Autoimmunity in PD-1-deficient mice

To delineate biological roles of PD-1 in vivo, Nishimura et al generated PD-1 deficient mice by a gene-targeting strategy.10–12 On the C57BL/6 background, the serum level of IgG3 in PD-1-knockout mice was markedly elevated compared with that in control wild-type mice. Aged PD-1 deficient mice developed lupus-like glomerulonephritis. Histological examination of PD-1-deficient mice revealed the deposition of IgG3 and C3 in glomeruli.

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Aged PD-1-deficient mice also developed arthritis with the proliferation of synovial cells histologically resembling rheumatoid arthritis.

When lpr mutation of the Fas gene was introduced into PD-1 deficient mice, they exhibited much more severe glomerulonephritis and arthritis. Moreover, massive lymphadenopathy took place in B6-lpr/lpr × PD-1−/− mice.

When PD-1-deficient mice were backcrossed on BALB/c background, PD-1-deficient mice began to die as early as 5 weeks of age. Two-thirds of PD-1-deficient mice died by 30 weeks of age. Histological examination of the autopsy specimens of PD-1-deficient mice demonstrated that both ventricles were dilated and the right ventricle wall was thin. Functional assessment revealed that the movement of walls of the left ventricles was significantly decreased. IgG was deposited on the cardiomyocytes. Thus, PD-1-deficient mice suffered from autoimmune dilated cardiomyopathy with severely impaired contraction.

These results on the PD-1 deficient mice with the several backgrounds indicated that loss of PD-1 led to the systemic autoimmunity in several organs. Moreover, also in humans, as discussed later in this paper, PD-1 blockade by anti-PD-1 antibody administrations leads to autoimmune adverse events, such as thyroiditis. Therefore, PD-1 is considered to play an important role in patrolling and preventing excessive immune reactions by negatively regulating the functions of autoreactive T-lymphocytes in a physiological state.

Expression of PD-L1 and PD-L2 in malignant tumors and PD-1 in tumor-infiltrating lymphocytes

As we mentioned above, in contrast to the limited expression of PD-1, PD-L1 is expressed in a wide variety of immune cells including activated T-lymphocytes, B-lymphocytes, dendritic cells, and macrophages. PD-L1 is also expressed in non-lymphoid cells or tissues, such as endothelial cells. To explore the expression level and pattern of PD-L1 in cancers, there have been several reports on the expression of PD-L1 in various types of malignant tumors using immunohistochemistry and other techniques.

There was a correlation between the expression of PD-L1 in renal cell carcinoma and stages and prognosis.13

The expression of PD-L1 in esophageal cancer and its association with prognosis was demonstrated by a Japanese group.14 PD-L2-positive patients also had a poorer prognosis than the negative patients.

Also, in gastric cancer, ovarian cancer, urothelial cancer, and pancreatic cancer, there is a correlation between the expression of PD-L1 and tumor stages and prognosis.15–18

Correlation between the expression of PD-L1 in malignant melanoma and tumor stages and prognosis was first studied by the group from Japan.19 Analysis of 59 clinical specimens of malignant melanoma revealed that patients with high PD-L1 immunoreactivity in tumors were at more advanced stage of malignant melanoma and were more likely to die from malignant melanoma than patients with low PD-L1 immunoreactivity. Another study by an Italian group also concluded that high PD-L1 expression in malignant melanoma indicates a poor prognosis.20

Taken together, the high expression of PD-L1 in tumors can be one of the prognostic factors in various malignant tumors indicating a worse survival rate. Moreover, in some tumors like esophageal cancer, high expression of PD-L2 in tumors is also correlated with poor prognosis.

The presence of PD-1-positive tumor-infiltrating lymphocytes is also associated with poor prognosis in various cancers, such as renal cancer and breast cancer.21,22

Anti-PD-1 antibody therapy in mice

To explore the possibility of modulation of immune checkpoint inhibitor PD-1/PD-L1 system for tumor treatment, a Japanese group injected the mouse tumor cell lines into mice and compared the frequencies of tumor formation between wild-type mice and PD-1-deficient mice.23–24 P815 is a murine mastocytoma cell line which does not express PD-L1 or PD-L2. P815 was transfected with an expression PD-L1 vector. Forced expression of PD-L1 in P815 cells made the cells less susceptible to the cytotoxic activity of cytotoxic T-lymphocytes. Moreover, P815 cells with high PD-L1 tended to form tumors when injected into mice.

Another cell line, myeloma J558L cell line, was injected into wild-type or PD-1-deficient mice in a subsequent study. In J558L-injected wild-type mice, tumors grew rapidly, while tumor formation was suppressed in PD-1-deficient mice. The same group also injected B16 melanoma cells into the murine spleen in wild-type or PD-1 transgenic mice and found that tumors grew more rapidly in PD-1 transgenic mice than in wild-type mice.24

These results on PD-1 and PD-L1 in tumor immunity led to a development of a new therapy for human malignant tumors (Figure 2).

(To view a larger version of Figure 2, click here.)