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, wherein the population of dual-specific lymphocytes target a plurality of TAAs and the strong antigen.
2. The purified populations of claim 1 , wherein the strong antigen is a pathogen, a pathogenic antigen, or an alloantigen.
3. The purified population of claim 2 , wherein the strong antigen is an antigen from a pathogen, 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 stomatitis virus, measles virus, influenza virus, and Newcastle disease virus.
4. The composition comprising the purified population of dual-specific lymphocytes of claim 1 and a pharmaceutically acceptable carrier.
5. 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.
6. The method of claim 5 , wherein the autologous dual-specific lymphocytes are injected into the patient intravenously and the strong antigen is injected intratumorally.
7. The method of claim 5 , wherein the strong antigen is selected from the group consisting of a viral antigen, a bacterial antigen, and an alloantigen.
8. The method of claim 7 , wherein the strong antigen is an antigen from a pathogen, 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 stomatitis virus, measles virus, influenza virus, and Newcastle disease virus.
9. 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, 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,
wherein the resulting population comprises dual-specific lymphocytes.
10. The method of claim 9 , wherein step (c) further comprises expanding the isolated tumor-specific lymphocytes in culture.
11. The method of claim 9 , wherein step (a) comprises isolating the lymphocytes from a tumor, peripheral blood or bone marrow of the patient.
12. The method of claim 9 , wherein the strong antigen is selected from the group consisting of a viral antigen, a bacterial antigen, and an alloantigen.
13. The method of claim 12 , wherein the strong antigen is an antigen from a pathogen, 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 stomatitis virus, measles virus, influenza virus, and Newcastle disease virus.
14. The method of claim 9 , wherein the genetically engineered receptor is a chimeric receptor.
15. The method of claim 9 , wherein the lymphocytes are selected from the group consisting of CD4+T cells, CD8+T cells, and natural killer (NK) cells.
16. The method of claim 9 , wherein the lymphocytes are stimulated in culture in the presence of an interleukin to stimulate growth.
17. The method of claim 16 , wherein the interleukin is selected from the group consisting of IL-2, IL-7 and IL-15.
18. The method of claim 9 , wherein the lymphocytes are stimulated with the strong antigen or TAA in culture to proliferate.
19. The method of claim 9 , wherein the genetic engineering comprises transducing the lymphocyte with a gene encoding the receptor to the strong antigen.