Method for selecting patients responsive for cancer treatments

1. A method of quantifying intracellular metabolite effluxes in permeabilized cancer cells for selecting cancer patients responsive for a cancer treatment, the method comprising the steps of:

a) providing a sample of cancer cells taken from a patient;

b) permeabilizing said cancer cells;

c) incubating said permeabilized cancer cells in a reaction medium for a period of time allowing biological activity of intracellular organelles and accumulation of one or more metabolites produced by said activity into the reaction medium in the presence of two or more substrates relating to a metabolite efflux or effluxes of interest, wherein the two or more substrates comprise at least glutamine and pyruvate;

d) determining a quantity of the one or more metabolites relating to said metabolite efflux or effluxes of interest accumulated in the reaction medium during step c); and

e) comparing the quantity of one or more metabolites determined in step d) to equivalent measurements performed on control samples of the same tissue type as the cancer cells; and

f) if a difference between the quantity of one or more metabolites in step d) and those of the control samples indicate elevated metabolic activity of the cancer cells, contacting the cancer cells with a drug affecting a metabolic pathway or pathways relating to said metabolite efflux or effluxes of interest to determine a treatment response of the cancer cells to the drug, wherein the drug comprises a glutaminase (GLS) inhibitor.

2. The method according to claim 1 , wherein the cells are permeabilized in step b) by electro-permeabilization or by contacting the cells with a permeabilization agent.

3. The method according to claim 2 , wherein said permeabilization agent is selected from the group consisting of: saponin and digitonin.

4. The method according to claim 1 , wherein the one or more metabolites determined in step d) are selected from the group consisting of: pyruvate, glutamine, glutamate, citrate, iso-citrate, alpha-ketoglutarate, aspartate, alanine, glycine, succinate, fumarate, malate, and oxaloacetate.

5. The method according to claim 4 , wherein the metabolites determined in step d) are pyruvate and glutamine.

6. The method according to claim 1 , wherein the two or more substrates relating to a metabolite efflux or effluxes of interest in step c) further comprise an additional substrate selected from the group consisting of: citrate, iso-citrate, glutamate, alpha-ketoglutarate, succinate, acylcarnitine, ornithine, arginine, formate, proline, and malate.

7. The method according to claim 6 , wherein the additional substrate comprises an acylcarnitine.

8. The method according to claim 1 , wherein, in a control sample, said substrate relating to a metabolite efflux of interest in step c) is an inhibitor of said metabolite efflux.

9. The method according to claim 1 , wherein the one or more metabolites determined in step d) are selected from the group consisting of: folate, ATP, ADP, citrulline, ornithine, δ-aminolevulinic acid, arginine, asparagine, cysteine, histidine, isoleucine, leucine, lysine, methionine, proline, serine, threonine, tryptophan, phenylalanine, tyrosine, valine, H 2 O 2 , carnitine, phosphoenolpyruvate, formate, urea, and 2-hydroxy-glutarate.

10. The method according to claim 1 , wherein said control samples in step e) are healthy tissue samples taken from said patient.

11. The method according to claim 1 , wherein said period of time in step c) is 0.1-180 minutes.

12. The method according to claim 1 , wherein said cancer cells are from breast cancer.

13. The method according to claim 1 , wherein the treatment response of the cancer cells to a drug affecting a metabolic pathway is assessed in step e) based on a metabolic dependency of the cancer cells shown by the results obtained in step d).

14. The method according to claim 13 , wherein said cancer cells are dependent on glutamine.

15. The method according to claim 1 , wherein an aggressiveness of the cancer cells is assessed in step e) based on the metabolic activity of the cancer cells shown by the results obtained in step d) so that metabolically more active cells correspond to an aggressive cancer, and metabolically less active cells correspond to a less aggressive cancer.

16. A method for selectively treating cancer patients comprising:

a) providing a sample of cancer cells taken from a patient;

b) permeabilizing said cancer cells;

c) incubating said permeabilized cancer cells in a reaction medium for a period of time allowing biological activity of intracellular organelles and accumulation of one or more metabolites produced by said activity into the reaction medium in the presence of two or more substrates relating to a metabolite efflux or effluxes of interest, wherein the two or more substrates comprise at least glutamine and pyruvate;

d) determining a quantity of the one or more metabolites relating to said metabolite efflux or effluxes of interest accumulated in the reaction medium during step c); and

e) comparing the quantity of one or more metabolites determined in step d) to equivalent measurements performed on control samples of the same tissue type as the cancer cells; and

f) if a difference between the quantity of one or more metabolites in step (d) and those of the control samples indicate elevated metabolic activity of the cancer cells, administering to the patient an amount of a drug affecting a metabolic pathway or pathways relating to said metabolite efflux or effluxes of interest, wherein the drug comprises a glutaminase (GLS) inhibitor.