Diagnostic biopsy does not commonly induce intratumoral CD8 T cell infiltration in Merkel cell carcinoma

Abstract

Background: Merkel cell carcinoma is a polyomavirus-associated cancer that is strongly linked with T lymphocyte immune suppression in epidemiologic studies. CD8+ T cell infiltration into MCC tumors (intratumoral) has recently been shown to be strongly predictive of improved survival. In contrast, the presence of CD8+ T cells at the border of the tumor (peritumoral) had no independent prognostic value. Spontaneous regression has been reported for MCC approximately one thousand times more often than would be expected given the frequency of this cancer. Many of these events began shortly after biopsy, and in some cases lymphocytic infiltration was described.

Methodology/principal findings: To determine whether CD8+ lymphocyte infiltration in MCC tumors is commonly altered by biopsy.33 MCC patients who had microscopic confirmation of MCC on both an initial biopsy and a re-excision specimen were included in this study. Intratumoral and peritumoral CD8 lymphocyte infiltration was quantitated using immunohistochemistry and compared using the paired t-test in biopsy versus re-excision samples. There was a trend toward increased CD8 infiltration after biopsy in a peritumoral ('stalled') pattern (p = 0.08), however, biopsy was not associated with a significant increase in CD8 T cells in the clinically more important intratumoral location (p = 0.58).

Conclusions/significance: The initial diagnostic biopsy for MCC does not commonly alter intratumoral CD8+ T cell infiltration, suggesting it does not directly induce immunologic recognition of this cancer. Because CD8 infiltration is typically stable after biopsy, this parameter may be useful to assess the efficacy of future immune therapies for this virus-associated, immunogenic, often-lethal cancer.

Figure 1. Peritumoral and intratumoral CD8 lymphocyte infiltration at biopsy and re-excision of a representative MCC tumor.

Top row: Biopsy specimen stained with H/E (a) and á-CD8 (b) from case # w313. Bottom row: Re-excision section stained with H/E (c) and á-CD8 (d) from the same case. Peritumoral and intratumoral CD8 lymphocytic infiltrates were each scored on a 0 to 5 scale as described. These images correspond to peritumoral CD8 score of 3 and intratumoral CD8 score of 0 at biopsy, and peritumoral CD8 score of 4 and intratumoral CD8 score of 1 at re-excision. CD8 scores represent average peri-/intra-tumoral infiltration across many more fields than are visible in the figure. Importantly, CD8 cells that are in contact with stroma are not considered to be intratumoral. The black dashed lines in á-CD8 images indicate the tumor/stromal interface (intratumoral: IT, peritumoral: PT). Brown cells indicated by arrows in panel d) are CD8+ cells in a true intra-tumoral location. Scalebar 100 uM. Abbreviations: H/E, hematoxylin and eosin.

Figure 2. CD8 cell infiltration of paired biopsy and re-excision specimens.

Each circle represents a pair of specimens from an MCC patient. Fig. 2a shows the extent of peritumoral CD8 infiltration (n = 31), and Fig. 2b represents intratumoral CD8 infiltration (n = 33; two cases had insufficient tissue to evaluate peritumoral infiltration). Cases for which the two specimens were identical in CD8 infiltration are depicted along the diagonal that is not shaded (marked with “no change after Bx”). Cases that had an increase in infiltration after biopsy are in the top left (green shaded) area. Cases with a decrease in infiltration after biopsy are in the lower right (red shaded) area. Abbreviations: Bx, biopsy. There was no statistically significant difference in CD8 infiltration for either location, intratumoral (P-value = 0.08) or peritumoral (P-value = 0.58) via paired-t test.