Supplementary Material s73

Supplementary Material

Supplementary Table 1. Dose modifications of RO4929097 and Temsirolimus

Agent Initial 1st 2nd reduction 3rd Dose reduction reduction RO4929097 20 mg 10 mg 5 mg Discontinue

10 mg 5 mg Discontinue -

Temsirolimus 37.5 mg 25 mg 20 mg Discontinue

25 mg 20 mg Discontinue -

Legend: In the event of toxicity, the doses of RO4929097 and temsirolims were adjusted. Patients requiring dose reductions could not have the dose re-escalated with subsequent treatments. Subjects with toxicities that were manageable with supportive therapy (e.g. nausea/vomiting) no dose reduction was mandated. Subjects in whom one agent was discontinued could continue receiving the other agent if, in the opinion of the treating physician, the patient may continue to benefit from treatment. In such cases, for the agent to be continued, the patient remained at the same dose as he/she was currently receiving, without dose escalation. Supplementary Table 2.Pharmacokinetic sampling schedules for Temsirolimus.

Cycle Temsirolimus PK Sampling Time-Points No. of Temsirolimus PK samples Cycle Day Time 1 Day 1 + 2 Before temsirolimus infusion 8 End of temsirolimus infusion, then 0.5, 1, 2, 4, 6 and 24 h after end of temsirolimus infusion Day 3 48 h after end of temsirolimus infusion 1 Day 8 + 9 Before temsirolimus and RO4929097 8 dosing End of temsirolimus infusion, then 0.5, 1, 2, 4, 6 and 24 h after end of temsirolimus infusion Day 10 48 h after end of temsirolimus infusion 1 Day 15 Before temsirolimus and RO4929097 1 dosing Day 22 + 23 Before temsirolimus and RO4929097 8 dosing End of temsirolimus infusion, then 0.5, 1, 2, 4, 6 and 24 h after end of temsirolimus infusion 2+ Day 1^ Before temsirolimus and RO4929097 1+ dosing ^Trough PK sample was collected on day 1 of cycle 2 onwards up to cycle 6 day 1. Supplementary Table 3.Pharmacokinetic sampling schedules for RO4929097.

Cycle RO4929097 PK Sampling Time-Points No. of RO4929097 PK samples Cycle Day Time 1 Day 8 + 9 Before temsirolimus and RO4929097 8 dosing End of temsirolimus infusion, then 0.5, 1, 2, 4, 6 and 24 h after end of temsirolimus infusion Day 10 48 h after end of temsirolimus infusion 1 Day 15 Before temsirolimus and RO4929097 1 dosing Day 22 + 23 Before temsirolimus and RO4929097 8 dosing End of temsirolimus infusion, then 0.5, 1, 2, 4, 6 and 24 h after end of temsirolimus infusion 2+ Day 1^ Before temsirolimus and RO4929097 1+ dosing Supplementary Table 4.Dose delays and reductions due to adverse events (AEs) per dose level and drug

Total Any dose Any Delay Dose Length of reduction Pt ID Main reason of delay Timing level delay Te RO (wk) Tem RO m 001 1 No Yes G3 mucositis, G2 rash 1 No No C1D15 002 1 No No n/a n/a No No n/a 003 1 Yes Yes G3 fatigue, G3 nausea 2 No No C2D8, C3D1, C3D15 004 1 Yes Yes Palliativeradiation 1 No No C9D1 005 1 No No n/a n/a No No n/a 006 1 No No n/a n/a No No n/a 007 1 Yes Yes G3 neutropenia 2 No No C1D8 008 1 Yes Yes G3 gastroenteritis ¶ 2 No No C3D1, C4D15 009 2 Yes Yes Investigation of n/a No No C1D28 disease progression 010 2 No No n/a n/a No* No C2D1 012⌃ 2 Yes Yes G2 1 No No C3D1 hypophosphatemia, G1 hypokaliemia 013 3 No No n/a n/a No No n/a 014 3 Yes Yes G2 eye infection, G1 1 No No C3D1, C4D15 hypocalcemia 015 3 Yes Yes G3 neutropenia 2 No No C2D1, C8D8 016 3 Yes Yes G2 rash 1 No No n/a 017 3 Yes Yes G3 abdominal pain¶ 1 No No C2D15 018 3 Yes Yes G2 urinary tract 1 No No C2D8 infection Legends: ⌃Patient 011 was not registered and did not receive treatment.*Temsirolimus was discontinued from C2D1 at the patient’s request. ¶ Non treatment-related AEs Abbreviations: n/a, not applicable. Supplementary Table 5. Notch pathway protein expression and TTP (N=14)

Protein HR (95% CI) P value Jagged-1 1.3(0.8-2.3) 0.3 NICD 1.0 (0.7-1.3) 0.7 Notch-3 1.1 (0.9-1.5) 0.3 Abbreviations: TTP, Time to progression; NICD, Notch Intracellular Domain; HR, Hazard Ratio; CI, Confidence Interval Supplementary Table 6. Mutational analysis and TTP (N=14)

Platform TTP Tumor Type Gene Mutation used^ (months) Melanoma NRAS Q61K MiSeq 2.1 Endometrial TP53 R249G MiSeq 1.4 NSCLC EGFR L858R MiSeq 5.7 GIST KIT K642E MiSeq 9.7 Uterine Sequenom JAK3 V722I 2.7 Leiomyosarcoma Abbreviations: NSCLC, Non-small cell lung cancer; GIST, gastrointestinal stromal tumor; TTP, time-to-progression Legends: ^ Genotyping was performed with either Sequenom MassARRAY (Sequenom) PMH v1.0 customized panel for solid tumors that includes 280 mutations in 23 genes or the MiSeq (Illumina) TruSeq Amplicon Cancer Panel (TSACP) panel that includes 212 amplicons in 48 genes in the CLIA-certified University Health Network Advanced Molecular Diagnostics Laboratory