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Supplementary materials - Cancer Chemotherapy and Pharmacology
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Computed determination of the in vitro optimal chemocombinations of
sphaeropsidin A with chemotherapeutic agents to combat melanomas
Aude Ingels1, Carina Dinhof2,3, Abhishek D. Garg4 , Lucia Maddau5, Marco Masi6, Antonio
Evidente6, Walter Berger 2,3 , Bieke Dejaegher7 and Véronique Mathieu1
1Laboratoire de Cancérologie et Toxicologie Expérimentale, Université Libre de Bruxelles,
Boulevard du Triomphe, Accès 2, 1050 Ixelles, Belgium.
2Department of Medicine I, Institute of Cancer Research, Medical University Vienna,
Spitalgasse 23, 1090 Vienna, Austria
3Comprehensive Cancer Center, Medical University Vienna, Spitalgasse 23, 1090 Vienna,
Austria
4Laboratory for Cell Death Research and Therapy (CDRT), Department of Cellular and
Molecular Medicine, KU Leuven University, Leuven, Belgium.
5Dipartimento di Agraria, Sezione di Patologia vegetale ed Entomologia, Università degli
Studi di Sassari, Viale Italia 39, 07100, Sassari, Italy.
6Dipartimento di Scienze Chimiche, Universita’ di Napoli Federico II, Complesso
Universitario Monte S. Angelo, Via Cintia 4, 80126 Napoli, Italy.
7Laboratoire d’Analyse Instrumentale et de Bioélectrochimie, Université Libre de Bruxelles,
Boulevard du Triomphe, Accès 2, 1050 Ixelles, Belgium
Correspondence to:
Véronique Mathieu, MD, PhD
Laboratoire de Cancérologie et Toxicologie Expérimentale – Faculté de Pharmacie –
Université Libre de Bruxelles (ULB)
Campus de la Plaine – Boulevard du Triomphe – 1050 Brussels – Belgium.
Tel: [phone] E-mail: [email protected]

Supplementary materials - Cancer Chemotherapy and Pharmacology
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Concentrations (µM)
N°Exp
Sph A
Cisplatin
Sph A
TMZ
1-3
0
0
0
0
4-6
6
0
6
0
7-9
0
100
0
1000
10-12
6
100
6
1000
13-15
0
50
0
500
16-18
6
50
6
500
19-21
3
0
3
0
22-24
3
100
3
1000
25-30
3
50
3
500
31
2
35
2
350
32
4
40
4
400
33
3
75
3
750
Table 1: Factorial experimental design performed to establish the model for
sphaeropsidin A (Sph A)/ cisplatin and Sph A/ temozolomide (TMZ) combinations and
tested on each cell line. 9 combinations of concentrations (µM) including the central point
and 3 test points permit to build the model.

Supplementary materials - Cancer Chemotherapy and Pharmacology
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Figs 1: Example of dissymmetric distribution of B16F10 dataset for sphaeropsidin A and
cisplatin. Boxplot for B16F10 data before (A) and after logarithm transformation (B). Table
C includes all absolute data visualized above.
Mean
Standard
Deviation
Minimum
Maximum
Centrum
B16F10
16.045
27.592
2.076
100.000
51.038
ln(B16F10)
2.021
1.085
0.731
4.605
2.668
A
B
C

Supplementary materials - Cancer Chemotherapy and Pharmacology
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Table 2: The second-order polynomial equations built from experimental data to predict
relationship between concentrations of each drug and their combined response. X1 and
X2 are the coded values for [sphaeropsidin A (Sph A)] and [Cisplatin/Temozolomide], b0 is
the intercept coefficient, b1/ b2 are the linear coefficients, b1-1/ b2-2 are the quadratic
coefficients, and b1-2 is the factor interaction coefficient.
Pre-treatment
Sph A and cisplatin
Y = 0.618 + 0.201 * X1 + 0.149 * X2 - 0.125 * (X1*X1) -
0.135 * (X2*X2) - 0.038 * (X1*X2)
Sph A and temozolomide
Y= 0.319 + 0.251 * X1 + 0.096 * X2 - 0.010 * (X1*X1) -
0.009 * (X2*X2) + 0.031 * (X1*X2)
Co-treatment
Sph A and cisplatin
Y= 0.740 + 0.148 * X1 + 0.189 * X2 - 0.106 * (X1*X1) -
0.156 * (X2*X2) - 0.105 * (X1*X2)
Sph A and temozolomide
Y= 0.631 + 0.252 * X1 + 0.093 * X2 - 0.212 * (X1*X1) -
0.055 * (X2*X2) - 0.033 * (X1*X2)

Supplementary materials - Cancer Chemotherapy and Pharmacology
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Pre-treatment
Sph A
Cisplatin
Y optimal
1
5
75
0.7247
2
5
70
0.7245
3
5
80
0.7223
4
5
65
0.7215
5
6
70
0.7172
6
5
85
0.7172
7
6
75
0.7162
8
5
60
0.7159
9
6
65
0.7155
10
6
80
0.7126
Pre-treatment
Sph A
TMZ
Y optimal
1
6
1000
0.68
2
6
950
0.67
3
6
900
0.66
4
6
850
0.65
5
6
800
0.63
6
6
750
0.62
7
6
700
0.61
8
6
650
0.60
9
5
1000
0.59
10
6
600
0.59
Tables 3: Summary tables of the ten best combinations of compounds (in µM) ranked
according to the best predicted response (Y optimal). Combinations of sphaeropsidin A
(Sph A) with cisplatin in pre-treatment and co-treatment conditions are presented in A and B
respectively. Combinations of Sph A with temozolomide (TMZ) in pre-treatment and co-
treatment conditions are presented in C and D respectively.
Co-treatment
Sph A
Cisplatin
Y optimal
1
4
75
0.8149
2
5
70
0.8139
3
4
70
0.8138
4
5
65
0.8131
5
4
80
0.8128
6
5
75
0.8114
7
4
65
0.8096
8
5
60
0.8092
9
4
85
0.8076
10
5
80
0.8058
Co-treatment
Sph A
TMZ
Y optimal
1
5
850
0.73
2
5
800
0.73
3
5
900
0.73
4
5
750
0.73
5
5
950
0.72
6
5
700
0.72
7
4
850
0.72
8
4
900
0.72
9
5
1000
0.72
10
5
650
0.72
A
B
C
D