Impact of Bcl-2 and Ha-ras on keratinocytes in organotypic culture.

In this study, the role of specific molecular alterations associated with multistep skin carcinogenesis was assessed using in vitro organotypic cultures of the spontaneously immortalized, nontumorigenic HaCaT keratinocyte cell line. HaCaT vector control clones and clones expressing bcl-2, activated Ha-ras, or both genes were generated. Clones were induced to stratify and differentiate by culturing on dermal equivalents for 2 wk at the air-medium interface. In parental and vector control HaCaT rafts the expression and distribution of cytokeratin K1, K14, involucrin, proliferating cell nuclear antigen, and p21cip1/waf1 were assessed using immunohistochemistry and immunoblotting and were similar to normal epidermis. Apoptosis was also examined using the TUNEL technique. HaCaT-bcl-2 rafts were similar to control rafts but exhibited lower spontaneous rates of apoptosis and a moderate increase in the rate of proliferation. Differentiation was significantly inhibited in HaCaT-ras organotypic cultures and was associated with high rates of proliferation and lower rates of spontaneous apoptosis. Additionally, HaCaT-ras rafts exhibited significantly higher rates of apoptosis following ultraviolet irradiation compared with vector control or HaCaT-bcl-2 rafts. Bcl-2 was able to largely restore normal differentiation, proliferation, and apoptosis in HaCaT-ras/bcl-2 organotypic cultures. Bcl-2 also abrogated apoptosis induction following ultraviolet irradiation in HaCaT-ras/bcl-2 organotypic cultures. Organotypic keratinocyte culture represents a valuable in vitro system to evaluate the impact of individual molecular genetic alterations on the coordinate regulation of cell proliferation, differentiation, and cell death.

Bax gene disruption alters the epidermal response to ultraviolet irradiation and in vivo induced skin carcinogenesis.

Bcl-2 family member proteins are differentially expressed in skin and in non-melanoma skin cancer (NMSC). To elucidate the contribution of bcl-2 and bax proteins to epidermal differentiation and skin carcinogenesis, we investigated keratinocyte proliferation, differentiation and tumourigenesis in bcl-2(-/-) and bax(-/-) mice. The rate and pattern of proliferation and spontaneous cell death in the bcl-2(-/-) and bax(-/-) mice were not different from control mice. The epidermis of bcl-2(-/-) and bax(-/-) expressed sightly higher levels of cytokeratin 1 and loricrin compared to control littermates. The apoptotic response to ultraviolet-induced genotoxic stress was assessed by quantitating TUNEL positive cells. Bax(-/-) keratinocytes showed a significant resistance to UV-induced cell death compared to control mice. The life-span of bcl-2(-/-) mice precluded an assessment of bcl-2 gene disruption on in vivo tumourigenesis. A significant increase in tumour incidence was observed in bax(-/-) mice compared to control mice in two-step chemical carcinogenesis studies. These findings suggest that bcl-2 and bax gene products may be important determinants of normal keratinocyte differentiation and response to genotoxic stress in vivo, and indicate that bax may provide a tumour suppressor effect during skin carcinogenesis.

Molecular determinants of cell death induction following adenovirus-mediated gene transfer of wild-type p53 in prostate cancer cells.

Adenoviral vectors expressing wild-type p53 (Ad-p53) induce apoptosis in different types of cancer cells. The therapeutic utility of Ad-p53 is now being evaluated in prostate-cancer patients. Bcl-2 is frequently expressed by prostate-cancer cells and has previously been shown to inhibit p53-mediated cell death following genotoxic stress. We studied the impact of bcl-2 on Ad-p53-induced cell death in human prostate-cancer cells. Human prostate-cancer cell lines LNCaP (p53 wt) and PC3 (p53 mut) were stably transfected with bcl-2. After p53 transduction, cell viability, apoptosis induction and modulation of specific apoptosis-regulatory proteins were assessed. LNCaP vector control and bcl-2-expressing cells underwent similar decreases in viability associated with apoptosis induction following Ad-p53 infection. Increased bcl-2 expression provided significant protection to PC3 cells transduced with Ad-p53. These findings are correlated with modulations in bax, bcl-2, bcl-x(L) and p21 protein levels. These data suggest that Ad-p53 may be useful in the treatment of some prostate cancers.

Bcl-2 accelerates multistep prostate carcinogenesis in vivo.

The impact of bcl-2 proto-oncogene expression on the pathogenesis and progression of prostate cancer was examined in a transgenic mouse model. Probasin-bcl-2 transgenic mice were crossed with TRAMP (TRansgenic Adenocarcinoma Mouse Prostate) mice that express the SV40 early genes (T/t antigens) under probasin control. Prostate size, cell proliferation, apoptosis, and the incidence and latency of tumor formation were evaluated. The double transgenic, probasin-bcl-2 X TRAMP F1 (BxT) mice exhibited an increase in the wet weight of the prostate. This was associated with an increase in proliferation, attributable to T/t antigens, and a decrease in apoptosis attributable to bcl-2. The latency to tumor formation was also decreased in the BxT mice compared to the TRAMP mice. The incidence of metastases was identical in both the TRAMP and BxT mice. Lastly, the incidence of hormone-independent prostate cancer was reduced in the BxT mice compared to the TRAMP mice. Together, these results demonstrate that bcl-2 can facilitate multistep prostate carcinogenesis in vivo.

The impact of bcl-2 expression and bax deficiency on prostate homeostasis in vivo.

Prostatic glandular epithelial cells undergo apoptosis in response to androgen-deprivation. The molecular determinants of androgen-responsiveness in these cells are incompletely understood. Recent evidence suggests that bcl-2 gene family members may be important in this context. We used the probasin promoter to target a human bcl-2 transgene specifically to the prostate in order to assess its impact on conferring resistance to androgen withdrawal in, otherwise sensitive, prostatic glandular epithelial cells in vivo. We examined the contribution of bax to mediating androgen-responsiveness in prostatic glandular epithelial cells using bax knockout mice. The histologic appearance of the prostates from probasin-bcl-2 transgenic mice or bax-/- mice did not differ from those of control littermates. There was no evidence of hyperplastic or neoplastic growth. There was no difference between probasin bcl-2 transgenic mice, bax-/- mice, and control littermates in steady-state levels of apoptosis. Following castration our findings suggest that both bax and bcl-2 may each contribute to the androgen-responsiveness of prostatic glandular epithelial cells. It is apparent from these results, however, that bax is not required to mediate cell death in prostatic glandular epithelial cells following castration. A comparison between the apoptotic indices in the ventral prostate from the probasin-bcl-2 and bax-/- mice following castration suggests that the presence of bcl-2 may be a more important indicator of androgen-sensitivity than a deficiency of bax.

Apoptosis and the Bcl-2 gene family -- patterns of expression and prognostic value in stage I and II follicular center lymphoma.

PURPOSE: The prognostic significance of spontaneous levels of apoptosis and Bcl-2, Bax, and Bcl-x protein expression in follicular center lymphoma (FCL) is unknown. The objectives of this retrospective study were (1) to investigate the relationship between pretreatment apoptosis levels and long-term treatment outcome in patients with Stage I and II FCL; (2) to define the incidence and patterns of Bax and Bcl-x protein expression in human FC; and (3) to determine the relationship of Bcl-2, Bax, and Bcl-x expression with spontaneous apoptosis levels and clinical outcome in localized FCL. METHODS AND MATERIALS: Between 1974 and 1988, 144 patients with Stage I or II FCL were treated. Hematoxylin and eosin (H & E) stained tissue sections of pretreatment specimens were retrieved for 96 patients. Treatment consisted of regional radiation therapy (XRT) for 25 patients, combined modality therapy (CMT) consisting of combination chemotherapy and XRT for 57 patients, and other treatments for 14 patients. Median follow-up for living patients was nearly 12 years. The apoptotic index (AI) was calculated by dividing the number of apoptotic cells by the total number of cells counted and multiplying by 100. Expression of Bcl-2, Bax, and Bcl-x proteins was assessed using immunohistochemistry. RESULTS: The mean and median AI values for the entire group were 0.53 and 0.4, respectively (range: 0-5.2). The AI strongly correlated with cytologic grade, with mean AI values of 0.25 for grade 1, 0.56 for grade 2, and 0.84 for grade 3 (p < 0.0005; Kendall correlation). A positive correlation was present between grouped AI and grouped mitotic index (MI) (p = 0.014). For patients treated with CMT, an AI < 0.4 correlated with improved freedom from relapse (FFR) p = 0.0145) and overall survival (OS) (p = 0.0081). An AI < 0.4 did not correlate with clinical outcome for the entire cohort or for patients receiving XRT only. Staining of tumor follicles for the Bcl-2 protein was positive, variable, and negative in 73%, 15%, and 12% of cases, respectively. Positive staining of tumor follicles was observed in 96% of cases for both the Bax and Bcl-x proteins. Expression of Bcl-2, Bax, or Bcl-x did not correlate with AI or clinical outcome. CONCLUSION: The level of spontaneous apoptosis in pretreatment specimens correlates with cytologic grade of FCL and is a significant predictor of FFR and OS for patients with localized FCL receiving CMT.

Altered expression of bcl-2 family member proteins in nonmelanoma skin cancer.

BACKGROUND: Differentiation, proliferation, and cell death are coordinated tightly within the epidermis. Alterations within keratinocytes that disrupt these processes are believed to contribute to the development of nonmelanoma skin cancers (NMSC). In the current study the authors examined the expression of selected members of the bcl-2 gene family in the skin and in case-matched samples of NMSC. METHODS: Immunohistochemistry was performed on tissue sections using antibodies against bcl-2, bcl-x, bax, and bak. Case-matched frozen nonneoplastic skin samples and tumor tissues were used for Western blot analysis. RESULTS: In normal epidermis, bcl-2 oncoprotein is expressed in keratinocytes of the basal layer but is down-regulated in suprabasal layers. The proapoptotic bax protein is expressed at low levels in basal keratinocytes and is up-regulated in suprabasal layers. The bcl-x and bak proteins both are expressed in the basal and spinous strata but are down-regulated in the granular cell layer. Both bcl-2 and bax were diffusely cytosolic whereas bcl-x and bak exhibited a distinct perinuclear distribution. Squamous cell carcinomas (SCC) were negative for bcl-2 whereas bcl-2 increased 5.5-fold in basal cell carcinomas (BCC). The distribution of bcl-x and bax proteins within BCC and SCC overlapped and were associated with squamous differentiation. Bax protein was increased twofold to threefold in NMSC. An increase in bak protein also was observed in SCC. However, bak was diffusely cytosolic within BCC in contrast to the perinuclear distribution in nonneoplastic keratinocytes. CONCLUSIONS: These findings suggest that altered expression of bcl-2 family members may play a role in the pathogenesis of NMSC.

Molecular correlates of bcl-2-enhanced growth following androgen-ablation in prostate carcinoma cells in vivo.

Androgen-independent growth of prostate cancer is correlated with expression of bcl-2. The impact of bcl-2 expression on the growth of prostate cancer cells following androgen ablation, was examined in the androgen-sensitive prostatic carcinoma cell line, LNCaP. Vector control and bcl-2 expressing LNCaP cells were grown subcutaneously in male nude mice. Tumor volume, apoptosis, and proliferation were assessed following castration. The levels of c-myc, p53, p21, bax, and bcl-2 protein were assessed by Western blotting. Bcl-2 expressing tumors exhibited a significant augmentation in growth compared to controls (p 0.01). No difference in the spontaneous rate of proliferation was observed between bcl-2 and control tumors, however, bcl-2 expressing tumors exhibited lower rates of apoptosis. Following orchiectomy the apoptotic index remained significantly lower in bcl-2 expressing tumors (p 0.002 at day 3). The proliferative index was maintained in bcl-2 expressing, but not control tumors following castration. This resulted in a significant growth advantage in bcl-2 tumors subsequent to androgen ablation (p 0.001). These changes were accompanied by alterations in the levels of gene products known to regulate the cell cycle and/or apoptosis. These results emphasize the significance of bcl-2 expression during prostate cancer progression and suggest possible mechanisms for the acquisition of androgen-independent tumor growth.

Bcl-2 expression causes redistribution of glutathione to the nucleus.

In this study we used HeLa cells transfected with a conditional Bcl-2 expression construct to study the effects of Bcl-2 on reduced glutathione (GSH) metabolism. Our previous work demonstrated that depletion of GSH by culturing cells in tissue culture medium lacking the amino acids cysteine and methionine, essential for GSH biosynthesis, caused cells overexpressing Bcl-2 to become sensitized to apoptotic induction. Here we report that Bcl-2 also dramatically alters GSH compartmentalization. Cellular distribution of GSH, assayed by confocal microscopy, revealed that when Bcl-2 expression was suppressed GSH was uniformly distributed primarily in the cytosol, whereas overexpression of Bcl-2 led to a relocalization of GSH into the nucleus. Isolated nuclei readily accumulated radiolabeled GSH and maintained higher nuclear GSH concentration in direct relation to Bcl-2 nuclear protein levels. Moreover, exogenous GSH blocked apoptotic changes and caspase activity in isolated nuclei exposed to the pro-apoptotic protease granzyme B. Our results indicate that one of the functions of Bcl-2 is to promote sequestration of GSH into the nucleus, thereby altering nuclear redox and blocking caspase activity as well as other nuclear alterations characteristic of apoptosis. We speculate that this mechanism contributes to the suppression of apoptosis in cells with elevated Bcl-2 levels.

Prostate carcinoma cell death resulting from inhibition of proteasome activity is independent of functional Bcl-2 and p53.

The ATP/ubiquitin-dependent 26S proteasome is a central regulator of cell cycle progression and stress responses. While investigating the application of peptide aldehyde proteasome inhibitors to block signal-induced IkappaBalpha degradation in human LNCaP prostate carcinoma cells, we observed that persistent inhibition of proteasomal activity signals a potent cell death program. Biochemically, this program included substantial upregulation of PAR-4 (prostate apoptosis response-4), a putative pro-apoptotic effector protein and stabilization of c-jun protein, a potent pro-death effector in certain cells. We also observed modest downregulation of bcl-XL, a pro-survival effector protein. However, in contrast to some recent reports stable, high level, expression of functional bcl-2 protein in prostate carcinoma cells failed to signal protection against cell death induction by proteasome inhibitors. Also in disagreement to a recent report, no evidence was found for activation of the JNK stress kinase pathway. A role for p53, a protein regulated by the proteasome pathway, was ruled out, since comparable cell death induction by proteasome inhibitors occurred in PC-3 cells that do not express functional p53 protein. These data signify that the ubiquitin/proteasome pathway represents a potential therapeutic target for prostate cancers irrespective of bcl-2 expression or p53 mutations.

Expression of bcl-2 oncoprotein and p53 protein accumulation in bone marrow metastases of androgen independent prostate cancer.

PURPOSE: We correlated the expression of bcl-2 with accumulation of p53 protein in bone marrow metastases from patients with androgen independent prostate cancer and a history of hormonal ablation therapy. These results were correlated with clinical parameters, including the extent of bone marrow metastases and patient survival. MATERIALS AND METHODS: All 43 patients studied had evidence of prostate cancer progression following androgen deprivation therapy and histologically confirmed bone marrow metastases. Decalcified tissue sections were used for immunohistochemical evaluation of bcl-2 protein and p53 protein accumulation. RESULTS: We previously established that p53 protein accumulation as detected by immunohistochemistry is a reliable indicator of p53 gene mutation in prostate cancer. Immunoreactivity was demonstrated for p53 protein in 22 of 43 cases and for bcl-2 protein in 14. A total of 28 cases (65%) exhibited immunohistochemical evidence of p53 and/or bcl-2 expression, and 15 (35%) were negative for p53 and bcl-2. The expression of bcl-2 and accumulation of p53 were independent events (p < 0.01). The expression of bcl-2 or accumulation of p53 protein in prostate cancer metastases did not significantly influence patient survival or the extent of metastatic disease. CONCLUSIONS: The presence or absence of p53 protein accumulation and/or bcl-2 expression did not correlate with tumor burden or patient survival in stage D androgen independent prostate cancer bone marrow metastases. The expression of bcl-2 protein occurs independently of and is inversely correlated with p53 mutations in advanced prostate cancer.

Bcl-2 inhibits p53 nuclear import following DNA damage.

Bcl-2 is an integral membrane oncoprotein that localizes to membranes of the mitochondria, endoplasmic reticulum, and nuclear envelope. Bcl-2 is a member of a family of cell death regulators and functions to inhibit apoptosis. Using confocal microscopy and immunoblotting we show that the ability of bcl-2 to suppress cell death following genotoxic damage can be a consequence of inhibiting nuclear import of induced wild-type p53 protein. Our data suggests that the ability of bcl-2 to modulate trafficking events is not cell type specific. These data support a 'gatekeeper' mechanism for cell death suppression by bcl-2.

bcl-2 expression confers androgen independence in androgen sensitive prostatic carcinoma.

Expression of the bcl-2 proto-oncogene is associated with the progression of prostate cancer to androgen-independence. Dunning R3327G (DG) cells were engineered to express high levels of bcl-2 protein. The parental DG (DG-P) cell line and bcl-2 transfectant (DG-B) clones were grown as subcutaneous tumor explants in male athymic nude mice. The rate of tumor growth after castration was significantly lower in DG-P tumors but was unaffected in DG-B tumors. The proliferative indices (PI) in DG-P and DG-B tumors were similar, however, apoptotic indices (ApI) were significantly lower in DG-B tumors before castration. Following castration the PI and ApI decreased significantly in DG-P but not DG-B tumors. Bax upregulation was not observed in the DG-P or DG-B tumors, but did occur in the ventral prostate, after castration. These findings support a role for bcl-2 expression in conferring androgen-independent growth during prostate cancer progression.

Bcl-2 suppresses apoptosis resulting from disruption of the NF-kappa B survival pathway.

A role has been delineated for both bcl-2 and NF-kappa B in mediating an adaptive survival response to the TNF-alpha signaling pathway for apoptosis. Additionally, we and others have demonstrated a role for bcl-2 upregulation during progression of prostate cancer and acquisition of androgen-independent growth (T. J. McDonnell et al., 1992, Cancer Res. 52, 6940-6944). Therefore, the relationship between bcl-2 and NF-kappa B in regulating TNF-alpha-induced apoptosis was investigated in prostate carcinoma cells. Enforced overexpression of bcl-2 protein in prostatic carcinoma cells impaired TNF-alpha-mediated cytotoxicity. Expression of bcl-2 did not impose a block to, or potentiate, TNF-alpha signaling of I kappa B alpha degradation, nuclear import of the RelA p65, or NF-kappa B-dependent transactivation. Expression of two dominant-negative I kappa B alpha mutant proteins significantly enhanced TNF-alpha-induced apoptosis in control cells but not in cells expressing high levels of bcl-2 protein. Similarly, PDTC, a strong antioxidant that interferes with activation of NF-kappa B in these prostate carcinoma cells, also potentiated TNF-alpha-stimulated apoptosis signaling through a bcl-2-regulated mechanism. These findings indicate that modulation of NF-kappa B survival signaling may be used to clinical advantage in the treatment of prostate cancer patients. The efficacy of strategies proposed to enhance TNF-alpha-mediated cytotoxicity by inhibiting NF-kappa B will likely be influenced by context-dependent variables such as bcl-2 expression.

Apoptosis suppression by bcl-2 is correlated with the regulation of nuclear and cytosolic Ca2+.

Bcl-2 expression is associated with the progression of prostate cancer from androgen-dependence to androgen-independence. Bcl-2 is an integral membrane protein which localizes to mitochondria, endoplasmic reticulum, and the nuclear envelope. Using spectrofluorometry and laser confocal microscopy, the ability of bcl-2 to modulate intracellular Ca2+ was examined in the Dunning G prostate carcinoma cell line following apoptosis induction by adriamycin. Adriamycin and thapsigargin, an endoplasmic reticulum Ca2+-pump inhibitor, were effective inducers of apoptosis in control, but not bcl-2 transfected, cells. Treatment with adriamycin was accompanied by a sustained rise in cytoplasmic Ca2+ in control and bcl-2 transfected cells. An increase in intranuclear Ca2+ was observed in control cells only. Apoptosis induction by thapsigargin was associated with an increase in cytoplasmic Ca2+ in control cells that was not detected in the resistant bcl-2 transfectants. Ca2+ was excluded from nuclei isolated from bcl-2 expressing cells, but was sequestered in control nuclei, following the addition of ATP. These findings suggest that bcl-2 may regulate levels of intranuclear Ca2+ independently of cytosolic Ca2+ levels. The ability of bcl-2 to modulate, directly or indirectly, sustained increases in both cytosolic and intranuclear Ca2+ may provide a common basis for bcl-2 function in different subcellular compartments.

Evidence that c-myc mediated apoptosis does not require wild-type p53 during lymphomagenesis.

Deregulation of c-myc, frequently implicated in oncogenesis, is associated with increased cell proliferation and also cell death. Similarly, the p53 tumor suppressor gene commonly mutated in human tumors, is known to induce apoptosis or cell cycle arrest in its wild-type conformation. Genetically altered mice simultaneously overexpressing c-myc and possessing a disrupted p53 gene were used to investigate whether c-myc mediated apoptosis requires wild-type p53. The accelerated development of malignant lymphomas in these mice was found to be a consequence of enhanced proliferation and not reduced apoptosis resulting from the synergistic effect of c-myc overexpression and p53 inactivation.

The expression and localization of bcl-2 protein in normal skin and in non-melanoma skin cancers.

Non-melanoma skin cancers are the most commonly diagnosed malignancies and are typically indolent in their clinical behavior. Although predisposing factors leading to the development of these cancers, such as ultraviolet irradiation, are well described, the molecular events involved in their pathogenesis are incompletely understood. The localization of bcl-2 expression within the skin was determined using immunohistochemical methodologies and an anti-bcl-2 monoclonal antibody. The cytoarchitectural distribution of bcl-2 protein in normal skin included basal keratinocytes, the dermal papillae of the hair follicle, the keratinized Huxley's and Henle's layers, and the keratinized outer root sheath cells of the isthmus and infundibulum of the hair follicle. In addition, intense immunoreactivity was noted in the secretory coil of eccrine sweat glands. The distribution of bcl-2 protein within normal skin did not correlate with the known histologic localization of stem cell compartments. Basal cell carcinomas expressed high levels of bcl-2 protein. In contrast, squamous cell carcinomas typically exhibited no immunohistochemically detectable bcl-2 protein. The findings suggest a potential contribution of bcl-2 gene deregulation to the pathogenesis of some types of non-melanoma skin cancer.

The functional basis of c-myc and bcl-2 complementation during multistep lymphomagenesis in vivo.

Oncogenes are known to be deregulated by chromosomal translocations occurring at high frequency in specific malignancies. Among the most well characterized of these are c-myc, associated with the t(8;14) in Burkitt's lymphomas, and bcl-2, associated with the t(14;18) in follicular lymphomas. In addition to their role in regulating rates of proliferation, it is known that oncogenes and tumor suppressor genes can also regulate rates of apoptotic cell death. The contribution of c-myc and bcl-2 to the regulation of cell death during lymphomagenesis in vivo is assessed using bcl-2-Ig and emu-myc trangenic mice and bcl-2/myc hybrid transgenic mice. Translocations between the endogenous c-myc gene and immunoglobulin loci, e.g., t(12;15), are common in lymphomas arising in the bcl-2-Ig mice. Furthermore, bcl-2/c-myc double transgenic mice exhibit accelerated lymphomagenesis, indicating cooperation between these two oncogenes. Genetic complementation of c-myc and bcl-2 during lymphomagenesis resulted from the suppression of c-myc-associated apoptosis. Other genes are likely involved in regulating cell death during multistep lymphomagenesis.

Apoptosis and expression of the bcl-2 proto-oncogene in the fetal and adult human kidney: evidence for the contribution of bcl-2 expression to renal carcinogenesis.

bcl-2 was identified as a transcript associated with the t(14;18) and imparts resistance to apoptotic cell death. bcl-2 is normally expressed in many tissues and exhibits remarkable structural and functional conservation. Using immunohistochemical and in situ DNA labeling techniques we examined the localization of bcl-2 in the developing human kidney. bcl-2 expression was rapidly upregulated in the induced metanephrogenic mesenchymal cells that differentiate into renal vesicles and nephrons. bcl-2 expression was undetectable in uninduced mesenchyme and in the renal ampullae and associated collecting system. The distribution of apoptotic cells within the developing kidney was inversely correlated with expression of bcl-2. The localization of bcl-2 protein in the adult human kidney also was examined. bcl-2 was expressed at high levels in all renal neoplasms examined providing a potential basis for the deregulation of apoptosis in the development and progression of these tumors.

The bcl-2 oncogene: apoptosis and neoplasia.

Cloning of the t(14;18) translocation breakpoint resulted in the identification of a new putative oncogene, which mapped to 18q21, termed bcl-2. The t(14;18) resulted in inappropriately high levels of bcl-2 expression in follicular lymphoma. Prospective studies using mice transgenic for a human bcl-2-immunoglobulin minigene, intended to recreate the molecular features of the t(14;18), demonstrated that bcl-2 gene deregulation was oncogenic. Interestingly, overexpression of bcl-2 showed no demonstrable influence on rates of cellular proliferation. Rather, bcl-2 was found to extend cellular viability by blocking apoptosis. Recent studies with other oncogenes and tumor suppressor genes, such as c-myc and p53, have demonstrated that the deregulation of apoptosis may be of general significance in the development of multiple types of cancer and appears to be a critical event during multistep carcinogenesis. The selective induction of apoptosis in tumor cell populations is now being considered in the design of novel therapeutic interventions.