Dramatic Reduction In Tumor Burden Research Articles

Potent Anticancer Activity of CXCR4-Targeted Nanostructured Toxins in Aggressive Endometrial Cancer Models

Patients with advanced endometrial cancer (EC) show poor outcomes. Thus, the development of new therapeutic approaches to prevent metastasis development in high-risk patients is an unmet need. CXCR4 is overexpressed in EC tumor tissue, epitomizing an unexploited therapeutic target for this malignancy. The in vitro antitumor activity of two CXCR4-targeted nanoparticles, including either the C. diphtheriae (T22-DITOX-H6) or P. aeruginosa (T22-PE24-H6) toxin, was evaluated using viability assays. Apoptotic activation was assessed by DAPI and caspase-3 and PARP cleavage in cell blocks. Both nanotoxins were repeatedly administrated to a subcutaneous EC mouse model, whereas T22-DITOX-H6 was also used in a highly metastatic EC orthotopic model. Tumor burden was assessed through bioluminescence, while metastatic foci and toxicity were studied using histological or immunohistochemical analysis. We found that both nanotoxins exerted a potent antitumor effect both in vitro and in vivo via apoptosis and extended the survival of nanotoxin-treated mice without inducing any off-target toxicity. Repeated T22-DITOX-H6 administration in the metastatic model induced a dramatic reduction in tumor burden while significantly blocking peritoneal, lung and liver metastasis without systemic toxicity. Both nanotoxins, but especially T22-DITOX-H6, represent a promising therapeutic alternative for EC patients that have a dismal prognosis and lack effective therapies.

Automated Generation and Phenotypic Characterization of Clinical Grade CD19 CAR-T Cells

BACKGROUNDChimeric antigen receptor (CAR) T cells have shown enormous promise in the treatment of certain B cell malignancies. Access to treatment is still limited due to a variety of issues, including pricing and centralized manufacturing models. Generation of CAR-T cells using an automated platform, followed by rigorous functional phenotyping, may contribute to the development of a robust long-lasting therapy. METHODSHere, we used the Miltenyi Prodigy (Miltenyi Biotech, Bergisch Gladbach, Germany) to automate the process of manufacturing genetically manipulated T cells in a closed system. The system obviates the need for clean room infrastructure. We tested the feasibility of utilizing the Miltenyi Prodigy to manufacture CAR-T cells using a CD19 scFV vector with the 4-1BB co-stimulatory domain. (Lentigen Technology, Inc, Gaithersburg, MD). The purity, differentiation capacity and effector function of the enriched CAR-T cells was studied using high-dimensional flow cytometry. Finally, the functional potential of these cells was tested in vitro and by treating NOD-SCID-gamma (NSG) mice infused with B cell lymphoma cells (Raji B cell), with the CAR-T cells. RESULTSStarting with 1 x 108 CD4 and CD8 cells from donor apheresis products, CAR-T cells were expanded for 12 days in culture media containing with TransAct (Miltenyi Biotech), IL7 and IL15. The mean fold-expansion at day 12 was 44 ± 5.6, range 39-50 (n=3). The mean transduction efficiency of CAR-T vector was 20%, range 10-25% (n=3), which is similar to other reported methods. The CD19 CAR-T product was enriched in both the CD4 and CD8 T cells subsets, and showed high-level of cytotoxicity against CD19+ cell lines in vitro and in vivo (Figure 1: Mice treated with the CD19-CAR T demonstrated a marked reduction in disease burden as compared to T cell control as measured by bioluminescence imaging and flow cytometric analysis). The CAR-T product was enriched in cell subsets with both effector (CD27-CCR7-; ~20% of total cells) and central memory phenotypes (CD27+CCR7+; ~30% of total T cells). The effector CD4 and CD8 T cells showed increased expression of major functional T cell differentiation transcription factors (i.e. T-bet and GATA3) critical for the development of anti-tumor responses. Whereas, the central CD4 and CD8 T cells were enriched for the expression of TCF7 (a stemness related member of the WNT signaling known to increase longevity of these cells). The frequencies and phenotypes of these cells were maintained in peripheral blood of NSG mice infused with B cell lymphoma cells (Raji B cells), 1 week after treatment. A significant expansion of CD8+ effector T cells and a dramatic reduction in tumor burden was observed over the next 4 weeks in all major organs. Interestingly, we observed that smaller proportion of central-memory like cells (with higher TCF7 levels) continued to persist 6 weeks post-treatment, potentially contributing to a long-lived recallable response. Based on these data we have initiated a phase 1 clinical trial to test the therapeutic potential of the CAR-T product in patients with advanced/refractory B cell lymphoma. The first clinical grade manufacturing run resulted in a CD19 + cell yield of 1.4 x109. CONCLUSIONOur data highlight that the automated CAR-T generation platform (i.e. Miltenyi Prodigy) is effective at the generating purified functionally competent CAR-T cells. Further, findings from our phenotyping analyses show that the CAR-T product is enriched in both effector and central memory subsets and is effective at tumor clearance in vivo. Thus far, we have treated one patient with CD19 CAR-T manufactured on this platform and 2 more have been enrolled. Though this initial study is based on CD19 CAR-T cells, the approach described here could easily be utilized to genetically engineer T cells with gene constructs that are more relevant for specific cancers and infectious diseases. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Adoptively transferred Vγ9Vδ2 T cells show potent antitumor effects in a preclinical B cell lymphomagenesis model.

A central issue for adoptive cellular immunotherapy is overcoming immunosuppressive signals to achieve tumor clearance. While γδ T cells are known to be potent cytolytic effectors that can kill a variety of cancers, it is not clear whether they are inhibited by suppressive ligands expressed in tumor microenvironments. Here, we have used a powerful preclinical model where EBV infection drives the de novo generation of human B cell lymphomas in vivo, and autologous T lymphocytes are held in check by PD-1/CTLA-4-mediated inhibition. We show that a single dose of adoptively transferred Vδ2+ T cells has potent antitumor effects, even in the absence of checkpoint blockade or activating compounds. Vδ2+ T cell immunotherapy given within the first 5 days of EBV infection almost completely prevented the outgrowth of tumors. Vδ2+ T cell immunotherapy given more than 3 weeks after infection (after neoplastic transformation is evident) resulted in a dramatic reduction in tumor burden. The immunotherapeutic Vδ2+ T cells maintained low cell surface expression of PD-1 in vivo, and their recruitment to tumors was followed by a decrease in B cells expressing PD-L1 and PD-L2 inhibitory ligands. These results suggest that adoptively transferred PD-1lo Vδ2+ T cells circumvent the tumor checkpoint environment in vivo.

Natural killer cell-based adoptive immunotherapy eradicates and drives differentiation of chemoresistant bladder cancer stem-like cells.

BackgroundHigh-grade non-muscle invasive bladder cancer (NMIBC) has a high risk of recurrence and progression to muscle-invasive forms, which seems to be largely related to the presence of tumorigenic stem-like cell populations that are refractory to conventional therapies. Here, we evaluated the therapeutic potential of Natural Killer (NK) cell-based adoptive immunotherapy against chemoresistant bladder cancer stem-like cells (CSCs) in a pre-clinical relevant model, using NK cells from healthy donors and NMIBC patients.MethodsCytokine-activated NK cells from healthy donors and from high-grade NMIBC patients were phenotypically characterized and assayed in vitro against stem-like and bulk differentiated bladder cancer cells. Stem-like cells were isolated from two bladder cancer cell lines using the sphere-forming assay. The in vivo therapeutic efficacy was evaluated in mice bearing a CSC-induced orthotopic bladder cancer. Animals were treated by intravesical instillation of interleukin-activated NK cells. Tumor response was evaluated longitudinally by non-invasive bioluminescence imaging.ResultsNK cells from healthy donors upon activation with IL-2 and IL-15 kills indiscriminately both stem-like and differentiated tumor cells via stress ligand recognition. In addition to cell killing, NK cells shifted CSCs towards a more differentiated phenotype, rendering them more susceptible to cisplatin, highlighting the benefits of a possible combined therapy. On the contrary, NK cells from NMIBC patients displayed a low density on NK cytotoxicity receptors, adhesion molecules and a more immature phenotype, losing their ability to kill and drive differentiation of CSCs. The local administration, via the transurethral route, of activated NK cells from healthy donors provides an efficient tumor infiltration and a subsequent robust tumoricidal activity against bladder cancer with high selective cytolytic activity against CSCs, leading to a dramatic reduction in tumor burden from 80 % to complete remission.ConclusionAlthough pre-clinical, our results strongly suggest that an immunotherapeutic strategy using allogeneic activated NK cells from healthy donors is effective and should be exploited as a complementary therapeutic strategy in high-risk NMIBC patients to prevent tumor recurrence and progression.Electronic supplementary materialThe online version of this article (doi:10.1186/s12916-016-0715-2) contains supplementary material, which is available to authorized users.

Abstract 5331: Targeting CA125 negative high grade serous ovarian cancer stem cells can result in a dramatic reduction in tumor burden

Abstract High-grade serous ovarian cancers (HGSOC) are deadly gynecologic malignancies that are often metastatic at presentation with a 5 year survival rate of 30-40%. While first-line treatment with carboplatin is often efficacious, disease usually recurs within 6-16 months. With successive rounds of chemotherapy a carboplatin resistant phenotype dominates. Emergence of carboplatin resistance is often thought to be an acquired property of HGSOC but we have recently demonstrated that carboplatin resistance is an innate property of all HGSOCs and inherent to a population of CA125 negative tumor cells with stem properties. The CA125 negative tumor cells comprise a minor population of HGSOCs but evade carboplatin therapy based on in vitro and in vivo drug assays and analysis of primary patient specimens obtained pre and post chemotherapy. We think that targeting the CA125 negative carboplatin resistant serous cancer stem cells (SCSC) will be essential for achieving long-term remissions in this disease. To explore mechanisms of therapy resistance in these cells, the transcriptome of SCSC isolated from 10 primary chemo-naïve HGSOCs was analyzed. An increase in the message levels of inhibitor of apoptosis proteins cIAP1 and cIAP2 was detected in the SCSC. Here we examine the efficacy of the drug Birinapant, a SMAC mimetic that targets cIAPs, in sensitizing human HGSOC to carboplatin therapy. In vitro analysis of 9 primary chemo-naive human HGSOC specimens demonstrated that combined therapy with Birinapant and carboplatin eliminated the chemo-resistant SCSC in 6 specimens, all of which had high levels of cIAP in their SCSC. Next, subcutaneous xenografts were established from 3 independent low-passaged cell lines derived from primary chemo-naive human HGSOCs. In vivo co-treatment of these tumors with Birinapant and carboplatin resulted in significantly diminished tumor burden and a durable response to treatment compared to either therapy alone. These results were further confirmed in a more physiologic intra-peritoneal tumor model. The combination of Birinapant and carboplatin was then tested in targeting in vivo xenografts derived directly from two chemo-resistant human HGSOCs. This co-therapy was efficacious in significantly decreasing tumor burden in the chemo-resistant HGSOC with high cIAP in its SCSC but ineffective in eliminating the tumor where a lower level of cIAP (the Birinapant target) was found in its cancer stem population. Collectively, these results provide proof-of-principle that pharmacologic targeting of the SCSC can be an effective strategy for sensitizing them to carboplatin resulting in a potentially durable therapeutic response in primary and recurrent HGSOC. We think these pre-clinical findings merit testing the efficacy of this combination therapy in clinical trials treating patients with HGSOC with high levels of cIAP in their tumor stem cells. Note: This abstract was not presented at the meeting. Citation Format: Deanna Janzen, Ekaterian Tiourin, Justin Salehi, Jing Lu, Matteo Pellegrini, Sanaz Memarzadeh. Targeting CA125 negative high grade serous ovarian cancer stem cells can result in a dramatic reduction in tumor burden. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5331. doi:10.1158/1538-7445.AM2015-5331

CD24 knockout prevents colorectal cancer in chemically induced colon carcinogenesis and in APCMin/CD24 double knockout transgenic mice

Increased expression of CD24 is seen in a large variety of solid tumors, including up to 90% of gastrointestinal (GI) tumors. Stable derivatives of SW480 colorectal cancer (CRC) cells that overexpress CD24 proliferate faster, and increase cell motility, saturation density, plating efficiency, and growth in soft agar. They also produce larger tumors in nude mice as compared to the parental SW480 cells. Most significantly, even depletion of one copy of the CD24 allele in the APC(Min/+) mice of a transgenic mouse model led to a dramatic reduction in tumor burden in all sections of the small intestine. Homozygous deletion of both CD24 alleles resulted in complete abolishment of tumor formation. Moreover, CD24 knockout mice exhibited resistance to chemically induced inflammation-associated CRC. Finally, a new signal transduction pathway is suggested: namely, CD24 expression downstream to COX2 and PGE2 synthesis, which is directly regulated by β-catenin. CD24 is shown in vitro and in vivo as being an important oncogene in the gut, and one that plays a critical role in the initiation and progression of carcinogenesis.

Highlights from Recent Cancer Literature

Highlights from Recent Cancer Literature

Abstract 945: Inhibition of T-type calcium channels sensitizes ovarian cancer cell growth and metastasis to platinum-based chemotherapy.

Abstract The inability to successfully treat women with ovarian cancer is due in large part to the advanced stage of disease at diagnosis and the development of platinum resistance. T-type calcium channels have recently garnered interest as potential targets for the treatment of many cancers. We investigated the hypothesis that inhibiting T-type calcium channels would sensitize ovarian cancer cells to platinum based chemotherapy using cell culture and mouse models. Sequential treatment of cultured ovarian cancer cell lines or primary cells obtained from patients with mibefradil followed by carboplatin showed a dose-dependent decrease in cell number that was greater than observed with either drug alone. Importantly, treatment of platinum-resistant tumor cells with mibefradil followed by carboplatin also decreased cell number. Moreover, either simultaneous administration of both drugs or treatment with carboplatin followed by mibefradil did not alter cellular response to carboplatin. Based on these findings, we tested the effect of mibefradil and carboplatin in a mouse model of peritoneal ovarian cancer metastasis with platinum-resistant ovarian cancer cells. Tumor-bearing mice were subjected to 3 cycles of Interlaced TherapyTM, which consists of 5 days of mibefradil treatment followed by one dose of carboplatin and 2 days off. We found that while neither drug alone significantly affected tumor growth compared to control, the mice that received the dual treatment had a dramatic reduction in tumor burden. Together, the data provide pre-clinical evidence supporting the use of T-type calcium channel blockers with platinum-based chemotherapy to treat ovarian cancer. Citation Format: Eli V. Casarez, Lloyd Gray, Amir A. Jazaeri, Jill K. Slack-Davis. Inhibition of T-type calcium channels sensitizes ovarian cancer cell growth and metastasis to platinum-based chemotherapy. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 945. doi:10.1158/1538-7445.AM2013-945

Highlights from Recent Cancer Literature

Highlights from Recent Cancer Literature

TLR5 mediated regulation of tumor growth in an inducible model of cancer (162.48)

Abstract It has been reported that up to 25% of the human population carries a dominant point mutation in TLR5 resulting in the loss of TLR5 signaling. TLR5 expression is paramount in modulating the balance of regulatory FoxP3+ and proinflamatory IL-17+ T cell lineages, two cellular subsets with potentially divergent roles in cancer. However, in the context of cancer, the significance of this mutation remains unknown. Sequencing of cDNA isolated from human ovarian tumor specimens revealed that 6% of tumor specimens harbor a functional mutation in TLR5, suggesting that a loss in TLR5 signaling results in “resistance” to cancer. To further investigate the role of TLR5 signaling in cancer progression, we utilized a novel p53 and KRas dependent model of inducible cancer. TLR5-/- mice had a remarkable delay in tumor onset and a dramatic reduction in tumor burden. Corresponding to the delayed tumor onset, we observed that T cells sorted from the tumor draining lymph nodes of TLR5-/- mice were able to vigorously recall against tumor antigens compared to a profound paralysis of effector function in T cells sorted from the draining lymph nodes of WT mice. These data indicate for the first time that pattern recognition receptors may influence the elimination and/or equilibrium phases of tumor progression at extraintestional locations, pinpointing potentially novel pathways to target for immunotherapy.

Dramatic Reduction in Tumor Burden With Neoadjuvant Sunitinib Prior to Bilateral Nephron-sparing Surgery

Neoadjuvant sunitinib has recently been described for the management of renal cell carcinoma. We present the pre and posttreatment images of a 49-year-old male with bilateral biopsy-proven clear cell renal cell carcinoma who underwent treatment with sunitinib prior to nephron-sparing surgery. After four four-week cycles of daily 50 mg sunitinib, the patient demonstrated a dramatic reduction in tumor burden allowing for successful bilateral partial nephrectomy.

Abstract 3110: RNA Polymerase I: A novel target in the treatment of Myc-driven malignancy

Abstract c-MYC plays a prominent role in cancer. Intriguingly, many of the genes regulated by this oncoprotein are associated with ribosome biogenesis and we have previously demonstrated that Myc regulates a major rate limiting step in this pathway, transcription of the 45S rRNA genes by RNA Polymerase I (Pol I). These observations lead us to hypothesize that cMyc's regulation of rRNA synthesis may contribute to its oncogenic properties. We have tested this hypothesis in a mouse model of Myc-driven lymphoma, the EμMyc transgenic mouse. B-cells purified from EμMyc mice display an increased growth rate in comparison to their wild-type littermates, with increased cell volume, total RNA and protein per cell. This phenotype is characterized by higher rates of 45S rRNA transcription and increased expression of factors specific for Pol I transcription. Knockdown of one of these factors, UBF, by RNAi in EμMyc lymphoma cell lines results in a selective proliferative disadvantage of cells in vitro, in a competition assay, and in vivo, in a transplant model. This phenotype is driven by an increased rate of apoptosis associated with a reduction in 45S rRNA transcription. Based on these findings we explored the potential therapeutic effectiveness in this model of a novel specific small molecule inhibitor of Pol I, CX-5461, currently in preclinical development. Transplanted EμMyc tumors showed marked sensitivity to CX-5461 in vivo, with a dramatic reduction in tumor burden in the peripheral blood (97.54%±0.56) and lymph nodes (94.96%±0.90) due to induction of apoptosis 24hrs following a single oral dose at 75mg/kg. Importantly a normal B-cell population was preferentially maintained in treated mice (13%±1.39 wt B220+ cells versus 1.04%±0.24 tumor B220+ cells, as a percentage of total WBC) indicating specificity of the compound for tumor cells. Four doses of CX-5461, 75mg/kg orally every third day, significantly delayed time to endpoint by 9.5 days (P<0.0001) compared to untreated animals. This delay was accompanied by a period of complete remission with normal white blood cell counts (6.76±0.48 ×10^9cells/L) and no identifiable tumor cells in the peripheral blood. Interestingly, in vitro dose curves indicate a dependence of CX-5461 sensitivity on wild-type p53 function (p53 wt and ARF−/− cell line IC50=9.28nM±1.53 in comparison to p53 mutant and p53−/− IC50=1.70uM±0.03), which can be reduced with over expression of Bcl2 (Bcl2 IC50=2.33uM±1.3). Notably even in the more resistant p53 mutant and p53−/− cell lines, cell death also occurred via apoptosis, suggesting p53-dependent and independent mechanisms to be involved in CX-5461 mediated cell death. In summary, this work with UBF RNAi and CX-5461 identifies inhibition of RNA Pol I transcription as a novel and effective target in the treatment of cMyc-driven malignancies, and for the first time establishes that dysregulation of rDNA transcription can directly contribute to malignant transformation. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3110.

Adoptively Transferred Expanded Natural Killer Cells Inhibit Myeloma Tumor Growth In Vivo.

Abstract 953Recently developed culture conditions for expanding and activating natural killer (NK) cells may improve immunotherapeutic options for patients with multiple myeloma. We have previously shown that co-incubation of NK cells from multiple myeloma patients and healthy donors with K562 cells genetically modified to express membrane-bound interleukin 15 and the co-stimulatory molecule 41BBL (K562-mb15-41BBL) leads to a dramatic increase in both NK cell number and anti-myeloma activity in vitro. In this study, we tested the anti-myeloma activity of these expanded NK cells in vivo using a murine model, which supports the growth of myeloma cells in a microenvironment that reproduces that of the human bone marrow. We first implanted human fetal bones subcutaneously into NOD/SCID/IL2Rγ null mice and allowed them to engraft. Luciferase transfected OPM2 myeloma cells were then injected into the human bone fragment. Tumor burden was followed by bio-imaging and ELISA for human Ig. NK cells were expanded from healthy donor PBMC by co-culture with irradiated K562-mb15-41BBL cells in the presence of 300U/ml IL2 for 10-12 days. Myeloma tumor bearing mice were dosed 6-7 days after OPM2 injection (bio-imaging confirmed myeloma engraftment) with expanded NK cells via tail vein injection followed by subcutaneous IL2 to prolong NK cell survival. Flow cytometry was used to track the human NK cells in blood. Histology was performed by H&E staining of formalin-fixed, paraffin embedded tissues harvested at the end of the study. Sufficient NK cells (3×109) for dosing mice were obtained from one unit of healthy donor blood and their ability to kill OPM2 myeloma was confirmed in vitro by chromium release assays. In experiment 1, OPM2-bearing mice received two tail vein injections, 48h apart, comprising PBS, 4 ×107, or 16 ×107 (total dose) expanded NK cells with 100U IL2 given twice per week. We observed significant myeloma growth inhibition in the cohort given 16 ×107 NK cells (p<0.04) and circulating human NK cells could be detected up to day 21 post-administration. Measurement of tumor burden by bio-imaging and ELISA were highly concordant (correlation coefficient = 0.995). Histologic analyses confirmed a dramatic reduction in tumor burden in the mice treated with expanded NK cells and revealed that bone loss was more pronounced in the implanted fetal bones of the control cohort not receiving NK cells. We observed in a subsequent experiment that increasing the IL2 dose to 1000U daily led to in vivo expansion of CSFE-labeled NK cells of up to 10 generations by day 6 post-infusion, which was associated with an enhanced anti-tumor effect in the 16 ×107 dose cohort. Furthermore, we found that multiple injections of NK cells (four injections of 4 ×107 NK cells versus two injections of 8 ×107) were better tolerated in the highest dose cohort (3/3 surviving versus 6/12 surviving). In conclusion, we observed a significant anti-myeloma effect against the aggressive plasma cell leukemia cell line OPM2 with expanded NK cells and low dose IL2. Adjusting the IL2 dose to 1000U daily led to in vivo expansion of NK cells, longer term NK cell persistence, and an increased anti-myeloma effect. Importantly, this IL2 dose is comparable to the 3×106 U daily dose well tolerated in a previous trial with non-expanded NK cells in humans. Single doses of NK cells >8×107 led to high morbidity in this model while multiple doses of 4×107 were well tolerated. Experiments are in progress testing the in vivo effect of expanded NK cells on primary myeloma cell tumors in this model. Our findings support the planned clinical application of ex vivo expanded NK cells. Disclosures:No relevant conflicts of interest to declare.

Intralesional Interferon Alpha-2b Therapy for Adnexal Kaposi Sarcoma

To describe a case of adnexal Kaposi's sarcoma successfully treated with intralesional interferon alpha 2b injections. Case Report. A 31 year old black Haitian male presents with severe nodular induration and erythema of both eyelids with hemorrhagic infiltration of the bulbar and palpebral conjunctivae, advanced on the right. Clinical, histological, and immunohistochemical examination yielded the diagnosis of Kaposi's sarcoma. The patient was subsequently diagnosed with AIDS, started on highly active anti-retroviral therapy (HAART) and systemic doxorubicin, and initially received topical 3 million IU per mL of interferon alpha-2b eyedrops (Intron-A(R), Schering Corporation, Kenilworth, NJ) every 6 hours to the right eye for 9 weeks. This systemic therapy resulted in minimal clinical response with a CD4 count increase from 12 to 61 cells per microliter. Progressive entropion and corneal compromise on the right prompted a trial of intralesional injections of interferon alpha-2b. Over a 4 week period, 3 injections (3 million IU in 0.5mL, 1.5 million IU in 0.5mL, and 1.5 million IU in 0.5mL) were given. This resulted in a dramatic reduction in tumor burden and lid edema, an improvement in lid position, and corneal healing. Ocular adnexal and conjunctival Kaposi's sarcoma was successfully treated with 3 adjuvant intralesional interferon alpha-2b injections leading to a dramatic decrease in tumor mass.

Suppression of Peutz—Jeghers polyposis by inhibition of cyclooxygenase-2

Peutz-Jeghers syndrome (PJS) is typically manifested as severe gastrointestinal polyposis. Polyps in PJS patients and in Lkb1(+/-) mice that model PJS polyposis are frequently characterized by elevated cyclooxygenase-2 (COX-2). This study was designed to determine whether COX-2 inhibition would reduce tumor burden in Lkb1(+/-) mice or Peutz-Jeghers patients. Genetic interactions between Cox-2 and Lkb1 in polyp formation were analyzed in mice with combined deficiencies in these genes. Pharmacologic inhibition of COX-2 was achieved by supplementing the diet of Lkb1(+/-) mice with 1500 ppm celecoxib between 3.5-10 and 6.5-10 months. In PJS patients, COX-2 was inhibited with a daily dose of 2 x 200 mg celecoxib for 6 months. Total polyp burden in Lkb1(+/-) mice was significantly reduced in a Cox-2(+/-) (53%) and in a Cox-2(-/-) (54%) background. Celecoxib treatment initiating before polyposis (3.5-10 months) led to a dramatic reduction in tumor burden (86%) and was associated with decreased vascularity of the polyps. Late treatment (6.5-10 months) also led to a significant reduction in large polyps. In a pilot clinical study, a subset of PJS patients (2/6) responded favorably to celecoxib with reduced gastric polyposis. These data establish a role for COX-2 in promoting Peutz-Jeghers polyposis and suggest that COX-2 chemoprevention may prove beneficial in the treatment of PJS.

Regression of upper gastric cancer in mice by FHIT gene delivery.

Fhit expression is reduced in most cancers, and Fhit replacement by FHIT expression viruses in lung, esophageal, pancreatic, and cervical cancers induces apoptosis in the cancer cells. Mice carrying one or two inactivated Fhit alleles are hypersensitive to development of N-nitrosomethylbenzylamine (NMBA)-induced forestomach tumors. In the present study, we investigated the kinetics and mechanism of tumor reversal and intervention by oral delivery of FHIT expression viruses. Tumor analysis showed that: a) by 37 days post-NMBA, control mice showed approximately 7 tumors and by 84 days approximately 10 tumors/forestomach; b) mice receiving FHIT virus at 2 or 42 days post-NMBA showed significantly reduced tumor burdens; c) Fhit was still expressed at 82 days postinfection; d) control viral infection had no effect on tumor development; and e) reduced Bcl2, increased Bax expression, and increased TUNEL-positive apoptotic nuclei characterized the restored epithelia of FHIT transduced forestomachs. Thus, FHIT viral gene delivery prevents or retards development of carcinogen-induced forestomach tumors and reverses development of established tumors by 60-70% through an apoptotic pathway. This dramatic reduction in tumor burden emphasizes the efficacy of targeting the FHIT apoptotic pathway for tumor eradication.