥1 (daratumumab igg, daratumumab + anti-idiotype IgG complex, daratumumab Igκ, daratumumab + anti- Igκ anti-idiotype; daratumumab or daratumumab + anti-idiotype by SPEP).

Then we guarantee the possibility of reproduction. We performed three independent runs of 10 samples from daratumumab-treated patients who achieved a PR or better and an M protein ≤5 g/dL. These runs were performed with DIRA, and the results (DIRA positive or DIRA negative) were then evaluated for reproducibility. Finally, two independent reviewers check and interpret all results to check for compatibility.

Once you have completed the DIRA assessment, what do you expect to find in the results?

In one specific case, the DIRA template used daratumumab-anti-idiotype as transmission controls for therapeutic antibody and daratumumab-anti-idiotype-transfected pairs.

Post-treatment serotype and baseline antibody profile were then compared to determine whether M protein remained after daratumumab change. DIRA-positive results showed, in this case, M protein, while DIRA-negative results showed only shift in daratumumab but no residual M protein (lanes 8, 12; Figure 4).

We also determined the lower limit of detection (LOD) in MM serum samples, using an IFE of 100 μg/mL in 9 of 10 samples with a factor ≥1 and 200 μg/mL in 10 of 10 samples. In the same samples analyzed by SPEP, either daratumumab and daratumumab plus the anti-autologous complex could be determined at 100 μg/mL in 3 of 10 samples and at 200 μg/mL in 10 of 10 samples.

In 10 of 10 (100%) daratumumab-treated patient samples, results were consistent across all three independent runs, demonstrating a sufficient level of DIRA reproducibility and concordance.

What makes DIRA a good method for assessing clinical response in multiple myeloma?

DIRA enables Determine clinical responses. In one study, a total of 33 daratumumab-treated patient samples from several different studies were evaluated for clinical response with DIRA; Of the 13 patients (39%) who were DiRA patients, 10 were confirmed to have achieved CR based on bone marrow and FLC while 20 (61%) were DIRA positive, meaning they will continue to be monitored.

As mentioned, DIRA is a specific and reproducible method that can confirm the interference of daratumumab in serum IFE at 100 to 200 μg/mL. Furthermore, DIRA-negative status signals that require additional testing to validate CR/sCR and IMWG response criteria may require modification as mAbs receive approval for MM treatment.

Figure 1. MM cells secrete high levels of M protein that can be detected by SPEP. Image credit: Cerba Research

Figure 2. Therapeutic antibodies may interfere with the ability to confirm clinical outcomes more deeply than very good partial responses. Image credit: Cerba Research

Figure 3. Schematic view of DIRA-positive and DIRA-negative treated patient samples. Image credit: Cerba Research

Figure 4. Example of DIRA-positive and DIRA-negative treated patient samples. Image credit: Cerba Research

Figure 5. Privacy Anti-Stupid Pattern MAb. Image credit: Cerba Research

Figure 6. Reproducibility of DIRA results between independent experiments. Image credit: Cerba Research

Table 1. The reviewers’ ratings agreed between trials. Source: Cerba Research

Dara, daratumab; Anti-identity, anti-identity. yes, n, no; VgPR, very good partial response.

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