Methods of preparing anti-human papillomavirus antigen T cells
Disclosed are methods of preparing an isolated population of human papillomavirus (HPV)-specific T cells comprise dividing an HPV-positive tumor sample into multiple fragments; separately culturing the multiple fragments; obtaining T cells from the cultured multiple fragments; testing the T cells for specific autologous HPV-positive tumor recognition; selecting the T cells that exhibit specific autologous HPV-positive tumor recognition; and expanding the number of selected T cells to produce a population of HPV-specific T cells for adoptive cell therapy. Related methods of treating or preventing cancer using the T cells are also disclosed.
1 . A method of expanding a population of T cells, the method comprising:
(a) culturing multiple fragments of an HPV + tumor sample from a cancer patient in the presence of at least one cytokine;
(b) obtaining T-cells from the cultured fragments;
(c) expanding the number of T-cells to produce an expanded population of HPV-specific T-cells using one or both of (i) irradiated allogenic feeder cells and (ii) irradiated autologous feeder cells; and one or both of (iii) OKT3 antibody and (iv) interleukin-2, wherein the T-cells have not been depleted of CD4 + cells; and
(d) optionally, adding a second culturing step following (c).
2 . The method of claim 1 , wherein the second culturing step is added after step (c) to further expand the number of selected T-cells.
3 . The method of claim 1 , wherein the cytokine in step (a) is IL-2.
4 . The method of claim 3 , wherein the IL-2 in step (a) is present at 6000 IU.
5 . The method of claim 1 , further comprising, prior to step (a), obtaining HPV + tumor samples from the patient and fragmenting said tumor samples.
6 . The method of claim 5 , wherein the tumor samples are fragmented mechanically or enzymatically.
7 . The method of claim 1 , wherein the expanded population of HPV-specific T-cells recognizes HPV 16-positive cervical cancer cells.
8 . The method of claim 1 , wherein the expanded population of HPV-specific cells recognizes an HPV antigen selected from the group consisting of HPV 16 E6 and HPV 16 E7.
9 . The method of claim 1 , wherein the expanded population of HPV-specific cells recognizes an HPV antigen selected from the group consisting of HPV 18 E6 and HPV 18 E7.
10 . The method of claim 1 , wherein the expanded population of cells obtained in (c) comprises multiple T-cells having different HPV specificities.
11 . The method of claim 1 , wherein the expanded population of cells obtained in (c) secretes at least about 200 pg/mL of interferon-gamma.
12 . The method of claim 1 , further comprising combining the expanded population of HPV-specific T-cells with a pharmaceutically acceptable carrier.
13 . The method of claim 1 , wherein the expanded population of HPV-specific T-cells comprises about 1.0×10 10 to 13.7×10 10 HPV-specific T-cells.
14 . The method of claim 12 , wherein the pharmaceutically acceptable carrier is saline, 5% dextrose in water, Ringer's lactate, an electrolyte solution, or PLASMA-LYTE A.
15 . The method of claim 1 , wherein the expanded population of HPV-specific T-cells is enriched for CD4 + T-cells.
16 . The method of claim 1 , wherein the culturing step (a) is performed for at least about 12 days.
17 . The method of claim 1 , wherein the cancer is an HPV + cancer is selected from head and neck squamous cell carcinoma (HNSCC) and cervical cancer.
18 . The method of claim 1 , wherein the IL-2 in step (c) is present at 6000 IU.