A number of inflammatory cytokines, peptide growth factors, HIV encoded Tat protein, and KSHV/HHV8 gene products contribute to KS cell growth and development.42 HHV8 antigens target cell signaling pathways and deregulate apoptosis and immune response through vCyclin, vFLIP, bcl-2 oncogene, viral interferon regulating factor, and vIL-6.33,43–47

The alterations of immune cells (lymphocytes, monocytes, histiocytes, and dendritic cells) have been suggested to play a role in the neoplastic process.48 Immune activation can cooperate with some growth factors and HIV-1 Tat protein in the development and progression of KS.49

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HIV-KS cells have been shown to produce angiogenic growth factors and cytokines such as fibroblast growth factors (FGFs), tumor necrosis factor-α (TNF-α), interleukin 1 (IL-1), IL-6, Tat, and oncostatin M, and express high affinity receptors for several cytokines.50,51 Elevated levels of IL-1, IL-6, and TNF-α have been reported in patients with HIV-KS.52

Oncostatin M, a cytokine produced by microphages and activated T-lymphocytes, has been shown to be a mitogen for HIV-KS derived spindle cells.53 Oncostatin M appears to be a major cytokine responsible for maintaining the long-term growth of HIV-KS in cell cultures.54

In addition, inflammatory cytokines induce the production of a potent autocrine growth factor for spindle cells known as basic fibroblast growth factor (bFGF).

The autologous production of bFGF is an important stage in KS tumorigenesis since antisense bFGF or anti bFGF antibodies interfere in KS cell growth in tissue culture.42 It has been shown that oncostatin M, IL-1, and TNF-α induce KS cell growth by inducing the expression of various bFGF isoforms.

HIV-associated NHL and its Problems in Diagnosis

NHL refers to a heterogeneous group of hematopoietic malignancies originating in the lymphocytes.55–57The majority of NHL cases (85–90%) arises from B-cell progenitors and develops into the various entities largely grouped into low, intermediate, and high-grade NHL based on the treated natural history and survival patterns.58 NHL comprises many subtypes, each with distinct epidemiology, etiology, and features (ie, morphology, immunophenotype, and clinical manifestations).59,60

Epstein–Barr virus (EBV) has been implicated in the development of many NHL subtypes in HIV-infected individuals.61 NHL is the second most common malignancy in HIV-infected patients, with diffuse large B-cell lymphoma (DLBCL) as the most common subtype of HIV-associated NHL (HIV-NHL) followed by Burkitt’s lymphoma (BL).62

DLBCLs are heterogeneous diseases that differ in nature of the genetic abnormalities, morphologic appearance, clinical features, and patients respond differently to treatment and vary in prognosis.63,64 Most DLBCLs are thought to arise from normal antigen exposed B-cells that have migrated to or through germinal centers.65

Gene expression profiling has identified two broad subgroups: those of germinal center origin, known as germinal centre B-cell like (GCB) lymphomas (typically CD10+ and BCL6+); and those arising from cells resembling activated B-cells (non-GCB) (typically IRF4/MUM-1+/− and CD13+).56,66 It has been shown that patients with GCB DLBCL have a better progression free and overall survival than those with non-GCB DLBCL, irrespective of the international prognostic index (IPI) score.67–71

Therefore, the subclassifications of DLBCL into GCB and non-GCB may serve as important predictive prognostic factors. BL is an aggressive form of NHL derived from germinal center B-cells.72 HIV-associated BL is characterized by cMYC translocations and overexpression;73 however, EBV infection is not necessarily a precursor to transformation.74

NHL is a very complex malignancy consisting of several types that are also divided into subclasses that differ in treatment response and prognosis. This may pose problems in the initial diagnosis of NHL. Biomarkers are necessary in the initial evaluation of the patients with newly diagnosed NHL, which must establish the precise histologic subtype, the extent, and site of disease (localized or advanced, nodal or extranodal). This is important in the determination of treatment approach and predicting the response to chemotherapy.

Biomarkers Used in HIV-NHL Diagnosis/Prognosis

The first step in the diagnosis of NHL is to obtain good quality and adequate sample of tissue by excisional biopsy of an affected lymph node or other mass lesion for assessment of cellular morphology and nodal architecture (Table 2).75–77

After the initial tissue biopsy provides a diagnosis of NHL, the following laboratory tests are performed: complete blood count, white blood cell differential, platelet count, and examination of the peripheral smear for the presence of atypical cells, suggesting peripheral blood and bone marrow involvement; biochemical tests including blood urea nitrogen (BUN), creatinine, alkaline phosphatase, aspartate aminotransferase (AST), alanine aminotransferase (ALT), lactate dehydrogenase (LDH), and albumin; serum calcium, electrolytes, and uric acid; serum protein electrophoresis; HIV, hepatitis B, and C serology; and beta-2 microglobulin levels (in patients with indolent lymphomas).78

TABLE 2. Summary of current biomarkers in HIV-NHL.

LDH Elevated 101,105,106
Ki-67/MIB-1 Elevated 61,79,80
CD19, CD 20, CD 22 Elevated 63,89,90
CD79a Elevated 63,90
PAX-5 Elevated 63,81
CD10 Elevated 61,62
bcI6 Elevated 61,62,65,87
MUM-1 Elevated 61,62,65
cMYC Translocation and Elevated  73,86,87
IL-6 Elevated 97,99
IL-10 Elevated 95,97,99
TNF-α Elevated 87,98
CRP Elevated 99,101
sCD23, sCD27, sCD30, sCD44 Elevated 48,92,99
B2M Elevated 105,106
CXCL13 Elevated 48,92
EBV DNA Elevated 108
FLC Elevated 97,100
FOXP1 Elevated 83,85
Abbreviations: LDH, lactate dehydrogenase; MIB-1, E3 ubiquitin-protein ligase MIB1; PAX-5, paired box protein 5; bcl6, B-cell lymphoma 6; MUM-1, multiple myeloma oncogene 1; IL-6, interleukin 6; IL-10, interleukin 10; TNF-α, tumor necrosis factor-α; CRP, C-reactive protein; sCD23, soluble CD23; sCD44, soluble CD44; B2M, beta-2 microglobulin; CXCL13, C-X-C motif chemokine 13; EBV DNA, Epstein–Barr virus deoxyribonucleic acid; FLC, free immunoglobulin light chains; FOXP1, forkhead box protein P1. 

This is followed by pathological evaluations, which include flow cytometry or immunohistochemical staining for immunophenotype.75

For aggressive lymphomas, this includes evaluation of proliferative fraction using Ki-67 or MIB-1 staining as a more aggressive regimen may be indicated for high growth fraction tumors.77 The expression of Ki-67 has been associated with poor outcome and survival in DLBCL patients.79,80

Immunophenotypic expression patterns of DLBCL include positivity for various pan B-cell markers such as CD19, CD20, CD22, CD79a, PAX-5, and demonstration of immunoglobulin surface light chain restriction by flow cytometry in the majority of cases.63

The presence of positive PAX-5 immunostaining has been strongly associated with B-cell differentiation as PAX-5 is a B-cell restricted transcription factor.81,82 Staining for CD10, bcl-6, and MUM-1 are usually routinely performed in order to distinguish GCB from non-GCB DLBCL.

Fork box protein P1 (FOXP1), an essential transcription regulator of B-cell development, has been shown to be overexpressed in non-GCB DLBCL as compared with GCB DLBCL.71,83,84 FOXP1 has also been associated with poor survival and prognosis.85

It is now recognized that FOXP1 may serve as an additional biomarker for distinguishing non-GCB from GCB DLBCL and should be included in the diagnosis/prognosis of DLBCL.

In addition, fluorescence in situ hybridization (FISH) analysis for cMYC is performed as translocations involving the cMYC occurs in 10–15% of DLBCL lymphomas and is associated with a worse outcome.86 MYC translocations confers a worse prognosis in patients treated with cyclophosphamide, hydroxydaunorubicin, oncovin, and prednisone (CHOP), and CHOP plus rituximab (R-CHOP) regimens.87

BL expresses a germinal center B-cell phenotype,88 and the immunophenotypic expression include B-cell antigens CD19, CD20, CD22, CD79a, and PAX-5 along with CD10, bcl-6, CD77, Ki-67 or MIB-1 and monotypic surface light chains such as IgM.89 BL also expresses CD43, TCL1, and CD38 but is negative for CD5, CD23, CD44, CD138, CD34, and TdT.90,91