The tumour microenvironment influences survival and time to transformation in follicular lymphoma in the rituximab era.

The tumour microenvironment influences outcome in patients with follicular lymphoma (FL), but its impact on transformation is less studied. We investigated the prognostic significance of the tumour microenvironment on transformation and survival in FL patients treated in the rituximab era. We examined diagnostic and transformed biopsies from 52 FL patients using antibodies against CD3, CD4, CD8, CD21 (CR2), CD57 (B3GAT1), CD68, FOXP3, TIA1, PD-1 (PDCD1), PD-L1 (CD274) and PAX5. Results were compared with a second cohort of 40 FL patients without signs of transformation during a minimum of five years observation time. Cell numbers and localization were semi-quantitatively assessed. Better developed CD21+  follicular dendritic cell (FDC) meshworks at diagnosis was a negative prognostic factor for overall survival (OS), progression-free survival (PFS) and time to transformation (TTT) in patients with subsequently transformed FL. Remnants of FDC meshworks at transformation were associated with shorter OS and PFS from transformation. High degrees of intrafollicular CD68+ and PD-L1+  macrophage infiltration, high total area scores and an extrafollicular/diffuse pattern of FOXP3+  T cells and high intrafollicular scores of CD4+  T cells at diagnosis were associated with shorter TTT. Scores of several T-cell subset markers from the combined patient cohorts were predictive for transformation, especially CD4 and CD57.

Higher World Health Organization grades of follicular lymphoma correlate with better outcome in two Nordic Lymphoma Group trials of rituximab without chemotherapy.

Abstract A common treatment for follicular lymphoma is rituximab monotherapy. To identify patients for whom this regimen is adequate as first-line therapy, we applied the World Health Organization (WHO) classification for grading follicular lymphoma in a prospective central pathology review of the biopsies of previously untreated patients in two randomized trials of rituximab without chemotherapy. In the first trial (n1 = 53), higher WHO grades correlated with longer time to next treatment, independently of clinical prognostic factors (p = 0.030); the finding was replicated in the second trial (n2 = 221; p = 0.019). Higher grades were associated with better treatment responses (p = 0.018). Furthermore, also grades externally confirmed by independent local pathologists correlated with time to next treatment (p = 0.048). Flow cytometry in a separate patient series showed that the intensity of CD20 increased with the malignant cell size (p < 0.00005). In conclusion, WHO grade 1 follicular lymphoma correlates with inferior outcome after rituximab monotherapy. WHO grading might provide a clinically useful tool for personalized therapy.

Genomic alterations reveal potential for higher grade transformation in follicular lymphoma and confirm parallel evolution of tumor cell clones.

Our aim was to examine the genetics of clonal evolution in follicular lymphoma (FL) and to identify genetic alterations associated with disease progression. A total of 100 biopsies from 44 patients diagnosed with t(14;18)-positive FL were examined by array comparative genomic hybridization. In 20 patients the patterns of somatic hypermutations (SHMs) in the variable region of heavy chain gene were additionally analyzed. Gain of chromosome X in male samples was a marker for poor outcome (P < .01). Gains involving chromosome 2, 3q, and 5 were exclusively present in FL biopsies from cases with higher grade transformation and were among the copy number alterations (CNAs) associated with inferior survival. Although we noted a trend for increasing genomic complexity in initial versus late FL samples, the overall frequencies of CNAs in initial and late FL biopsies showed a surprisingly stable pattern through the course of the disease. In 27 of cases the initial samples harbored CNAs that were absent in relapse samples, indicating that tumor cell clones at relapse were not direct descendants of initially dominating clones. The pattern of SHMs confirmed parallel development of tumor cell clones in 14 cases. Our findings support the hypothesis of common progenitor cells in FL.

PI3K/Akt-dependent Epo-induced signalling and target genes in human early erythroid progenitor cells.

Erythropoietin (Epo) is the major regulator of differentiation, proliferation and survival of erythroid progenitors, but the Epo-induced changes in gene expression that lead to these effects are not fully understood. The aim of this study was to examine how Epo, via activation of phosphatidylinositol 3-kinase (PI3K)/Akt, exerts its role in the development of erythroid progenitors from CD34+ cells, and to identify early Epo target genes in human erythroid progenitors. In CD34+ progenitor cells, Epo alone was able to induce cell cycle progression as demonstrated by upregulation of cyclin D3, E and A leading to hyperphosphorylation of the retinoblastoma protein (RB). These effects were completely counteracted by the PI3K inhibitor LY294002. Furthermore, enforced expression of an activated form of Akt kinase highly augmented Epo-induced erythropoiesis. Fluorescent-activated cell sorting (FACS)-sorted CD34+CD71+CD45RA-GPA- erythroid progenitors stimulated with Epo in the presence or absence of LY294002 were subjected to gene expression profiling. Several novel target genes of Epo were identified, and the majority were regulated in a PI3K-dependent manner, including KIT (CD117) and CDH1 (E-cadherin). FACS analysis of Epo-stimulated erythroid progenitors showed that the increased mRNA expression of KIT and CDH1 was accompanied by an induction of the corresponding proteins CD117 and E-cadherin.