1. Paraproteins and response assessments
– what have we learnt from MRC trials?
Professor Mark Drayson, University of Birmingham 19th March 2015

2. Paraprotein type at presentation: 2,789 Myeloma 11th trial patients.
Number patients
Percent total
Kappa : lambda ratio
IgG
1,665 60
2.25
IgA
699 25
1.78
Light Chain Only
361 13
1.34
IgD 35
1.3
0.46
Non Secretor 23
0.8
n/a
IgM 6
0.2
2.00
Light Chain Only (6% urine fix negative)
serum FLC dif >100mg/l
(>500mg/l in 85%)
Non Secretor
Immunofixation negative and
serum FLC dif <100mg/l
(15/23 pts <10mg/l; 5/23pts >50mg/l))
Same findings in Myeloma 9 and in
Myeloma 4 - 8

3. Paraprotein level at presentation:
32.7% < 30 g/L
42.3% < 30 g/L

4. Difficult to monitor paraproteins starting at <10g/l
Paraprotein level g/l
IgG
% of 1,665 pts
IgA
% of 699 pts
IgD
% of 35 pts
IgM
% of 6 pts
<5
3.7
6.4
54.2
16.7
5 - <10
4.4
5.7
17.1
9.8
14.3
49.0
42.5
11.4
>40
33.1
31.0
66.7
No difference between kappa and
lambda light chain types
Patients who had a whole paraprotein on serum immunofixation

5. FLC enable monitoring of 75% patients with PP <10g/l
FLC Level k/l
difference
mg/l
IgG <10g/l %
134pts
IgG >10g/l
1,531pts
IgA <10g/l
85pts
IgA >10g/l
614pts
IgD <10g/l
25pts
IgD >10g/l
10pts
<100 28 32 26 37 4 20 34 25 24 30
>500 52 49 35 72 70
FLC are higher when paraprotein <10g/l (no variation above 10g/l)
IgD patients have low paraprotein and high FLC levels – usually lambda

6. Suppression of polyclonal immunoglobulin
Normal range B
Normal range C
Normal range
n (%)
IgG
(n =1302)
IgA
(n = 2471)
IgM
(n = 3228)
Below NR
1049 (80.6)
1981(80.2)
2871 (88.9)
Within NR
253 (19.4)
479 (19.4)
349 (10.8)
Above NR
0 (0.0)
11 (0.4)
8 (0.2)
Median (range) g/L
3.90 (0–15.65)
0.34 (0–5.60)
0.16 (0–3.56)

7. FLC levels are higher when the light chain type is lambda
FLC Level k/ldifference
mg/l
IgG kappa %
1,153pt
IgG lambda
512pts
IgA kappa
448pts
IgA lambda
251pts
LCO
kappa
207pts
lambda
154pts
<100 32 30 40 25 34 29 27 15
>500 41 33 46 85
Lambda higher than kappa analysing as continuous variables. Equally nephrotoxic

8. eGFR by difference between involved and uninvolved FLC (dFLC) clone at presentation in mg/L
Dr Punit Yadav analysis of 1,595 Myeloma 9 patients

9. Immunoglobulin half lives and response
IgG 21 days
IgA 5 days
FLC 12 hours
Short half lives of serum FLC allow
real time evaluation of tumour kill

10. Paraprotein response to 3 week cycles of myeloma therapy
100
40 g/l 80 60
IgG paraprotein g/l
20 g/l
8 g/l 40
10 g/l
8g/l
5.0 g/l 20
4 g/l
2.0 g/l
1.6 g/l
0.8 g/l
0.3 g/l
1st cycle
80% kill
2nd cycle
80% kill
3rd cycle
80% kill
4th cycle
80% kill (0.16% residue)

11. FLC response to 3 week cycles of myeloma therapy
5000
4000
3000
2000
1000
1st cycle
80% kill
2nd cycle
3rd cycle
4th cycle
80% kill (0.16% residue)
Dif FLC mg/l
4000 mg/l
800 mg/l
160 mg/l
normalise

12. Does a reducing kill rate indicate clonal heterogeneity and predict poor survival?
5000
4000
3000
2000
1000
1st cycle
80% kill
2nd cycle
20% kill
3rd cycle
4th cycle
0% kill (3.2% residue)
Dif FLC mg/l
4000 mg/l
200 mg/l
160 mg/l
normalise

13. FLC response at end of cycle 1 predicts later response
Correlation coefficient=0.588
Correlation coefficient=0.155
227 Myeloma 11 patients with
IgG and IgA paraproteins >10g/l
and dFLC >100mg/l

14. Early response in myeloma 11 and the importance of Ig half life
Unpaired T-test
(N)
531
117
211
756
328 58
225
% Reduction in paraprotein/FLC post -cycle 1 - three weeks
p=0.99
p=0.07
p= 5.1E-15
p= 0.005
p= 1.4E-07
FLC
IgG
IgA

15. IgG level does not appear to affect IgG half-life in myeloma 11
IgG level does not appear to affect IgG half-life in myeloma 11. (FcR neonatal receptor saturated at about 30g/l)
IgG
FLC
IgG
FLC
IgG
FLC
IgA
FLC
FLC

16. No difference in early paraprotein and FLC responses between Intensive and Non-intensive patients 11th trial
Figure 1: Percentage reduction in paraprotein and sFLC light chain levels for patients treated in the MRC Myeloma XI trial at the end of the first cycle of induction chemotherapy. Whisker boxplots showing the median, 25th and 75th centiles. Tails represent the 5th and 95th centiles. The diamonds represent the means. Mann Whitney U tests were performed to assess the statistical significance of the differences between the groups. RCD=lenalidomide, cyclophosphamide and dexamethasone (320mg/cycle); CTD=cyclophosphamide, thalidomide and dexamethasone (320mg/cycle); RCDa=lenalidomide, cyclophosphamide and dexamethasone (160mg/cycle) and CTDa=cyclophosphamide, thalidomide and dexamethasone (160mg/cycle).
p=0.11
p=0.30
p=0.38
p=0.40
p=0.24

17. No difference in maximum responses to induction therapy between Intensive and Non-intensive patients 9th & 11th trial
p=0.07
p=0.77
p=0.70
p=0.13
Figure 2: Percentage reduction in paraprotein and sFLC light chain levels for patients treated with CTD or CTDa in the MRC Myeloma IX trial and RCD/CTD or RCDa/CTDA in the MRC Myeloma XI trial at the end of induction chemotherapy. Whisker boxplots showing the median, 25th and 75th centiles. Tails represent the 10th and 95th centiles. The diamonds represent the means. Mann Whitney U tests were performed to assess the statistical significance of the differences between the groups. RCD=lenalidomide, cyclophosphamide and dexamethasone (320mg/cycle); CTD=cyclophosphamide, thalidomide and dexamethasone (320mg/cycle); RCDa=lenalidomide, cyclophosphamide and dexamethasone (160mg/cycle) and CTDa=cyclophosphamide, thalidomide and dexamethasone (160mg/cycle).

18. Response at end of induction versus 100d post HDM HDM deepens response but that is not completely evident by 100 d for IgG paras
FLC Response
Paraprotein Response
IgA 45
IgG
117
LCO 44
Percentage reduction in sFLC and paraprotein levels at the end of induction chemotherapy compared to
at 100 days post high dose melphalan for patients treated in the MRC Myeloma IX trial.
Whisker boxplots showing the median, 25th and 75th centiles. Tails represent the 10th and 95th centiles.
The diamond represents the mean.
Mann Whitney U tests p<0.01 for all pairs

19. Response at end of induction versus 100d post HDM HDM deepens response but that is not completely evident by 100 d for IgG paras
FLC Response
Paraprotein Response
LCO
105
IgA 97
IgG
249
IgA 97
IgG
249
Percentage reduction in sFLC and paraprotein levels at the end of induction chemotherapy compared to
at 100 days post high dose melphalan for patients treated in the MRC Myeloma XI trial.
Whisker boxplots showing the median, 25th and 75th centiles. Tails represent the 10th and 95th centiles.
The diamond represents the mean.
Mann Whitney U tests p<0.01 for all pairs

20. At relapse there were three types of relapse:-
Serum free immunoglobulin light chain evaluation as a marker of impact from intraclonal heterogeneity on myeloma outcome Blood 123(22):
The first 520 patients in myeloma 9 who had an IgA or IgG paraprotein at presentation
At relapse there were three types of relapse:-
183/520 (35.2%) patients had a significant increase in both paraprotein and FLC levels
258/520 (49.6%) patients only had a significant increase in their paraprotein levels
Paraprotein only relapse
/520 (10.4%) patients relapsed with only an increase in their FLC levels:
Free light chain escape
24/369 (6.5%) IgG and 30/151 (19.9%) IgA patients relapsed with FLC escape

21. Effect of Relapse Type on Survival

22. Model of Darwinian Evolution in Multiple Myeloma

23. Is this type of clonal heterogeneity present in response?
IgG and FLC>100mg/l
Post cycle one results
IgA and FLC>100mg/l
Post cycle one results

24. Key findings
IgG 60%; IgA 25%; LCO 13%; IgD 1.3%; NS 0.8% (LCO missed in older pts?) In 10% of patients whole paraprotein <10g/l but can be monitored by FLC Polyclonal Igs are below normal level in 80% and above normal level in <0.4% (3,228pts) FLC levels are higher when type is lambda but kappa and lambda FLC are equally likely to be nephrotoxic at the same level. Many patients have non-toxic FLC. FLC levels need to be >500mg/l to have nephrotoxicity. Ig half life is important when assessing response IgG 21days (7-10d on dex); IgA 5 days FLC <12 hrs Early FLC response indicates kill rate and can predict max response. Changing kill rate with subsequent cycles of therapy may indicate clonal heterogenity and poor prognosis No difference in speed of response or final depth of response between patients on 20 and on 40 mg dex Light chain escape in 6% IgG and 20% IgA patients. Different relapse patterns indicate clonal heterogeneity. Patients relapsing with FLC survive worse from relapse than paraprotein only relapses Some patients have asynchronous FLC and paraprotein response which might also indicate clonal heterogeneity and poor survival