Optimal sequencing of ibrutinib, idelalisib, and venetoclax in chronic lymphocytic leukemia: results from a multicenter study of 683 patients.

Background: Ibrutinib, idelalisib, and venetoclax are approved for treating CLL patients in the United States. However, there is no guidance as to their optimal sequence. Patients and methods: We conducted a multicenter, retrospective analysis of CLL patients treated with kinase inhibitors (KIs) or venetoclax. We examined demographics, discontinuation reasons, overall response rates (ORR), survival, and post-KI salvage strategies. Primary endpoint was progression-free survival (PFS). Results: A total of 683 patients were identified. Baseline characteristics were similar in the ibrutinib and idelalisib groups. ORR to ibrutinib and idelalisib as first KI was 69% and 81%, respectively. With a median follow-up of 17 months (range 1-60), median PFS and OS for the entire cohort were 35 months and not reached. Patients treated with ibrutinib (versus idelalisib) as first KI had a significantly better PFS in all settings; front-line [hazard ratios (HR) 2.8, CI 1.3-6.3, P = 0.01], relapsed-refractory (HR 2.8, CI 1.9-4.1, P < 0.001), del17p (HR 2.0, CI 1.2-3.4, P = 0.008), and complex karyotype (HR 2.5, CI 1.2-5.2, P = 0.02). At the time of initial KI failure, use of an alternate KI or venetoclax had a superior PFS when compared with chemoimmunotherapy. Furthermore, patients who discontinued ibrutinib due to progression or toxicity had marginally improved outcomes if they received venetoclax (ORR 79%) versus idelalisib (ORR 46%) (PFS HR .6, CI.3-1.0, P = 0.06). Conclusions: In the largest real-world experience of novel agents in CLL, ibrutinib appears superior to idelalisib as first KI. Furthermore, in the setting of KI failure, alternate KI or venetoclax therapy appear superior to chemoimmunotherapy combinations. The use of venetoclax upon ibrutinib failure might be superior to idelalisib. These data support the need for trials testing sequencing strategies to optimize treatment algorithms.

Experimental study of wet granulation in fluidized bed: impact of the binder properties on the granule morphology.

In this work, the effect of the physicochemical properties of aqueous hydroxypropyl-cellulose (HPC) binder solutions and different pharmaceutical excipients (mannitol and anhydrous CaHPO(4)) on the agglomeration kinetics and granule properties were investigated. First, a particle size distribution (PSD) analysis together with a detailed analysis of morphological properties of the excipient particles were performed. Second, the viscosity, density, surface tension and size of the spray droplets of binder solutions with different HPC concentrations were determined and wetting characteristics of the binders on the excipients were measured. Third, several fluid bed wet granulation experiments were conducted for pure excipients and their blends with binder solution of different HPC concentrations in a pilot plant Wurster granulator. The observed granule growth for different binder concentrations was a strong function of the binder concentration and the excipient solubility. For mannitol, a significant "coating" period followed by a slow granule growth was observed for the case with the diluted 5% binder. The "coating" period was significantly shorter for the 10% HPC binder and did not exist for the 15% HPC for which immediate and fast granule growth was observed. For anhydrous CaHPO(4) (trademark A-TAB), no growth was observed for the 10% HPC binder and a long coating period followed by fast granule growth was observed for the 15% HPC. Simple physically based criteria were also evaluated, which employ the morphological properties of excipients (size and surface roughness) together with physical properties of the used binder for prediction of the coating versus agglomeration regime at given flow conditions (collision velocity). As expected, a preferential coalescence and growth of the mannitol granules from the blend of mannitol+A-TAB was observed. Finally, the mechanical and morphological properties of the produced granules were measured and correlated to the HPC concentration of the binder used in the experiments. A clear correlation between the granule porosity (evaluated by X-ray tomography) and the binder concentration was found for the mannitol granules.

Preclinical screening for drugs effective against 5-fluorouracil-resistant cells with a murine L5178Y cell line in vitro.

A subline of L5178Y cells has been established in vitro that exhibits a fiftyfold order of resistance to 5-fluorouracil (FUra) as compared to that of the parent line. The cytotoxic effects of 24-hour exposures to 23 antitumor drugs and to radiation were compared in the two cell lines. Four patterns of response were identified: 1) Only two drugs, mitomycin C and adriamycin, proved significantly more cytotoxic to FUra-resistant cells. 2) Four other drugs--anguidine, 4'-(9-acridinylamino)-methanesulfon-m-anisidide, melphalan, and quelamycin--showed marginal superiority against resistant cells. 3) X-radiation and the majority of drugs tested--including 5-azacytidine, 1,3-bis(2-chloroethyl)-1-nitrosourea, cisplatin, bleomycin, dibromodulcitol, razoxane, hydroxyurea, methotrexate, teniposide, etoposide, and three experimental agents, metoprine, spirogermanium HCl, and ellipticinum--proved equally cytotoxic to both cell lines. 4) Cross-resistance with FUra was exhibited with vincristine, vindesine, pyrazofurin, and indicine-N-oxide. This experimental system provides a simple method of testing agents for activity against FUra-resistant cells before phase 1 clinical studies.

Murine L5178Y cells resulting in altered drug sensitivities from fractionated radiation exposure in vitro.

Exposure of murine L5178Y cells in vitro to fractionated X-irradiation (DXR) produced a subline that exhibited significantly altered responses to the cytotoxic effects of a range of antitumor drugs. A DXR schedule of 2 Gy per fraction for 10 fractions (total dose, 20 Gy) was used. Characterization of this subline revealed no marked differences when compared with the parent line in terms of growth rate, cloning efficiency, cell volume, chromosome number, macromolecular content, cell cycle distribution, or response to acute DXR exposure. Drug sensitivities were assessed following 24-hour exposures by colony formation in soft agar. Three distinct patterns of response were found: increased sensitivity of DXR-pretreated cells to bleomycin, cis-diamminedichloroplatinum (II), or dibromodulcitol; comparable responses in both lines to doxorubicin, 5-fluorouracil, or hydroxyurea; and decreased sensitivity of DXR-pretreated cells to vincristine or VP 16213.

Comparative cell killing and kinetic effects of vincristine or vindesine in mammalian cell lines.

Survival curves for several monolayer and suspension cultures following 24-hour exposures to various vincristine (VCR) or vindesine (VDS) concentration were all of the exponential-plateau type. Cytotoxicity increased with duration of exposure in hamster NIL8 cells and was comparable for both drugs. Murine L5178Y cells, although 200 times more sensitive than NIL8 cells, also showed similar sensitivities to VCR and VDS. Murine neuroblastoma cells proved approximately fivefold less sensitive to VDS than VCR, whereas qualitative and quantitative differences in the activity of these two drugs were noted in human neuroblastoma cells. The lethal effects of VCR and VDS were exerted solely on logarithmically growing NIL8 cells in the S-phase. Plateau-phase human neuroblastoma cells were also significantly less sensitive to both drugs than were logarithmically growing populations, were cell kill occurred during the S-phase. Complete mitotic arrest, induced by a 24-hour exposure to VCR or VDS, was only partially reversible, whereas rapid and complete reversibility occurred after only 5 hours of drug exposure.

Superiority of adriamycin-14-octanoate over adriamycin in reducing viability of methotrexate-resistant L5178Y cells: brief communication.

Adriamycin-14-octanoate (ADR-OCT) was superior to adriamycin (ADR) in reducing the viability of L5178Y cells resistant to methotrexate (MTX). This effect was seen in logarithmically growing and plateau-phase cultures and increased both with dose and duration of exposure. Both ADR-OCT and ADR were effective inhibitors of the exogenous Escherichia coli DNA-dependent RNA polymerases in vitro and of the endogenous polymerase in mammalian cultured cells. Drug concentrations required for approximately 50% enzyme inhibition in both systems were comparable for both agents, being of the order of 10(-5) M. These experimental studies suggested that ADR-OCT may be a valuable agent for treating neoplasms resistant to MTX.

Dominant expression of multiple drug resistance after in vitro X-irradiation exposure in intraspecific Chinese hamster ovary hybrid cells.

BACKGROUND: Exposure of Chinese hamster ovary (CHO) cells to fractionated x irradiation in vitro has led to expression of a distinctive multiple-drug-resistant phenotype. This phenotype is characterized by overexpression of P-glycoprotein without an increase in P-glycoprotein messenger RNA or gene amplification; increased glutathione S-transferase (GST) activity; and resistance to vincristine, colchicine, and etoposide but not to doxorubicin. PURPOSE: To investigate whether this phenotype is dominant or recessive, we established intraspecific hybrids by fusion of x-ray-treated, drug-resistant CHO cells (DXR-10I or DXR-10II) with drug-sensitive CHO cells (E29). METHODS: Drug resistance levels were determined in the wild-type CHO cell line AuxB1, the drug-sensitive E29 line, the x-ray-pretreated lines, and the hybrid lines by colony-forming assay of cells grown in increasing concentrations of colchicine, vincristine, or doxorubicin. The hybrids were characterized by analysis of DNA content, P-glycoprotein expression by Western blotting, GST activity by use of 1-chloro-2,4-dinitrobenzene as substrate, and sensitivity to reversal of resistance to vincristine by exposure to verapamil. RESULTS: These hybrids proved resistant to colchicine (two-fold) and vincristine (five- to seven-fold) but not to doxorubicin. After the hybrids were exposed to verapamil, vincristine cytotoxicity was increased 10- to 12-fold. The hybrid lines exhibited levels of P-glycoprotein comparable to those of the unfused x-ray-treated parent cell line, suggesting that P-glycoprotein overexpression is a dominant trait in these hybrid lines. Interpretation of the role of increased GST activity in these hybrids was inconclusive because of the very high levels of GST in the drug-sensitive cell-fusion partner. CONCLUSIONS: The multiple-drug-resistant phenotype following x-ray treatment of CHO cells in vitro was dominantly expressed. Overall, these data are consistent with the hypothesis that this phenotype is a consequence of the dominant genetic alteration resulting from exposure to x irradiation. IMPLICATIONS: This work adds weight to our hypothesis that there is a biological basis for the expression of clinical drug resistance in certain patients whose tumors have been previously irradiated.

Overexpression of P-glycoprotein in mammalian tumor cell lines after fractionated X irradiation in vitro.

We observed that in vitro exposure of mammalian tumor cells to fractionated x irradiation results in the expression of drug resistance. The cause of this resistance was investigated in a series of Chinese hamster ovary cell lines that had survived exposure to multiple lethal doses of radiation. These cell lines had increased levels of P-glycoprotein (Pgp), the multidrug-resistance-associated membrane glycoprotein. Consistent with the classic multidrug resistance phenotype, they exhibited cross-resistance to multiple drugs, as well as sensitivity to reversal of vincristine resistance by verapamil. However, the cell lines showed no change in their sensitivity to x rays. Pgp overexpression occurred in these cells, despite a lack of Pgp gene amplification or of significant alteration in Pgp messenger RNA levels. Although the cause of increased Pgp levels is not yet known, these data suggest a biological basis for the clinical problem of drug resistance that can occur in previously irradiated tumors.

Characteristics of four new human cell lines derived from squamous cell carcinomas of the head and neck.

Four human cell lines were established from biopsy specimens of squamous cell carcinomas of the larynx (TR131 and TR138), tongue (TR126), and buccal mucosa that had infiltrated a lymph node (TR146). All 4 lines readily formed colonies on a plastic substratum, but they were virtually incapable of forming colonies in an anchorage-independent semisolid support system of soft agar (cloning efficiencies, less than 0.02%). The proliferation of this group of tumor-derived cell lines, therefore, appeared to be highly anchorage dependent. Keratin filaments could be visualized in each line by indirect immunofluorescence with the use of polyclonal or monoclonal antibodies to keratins; staining with monospecific antibodies indicated that 3 of the 4 lines expressed simple epithelial keratins 8 and 18, whereas 1 of the 4 also expressed keratin 19. A panel of lectins revealed characteristic localization patterns distinct from those observed on other epithelial cell lines. Cells from 3 lines (TR131, TR138, and TR146) inoculated into nude mice (nu/nu) produced cystic nodules or unequivocal tumors having a histology indicating a squamous cell origin for the injected cells. Electron microscopy demonstrated that the cell lines covered a spectrum of differentiation capability ranging from the undifferentiated monolayer cultures of TR126 to the rather well differentiated, stratified cultures of TR131.

Anomalous relationship between cisplatin sensitivity and the formation and removal of platinum-DNA adducts in two human ovarian carcinoma cell lines in vitro.

Two human ovarian tumor cell lines (SK-OV-3 and TR175), established from patients previously treated with alkylating agents, but not with cisplatin, expressed greater than 23-fold differences in cisplatin sensitivities in vitro. Cisplatin resistance in SK-OV-3 cells appeared to be associated with increased levels of glutathione and activities of glutathione reductase and glutathione peroxidase, with reduced catalase activity. No significant modification of drug uptake was noted and there was only marginally lower (16%) total platination of DNA, measured immunochemically, in these cells compared with the more sensitive TR175 cell line. SK-OV-3 cells, however, showed a significantly lower overall ability to remove drug-induced DNA damage, with an apparent inability to remove either the major DNA-DNA intrastrand cross-links in the sequence pGpG or the adducts cis-Pt(NH3)2d(GMP)2, although by alkaline elution repair of DNA-DNA interstrand cross-links was demonstrated. Significantly more of these interstrand cross-links were induced in these resistant cells. These data provide evidence for the involvement of altered glutathione metabolism and increased tolerance of certain types of drug-induced DNA damage as factors associated with the resistance phenotype of SK-OV-3 cells. Paradoxically, however, although the highly cisplatin-sensitive TR175 cells had lower glutathione levels this was not reflected in significantly higher total platination of DNA, and these cells appeared to be proficient in removing all the major platinum-DNA adducts quantitated in this study. Mechanisms responsible for this relative sensitivity to cisplatin remain to be identified.

Novel actions of the antitumor drugs vinflunine and vinorelbine on microtubules.

Vinflunine is a novel Vinca alkaloid presently in Phase I clinical trials. In preclinical studies, it exhibited superior antitumor activity to that of other Vinca alkaloids, including vinorelbine from which it was synthetically derived. Vinca alkaloids appear to inhibit cell proliferation by affecting the dynamics of spindle microtubules. Here we have analyzed the effects of vinflunine and vinorelbine on microtubule dynamic instability and treadmilling and found that these newer drugs exert effects on microtubule dynamics that differ significantly from those of the classic Vinca alkaloid, vinblastine. The major effects of vinflunine and vinorelbine on dynamic instability were a slowing of the microtubule growth rate, an increase in growth duration, and a reduction in shortening duration. In marked contrast to the action of vinblastine, they neither reduced the rate of shortening nor increased the percentage of time the microtubules spent in an attenuated state, neither growing nor shortening detectably. In addition, vinflunine and vinorelbine suppressed treadmilling, but less strongly than vinblastine. The diverse actions of these drugs on microtubules are likely to produce different effects on mitotic spindle function, leading to different effects on cell cycle progression and cell killing. Nontumor cells with normal checkpoint proteins may tolerate the relatively less powerful inhibitory effects of vinflunine and vinorelbine on microtubule dynamics better than the more powerful effects of vinblastine. Thus the unique constellation of effects of vinflunine and vinorelbine on dynamic instability and treadmilling may contribute to their superior antitumor efficacies.

Cytotoxic effects and biological activity of 2-aza-8-germanspiro[4,5]-decane-2-propanamine-8,8-diethyl-N,N-dimethyl dichloride (NSC 192965; spirogermanium) in vitro.

Lethal and other biological effects of 2-aza-8-germanspiro[4,5]decane-2-propanamine-8,8-diethyl-N,N-dimethyl dichloride (NSC 192965; spirogermanium), representing a new chemical class of compound exhibiting antitumor activity, have been studied in vitro. Survival curves for NIL 8 hamster cells were exponential with greater kill occurring with increasing drug concentrations and longer exposure times. Cytotoxicity was temperature dependent. "Quiescent" cultures were significantly less sensitive to spirogermanium than were logarithmically growing cells. These lethal effects showed no phase specificity. There was no evidence of progression delay through the cycle following spirogermanium treatment. When spirogermanium was tested against a range of human cell lines, the consistency of the values for the drug concentration required to reduce survival by 50% on the exponential part of the survival curve, derived from colony-forming assays, was most marked. The survival curves, characterized by an initial shoulder, were steep and exponential with measurements possible over only a narrow concentration range since complete cell lysis occurred at levels causing a greater than 2-log kill. Cell membrane damage by spirogermanium, as judged by dye exclusion, was progressive with time and increasing drug concentrations. Protein synthesis proved most susceptible to the drug. Spirogermanium concentrations cytotoxic to tumor cells were also toxic to cultured rat neurons, confirming the clinical neurological toxicity encountered. The precise mode of action of spirogermanium remains to be established, and these data further illustrate its apparent lack of specificity.

Differential repair of platinum-DNA adducts in human bladder and testicular tumor continuous cell lines.

The formation and removal of four platinum-DNA adducts were immunochemically quantitated in cultured cells derived from a human bladder carcinoma cell line (RT112) and from two lines derived from germ cell tumors of the testis (833K and SUSA), following exposure in vitro to 16.7 microM (5 micrograms/ml) cisplatin. RT112 cells were least sensitive to the drug and were proficient in the repair of all four adducts, whereas SUSA cells, which were 5-fold more sensitive, were deficient in the repair of DNA-DNA intrastrand cross-links in the sequences pApG and pGpG. Despite expressing a similar sensitivity to SUSA cells, 833K cells were proficient in the repair of all four adducts, although less so than the RT112 bladder tumor cells. In addition, SUSA cells were unable to repair DNA-DNA interstrand cross-links whereas 50-85% of these lesions were removed in RT112 and 833K cells 24 h following drug exposure. It is possible that the inability of SuSa cells to repair platinated DNA may account for their hypersensitivity to cisplatin.

Characteristics of a new human neuroblastoma cell line which differentiates in response to cyclic adenosine 3':5'-monophosphate.

A new human cell line, TR14 , has been established in tissue culture from biopsy material of a primary neuroblastoma tumor. Most TR14 cells have short processes and grow mainly in clumps adhering to cells attached to the substratum. TR14 cells form colonies in soft agar demonstrating anchorage independence of growth and produce tumors in nude mice with histologies similar to that of the patient's tumor. The neurotransmitter-synthesizing activity of these cells is predominantly cholinergic with only a minor adrenergic component, since the activity of choline acetyltransferase is about 20-fold greater than that of tyrosine hydroxylase. Treatment with N6,O2'-dibutyryl cyclic adenosine 3':5'-monophosphate induces TR14 neuroblastoma cells to extend fine, long processes or neurites. This morphological change is accompanied by elevated numbers of cytoplasmic dense-core vesicles observed by electron microscopy and an increase in the activities of neurotransmitter-synthesizing enzymes. Differentiation therefore occurs at the levels of cellular morphology, ultrastructure, and biochemistry. Prostaglandin E1 and cholera toxin can also induce differentiation, but a range of other agents including dimethyl sulfoxide, nerve growth factor, butyrate, corticosteroids, and 5-bromodeoxyuridine is ineffective. The concomitant induction of both morphological and biochemical differentiation therefore appears to be exclusively a cyclic adenosine 3':5'-monophosphate-mediated event in this cell line.

Characterization of a cisplatin-resistant human ovarian carcinoma cell line expressing cross-resistance to 5-fluorouracil but collateral sensitivity to methotrexate.

In vitro exposure of the TR170 ovarian carcinoma cell line to six intermittent 24-h treatments with a 90% inhibitory concentration of cisplatin (CDDP) (0.15 micrograms/ml; 0.5 microM) resulted in a 2-fold stably resistant subline designated TR170/CP+ (B.T. Hill et al., Int. J. Cancer, 39: 219-225, 1987). Resistance to CDDP in these CP+ cells has now been associated with reduced uptake of 195mCDDP (2-fold; P less than 0.01) and decreased removal of specific Pt-DNA adducts, quantitated immunochemically, indicative of an apparent increased tolerance of CDDP-induced DNA damage. Specifically these resistant cells appeared deficient in removal of the major cis-Pt-(NH3)2d(pGpG) adduct and the difunctional cis-Pt(NH3)2d(GMP)2 lesion, showed less efficiency in removing cis-Pt(NH3)2d(pApG) adducts, but proved as proficient as the parental cell line in removing DNA-DNA interstrand cross-links. Activities of DNA polymerase-alpha and -beta were comparable in both lines, and no significant alterations in glutathione metabolism were identified. Response to acute X-irradiation was not modified in these TR170/CP+ cells, but they showed marked (10-fold) cross-resistance to 5-fluorouracil and, unusually, proved collaterally sensitive (12-fold) to methotrexate. Resistance to 5-fluorouracil was associated with significantly increased thymidylate synthase activity (P less than 0.01), but this was not reflected in altered gene expression, while increased sensitivity to methotrexate was accompanied by increased drug uptake but by unaltered activity and expression of dihydrofolate reductase. These results indicate that exposure to CDDP can result in numerous alterations, both intracellularly and at the cellular membrane, reflected in significant changes in the tumor cells' responses to the cytotoxic effects of a range of antitumor drugs. The clinical relevance of these observations remains to be established.

Tissue culture model of transitional cell carcinoma: characterization of twenty-two human urothelial cell lines.

Twenty-two continuous cell lines derived from normal and neoplastic urothelium, maintained under identical culture conditions, were characterized in terms of isozyme phenotype, tumorigenicity, and xenograft morphology following xenotransplantation to nude mice, cytological appearance, in vitro growth rate, labelling index, and colony-forming efficiency, in parallel with separate studies of in vitro drug sensitivities and monoclonal antibody reactivities. Three groups were identified: (a) distinct lines with differing isozyme patterns, a broad spectrum of growth characteristics, and xenograft morphologies similar to the histopathology of the parent tumors after periods of up to 17 yr following establishment in vitro; (b) cross-contaminated sublines (maintained separately in different laboratories for periods of up to 10 yr), with identical isozyme patterns and similar growth characteristics, but differing markedly in tumorigenicity and xenograft morphology; and (c) lines derived from normal urothelium which were nontumorigenic and had an isozyme pattern usually only encountered in untransformed cells. These data indicate that cell lines representative of human transitional cell carcinomas can be selected on the basis of xenograft morphology and isozyme patterns, and that a panel of lines derived from normal and neoplastic urothelium could provide a model system to study the biology and treatment of this disease.

Defining AML and MDS second cancer risk dynamics after diagnoses of first cancers treated or not with radiation.

Risks of acute myeloid leukemia (AML) and/or myelodysplastic syndromes (MDS) are known to increase after cancer treatments. Their rise-and-fall dynamics and their associations with radiation have, however, not been fully characterized. To improve risk definition we developed SEERaBomb R software for Surveillance, Epidemiology and End Results second cancer analyses. Resulting high-resolution relative risk (RR) time courses were compared, where possible, to results of A-bomb survivor analyses. We found: (1) persons with prostate cancer receiving radiation therapy have increased RR of AML and MDS that peak in 1.5-2.5 years; (2) persons with non-Hodgkin lymphoma (NHL), lung and breast first cancers have the highest RR for AML and MDS over the next 1-12 years. These increased RR are radiation specific for lung and breast cancer but not for NHL; (3) AML latencies were brief compared to those of A-bomb survivors; and (4) there was a marked excess risk of acute promyelocytic leukemia in persons receiving radiation therapy. Knowing the type of first cancer, if it was treated with radiation, the interval from first cancer diagnosis to developing AML or MDS, and the type of AML, can improve estimates of whether AML or MDS cases developing in this setting are due to background versus other processes.

Association of socioeconomic status with autologous hematopoietic cell transplantation outcomes for lymphoma.

Socioeconomic status (SES) is an important determinant of disparities in health care. The association of SES with outcomes in autologous hematopoietic cell transplantation (AHCT) has not been described previously. We conducted a retrospective cohort study of 687 AHCT recipients with lymphoma transplanted between 2003 and 2013. Patients were categorized into low (<$50 000/year) and high SES (⩾$50 000/year). A greater proportion of low SES patients lived farther away from our center (median 54 vs 28 miles), belonged to a racial minority (12 vs 3%), had poorer performance status (4 vs 1%) and had high-risk disease at AHCT (9 vs 5%). Median follow-up was 53 months. In univariable analysis, low SES patients had significantly higher relapse mortality and lower OS and PFS. This was confirmed on multivariable analysis for relapse mortality (HR for high vs low SES: 0.74 (95% confidence interval (CI), 0.54-0.99), P=0.05), OS (HR 0.74 (0.58-0.95), P=0.02) and PFS (HR 0.77 (0.63-0.95), P=0.02). In multivariable analysis of ⩾1-year progression-free survivors, high SES patients had better OS (HR 0.73, P=0.05 vs low SES) that was primarily driven by a trend toward lower risk of non-relapse mortality (HR 0.62, P=0.06). SES is associated with outcomes of AHCT in patients with lymphoma.