Primary effect of chemotherapy on the transcription profile of AIDS-related Kaposi's sarcoma

Abstract

Background: Drugs & used in anticancer chemotherapy have severe effects upon the cellular transcription and replication machinery. From in vitro studies it has become clear that these drugs can affect specific genes, as well as have an effect upon the total transcriptome.

Methods: Total mRNA from two skin lesions from a single AIDS-KS patient was analyzed with the SAGE (Serial Analysis of Gene Expression) technique to assess changes in the transcriptome induced by chemotherapy. SAGE libraries were constructed from material obtained 24 (KS-24) and 48 (KS-48) hrs after combination therapy with bleomycin, doxorubicin and vincristine. KS-24 and KS-48 were compared to SAGE libraries of untreated AIDS-KS, and to libraries generated from normal skin and from isolated CD4+ T-cells, using the programs USAGE and HTM. SAGE libraries were also compared with the SAGEmap database.

Results: In order to assess the primary response of AIDS-related Kaposi's sarcoma (AIDS-KS) to chemotherapy in vivo, we analyzed the transcriptome of AIDS-KS skin lesions from a HIV-1 seropositive patient at two time points after therapy. The mRNA profile was found to have changed dramatically within 24 hours after drug treatment. There was an almost complete absence of transcripts highly expressed in AIDS-KS, probably due to a transcription block. Analysis of KS-24 suggested that mRNA pool used in its construction originated from poly(A) binding protein (PABP) mRNP complexes, which are probably located in nuclear structures known as interchromatin granule clusters (IGCs). IGCs are known to fuse after transcription inhibition, probably affecting poly(A)+RNA distribution.Forty-eight hours after chemotherapy, mRNA isolated from the lesion was largely derived from infiltrating lymphocytes, confirming the transcriptional block in the AIDS-KS tissue.

Conclusions: These in vivo findings indicate that the effect of anti-cancer drugs is likely to be more global than up- or downregulation of specific genes, at least in this single patient with AIDS-KS. The SAGE results obtained 24 hrs after chemotherapy can be most plausibly explained by the isolation of a fraction of more stable poly(A)+RNA.

Figure 1

Hematoxylin-eosin (H&E) staining of the AIDS-KS lesions used for SAGE Two skin biopsies were taken from a single patient, respectively 24 hrs (top panel) and 48 hrs (lower panel) after start of chemotherapy with doxorubicin, bleomycin and vincristine. In both biopsies, endothelial cell proliferations are seen in the dermis consistent with plaque stage Kaposi's sarcoma. The elongated nuclei of these cells show moderate atypia. Within the spindle cell areas, narrow slits containing erythrocytes are visible. Immunohistochemistry demonstrated that in both lesions the atypical spindle cells are CD31 and HHV8 positive (not shown). Additionally, in the upper panel, many apoptotic keratinocytes with nuclear dust are found scattered over the basal layer of the epidermis. This histologically visible therapy effect was not conspicuous any more after 48 hours. Both lesions contain sparse inflammatory cell infiltrates.

Figure 2

Clustered display of SAGE gene expression data Cluster tree is based upon an average linkage analysis of the six SAGE libraries [56]. Only tags that appear at least twice in at least one of the libraries have been used.

Figure 3

Effect of chemotherapy in cycling cells. Drug-induced DNA damage, results in G2/M checkpoint-arrest and inhibition of RNA transcription. Twenty-four hours after drug treatment only long-lived mRNA's can be isolated, most likely residing in PABP-mRNP complexes.