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Metabolic reprogramming is a hallmark of cancer, crucial for malignant transformation and metastasis. Chronic lymphocytic leukaemia (CLL) and prostate cancer exhibit similar metabolic adaptations, particularly in glucose and lipid metabolism. Understanding this metabolic plasticity is crucial for identifying mechanisms contributing to metastasis. This review considers glucose and lipid metabolism in CLL and prostate cancer, exploring their roles in healthy and malignant states and during disease progression. In CLL, lipid metabolism supports cell survival and migration, with aggressive disease characterised by increased fatty acid oxidation and altered sphingolipids. Richter's transformation and aggressive lymphoma, however, exhibit a metabolic shift towards increased glycolysis. Similarly, prostate cell metabolism is unique, relying on citrate production in the healthy state and undergoing metabolic reprogramming during malignant transformation. Early-stage prostate cancer cells increase lipid synthesis and uptake, and decrease glycolysis, whereas metastatic cells re-adopt glucose metabolism, likely driven by interactions with the tumour microenvironment. Genetic drivers including TP53 and ATM mutations connect metabolic alterations to disease severity in these two malignancies. The bone microenvironment supports the metabolic demands of these malignancies, serving as an initiation niche for CLL and a homing site for prostate cancer metastases. By comparing these malignancies, this review underscores the importance of metabolic plasticity in cancer progression and highlights how CLL and prostate cancer may be models of circulating and solid tumours more broadly. The metabolic phenotypes throughout cancer cell transformation and metastasis, and the microenvironment in which these processes occur, present opportunities for interventions that could disrupt metastatic processes and improve patient outcomes., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2025 The Authors. Published by Elsevier B.V. All rights reserved.)

Chronic lymphocytic leukemia (CLL) is a malignancy of mature B cells, and it is the most frequent form of leukemia diagnosed in Western countries. It is characterized by the proliferation and accumulation of neoplastic B lymphocytes in the blood, lymph nodes, bone marrow and spleen. We report the synthesis and antiproliferative effects of a series of novel ethanoanthracene compounds in CLL cell lines. Structural modifications were achieved via the Diels-Alder reaction of 9-(2-nitrovinyl)anthracene and 3-(anthracen-9-yl)-1-arylprop-2-en-1-ones (anthracene chalcones) with dienophiles, including maleic anhydride and N -substituted maleimides, to afford a series of 9-( E )-(2-nitrovinyl)-9,10-dihydro-9,10-[3,4]epipyrroloanthracene-12,14-diones, 9-( E )-3-oxo-3-phenylprop-1-en-1-yl)-9,10-dihydro-9,10-[3,4]epipyrroloanthracene-12,14-diones and related compounds. Single-crystal X-ray analysis confirmed the structures of the novel ethanoanthracenes 23f , 23h , 24a , 24g , 25f and 27 . The products were evaluated in HG-3 and PGA-1 CLL cell lines (representative of poor and good patient prognosis, respectively). The most potent compounds were identified as 20a , 20f, 23a and 25n with IC 50 values in the ranges of 0.17-2.69 µM (HG-3) and 0.35-1.97 µM (PGA-1). The pro-apoptotic effects of the potent compounds 20a , 20f, 23a and 25n were demonstrated in CLL cell lines HG-3 (82-95%) and PGA-1 (87-97%) at 10 µM, with low toxicity (12-16%) observed in healthy-donor peripheral blood mononuclear cells (PBMCs) at concentrations representative of the compounds IC 50 values for both the HG-3 and PGA-1 CLL cell lines. The antiproliferative effect of the selected compounds, 20a , 20f, 23a and 25n, was mediated through ROS flux with a marked increase in cell viability upon pretreatment with the antioxidant NAC. 25n also demonstrated sub-micromolar activity in the NCI 60 cancer cell line panel, with a mean GI 50 value of 0.245 µM. This ethanoanthracene series of compounds offers potential for the further development of lead structures as novel chemotherapeutics to target CLL.

Chronic lymphocytic leukaemia (CLL) is a malignancy of the immune B lymphocyte cells and is the most common leukaemia diagnosed in developed countries. In this paper, we report the synthesis and antiproliferative effects of a series of ( E )-9-(2-nitrovinyl)anthracenes and related nitrostyrene compounds in CLL cell lines and also in Burkitt's lymphoma (BL) cell lines, a rare form of non-Hodgkin's immune B-cell lymphoma. The nitrostyrene scaffold was identified as a lead structure for the development of effective compounds targeting BL and CLL. The series of structurally diverse nitrostyrenes was synthesised via Henry-Knoevenagel condensation reactions. Single-crystal X-ray analysis confirmed the structure of ( E )-9-chloro-10-(2-nitrobut-1-en-1-yl)anthracene ( 19f ) and the related 4-(anthracen-9-yl)-1 H -1,2,3-triazole ( 30a ). The ( E )-9-(2-nitrovinyl)anthracenes 19a, 19g and 19i-19m were found to elicit potent antiproliferative effects in both BL cell lines EBV - MUTU-1 (chemosensitive) and EBV + DG-75 (chemoresistant) with >90% inhibition at 10 μM. Selected ( E )-9-(2-nitrovinyl)anthracenes demonstrated potent antiproliferative activity in CLL cell lines, with IC 50 values of 0.17 μM (HG-3) and 1.3 μM (PGA-1) for compound 19g . The pro-apoptotic effects of the most potent compounds 19a , 19g , 19i , 19l and 19m were demonstrated in both CLL cell lines HG-3 and PGA-1. The ( E )-nitrostyrene and ( E )-9-(2-nitrovinyl)anthracene series of compounds offer potential for further development as novel chemotherapeutics for CLL.

Peripheral blood and bone marrow aspirates are routinely obtained from blood cancer patients for diagnostic investigations and provide an accessible source of patient-specific cancer cells, as well as non-malignant cells, for research proposes. The simple and reproducible method presented here allows isolation of viable mononuclear cells, including malignant cells, from fresh peripheral blood or bone marrow aspirates using density gradient centrifugation. The cells obtained using the protocol described can be further purified for a variety of cellular, immunological, molecular, and functional assays. In addition, these cells can be cryopreserved and bio-banked for future research studies., (© 2023. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

This study was carried out to assess the prognostic power of low CD49d expression (≥10%) in newly diagnosed CLL patients using a previously described cohort. Eighty-five patients were included. Median age at diagnosis; 70 years (43-88); CD49d was expressed in 33/85 (38.8%); 23/33 (69.7%) at ≥30% referred to as 'HiCD49d' and 10/33 (30.3%) between 10 and 30% with a bimodal pattern on scatterplot analysis referred to as 'LoCD49d'. Eleven patients (12.9%) presented as Binet stage B, of whom 8 (72.7%) were CD49d+ (HiCD49d 7/8; LoCD49d 1/8). Seven of 81 patients (8.6%) were NOTCH1 mutated and all were CD49d+ (p ≤ .01). IgVH analysis was performed on 29 (87.8%) of the CD49d+ cases, of whom 21 (72.4%) were unmutated and 8 (27.6%) were mutated. CD38+/CD49d+ accounted for 11/20 (55%) (CD38+/HiCD49D: 9/11; CD38+/LoCD49D: 2/11). At 42 months, treatment had been initiated in 18/85 (21%) patients, of these 10/33 (30.3%) were CD49d+ versus 8/52 (15.4%) of the CD49d- group. The median treatment free interval for the CD49d+ group was 11 months (HiCD49d; 14.5 months, LoCD49d; 11 months) compared to 21.5 months for the CD49d- group. These findings suggest that the predictive value of CD49d expression is retained at expression levels down to 10%., (© 2022 The Authors. European Journal of Haematology published by John Wiley & Sons Ltd.)

Aims: We have recently described a novel guanidinium-based compound, VP79s, which induces cytotoxicity in various cancer cell lines. Here, we aim to investigate the activity of VP79s and associated mechanisms of action in multiple myeloma (MM) cells in vitro and ex vivo., Main Methods: The effects of VP79s on cell viability and induction of apoptosis was examined in a panel of drug-sensitive and drug-resistant MM cell lines, as well as ex vivo patient samples and normal donor lymphocytes and platelets. Cell signaling pathways associated with the biological effects of VP79s were analysed by immunoblotting and flow cytometry. Gene expression changes were assessed by quantitative real-time PCR analysis., Key Findings: VP79s was found to rapidly inhibit both constitutively active and IL-6-induced STAT3 signaling with concurrent downregulation of the IL-6 receptors, CD130 and CD126. VP79s induced a rapid and dose-dependent downregulation of anti-apoptotic Bcl-2 family member, myeloid cell leukaemia-1 (MCL-1). VP79s enhanced bortezomib induced cell death and was also found to overcome bone marrow stromal cell induced drug resistance. VP79s exhibited activity in ex vivo patient samples at concentrations which had no effect on peripheral blood mononuclear cells, lymphocytes and platelets isolated from healthy donors., Significance: As VP79s resulted in rapid inhibition of the key IL-6/STAT3 signaling pathway and downregulation of MCL-1 expression with subsequent selective anti-myeloma activity, VP79s may be a potential therapeutic agent with a novel mechanism of action in MM cells., (Copyright © 2021 Elsevier Inc. All rights reserved.)

We previously identified a guanidinium-based lead compound that inhibited BRAF through a hypothetic type-III allosteric mechanism. Considering the pharmacophore identified in this lead compound (i.e., "lipophilic group", "di-substituted guanidine", "phenylguanidine polar end"), several modifications were investigated to improve its cytotoxicity in different cancer cell lines. Thus, several lipophilic groups were explored, the di-substituted guanidine was replaced by a secondary amine and the phenyl ring in the polar end was substituted by a pyridine. In a structure-based design approach, four representative derivatives were docked into an in-house model of an active triphosphate-containing BRAF protein, and the interactions established were analysed. Based on these computational studies, a variety of derivatives was synthesized, and their predicted drug-like properties calculated. Next, the effect on cell viability of these compounds was assessed in cell line models of promyelocytic leukaemia and breast, cervical and colorectal carcinomas. The potential of a selection of these compounds as apoptotic agents was assessed by screening in the promyelocytic leukaemia cell line HL-60. The toxicity against non-tumorigenic epithelial MCF10A cells was also investigated. These studies allowed for several structure-activity relationships to be derived. Investigations on the mechanism of action of representative compounds suggest a divergent effect on inhibition of the MAPK/ERK signalling pathway.

The nitrostyrene scaffold was previously identified as a lead target structure for the development of effective compounds targeting Burkitt's lymphoma. The present study aimed to develop these compounds further in haematological malignancies, including chronic lymphocytic leukaemia (CLL). Cellular viability, flow cytometry and lactate dehydrogenase assays, amongst others, were used to examine the effects of nitrostyrene compounds on CLL cells, including a cell line representing disease with poor prognosis (HG‑3) and in ex vivo CLL patient samples (n=14). The results demonstrated that two representative nitrostyrene compounds potently induced apoptosis in CLL cells. The pro‑apoptotic effects of the compounds were found to be reactive oxygen species and caspase‑dependent, and had minimal effects on the viability of normal donor peripheral blood mononuclear cells. Nitrostyrene compounds exhibited synergistic augmentation of apoptosis when combined with the phosphatidylinositol 3‑kinase inhibitor idelalisib and demonstrated potent toxicity in ex vivo CLL cells, including those co‑cultured with bone marrow stromal cells, making them more resistant to apoptosis (n=8). These compounds also demonstrated activity in samples from patients with poor prognostic indicators; unmutated immunoglobulin heavy chain genes, expression of CD38 and deletions in chromosomes 17p and 11q. These results suggest that this class of pharmaceutically active compounds offer potential in the treatment of CLL.

Invariant natural killer T (iNKT) cells are cytotoxic T cells that respond to glycolipid antigens presented by CD1d. Therapeutic activation of iNKT cells with α-galactosylceramide (α-GalCer) can prevent and reverse tumor growth in mice and clinical trials involving α-GalCer-stimulated iNKT cells are ongoing in humans. B cells express CD1d, however, we show that CD1d expression is reduced on B cells from patients with chronic lymphocytic leukemia (CLL). B cells from CLL patients pulsed with α-GalCer failed to stimulate cytolytic degranulation by iNKT cell lines, but could present the more potent glycolipid analogue, 7DW8-5. Retinoic acid receptor-α (RAR-α) agonists induced CD1d expression by CLL B cells, restoring their ability to present α-GalCer to CD8α + iNKT cells, resulting in cytolytic degranulation. Thus, RAR-α agonists can augment the anti-tumor activities of iNKT cells against CLL cells in vitro. Their inclusion in iNKT cell-based therapies may benefit patients with CLL., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Although recent decades have seen an improved cure rate for newly diagnosed paediatric acute lymphoplastic leukaemia (ALL), the treatment options for adult ALL, T-cell ALL (T-ALL) and relapsed disease remain poor. We have developed a novel series of pyrrolo-1,5-benzoxazepine (PBOX) compounds and established their anticancer efficacy in a variety of human tumour cell types. Here, we demonstrate that PBOX-15 inhibits cell growth, and induces G2/M cell cycle arrest and apoptosis in both T-ALL and B-cell ALL (B-ALL) cells. In addition, prior to PBOX-15-induced apoptosis, PBOX-15 decreases ALL cell adhesion, spreading and migration. Concurrently, PBOX-15 differentially down-regulates β1-, β2- and α4-integrin expression in ALL cells and significantly decreases integrin-mediated cell attachment. PBOX-15 interferes with the lateral mobility and clustering of integrins in both B-ALL and T-ALL cells. These data suggest that PBOX-15 is not only effective in inducing apoptosis in ALL cells, but also has the potential to disrupt integrin-mediated adhesion of malignant lymphocytes, which represents a novel avenue for regulating leukaemic cell homing and migration.

Since the initial report of the BRAF V600E mutation in hairy cell leukemia, numerous investigators have demonstrated the presence of this activating mutation in nearly all cases of this disease. A case of hairy cell leukemia is documented with a classical clinical, morphological, immunophenotypic, and cytochemical profile in which the BRAF V600E was not detected. The diagnostic and therapeutic implications are discussed.

The spectrum of underlying molecular abnormalities of clinically and biologically heterogeneous chronic lymphocytic leukaemia (CLL) and prolymphocytic leukaemia (PLL) has yet to be identified. While whole genome sequencing has identified several genes implicated in the pathogenesis and progression of CLL, the molecular lesions in a substantial proportion of patients remain to be elucidated. The incidence of the BRAF V600E mutation, widely implicated in solid tumours and other B-cell malignancies, was sought in a cohort of patients with CLL and related disorders. One CLL patient and one patient with B-prolymphocytic leukaemia (PLL) were found to harbour this mutation. Although present at a low frequency, the finding of BRAF V600E has biological and clinical implications for CLL and PLL., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Background: We proposed to investigate the radiosensitizing properties of PBOX-15, a novel microtubule-disrupting agent, in a panel of cancer cell lines., Results: PBOX-15 treatment was associated with significant cell kill and increased radiosensitivity in all three cell lines tested. The number of surviving cells in response to the combined treatment was significantly less than PBOX -15 alone in 22Rv1 cells. In these cells, radiosensitisation correlated with induction of G2/M cell cycle arrest by PBOX-15. The compound sustained its activity and increased HIF-1Α expression under hypoxic conditions. PBOX-15 prevented onset of hypoxia-induced radioresistance in hypoxic prostate cells and reduced the surviving fraction of irradiated hypoxic cells to levels similar to those achieved under aerobic conditions., Methods: Clonogenic assays were used to determine sensitivity of a panel of cancer cell lines (22Rv1, A549, U87) to PBOX-15 alone or in combination with a single 2Gy dose fraction. Induction of cell cycle arrest and apoptosis was investigated in 22Rv1 prostate cancer cells. The cytotoxic properties of the compound under hypoxic conditions were correlated with Hypoxia Inducible Factor 1 alpha (HIF-1Α) gene and protein expression levels and its radiosensitisation potential was investigated in hypoxic 22Rv1 using clonogenic assays., Conclusions: This preliminary data identifies the potential of PBOX-15 as a novel radiosensitising agent for the management of solid tumours and eradication of hypoxic cells.

Background: In recent years, much progress has been made in the treatment of multiple myeloma. However, a major limitation of existing chemotherapeutic drugs is the eventual emergence of resistance; hence, the development of novel agents with new mechanisms of action is pertinent. Here, we describe the activity and mechanism of action of pyrrolo-1,5-benzoxazepine-15 (PBOX-15), a novel microtubule-targeting agent, in multiple myeloma cells., Methods: The anti-myeloma activity of PBOX-15 was assessed using NCI-H929, KMS11, RPMI8226, and U266 cell lines, and primary myeloma cells. Cell cycle distribution, apoptosis, cytochrome c release, and mitochondrial inner membrane depolarisation were analysed by flow cytometry; gene expression analysis was carried out using TaqMan Low Density Arrays; and expression of caspase-8 and Bcl-2 family of proteins was assessed by western blot analysis., Results: Pyrrolo-1,5-benzoxazepine-15 induced apoptosis in ex vivo myeloma cells and in myeloma cell lines. Death receptor genes were upregulated in both NCI-H929 and U266 cell lines, which displayed the highest and lowest apoptotic responses, respectively, following treatment with PBOX-15. The largest increase was detected for the death receptor 5 (DR5) gene, and cotreatment of both cell lines with tumour necrosis factor-related apoptosis-inducing ligand (TRAIL), the DR5 ligand, potentiated the apoptotic response. In NCI-H929 cells, PBOX-15-induced apoptosis was shown to be caspase-8 dependent, with independent activation of extrinsic and intrinsic apoptotic pathways. A caspase-8-dependent decrease in expression of Bim(EL) preceded downregulation of other Bcl-2 proteins (Bid, Bcl-2, Mcl-1) in PBOX-15-treated NCI-H929 cells., Conclusion: PBOX-15 induces apoptosis and potentiates TRAIL-induced cell death in multiple myeloma cells. Thus, PBOX-15 represents a promising agent, with a distinct mechanism of action, for the treatment of this malignancy.

Background: Imatinib is a direct and potent inhibitor of the constitutively active tyrosine kinase, breakpoint cluster region-Abelson (Bcr-Abl), which is central to the pathogenesis of chronic myeloid leukaemia (CML) patients. As such, imatinib has become the front-line treatment for CML patients. However, the recent emergence of imatinib resistance, commonly associated with point mutations within the kinase domain, has led to the search for alternative drug treatments and combination therapies for CML., Methods: In this report, we analyse the effects of representative members of the novel pro-apoptotic microtubule depolymerising pyrrolo-1,5-benzoxazepines or PBOX compounds on chemotherapy-refractory CML cells using a series of Bcr-Abl mutant cell lines, clinical ex vivo patient samples and an in vivo mouse model., Results: The PBOX compounds potently reduce cell viability in cells expressing the E225K and H396P mutants as well as the highly resistant T315I mutant. The PBOX compounds also induce apoptosis in primary CML samples including those resistant to imatinib. We also show for the first time, the in vivo efficacy of the pro-apoptotic PBOX compound, PBOX-6, in a CML mouse model of the T315I Bcr-Abl mutant., Conclusion: Results from this study highlight the potential of these novel series of PBOX compounds as an effective therapy against CML.

Pyrrolo-1,5-benzoxazepine-15 (PBOX-15) is a novel microtubule depolymerization agent that induces cell cycle arrest and subsequent apoptosis in a number of cancer cell lines. Chronic lymphocytic leukemia (CLL) is characterized by clonal expansion of predominately nonproliferating mature B cells. Here, we present data suggesting PBOX-15 is a potential therapeutic agent for CLL. We show activity of PBOX-15 in samples taken from a cohort of CLL patients (n = 55) representing both high-risk and low-risk disease. PBOX-15 exhibited cytotoxicity in CLL cells (n = 19) in a dose-dependent manner, with mean IC(50) of 0.55 micromol/L. PBOX-15 significantly induced apoptosis in CLL cells (n = 46) including cells with poor prognostic markers: unmutated IgV(H) genes, CD38 and zeta-associated protein 70 (ZAP-70) expression, and fludarabine-resistant cells with chromosomal deletions in 17p. In addition, PBOX-15 was more potent than fludarabine in inducing apoptosis in fludarabine-sensitive cells. Pharmacologic inhibition and small interfering RNA knockdown of caspase-8 significantly inhibited PBOX-15-induced apoptosis. Pharmacologic inhibition of c-jun NH(2)-terminal kinase inhibited PBOX-15-induced apoptosis in mutated IgV(H) and ZAP-70(-) CLL cells but not in unmutated IgV(H) and ZAP-70(+) cells. PBOX-15 exhibited selective cytotoxicity in CLL cells compared with normal hematopoietic cells. Our data suggest that PBOX-15 represents a novel class of agents that are toxic toward both high-risk and low-risk CLL cells. The need for novel treatments is acute in CLL, especially for the subgroup of patients with poor clinical outcome and drug-resistant disease. This study identifies a novel agent with significant clinical potential.

The ordered, directional migration of T-lymphocytes is a key process during immune surveillance, immune response, and development. A novel series of pyrrolo-1,5-benzoxazepines have been shown to potently induce apoptosis in variety of human chemotherapy resistant cancer cell lines, indicating their potential in the treatment of both solid tumors and tumors derived from the hemopoietic system. Pyrrolobenzoxazepine 4-acetoxy-5-(1-naphtyl)naphtho[2,3-b]pyrrolo[1,2-d][1,4]-oxazepine (PBOX-15) has been shown to depolymerize tubulin in vitro and in the MCF7 breast cancer cell line. We hypothesized that this may suggest a role for this compound in modulating integrin-induced T-cell migration, which is largely dependent on the microtubule dynamics. Experiments were performed using human T lymphoma cell line Hut78 and peripheral blood T-lymphocytes isolated from healthy donors. We observed that human T-lymphocytes exposed to PBOX-15 have severely impaired ability to polarize and migrate in response to the triggering stimulus generated via cross-linking of integrin lymphocyte function associated antigen-1 receptor. Here, we show that PBOX-15 can dramatically impair microtubule network via destabilization of tubulin resulting in complete loss of the motile phenotype of T-cells. We demonstrate that PBOX-15 inhibitory mechanisms involve decreased tubulin polymerization and its post-translational modifications. Novel microtubule-targeting effects of PBOX-15 can possibly open new horizons in the treatment of overactive inflammatory conditions as well as cancer and cancer metastatic spreading.

A novel peptide antibody to UCP 3 is characterized which is sensitive and discriminatory for UCP 3 over UCP 2, UCP 1 and other mitochondrial transporters. The peptide antibody detects UCP 3 expression in E. coli, COS cells and yeast expression systems. The peptide antibody detects a single approximately 33 kDa protein band in mitochondria from isolated rat skeletal muscle, mouse and rat brown adipose tissue, and in whole muscle groups (soleus and extensor digitorum longus) from mice. No 33 kDa band is detectable in isolated mitochondria from liver, heart, brain, kidney and lungs of rats, or gastrocnemius mitochondria from UCP 3 knock-out mice. From our data, we conclude that the peptide antibody is detecting UCP 3 in skeletal muscle, skeletal muscle mitochondria and brown adipose tissue mitochondria. It is also noteworthy that the peptide antibody can detect human, mouse and rat forms of UCP 3. Using the UCP 3 peptide antibody, we confirm and quantify the increased (2.8-fold) UCP 3 expression observed in skeletal muscle mitochondria isolated from 48-h-starved rats. We show that UCP 3 expression is increased (1.6-fold) in skeletal muscle of rats acclimated over 8 weeks to 8 degrees C and that UCP 3 expression is decreased (1.4-fold) in rats acclimated to 30 degrees C. Furthermore, UCP 3 expression is increased (2.3-fold) in skeletal muscle from hyperthyroid rats compared to euthyroid controls. In addition, we show that UCP 3 expression is only coincident with the mitochondrial fraction of skeletal muscle homogenates and not peroxisomal, nuclear or cytosolic and microsomal fractions.

Maintenance of the extracellular matrix (ECM) is important for tissue integrity and cellular physiology. Normal ECM turnover is regulated by a balance between matrix metalloproteinases and their inhibitors, the tissue inhibitors of metalloproteinases (TIMPs). In metastasis, this balance favours increased ECM degradation. The objective of this study was to determine the effects of TIMP-1 overexpression on the metastatic process. To this end, we stably transfected a renal carcinoma cell line, Caki-1, with TIMP-1, using a pRc/CMV expression plasmid and LIPOFECTAMINE transfection reagent. The resultant clones displayed increased adhesion on the ECM substratum, including collagen type IV and laminin, and altered invasive capacity through fibronectin and Matrigel, dependent upon the level of TIMP-1 expression. These changes were not due to altered integrin expression, as assessed by flow cytometry. As well as protease inhibitory activity, TIMPs can influence cell proliferation and cell survival. The TIMP-1 clones displayed no changes in proliferation under normal growth conditions, compared with Caki-1 cells. However, under reduced serum conditions, the TIMP-1 clones had a greater percentage of cells in both S (P<0.05) and G(2)/M (P<0.005) phases and less cells in G(0)/G(1) (P<0.001) of the cell cycle than Caki-1 cells. The results confirm a dual role for TIMP-1 in invasion and metastasis, and provide further clues behind the molecular mechanisms in these processes.