Supplementary

A gene expression based predictor for myeloma patients at high risk of developing bone disease on bisphosphonate treatment

Ping Wu1, Brian A. Walker1, Daniel Brewer2, Walter M. Gregory3, John Ashcroft4, Fiona M. Ross5, Graham H Jackson6, Anthony J Child3,Faith E. Davies1,Gareth J. Morgan1

1Section of Haemato-Oncology, Institute of Cancer Research, UK; 2Section of Molecular Carcinogenesis, Institute of Cancer Research, UK;3Clinical Trials and Research Unit, University of Leeds, UK; 4 Mid Yorkshire Hospitals NHS Trust, Wakefield, UK ; 5 University of Southampton, Wessex Regional Genetics Laboratory, Salisbury, UK; 6Royal Victoria Infirmary, Newcastle-on-Tyne, UK

Running title:Definingmyeloma patients at high risk of developing bone disease

Corresponding Author: Gareth Morgan, Section of Haemato-Oncology, Institute of Cancer Research, 15 Cotswold Road, Sutton, SurreySM2 5NG, United Kingdom. Telephone +44(0)20 87224130, Fax +44(0)20 87224432, Email:

Table A1. Patient characteristics at presentation: the patients included in the GEP analysis were representative of patients entered into the overall trial.

Characteristic / Expression
(n=261) / Myeloma IX
(n=1960)
Age, years
Mean
SD / 64.6
10.1 / 64.6
10.2
Serum β2microglobulin, mg/l
Mean
SD
Total patients / 5.9
4.2
193 / 6.1
5.9
1789
Ca, mmol/l
Mean
SD
Total patients / 2.46
0.65
254 / 2.46
2.27
1884
Haemoglobin, g/dl
Mean
SD
Total patients / 10.5
1.8
261 / 10.8
4.3
1880
Serum albumin, g/l
Mean
SD
Total patients / 34.2
7.1
260 / 34.7
7.0
1858
Deletion 13q
%
No. patients
Total patients / 48.7
116
238 / 45.3
473
1043
t(4;14)
%
No. patients
Total patients / 18.5
45
243 / 11.4
120
1052
t(11;14)
%
No. patients
Total patients / 19
46
242 / 13.9
146
1047
Hyperdiploidy
%
No. patients
Total patients / 54.8
131
239 / 57.7
578
1002
Baseline BD
%
No. patients
Total patients / 69.7
182
261 / 72.1
1401
1941
On-treatment SRE
%
No. patients
Total patients / 29.3
76
259 / 28.3
545
1924
Chemotherapy, % of patients
CVAD
CTD
MP
CTDa / 25.4
29.8
22.4
22.4 / 28.3
28.2
21.6
21.8
Biphosphonate, % of patients
Sodium clodronate
Zoledronic acid / 44.4
55.6 / 49.9
50.1

Table A2. Associations of FISH abnormalities with presenting BD and on-treatment SREs in the expression training set (N=205) were consistent with overall trial data.

Baseline BD % (no./total no.) / p value / On-treatment SRE % (no./total no.) / p value
hyperdiploidy / 77.9%
(81/104) / 0.01 / 28.1%
(25/89) / 0.05
non-hyperdiploidy / 60.8%
(48/79) / 13.8%
(9/65)
t(4;14) / 51.9%
(14/27) / 0.02 / 12.5%
(3/24) / NS
non t(4;14) / 74.2%
(118/159) / 23.3%
(31/133)
t(11;14) / 70.6%
(24/34) / NS / 21.4%
(6/28) / NS
non t(11;14) / 70.9%
(107/151) / 21.1%
(27/128)
MAF translocations / 50%
(4/8) / NS / 14.3%
(1/7) / NS
non MAF translocations / 71.8%
(127/177) / 21.5%
(32/149)

Table A3.50 differentially expressed genes were identified by SAM between patients with BD at diagnosis and those without (ranked by SAM score).

Affymetrix Probeset / Gene Symbol / SAM Score / Pathway/Function / Cytoband
204602_at / DKK1 / 4.45 / WNT signalling inhibitor / 10q11.2
219056_at / RNASEH2B / 3.90 / DNA replication / 13q14.3
225858_s_at / XIAP / -3.83 / Apoptosis / Xq25
210564_x_at / CFLAR / -3.74 / Apoptosis / 2q33-q34
221486_at / ENSA / -3.63 / Modulate insulin secretion / 1q21.3
215075_s_at / GRB2 / -3.51 / Growth factor signalling / 17q24-q25
211992_at / WNK1 / -3.47 / Growth factor signalling / 12p13.3
223049_at / GRB2 / -3.46 / Growth factor signalling / 17q24-q25
225859_at / XIAP / -3.46 / Apoptosis / Xq25
1564154_at / ---- / -3.46 / Not known / ---
227697_at / SOCS3 / -3.34 / Growth factor signalling / 17q25.3
225764_at / ETV6 / -3.31 / Transcriptional regulator / 12p13
233841_s_at / SUDS3 / -3.31 / Transcriptional regulator / 12q24.23
219297_at / WDR44 / -3.28 / Cell migration / Xq24
224683_at / FBXO18 / -3.28 / DNA repair / 10p15.1
204980_at / CLOCK / -3.28 / Transcriptional regulator / 4q12
1564155_x_at / ---- / -3.23 / Not known / ---
200605_s_at / PRKAR1A / -3.20 / Growth factor signalling/Apoptosis / 17q23-q24
227748_at / RBMXL1 / -3.19 / Nucleic acid binding / 1p22.2
211994_at / WNK1 / -3.19 / Growth factor signalling / 12p13.3
224778_s_at / ---- / -3.18 / Not known / ---
218312_s_at / ZSCAN18 / -3.17 / Transcriptional regulator / 19q13.43
208686_s_at / BRD2 / -3.16 / Transcriptional regulator / 6p21.3
210649_s_at / ARID1A / -3.14 / Transcriptional regulator / 1p35.3
211986_at / AHNAK / -3.14 / Nervous system development / 11q12.2
211862_x_at / CFLAR / -3.13 / Apoptosis / 2q33-q34
210555_s_at / NFATC3 / -3.13 / Transcriptional regulator / 16q22.2
228628_at / SRGAP2P1 / -3.12 / Not known / 1p11.2
209048_s_at / ZMYND8 / -3.12 / Transcriptional regulator / 20q13.12
1561167_at / ---- / -3.09 / Not known / ---
220169_at / TMEM156 / -3.09 / Not known / 4p14
212152_x_at / ARID1A / -3.08 / Transcriptional regulator / 1p35.3
217550_at / ATF6 / -3.08 / Transcriptional regulator / 1q22-q23
217936_at / ARHGAP5 / -3.07 / Cell migration / 14q12
211316_x_at / CFLAR / -3.07 / Apoptosis / 2q33-q34
224513_s_at / UBQLN4 / -3.06 / Protein degradation / 1q21
220532_s_at / TMEM176B / -3.06 / Cell differentiation / 7q36.1
201715_s_at / ACIN1 / -3.06 / Cell differentiation/Apoptosis / 14q11.2
208704_x_at / APLP2 / -3.05 / Growth factor signalling / 11q23-q25|11q24
201059_at / CTTN / -3.05 / Cell migration / 11q13
219429_at / FA2H / -3.05 / Cell death / 16q23
212956_at / TBC1D9 / -3.04 / Growth factor signalling / 4q31.21
213329_at / SRGAP2 / -3.03 / Growth factor signalling / 1q32.1
211317_s_at / CFLAR / -3.03 / Apoptosis / 2q33-q34
218004_at / BSDC1 / -3.03 / Not known / 1p35.1
238559_at / ---- / -3.02 / Not known / ---
208248_x_at / APLP2 / -3.02 / Growth factor signalling / 11q23-q25|11q24
210616_s_at / SEC31A / -2.99 / Protein transport / 4q21.22
225289_at / STAT3 / -2.99 / Growth factor signalling / 17q21.31
211985_s_at / CALM1 / -2.99 / Cell cycle / 14q24-q31

Table A4. The risk of SRE development within 2 years predicted by the 7-gene plus Ca model and the actual risk in our cohort of patients (n=166, 6 patients with missing Ca value excluded). The range of the risks under the same number of risk factors reflects the different combinations of the factors for individual patients.

No. of risk factors / Number of patients (%) / Model predicted SRE risk, average, range (%) / Actual SRE risk
(%)
8 / 2 (1.2%) / 97.1 / 100%
7 / 1 (0.6%) / 90.7 / 100%
6 / 3 (1.8%) / 84.2 (80.5 - 87.1) / 100%
5 / 6 (3.6%) / 68.5 (62.5 – 74.5) / 66.7%
4 / 19 (11.4%) / 50 (38.5 – 60.4) / 57.9%
3 / 22 (13.3%) / 28.3 (20.1 – 40.2) / 27.3%
2 / 30 (18.1%) / 14.9 (10.5 – 22.1) / 10%
1 / 45 (27.1%) / 7.7 (5.4 – 10.2) / 8.9%
0 / 38 (22.9%) / 3.2 / 7.9%

Figure A1. Examples of cumulative incidence curve show that the genes forming the SRE signature could significantly discriminate the risk developing an on-treatment SRE.

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