Method of manufacturing dual-specific T-cells for use in cancer immunotherapy
The present invention relates to autologous dual-specific lymphocytes, methods of making and uses for the treatment of tumors. In particular, the invention relates to methods producing autologous dual-specific lymphocytes comprising an endogenous receptor for at least one tumor associated antigen and an exogenous receptor for a strong antigen.
1. A purified population of autologous dual-specific lymphocytes which have specificity for two or more antigens, wherein a population of lymphocytes is isolated from a patient and each lymphocyte expresses an endogenous receptor for a tumor associated antigen (TAA) and is genetically engineered to express an additional receptor for a strong antigen derived from a pathogen, wherein the population of dual-specific lymphocytes target a plurality of TAAs and the strong antigen derived from a pathogen.
2. The purified population of autologous lymphocytes of claim 1 , wherein the lymphocytes are selected from the group consisting of CD8+ T cells, CD4+ T cells, and natural killer (NK) cells.
3. The purified population of claim 1 , wherein the autologous lymphocytes are tumor-infiltrating lymphocytes.
4. The purified populations of claim 1 , wherein the lymphocytes are CD8+ T cells.
5. The purified population of claim 1 , wherein the pathogen is selected from the group consisting of listeria monocytogenes, Bacillus Calmette-Guérin, tetanus, diphtheria, adenovirus, herpes simplex virus, vaccinia virus, myxoma virus, poliovirus, vesicular stamatis virus, measles virus, influenza virus, and Newcastle disease virus.
6. The purified population of claim 1 , wherein the strong antigen is an antigen from listeria monocytogenes.
7. The purified population of claim 1 , wherein the lymphocytes are isolated from the tumor of the patient.
8. The purified population of claim 1 , wherein the lymphocytes are isolated from the peripheral blood and optionally are PD-1 positive.
9. The purified population of claim 1 , wherein the genetically engineered additional exogenous receptor is a chimeric receptor.
10. A composition comprising the purified population of dual-specific lymphocytes of claim 1 and a pharmaceutically acceptable carrier.
11. A method of producing an autologous population of dual-specific lymphocytes that can target a plurality of tumor associated antigens and at least one strong antigen derived from a pathogen, the method comprising the steps of:
(a) isolating lymphocytes from a patient;
(b) purifying the tumor-specific lymphocytes from the isolated lymphocytes; and
(c) genetically engineering the purified lymphocytes to express a second receptor specific to a strong antigen derived from a pathogen,
wherein the resulting population comprises dual-specific lymphocytes.
12. The method of claim 11 , wherein step (c) further comprises expanding the isolated tumor-specific lymphocytes in culture.
13. The method of claim 11 wherein step (a) comprises isolating the lymphocytes from a tumor, peripheral blood, or bone marrow of the patient.
14. The method of claim 11 , wherein the pathogen is selected from the group consisting of group of listeria monocytogenes, Bacillus Calmette-Guérin, tetanus, diphtheria, adenovirus, herpes simplex virus, vaccinia virus, myxoma virus, poliovirus, vesicular stamatis virus, measles virus, influenza virus, and Newcastle disease virus..
15. The method of claim 11 , wherein the genetically engineered receptor is a chimeric receptor.
16. The method of claim 11 , wherein the lymphocytes are selected from the group consisting of CD4+ T cells, CD8+ T cells, and natural killer (NK) cells.
17. The method of claim 16 , wherein the lymphocytes are CD8+ T cells.
18. The method of claim 11 , wherein the lymphocytes are stimulated in culture in the presence of an interleukin to stimulate growth.
19. The method of claim 18 , wherein the interleukin is selected from the group consisting of IL-2, IL-7 and IL-15.
20. The method of claim 11 , wherein the lymphocytes are stimulated with the strong antigen or TAA in culture to proliferate.
21. A method of treating a patient with a tumor comprising:
(a) administering to the patient an effective amount of the autologous dual-specific lymphocytes of claim 1 , and (b) injecting the patient with a strong antigen derived from a pathogen, wherein the strong antigen derived from a pathogen activates the autologous dual-specific lymphocytes, thereby treating the patient with the tumor.
22. The method of claim 21 , wherein the autologous dual-specific lymphocytes are injected into the patient intravenously and the strong antigen derived from a pathogen is injected intratumorally.
23. The method of claim 21 , wherein the strong antigen derived from a pathogen is selected from the group consisting of a viral antigen and a bacterial antigen.
24. The method of claim 23 , wherein the pathogen is selected from the group consisting of group of listeria monocytogenes, Bacillus Calmette-Guérin, tetanus, diphtheria, adenovirus, herpes simplex virus, vaccinia virus, myxoma virus, poliovirus, vesicular stamatis virus, measles virus, influenza virus, and Newcastle disease virus.