Youjin Lee

Therapeutic effects of ablative radiation on local tumor require CD8+ T cells: changing strategies for cancer treatment

Patients with locally advanced cancer or distant metastasis frequently receive prolonged treatment with chemotherapy and/or fractionated radiotherapy (RT). Despite the initial clinical response, treatment resistance frequently develops and cure in these patients is uncommon. Developments in RT technology allow for the use of high-dose (or ablative) RT to target local tumors, with limited damage to the surrounding normal tissue. We report that reduction of tumor burden after ablative RT depends largely on T-cell responses. Ablative RT dramatically increases T-cell priming in draining lymphoid tissues, leading to reduction/eradication of the primary tumor or distant metastasis in a CD8(+) T cell-dependent fashion. We further demonstrate that ablative RT-initiated immune responses and tumor reduction are abrogated by conventional fractionated RT or adjuvant chemotherapy but greatly amplified by local immunotherapy. Our study challenges the rationale for current RT/chemotherapy strategies and highlights the importance of immune activation in preventing tumor relapse. Our findings emphasize the need for new strategies that not only reduce tumor burden but also enhance the role of antitumor immunity.

Intratumor depletion of CD4+ cells unmasks tumor immunogenicity leading to the rejection of late-stage tumors

Tumor environment can be critical for preventing the immunological destruction of antigenic tumors. We have observed a selective accumulation of CD4+CD25+ T cells inside tumors. In a murine fibrosarcoma Ld-expressing Ag104, these cells made up the majority of tumor-infiltrating lymphocytes at the late stage of tumor progression, and their depletion during the effector phase, rather than priming phase, successfully enhanced antitumor immunity. We show here that CD4+CD25+ T cells suppressed the proliferation and interferon-γ production of CD8+ T cells in vivo at the local tumor site. Blockade of the effects of IL-10 and TGF-β partially reversed the suppression imposed by the CD4+ cells. Furthermore, local depletion of CD4+ cells inside the tumor resulted in a change of cytokine milieu and led to the eradication of well-established highly aggressive tumors and the development of long-term antitumor memory. Therefore, CD4+CD25+ T cells maintained an environment in the tumor that concealed the immunogenicity of tumor cells to permit progressive growth of antigenic tumors. Our study illustrates that the suppression of antitumor immunity by regulatory T cells occurs predominantly at the tumor site, and that local reversal of suppression, even at a late stage of tumor development, can be an effective treatment for well-established cancers.

Priming of naive T cells inside tumors leads to eradication of established tumors

The tumor barrier comprised of nonantigenic stromal cells may contribute to the failure of tumor rejection. The tumor-necrosis factor superfamily member LIGHT (also known as TNFSF-14) is a ligand of stromal cell–expressed lymphotoxin-β receptor and T cell–expressed herpes viral entry mediator (HVEM). Here we show that forced expression of LIGHT in the tumor environment induces a massive infiltration of naive T lymphocytes that correlates with an upregulation of both chemokine production and expression of adhesion molecules. Activation of these infiltrating T cells, possibly through HVEM, leads to the rejection of established, highly progressive tumors at local and distal sites. Our study indicates that targeting the tumor barrier may be an effective strategy for cancer immunotherapy.

The Efficacy of Radiotherapy Relies upon Induction of Type I Interferon–Dependent Innate and Adaptive Immunity

The most widely held explanation for the efficacy of local radiotherapy (RT) is based on direct cytotoxicity to cancer cells through the induction of lethal DNA damage. Recent studies have shown that local ablative radiation of established tumors can lead to increased T-cell priming and T-cell-dependent tumor regression, but the underlying mechanism remains unclear. Here, we describe an essential role for type I IFN in local RT-mediated tumor control. We show that ablative RT increases intratumoral production of IFN-β and, more surprisingly, the antitumor effect of RT is abolished in type I IFN nonresponsive hosts. Furthermore, the major target of RT-induced type I IFN is the hematopoietic compartment. RT drastically enhances the cross-priming capacity of tumor-infiltrating dendritic cells (TIDC) from wild-type mice but not type I IFN receptor-deficient mice. The enhanced cross-priming ability of TIDCs after RT was dependent on autocrine production of type I IFNs. By using adenoviral-mediated expression of IFN-β, we show that delivery of exogenous IFN-β into the tumor tissue in the absence of RT is also sufficient to selectively expand antigen-specific T cells leading to complete tumor regression. Our study reveals that local high-dose RT can trigger production of type I IFN that initiates a cascading innate and adaptive immune attack on the tumor.

Differential regulation of CCL21 in lymphoid/nonlymphoid tissues for effectively attracting T cells to peripheral tissues

CC chemokine ligand 21 (CCL21)/secondary lymphoid chemokine (SLC), a ligand for CC chemokine receptor 7 (CCR7), has been demonstrated to play a vital role in the homing and localization of immune cells to lymphoid tissues, but its role in nonlymphoid tissues largely remains undefined. Here, we provide evidence that CCL21 in lymphoid and nonlymphoid tissues is differentially regulated by lymphotoxin-dependent (LT-dependent) and -independent mechanisms, respectively. This differential regulation is due to the selective regulation of the CCL21-Ser/CCL21a but not the CCL21-Leu/CCL21b gene by the LT and noncanonical NF-kappaB pathways. This alternate pathway, not dependent on LT or lymphocytes, leading to constitutive expression of CCL21 in nonlymphoid tissues, is critical for the initial recruitment of T lymphocytes to peripheral effector sites. CCL21 expression is subsequently further enhanced in a LT-dependent fashion following airway challenge, potentially facilitating a positive feedback loop to attract additional CCR7+ effector cells. These findings establish an essential role for CCL21 in the recruitment of effector T cells to peripheral tissues and suggest that LT-dependent and -independent regulation of CCL21 plays a role in balancing the central and peripheral immune responses between lymphoid and nonlymphoid tissues.

Targeting the Primary Tumor to Generate CTL for the Effective Eradication of Spontaneous Metastases

Metastatic disease is the major cause of morbidity and mortality in cancer. Although surgery, chemotherapy, or radiation can often control primary tumor growth, successful eradication of disseminated metastases remains rare. We have now tested whether direct targeting tumor tissues to generate antitumor immune response before surgical excision produces sufficient CTL against micrometastases. One unsolved problem is whether such response allows coming CTL to be educated and then exit the tumor site. Another unsolved problem is whether these CTL can then patrol and effectively eliminate spontaneously metastasized tumor cells in the periphery. In this study, we have shown that adenovirus-expressing TNFSF14 [LIGHT (name derived from homologous to lymphotoxins, shows inducible expression, and competes with herpes simplex virus glycoprotein D for herpes virus entry mediator, a receptor expressed by T lymphocytes); Ad-LIGHT] inoculated directly into primary 4T1 tumor, a highly aggressive, spontaneously metastasizing mammary carcinoma, followed by surgical removal of the primary tumor can eradicate established and disseminated metastatic tumor cells in the peripheral tissues. Furthermore, we clearly show with a fibrosarcoma model Ag104Ld that local treatment can generate plenty of tumor-specific CTL that exit the primary tumor and infiltrate distal tumors to completely eradicate distal tumors. Therefore, targeting the primary tumor with Ad-LIGHT before surgical excision is a new strategy to elicit better immune response for the eradication of spontaneous metastases.

Lymphotoxin is required for maintaining physiological levels of serum IgE that minimizes Th1-mediated airway inflammation.

Although elevated levels of IgE in asthmatic patients are strongly associated with lung infiltration by activated T helper (Th) 2 cells, the physiological role of immunoglobulin E (IgE) in the airway remains largely undefined. Lymphotoxin-deficient α (LTα−/−) mice exhibit increased airway inflammation, paradoxically accompanied by diminished levels of IgE and reduced airway hyperresponsiveness in response to both environmental and induced antigen challenge. The severe lung inflammation in LTα−/− mice is Th1 in nature and can be alleviated by IgE reconstitution. Conversely, depletion of IgE in wild-type mice recapitulates the lung pathologies of LTα−/− mice. Therefore, this work has revealed that lymphotoxin is essential for IgE production, and a physiological role of IgE in the airway may consist of maintaining the balance of Th1 and Th2 responses to prevent aberrant inflammation.

B and T Lymphocyte Attenuator Tempers Early Infection Immunity

Coinhibitory pathways are thought to act in later stages of an adaptive immune response, but whether coinhibition contributes to early innate immunity is unclear. We show that engagement of the newly discovered coinhibitory receptor B and T lymphocyte attenuator (BTLA) by herpesvirus entry mediator (HVEM) is critical for negatively regulating early host immunity against intracellular bacteria. Both HVEM−/− and BTLA−/−, but not LIGHT−/−, mice are more resistant to listeriosis compared with wild-type mice, and blockade of the BTLA pathway promotes, while engagement inhibits, early bacterial clearance. Differences in bacterial clearance were seen as early as 1 day postinfection, implicating the initial innate response. Therefore, innate cell function in BTLA−/− mice was studied. We show that innate cells from BTLA−/− mice secrete significantly more proinflammatory cytokines upon stimulation with heat-killed Listeria. These results provide the first evidence that a coinhibitory pathway plays a critical role in regulating early host innate immunity against infection.

Inhibition of Th2-Mediated Allergic Airway Inflammatory Disease by CD137 Costimulation

The engagement of CD137 (4-1BB), an inducible T cell costimulatory receptor and member of the TNF receptor superfamily, by agonistic Abs can promote strong tumor and viral immunity mediated by CD8+ T cells and stimulate IFN-γ production. However, its role in Th2-mediated immune responses has not been well defined. To address this issue, we studied the function of CD137 engagement using an allergic airway disease model in which the mice were sensitized with inactivated Schistosoma mansoni eggs followed by S. mansoni egg Ag challenge directly in the airways and Th1/2 cytokine production was monitored. Interestingly, treatment of C57BL/6 mice with agonistic anti-CD137 (2A) during sensitization completely prevents allergic airway inflammation, as shown by a clear inhibition of T cell and eosinophil infiltration into the lung tissue and airways, accompanied by diminished Th2 cytokine production and reduced serum IgE levels, as well as a reduction of airway hyperresponsiveness. At various time points after immunization, restimulated splenocytes from 2A-treated mice displayed reduced proliferation and Th2 cytokine production. In accordance with this, agonistic Ab to CD137 can directly coinhibit Th2 responses in vitro although it costimulates Th1 responses. CD137-mediated suppression of Th2 response is independent of IFN-γ and T regulatory cells. Our study has identified a novel pathway to inhibit Th2 responses in a CD137-dependent fashion.