An in vitro model demonstrates the potential of neoplastic human germ cells to influence the tumour microenvironment

Abstract

Testicular germ cell tumours (TGCT) typically contain high numbers of infiltrating immune cells, yet the functional nature and consequences of interactions between GCNIS (germ cell neoplasia in situ) or seminoma cells and immune cells remain unknown. A co-culture model using the seminoma-derived TCam-2 cell line and peripheral blood mononuclear cells (PBMC, n = 7 healthy donors) was established to investigate how tumour and immune cells each contribute to the cytokine microenvironment associated with TGCT. Three different co-culture approaches were employed: direct contact during culture to simulate in situ cellular interactions occurring within seminomas (n = 9); indirect contact using well inserts to mimic GCNIS, in which a basement membrane separates the neoplastic germ cells and immune cells (n = 3); and PBMC stimulation prior to direct contact during culture to overcome the potential lack of immune cell activation (n = 3). Transcript levels for key cytokines in PBMC and TCam-2 cell fractions were determined using RT-qPCR. TCam-2 cell fractions showed an immediate increase (within 24 h) in several cytokine mRNAs after direct contact with PBMC, whereas immune cell fractions did not. The high levels of interleukin-6 (IL6) mRNA and protein associated with TCam-2 cells implicate this cytokine as important to seminoma physiology. Use of PBMCs from different donors revealed a robust, repeatable pattern of changes in TCam-2 and PBMC cytokine mRNAs, independent of potential inter-donor variation in immune cell responsiveness. This in vitro model recapitulated previous data from clinical TGCT biopsies, revealing similar cytokine expression profiles and indicating its suitability for exploring the in vivo circumstances of TGCT. Despite the limitations of using a cell line to mimic in vivo events, these results indicate how neoplastic germ cells can directly shape the surrounding tumour microenvironment, including by influencing local immune responses. IL6 production by seminoma cells may be a practical target for early diagnosis and/or treatment of TGCT.

High levels of CCL2 or CCL4 in the tumor microenvironment predict unfavorable survival in lung adenocarcinoma

“…In a mouse model of peritoneal dissemination of a colon tumor, intraperitoneal injection of hemagglutinating virus of Japan cationic liposomes containing the MIP‐1β gene resulted in local expression of MIP‐1β and local accumulation of CD4 + T lymphocytes and significantly increased survival of the cancer cell‐injected mice, suggesting that CCL4 might serve as a potential therapeutic target against peritoneal disseminated cancer . The properties of tumor microenvironments have a strong impact on cancer progression, response to therapy, and patient prognosis …”

Cytokines in Male Fertility and Reproductive Pathologies: Immunoregulation and Beyond

“…Further work by Klein et al ( 106 ) created a co-culture system to examine the contributions of different cell types to the testis cancer cytokine TME, using TCam-2 cells to represent neoplastic germ cells (seminoma) and peripheral blood monocytic cells (blood-derived immune cells), as a surrogate for testis cancer-infiltrating immune cells. The cytokine pattern observed after 24 h of co-culture was similar to that documented in ex vivo seminoma samples ( 106 ). Intriguingly, this appeared to be initiated by the TCam-2 cells only following direct contact with immune cells, with IL6 measured at significantly higher levels in TCam-2 cells than in PBMCs recovered from the co-cultures.…”

“…With regard to the high expression of IL6 associated with testicular germ cell neoplasia (i.e., seminoma), the possible diagnostic (and/or prognostic) value of corresponding levels in seminal plasma and/or peripheral blood remains to be elucidated ( 61 , 106 ). Proteomics of testicular fluids characterizing different pathologies ( 163 ) may help to identify new organ-specific immunoregulatory molecules or diagnostics to indicate the level of function within the seminiferous epithelium; these may also occur in seminal plasma and/or peripheral blood.…”

“…As discussed above (see section “ Testicular Cancer ”), the involvement of IL6 in testis cancer and the suggested immuno-editing by neoplastic germ cells ( 61 , 100 , 106 ) renders this pro-inflammatory and pro-proliferative molecule a putative therapeutic target. Additional cytokine pathways involved with establishment and maintenance of the tumor microenvironment may be strong candidates for adjunct therapeutics, but much work remains to be done to reach this outcome.…”

Immune mechanisms and possible immune therapy in testicular germ cell tumours

“…This finding corresponds with findings from the analysis of tissue from the previous study. Moreover, the production of IL‐6 was increased in co‐cultured and monocultured cells and did not significantly differ (Klein et al ., ). Based on these findings, IL‐6 may be the central pro‐inflammatory cytokine in the pathogenesis of GCTs.…”