PIP3 (38:4) pmol/mg (tissue) N=2を入れる　PIP3 (38:4) に入れる Supplementary Fig. S1. The mean baseline Levels of PIP3 (38:4) per mg tissue in xenograft models transplanted with SCLC cells. The red and blue bars indicate gedatolisib-sensitive H1048 tumors and gedatolisib-resistant SBC-5 tumors, respectively.

A B Increase Decrease PRPP ADP-Rib CM-Asp Val Lactate ATP AMP Met Ile Put B Increase Decrease PRPP ATP UA Guanine SA Put GSH His Ile Lys Phe Trp Val Supplementary Fig S2. Metabolic alteration in SCLC cells treated with gedatolisib in vitro and in vivo. Fold change in metabolite levels on comparison of DMSO control and gedatolisib-exposed sensitive SCLC cells and gedatolisib-exposed resistant SCLC cells (n = 3–5). (A) SCLC cells were harvested 6 hours post gedatolisib exposure and intracellular metabolites were measured using a metabolomic approach. Blue bars indicate sensitive SCLC cell lines and red bars indicate the resistant SCLC cell line. (B) SCLC tumors were harvested 6 hours post intravenous gedatolisib and intracellular metabolites were measured using a metabolomic approach. Blue bars indicate gedatolisib-sensitive H1048-xenografted tumors and red bars indicate gedatolisib-resistant SBC-5-xenografted tumors.

PLS-DA LAD cell lines Sensitive SCLC cell lines Resistant SCLC cell lines Supplementary Fig. S3. Comparative metabolic profiling in LAD and SCLC cell lines Partial Least Squares Discriminant Analysis (PLS-DA) score plots are shown. A total 116 of metabolites were analyzed. The scores plot: the blue dots denote the lung adenocarcinoma cell lines, red dots denote the sensitive SCLC cell lines, green dots denote the resistant SCLC cell lines.

A Increase CCNG2 KLHL24 CITED2 MXI1 BTG1 CDKN1B CCND1 AMD1 MAT2A SRM Decrease FOXO Pathway Polyamine B * * * * H1048 DMS114 H1694 H1048 DMS114 H1694 H1048 DMS114 H1694 Supplementary Fig. S4. Gene expression alteration after treatment with PI3K/mTOR inhibitor (A) RNA–Seq for transcriptome analyses. The gene expression level was calculated by reads per million mapped reads (RPKM). The y-axis represents the log2 ratio normalized by the value of the DMSO control. (B) RT-PCR for the expression level of genes related with polyamine biosynthesis and purine salvage pathway in sensitive H1048 and DMS114, and resistant H1694 cells. Error bars represent one s.d. *p < 0.05, t-test.

Polyamine Biosynthesis THF Cycle Folate 5-methylTHF Polyamine Biosynthesis Methionine Cycle Ornithine S-Adenosyl- L-homocysteine ODC1 SAH AMD1 Putrescine Homocysteine S-Adenosyl methionine dcSAM SAM SRM Methionine MAT Spermidine Methylthioadenosine (MTA) MTAP Adenine PRPP Purine Salvage Supplementary Fig. S5. Schematic diagram of metabolic linkage Metabolic linkage between the methionine cycle, polyamine biosynthesis, and purine salvage pathway.

A ** * Gedatolisib B ** Gedatolisib Supplementary Fig. S6. Addition of 5'-deoxy-5'-(methylthio)-adenosine (MTA) reversed drug sensitivity from a resistant to a sensitive phenotype. DMS273 (A) or SBC-5 (B) cell survival (%) was measured in the presence of 0.1 mM MTA treated together with 100 nM or 10 nM of gedatolisib. 　　　The data are shown as the mean ± s.d. (n = 8). Error bars represent one s.d. *p < 0.05; **p < 0.01, t-test.

A B siNC #1 #2 #3 siHPRT1 GAPDH HPRT1 siHPRT1 C HE HPRT1 H1048 SBC-5 Supplementary Fig. S7. HPRT1 expression was downregulated by an HPRT1-specific siRNA in SCLC cells. (A) siRNA targeting HPRT1 successfully knocked-down HPRT1 mRNA as much as 80% in DMS273 cells compared with a negative control construct expressing siNC. (B) Western blot (WB) analysis of siHPRT1-treated SCLC cells. siRNA targeting HPRT1 knocked down HPRT1 levels in DMS273 cells. The size signal for HPRT1 was approximately detected at 25 kDa. GAPDH was used as a loading control. (C) Representative images from IHC analysis using an antibody against HPRT1. Tumors derived from H1048 or SBC-5 cells were removed, and HPRT1 were visualized as brown. Scale bar indicates 100 μm.