PMC8620941_01
Target antigen expression at various treatment time points.. CD20, CD30 and CD19 expression at diagnosis, post-rituximab or post-brentuximab/pre-CART19 and Post-CART19 infusion are shown. CD20 and CD30 are lost after rituximab and brentuximab respectively while CD19 expression changes after CART-19 therapy. The cytology and morphology of tumor was similar across the three time points.
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F1
Biopsy showed diffuse infiltration by large mature B cells (CD20+, CD19+, CD30subset+) with abundant cytoplasm and delicate fibrosis consistent with involvement by PMLBCL (Figure 1; column a). Follow up imaging revealed new renal and pancreatic lesions that were confirmed by biopsy to be additional involvement by PMLBCL which was now negative for CD20 (Figure 1; column b, row i). A subsequent biopsy showed predominantly CD30-negative PMLBCL (Figure 1; column b, row ii). Interestingly, CD20 and CD30 were positive in a subset of these tumor cells (Figure 1 column c row ii). Sequencing of the CD19 gene in the tumor identified a novel heterozygous missense mutation (G210D) in exon 4 that substitutes glycine, a highly conserved amino acid localized in the extracellular domain of CD19, with a negatively charged aspartic acid (supplemental figure 1A and 1B). Fig. 1C and 1D).
Loss of membrane expression of CD19 post CART19 therapy.. CD19 immunostain and flow cytometric data from pre and post-CART therapy biopsies are shown. CD19 expression changes from strong membranous pattern to dim cytoplasm pattern (panel a). Flow cytometric evaluation of surface CD19 reveals loss of CD19 but intact CD20 after CART therapy (panel b).
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F2
However, two months after CART-19 infusion, she developed renal insufficiency and a renal biopsy revealed persistent involvement by PMLBCL that was remarkable for absence of surface CD19 by flow cytometry but positive for cytoplasmic expression by immunohistochemistry (Figure 2). Immunostaining for mismatch repair proteins confirmed the loss of MSH2 and MSH6 expression with intact MLH1 and PMS2 expression (supplemental figure 2).