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Kaposi sarcoma (KS)-associated herpesvirus (KSHV)–associated multicentric Castleman disease (MCD) is a relapsing and remitting systemic lymphoproliferative disorder characterized by severe inflammatory symptoms most common among people living with HIV (PLWH). Patients with KSHV-MCD may present with concurrent KSHV-associated diseases, such as KS and/or primary effusion lymphoma (PEL). We evaluated clinical and immunologic characteristics, the effects of concurrent KSHV malignancies, and treatments from the largest prospective natural history study of participants with KSHV-MCD within the United States. Treatment options administered at investigator discretion included high-dose zidovudine with valganciclovir (AZT/VGC), rituximab, or rituximab with liposomal doxorubicin (R-Dox) during KSHV-MCD flares. Survival analyses and prognostic factors were explored for all participants. Sixty-two participants with HIV were enrolled, including 20 with KSHV-MCD alone, 34 with KSHV-MCD and KS, 1 with KSHV-MCD and PEL, and 7 with all KSHV-associated diseases. Forty-four percent of KSHV-MCD diagnoses were made at our institution. Forty-four participants received rituximab-based therapies, 20 of whom had maintenance AZT/VGC or interferon. Participants receiving R-Dox and then maintenance AZT/VGC had the highest 5-year progression-free survival (89%). Cytokine profiles during KSHV-MCD flares did not differ by the presence of concurrent KSHV-associated diseases. The 10-year survival was 71% (95% confidence interval [CI], 56% to 82%) for all participants. A concurrent diagnosis of PEL negatively impacted survival (PEL hazard ratio, 5.4; 95% CI, 1.8 to 16.8). KSHV-MCD is an underdiagnosed condition among PLWH, including those with KS. KSHV-MCD has an excellent prognosis with appropriate treatment. Physicians should be alert for patients with multiple KSHV diseases, which impact optimal treatment and survival outcomes. This study was registered at www.clinicaltrials.gov as #NCT00099073.

Background: Primary effusion lymphoma (PEL) is a rare, B cell non-Hodgkin lymphoma (NHL) caused by Kaposi sarcoma herpesvirus (KSHV), also known as human herpesvirus 8, that is strongly associated with HIV infection. PEL often presents as malignant body cavity effusions but can also present as extracavitary masses. Patients may also have concurrent Kaposi sarcoma (KS) or multicentric Castleman disease (MCD). 80% of PEL is also positive for EBV, and in a cohort of 20 patients we previously showed EBV + PEL had improved overall survival (OS) (Lurain et al. Blood 2019). PEL prognosis is poor compared to other HIV-associated NHL with median survival of 6-10 months. While patients are often treated with chemotherapy regimens used in other NHL, the optimal front-line and subsequent-line treatments are unknown. In this largest cohort of patients with PEL in the United States, we evaluated survival, prognostic markers, and second-line treatments in relapsed/refractory disease. Methods: We analyzed clinical records of all patients with PEL treated in the HIV and AIDS Malignancy Branch at the National Cancer Institute from 2000-2021. We collected demographics, disease characteristics, concurrent KS and MCD, EBV status of tumor by EBER staining, and laboratory data at diagnosis as well as at progression and subsequent lines of therapy. We calculated overall survival (OS) of all patients and progression-free survival (PFS) in those who received front-line EPOCH by Kaplan-Meier method. The survival of extracavitary vs. cavitary PEL was compared using an exact log-rank test. A univariate Cox proportional hazard model was performed to determine prognostic variables associated with OS. Results: We identified 40 patients with PEL: 38 cisgender men, 2 cisgender women; 18 non-Hispanic Black, 6 Hispanic, 16 non-Hispanic White; median age 41 years. At diagnosis of PEL, 29 patients had concurrent KS, and 7 had concurrent MCD. Eight patients had only extracavitary disease, and 5 had leptomeningeal disease. All but 1 had HIV, and all with HIV received antiretroviral therapy. At diagnosis, median HIV viral load (VL) was 50 (IQR: 27-8323) copies/mL, median CD4 + cell count was 100 (IQR:44-395) cells/µL, and median KSHV VL was 4147 copies/10 6 cells (IQR:178-68,526). One patient died prior to receiving treatment. Of the 39 patients who received front-line treatment, 38 received combination chemotherapy (EPOCH [36], CHOP [1], ICE [1]) with rituximab added in 34 patients to treat concurrent MCD or deplete the KSHV-infected B cell reservoir. One patient received pembrolizumab as up-front therapy due to concurrent progressive multifocal leukoencephalopathy. Twenty-three (57%) had complete remission (CR) by Lugano criteria. Median PFS after front-line EPOCH was 16.4 months (Figure 1a), and median OS in all patients was 35.6 months (Figure 1b), with 5-year OS of 44%. Median OS for patients with extracavitary disease was significantly higher than in patients with cavitary disease (p=0.023, Figure 1c). EBER-positive tumors (HR=0.21, p=0.005) were associated with improved survival, while concurrent KS, MCD, leptomeningeal disease, and baseline CD4 and KSHV VL were not associated with OS outcomes. Eleven patients with relapsed/refractory disease received second-line therapy, including combination chemotherapy (3); brentuximab (3); pembrolizumab (1); pembrolizumab and pomalidomide (1); interferon-alpha, ganciclovir, and bevacizumab (1); bortezomib, ganciclovir, and zidovudine (1); and radiation to pelvic lymph nodes (1). Five of these patients received subsequent treatment after disease progression, including 2 patients with leptomeningeal PEL, 1 who received daratumumab and 1 who received pomalidomide and pembrolizumab with ongoing CRs at 12 months and 33 months, respectively. Conclusion: Patients with PEL had a median OS of 35.6 months, which is improved from previous reports. Patients with extracavitary PEL appear to have improved OS, and we confirmed our prior study noting EBER-positive PEL is a positive prognostic marker. Promising chemotherapy-sparing treatment regimens in relapsed/refractory PEL, include pembrolizumab, pomalidomide, and daratumumab. Figure 1 Figure 1. Disclosures Uldrick: Regeneron: Current Employment; Roche: Research Funding; Merck: Other: Receives study drug; Celgene: Research Funding. Yarchoan: Celgene: Research Funding; Merck: Other: Clinical Trial Support; EMD-Serano: Other: Clinical Trial Support; Eli Lilly: Other: Clinical Trial Support; CTI BioPharma: Other: Clinical Trial Support; Janssen Pharmaceuticals: Other: Clinical Trial Support; U.S. Dept of Health and Human Services: Patents & Royalties. Lurain: EMD-Serrono: Research Funding; BMS-Celgene: Research Funding; CTI Biopharma: Research Funding; Merck: Research Funding; Janssen: Research Funding.

Background: Primary effusion lymphoma (PEL) is a rare, aggressive B-cell neoplasm strongly associated with HIV infection. It generally presents with malignant effusions but may also present with extracavitary (EC) masses. It is caused by the gammaherpesvirus Kaposi sarcoma herpesvirus (KSHV, also called human herpesvirus 8), which is also responsible for the development of Kaposi sarcoma (KS) and a form of multicentric Castleman disease (MCD). There are no prospective studies in PEL and no standard therapy. Prognosis is poor compared to other HIV-associated lymphomas with median survival of 10-22 months when treated with conventional lymphoma chemotherapy regimens. In vitro data demonstrate lenalidomide (LEN), an immunomodulatory agent, downregulates IRF4, a cell activation marker overexpressed in PEL, and can reverse KSHV-induced downregulation of MHC-1 and ICAM-1. Dose-adjusted infusional etoposide, vincristine, and doxorubicin with cyclophosphamide and prednisone combined with rituximab (EPOCH-R) has been shown to be safe and effective for CD20+ HIV-associated lymphomas. While PEL is generally CD20 negative, rituximab eradicates the KSHV-infected B-cell reservoir, a source of inflammatory cytokines that drive the natural history of PEL; moreover, many patients have concurrent MCD for which rituximab is a standard treatment. Methods: In a prospective phase 1/2 study conducted in the HIV and AIDS Malignancy Branch at the National Cancer Institute (NCI), we are evaluating EPOCH combined with rituximab and LEN (EPOCH-R2) in participants (pts) with untreated PEL. The primary objective was to evaluate safety and tolerability of EPOCH-R2 and determine the recommended phase 2 dose (RP2D) of LEN. PEL was diagnosed/confirmed in the NCI Laboratory of Pathology via cytology and/or flow cytometry of effusions or via biopsy in EC disease. Pts received EPOCH-R days 1-5 and every 21 days for 6 cycles. LEN was administered orally days 1-10 of each cycle starting at 25 mg (with dose de-escalation if toxicity occurred). Dose-limiting toxicities were evaluated during cycles 1-2. Pts with leptomeningeal PEL (CSF-PEL), determined by cytology and/or flow cytometry of cerebrospinal fluid, received intrathecal chemotherapy; intrathecal prophylaxis was given to pts without CSF-PEL. All pts received thromboprophylaxis (aspirin 81 mg daily), opportunistic infection prophylaxis, and antiretroviral therapy (ART). Adverse events (AEs) were evaluated using CTCAEv5 and treatment response was evaluated by Lugano criteria after cycles 2 and 6. Response to treatment, immune reconstitution, and overall survival (OS) were evaluated using descriptive statistics, Wilcoxon signed-rank test and Kaplan-Meier methodology. Results: We are reporting results of the completed Phase I portion of the trial. 6 HIV+ cisgender men (5 Black, 1 White) with stage 4 PEL were enrolled July 2017-August 2019. All received non-boosted integrase inhibitor-based ART. 3 had pleural effusions, 2 had EC disease without effusions, and 1 had both effusions and EC disease. 3 had CSF-PEL and 1 had bone marrow involvement. 4 had concurrent KS; 1 also had concurrent MCD. Median CD4+was 231 cells/µL (IQR: 10, 310) at baseline and 189.5 cell/µL (IQR: 56, 224) at end-of-treatment, which was not a significant decline (p=0.46). The most common AEs were hematologic, including grade (G) 4 neutropenia (100%), leukopenia (100%), thrombocytopenia (67%) and CD4+ lymphopenia (67%). 3 pts (50%) developed G3 pulmonary emboli despite thromboprophylaxis. There were 7 episodes of G3 febrile neutropenia and 2 episodes of sepsis. No pts developed opportunistic infections. There were no dose-limiting toxicities and LEN 25 mg is the RP2D. 4 pts completed all 6 cycles. 2 pts completed 5 cycles: 1 died due to progressive disease and 1 had progressive disease and went on to receive additional therapy. 1 pt who completed all 6 treatment cycles died due to HIV-related complications 5 months after EPOCH-R2 with no evidence of PEL at autopsy. The response rate was 83.3% (95% CI: 36.8-99.6) and 50% (95% CI: 11.8-88.1) after cycles 2 and 6, respectively. 2-year overall survival was 66.7% (95% CI:19.5-90.4). Conclusions: Front-line PEL treatment with EPOCH-R2 is safe, and the most common toxicities were hematologic. This regimen showed preliminary evidence of activity and good OS, which will be further evaluated in the ongoing phase 2. Disclosures Lurain: Celgene: Research Funding; EMD-Serrono: Research Funding; Merck: Research Funding. Ramaswami:Celgene: Research Funding; EMD-Serrono: Research Funding; Merck: Research Funding. Whitby:Patent: Patents & Royalties: Co-inventor on US Patent 10,001,483 entitled "Methods for the treatment of Kaposi's sarcoma or KSHV-induced lymphoma using immunomodulatory compounds, and uses of biomarkers.. Uldrick:Patent: Patents & Royalties: Co-inventor on US Patent 10,001,483 entitled "Methods for the treatment of Kaposi's sarcoma or KSHV-induced lymphoma using immunomodulatory compounds, and uses of biomarkers."; Celgene: Research Funding; Merck: Research Funding; Roche: Research Funding. Yarchoan:Patent: Patents & Royalties: Coinventor on patents on a peptide vaccine for HIV and on the treatment of Kaposi sarcoma with IL12; EMD-Serrono: Patents & Royalties; Patent: Patents & Royalties: Co-inventor on US Patent 10,001,483 entitled "Methods for the treatment of Kaposi's sarcoma or KSHV-induced lymphoma using immunomodulatory compounds, and uses of biomarkers.; Patent: Patents & Royalties: An immediate family member of R. Yarchoan is a co-inventor on patents related to internalization of target receptors, on KSHV viral IL-6, and on the use of calreticulin and calreticulin fragments to inhibit angiogenesis; Celgene: Research Funding; Merck: Research Funding. OffLabel Disclosure: Dose-adjusted infusional etoposide, vincristine, and doxorubicin with cyclophosphamide and prednisone combined with rituximab and lenalidomide for the use of primary effusion lymphoma will be discussed.

Kaposi sarcoma-associated herpesvirus (KSHV) is a principal causative agent of primary effusion lymphoma (PEL) with a poor prognosis in immunocompromised patients. However, it still lacks effective treatment which urgently requires the identification of novel therapeutic targets for PEL. Here, we report that the hepatocyte growth factor (HGF)/c-MET pathway is highly activated by KSHV in vitro and in vivo. The selective c-MET inhibitor, PF-2341066, can induce PEL apoptosis through cell cycle arrest and DNA damage, and suppress tumor progression in a xenograft murine model. By using microarray analysis, we identify many novel genes that are potentially controlled by HGF/c-MET within PEL cells. One of the downstream candidates, ribonucleoside-diphosphate reductase subunit M2 (RRM2), also displays the promising therapeutic value for PEL treatment. Our findings provide the framework for development of HGF/c-MET-focused therapy and implementation of clinical trials for PEL patients.

Kaposi sarcoma (KS) herpesvirus-associated multicentric Castleman disease (KSHV-MCD) is a lymphoproliferative disorder, most commonly seen in HIV-infected patients, that has a high mortality if untreated. Concurrent KS is common. Although rituximab has reported activity in KSHV-MCD, its use is often associated with KS progression. Within a natural history study of KSHV-MCD, we prospectively evaluated rituximab 375 mg/m(2) combined with liposomal doxorubicin 20 mg/m(2) (R-Dox) every 3 weeks in 17 patients. Patients received a median of 4 cycles (range 3-9). All received antiretroviral therapy, 11 received consolidation interferon-α, and 6 received consolidation high-dose zidovudine with valganciclovir. Using NCI KSHV-MCD response criteria, major clinical and biochemical responses were attained in 94% and 88% of patients, respectively. With a median 58 months' potential follow-up, 3-year event-free survival was 69% and 3-year overall survival was 81%. During R-Dox therapy, cutaneous KS developed in 1 patient, whereas 5 of 6 patients with it had clinical improvement. R-Dox was associated with significant improvement in anemia and hypoalbuminemia. KSHV viral load, KSHV viral interleukin-6, C-reactive protein, human interleukin-6, and serum immunoglobulin free light chains decreased with therapy. R-Dox is effective in symptomatic KSHV-MCD and may be useful in patients with concurrent KS. This trial was registered at www.clinicaltrials.gov as #NCT00092222.

Primary effusion lymphoma (PEL) is a rare, aggressive B-cell neoplasm with a unique clinical profile that is most frequently associated with HIV infection. It generally presents with malignant effusions but may also present with extracavitary masses. It is caused by the gamma-herpesvirus Kaposi sarcoma herpesvirus (KSHV, also called human herpesvirus-8 [HHV-8]), which is also the etiologic agent of Kaposi sarcoma and the plasmablastic form of multicentric Castleman disease (KSHV-MCD). KSHV is known to cause cytokine related severe inflammation associated with elevated human interleukin (IL)-6 and IL-10. There have been no prospective studies in PEL and there is currently no standard therapy for this disease. The prognosis of PEL is poor compared to other HIV-associated lymphomas. PEL has a median survival of 10-22 months when treated with conventional chemotherapy regimens for non-Hodgkin lymphoma according to the most recent retrospective reports. Lenalidomide, a derivative of thalidomide with immunomodulatory activity, has been shown to have in vitro antitumor effects in PEL cell lines. KSHV itself may cause downregulation of immune surface markers as a way to avoid immune detection. Lenalidomide can reverse this and enhance surface expression of major histocompatibility complex-1 and intercellular adhesion molecule-1 in PEL cell lines. Dose-adjusted infusional etoposide, vincristine, and doxorubicin with cyclophosphamide and prednisone (DA-EPOCH) is an anthracycline-based regimen that allows for personalization of dose-intensity, and prior anecdotal experience indicates it has activity in PEL. In combination with rituximab, an anti-CD20 monoclonal antibody, DA-EPOCH has been shown to be safe and effective for HIV-associated diffuse large B-cell lymphoma and Burkitt lymphoma. PEL is a CD20 negative tumor; however, many patients have concurrent KSHV-MCD for which rituximab is the standard treatment. In addition, rituximab may aid in eradication of the KSHV B-cell reservoir, a source of inflammatory cytokines that drive the natural history of PEL. Patients of any HIV serostatus with both effusion and extracavitary presentations, including those with concurrent KSHV-associated diseases, such as Kaposi sarcoma and KSHV-MCD, are eligible. Patients with ECOG performance status of ≤4 are eligible for enrollment. The phase I portion of the study will evaluate the safety, tolerability, and maximum tolerated dose of lenalidomide combined with DA-EPOCH and rituximab (DA-EPOCH-R2). The phase II component will evaluate the activity and overall survival. Patients will receive 6, 21-day cycles of DA-EPOCH-R2. Lenalidomide, initially at 25 mg, will be given on days 1 to 10. Patients will receive rituximab on day 1 and DA-EPOCH on days 1 to 5. Patients with HIV will be prescribed antiretroviral therapy as this is central to controlling HIV viremia and managing KSHV-associated malignancies. The lenalidomide dose will be de-escalated in a second dose group if 2 of 6 patients in phase 1 experience a dose-limiting toxicity. In the phase II portion of the study, 15 evaluable patients will be enrolled at the maximal tolerated dose of lenalidomide. At 12 months follow-up after the last patient has enrolled, a 1-tailed 0.10 alpha level test would have 80% power to determine if the overall survival curve would demonstrate a 1-year overall survival consistent with 45% or better and ruling out 20% or worse survival. Secondary outcomes include the evaluation of the pharmacokinetics of lenalidomide in blood, effusions, and CSF and the effect of DA-EPOCH-R2on concurrent KS, KSHV-MCD, and the KSHV-associated inflammatory cytokine syndrome. The study will also examine the effect of lenalidomide alone and in combination with rituximab and DA-EPOCH on the KSHV viral load, serum cytokines, lymphocyte subset reconstitution, HIV latency reversal, cellular measures of HIV, and the pharmacokinetics of tenofovir and tenofovir-diphosphate in plasma and peripheral blood mononuclear cells. The study began enrollment in July 2017, and 5 patients have been enrolled on the phase I portion. All 5 patients have completed treatment without dose limiting toxicities. Disclosures Lurain: Celgene: Other: I receive research support from Celgene through a CRADA at the NCI; Merck: Other: I receive drug for a clinical trial from Merck through a CRADA with the NCI. Ramaswami:Celgene: Other: I receive research support from Celgene through a CRADA at the NCI; Merck: Other: I receive drug for a clinical trial from Merck through a CRADA with the NCI. Whitby:Patent: Patents & Royalties: co-inventor on US Patent 10,001,483 entitled "Methods for the treatment of Kaposi's sarcoma or KSHV- induced lymphoma using immunomodulatory compounds, and uses of bio- markers.". Uldrick:Merck: Other: drug for a clinical trial from Merck through a CRADA with the NCI; Roche: Other: commercial research support through a CTA with Fred Hutchinson Cancer Research Center; Celgene: Other: research support from Celgene through a CRADA at the NCI; Patent: Patents & Royalties: co-inventor on US Patent 10,001,483 entitled . Yarchoan:Celgene: Other: I research support from Celgene through a CRADA at the NCI; Merck: Other: I receive drug for a clinical trial from Merck through a CRADA with the NCI; Patent: Patents & Royalties: coinventor on US Patent 10,001,483 entitled ; Patent: Patents & Royalties: coinventor on patents on a peptide vaccine for HIV and on the treatment of Kaposi sarcoma with IL12; Patent: Patents & Royalties: immediate family member is co-inventor on patents related to internal- ization of target receptors, on KSHV viral IL6, and on the use of calreticulin and calreticulin fragments to inhibit angiogenesis.

BACKGROUND: Kaposi sarcoma herpesvirus-associated multicentric Castleman disease (KSHV-MCD) is a rare lymphoproliferative disorder that occurs primarily in HIV-infected patients and is characterized by inflammatory symptoms with a waxing and waning course. If untreated, it leads to multiorgan failure and death, usually within 2 years. KSHV-MCD symptoms are caused by increased levels of inflammatory cytokines, including human interleukins (IL) 6 and 10, and a KSHV-encoded IL-6 analog (vIL-6). Rituximab is an effective therapy in KSHV-MCD but can result in the development or worsening of Kaposi sarcoma (KS) in those with concurrent diagnoses. Other therapies include rituximab plus liposomal doxorubicin and zidovudine (AZT) plus valganciclovir (VGC). Tocilizumab, a humanized anti-IL-6 receptor (gp80) antibody, has demonstrated benefit in KSHV-negative MCD, although its utility in patients with KSHV-MCD is unknown. We explored the safety and efficacy of tocilizumab in KSHV-MCD and the effect of the addition of AZT and VGC in cases where tocilizumab alone does not lead to clinical benefit. PATIENTS & METHODS: Patients enrolled on this prospective pilot study had symptomatic KSHV-MCD. They received 8mg/kg of tocilizumab every 2 weeks for up to 12 weeks, or 6 cycles. Patients with HIV were required to continue antiretroviral therapy. Treatment responses were assessed using a KSHV-MCD clinical benefit response (CBR) criteria, which consists of 8 indicator abnormalities (4 symptom groups and 4 laboratory parameters) that are closely associated with disease activity. If patients had evidence of progression or lack of improvement with tocilizumab monotherapy, AZT 600mg orally every 6 hours and VGC 900mg orally every 12 hours on days 1-5 of a 14-day cycle could be administered with tocilizumab. Peripheral blood mononuclear cell (PBMC)-associated KSHV viral loads were quantified by PCR using primers to the KSHV gene K6 after each cycle of treatment. The primary objective of the study was to estimate the clinical benefit of tocilizumab in patients with symptomatic KSHV-MCD using a modified KSHV-MCD CBR criteria. RESULTS: Eight HIV positive patients (7 male, 1 female) with a median age of 48.8 years were enrolled. All patients were taking combined antiretroviral therapy and had a baseline CD4+ T cell count of 254 cells/mL. Four patients (50%) had prior KSHV-MCD therapy with rituximab and 3 patients (38%) had KS at the time of enrollment. With tocilizumab alone, the overall response rate (partial and complete responses using the CBR criteria for all patients was 63% (95% confidence interval 25% to 92%). Three patients required the addition of AZT and VGC; one patient required AZT and VGC after 1 cycle and 2 required the combination treatment after 3 cycles of tocilizumab. Two out of 3 patients had a complete response after the addition of AZT and VGC to tocilizumab. Among all patients, 3 patients stopped therapy prior to 6 cycle due to progression of symptoms despite combination therapy, worsening pulmonary KS symptoms and deterioration in performance status due to progressive KSHV-MCD. Two remaining patients with concurrent KS had stable disease at the end of treatment. Among all patients, the median time to next therapy for symptomatic KSHV-MCD was 3.2 months (range 1 - 37 months); subsequent therapies included rituximab alone (3 patients), or in combination with liposomal doxorubicin (2 patients), or treatment with pomalidomide and liposomal doxorubicin (2 patients). One patient had a durable complete response over 3 years of follow-up. Treatment was well tolerated; the most common grade 3 and 4 adverse events included thrombocytopenia and neutropenia as well as hyperuricaemia, which were attributed to KSHV-MCD rather than study therapies. Furthermore, tocilizumab alone or in combination with AZT and VGC did not interfere with the CD4+ T cell count (net increase of 16 cells cells/mL) or HIV viral load. There was an overall decrease in KSHV viral load of 4577 copies/mL from baseline to the final cycle, reflecting clinical response. CONCLUSION: Tocilizumab is safe in patients with HIV and may have a role in the management of symptoms associated with KSHV-MCD. Incomplete responses to tocilizumab may occur because tocilizumab binds to the gp80 IL-6 receptor and KSHV vIL-6 can bind to the gp130 receptor subunit without the requirement for gp80. The addition of AZT and VGC to tocilizumab also showed clinical benefit. Disclosures Uldrick: Celgene: Research Funding; Celgene: Patents & Royalties: 10,001,483 B2; Merck: Research Funding. Yarchoan:NIH: Patents & Royalties: Patents on IL-12 for KS and cereblon-binding drugs for KSHV diseases. Spouse has patent on KSHV IL-6. Patents assigned to DHHS/NIH.; Celgene Corp.: Research Funding.

BACKGROUND: Kaposi sarcoma herpesvirus (KSHV, also called human herpesvirus-8) is the causative agent of 3 disorders: primary effusion lymphoma (PEL), Kaposi sarcoma (KS), and a plasmablastic form of multicentric Castleman disease (KSHV-MCD). It also causes KSHV inflammatory cytokine syndrome (KICS), which is characterized by inflammatory symptoms and an elevated KSHV viral load. Multiple KSHV-associated diseases, which usually develop in HIV-infected patients, can present together in the same patient. Effusions can occur in each of these diseases, thereby presenting a diagnostic challenge. Identifying PEL is especially crucial as prompt treatment with multi-agent chemotherapy can be curative. We analyzed effusions from patients with KICS, PEL, and KSHV-MCD to identify distinct immunologic characteristics and virologic profiles that may aid diagnosis, inform treatment and elucidate pathogenesis. PATIENTS AND METHODS: We identified 22 HIV-infected patients with effusions [pleural effusions (20), ascites (1) and pericardial effusion (1)] with diagnoses of PEL (9 patients), KICS (8 patients), or KSHV-MCD (5 patients). All patients had a concurrent diagnosis of KS. We obtained clinical and immunologic characteristics from effusions and paired serum samples at the same timepoint for each patient. Serum and effusion cytokine levels of interleukin (IL)-1ß, IL-2, IL-4, IL-6, IL-8, IL-10, IL-13, and IL-12p70; interferon gamma (IFN-g); tumor necrosis factor alpha (TNF-a); vascular endothelial growth factor (VEGF); and inducible protein 10 (IP-10) were evaluated using a commercial multiplex assay. Peripheral blood mononuclear cell (PBMC) and effusion- associated KSHV and Epstein Barr virus (EBV) viral DNA (KSHV-VL, EBV-VL) in PBMC and cells were quantified using PCR with primers to KSHV K6 and EBV pol. Effusion and serum immunologic and virologic characteristics were compared within each disease and separately between diseases using the Wilcoxon sign rank test and Wilcoxon rank sum test, respectively. In these exploratory analyses with few patients, no correction was made for multiple comparisons. RESULTS: In patients with PEL, the median (med) age was 42 years with med CD4+ count of 54 T-cells/μL and HIV viral load (VL) of 325 copies/mL. In those with KICS, the med age was 32 years, with a med CD4+ count of 119 T-cells/μL and HIV VL of 48 copies/mL. In patients with KSHV-MCD, the med age was 31 years, med CD4+ count of 118 T-cells/μL and HIV VL was 75 copies/mL. IL-13 was substantially higher in PEL effusions as compared to serum levels (med 16.9 vs. In analyses comparing serum and cytokine differences among diseases, effusions from patients with PEL had detectable levels of IL-13 (med 16.9 ng/ml; interquartile range 9.7-26.9 ng/ml) compared to patients with KSHV-MCD (med CONCLUSIONS: In HIV-infected patients, effusions can indicate a diagnosis of PEL, KSHV-MCD and/or KICS. Quantifying KSHV VL in the effusion may be useful in the diagnosis of PEL. Effusions in PEL had a distinct profile compared to the circulation or other KSHV-associated diseases, particularly with regard to elevated IL-13, which may aid in diagnosis. In contrast, the inflammatory and virologic findings in KSHV-MCD and KICS effusions roughly paralleled the levels seen in the circulation. This study suggests that KSHV upregulation of IL-13 is a unique feature of PEL, which may contribute to PEL pathogenesis through STAT6 activation and be a potential future therapeutic target. Disclosures Uldrick: Celgene: Patents & Royalties: 10,001,483 B2; Celgene: Research Funding; Merck: Research Funding. Yarchoan:Celgene Corp.: Research Funding; NIH: Patents & Royalties: Patents on IL-12 for KS and cereblon-binding drugs for KSHV diseases. Spouse has patent on KSHV IL-6. Patents assigned to DHHS/NIH..

Introduction: Primary effusion lymphoma (PEL) and large cell lymphoma arising in Kaposi sarcoma-associated herpesvirus associated multicentric Castleman disease (KSHV-MCD) are KSHV-associated non-Hodgkin lymphomas (together, KSHV-NHL) with overlapping clinical and pathologic features. KSHV-NHL have biologic profiles distinct from other AIDS-related lymphomas, a poor prognosis and no established therapy. Immune dysregulation, including KSHV-associated interleukin (IL)-6 syndromes, contribute to pathobiology and may affect outcomes. We hypothesized that biomarkers of KSHV-associated immune activation and lymphoproliferation could predict overall survival (OS) in patients with KSHV-NHL treated with modified dose-adjusted (DA)-EPOCH (continuous infusional etoposide, vincristine, and doxorubicin with cyclophosphamide and prednisone). Methods:We analyzed OS in a retrospective series of 19 patients with pathology-confirmed KSHV-NHL diagnosed 2000-2013 who were evaluated in the HIV & AIDS Malignancy Branch Clinic and received curative intent modified DA-EPOCH regimens. Therapies combined with DA-EPOCH included rituximab, bevacizumab, and/or high dose methotrexate. All patients received combination antiretroviral therapy. Baseline clinical laboratories and immunologic biomarkers as well as treatment factors were evaluated for prognostic value. Continuous parameters were dichotomized near the median (med). OS was evaluated using Kaplan-Meier methods and 2-sided log-rank test. Multivariable analysis employed Cox regression with backward selection, initially including variables with a univariate p ² 0.10. Correlations between significant prognostic markers were evaluated by Spearman's rank correlation. Results: Med age was 44 years (22-69); 1 woman, 18 men; 17 had PEL, with extracavitary involvement in 11; 2 had large cell lymphoma arising in KSHV-MCD. 42% had HIV viral load (VL) < 100 copies/mL. Med CD4 was 120 cells/uL (Table 1). Med number of chemotherapy cycles was 6 (range 1-13). Best response: 10 patients (53%) had complete response, 2 achieved stable disease and 7 progressed. With 3.3 years med potential follow up, med OS was 1.8 years and 3-year OS was 42.8%. Baseline predictors for inferior OS in univariate analysis, were elevated interferon-gamma (IFNg), IL-10, IL-6, immunoglobulin E (IgE), and KSHV VL (Figure 1). Elevated ferritin (p=0.07) and decreased platelets (p = 0.1) also predicted OS. Hemoglobin, albumin, C-reactive protein, serum free light chains (FLC), LDH, CD4 count, HIV viremia, IL-8, tumor necrosis factor-a (TNFa), and treatment factors did not predict OS. Moderately strong to strong correlation between IL-10, IL-6 and IFNg (r=0.64 - 0.91); IL6 and IL-10 with ferritin (r=0.51- 0.59); and IL-10 with IgE (r=0.54) but not ferritin with IgE (r=0.11) were observed and affected selection models. In a Cox model with backward selection, levels above the med for IgE (HR 30, p=0.01) and ferritin (HR=13, p=0.03) were jointly predictive of OS. Excluding IgE from the model, elevated IL-10 was the best predictor of OS. Conclusion: Modified DA-EPOCH resulted in 3-year OS of 43%. Improved targeted therapies are needed. Hyperferritinemea, elevated IgE and inflammatory human cytokine profiles associated with dysregulated KSHV-lytic activation (IL-10, IL-6 and KSHV-VL) and macrophage activation syndromes (ferritin, IFNg, IL-6 and IL-10) suggest that life-threatening abnormalities in both innate and acquired immunity are common in KSHV-NHL and contribute to pathogenesis and prognosis. IgE and ferritin require further evaluation in KSHV-associated diseases. Table 1. Baseline characteristics Characteristic Value Demographics Age, med (range)(IQR) 44(22-69) Sex, M:F 18:1 Race n(%) White 9 (47) Hispanic 4 (21) African American 4 (21) African Immigrant 2 (11) Clinical Characteristics n(%) KS 14(74) KSHV-MCD 5(26) HIV VL Figure 1: Overall survival by immune parameters and KSHV viral load Figure 1:. Overall survival by immune parameters and KSHV viral load Disclosures Uldrick: Celgene Corporation: Research Funding. Off Label Use: Bevacizumab use in primary effusion lymphoma. Tosato:NIH: Has patent for KSHV-viral IL-6 assay Patents & Royalties. Yarchoan:Celgene: Research Funding.

Abstract 2708 Background: KSHV, also known as human herpesvirus-8, is the etiologic agent of a lymphoproliferative disorder, KSHV-associated multicentric Castleman disease (KSHV-MCD), two lymphomas: primary effusion lymphoma (PEL) and large cell lymphoma arising in KSHV-MCD (together, KSHV-NHL) and Kaposi sarcoma (KS). KSHV-associated diseases mainly occur in patients with HIV. KSHV is notable for its modulation of host immune response, including induction of IL6 and IL10; and an IL6-related KSHV-associated inflammatory cytokine syndrome (KICS) has been described in HIV/KSHV infected patients without KSHV-MCD. The clinical features and natural history of KSHV-NHL are not well understood. We hypothesize KSHV-NHL is associated with pathophysiologic features similar to those of KSHV-MCD, and is unique among HIV-associated lymphomas. Methods: Clinical records of patients with KSHV-NHL diagnosed 2000-12, treated in the HIV and AIDS Malignancy Branch Clinic were reviewed, and evaluated for clinical, and laboratory abnormalities using our criteria for KICS (NCT01419561): at least 2 select clinical and laboratory manifestations, not otherwise explained; elevated c-reactive protein [CRP]; and KSHV viral load greater than 100 copies/106 PBMC; Additionally, serum inflammatory cytokines, (IFN-gamma, TNF-alpha, IL-1beta, IL-6, IL-8, IL-10, IL12p70; Mesoscale Discovery, Gathersburg, MD) as well as PBMC-associated KSHV viral load, platelets, hemoglobin, and albumin were evaluated in KSHV-NHL patients, and compared to patients with A) Symptomatic KSHV-MCD and no KSHV-NHL, and B) Other HIV-associated lymphomas. Comparisons used exact two-tailed Wilcoxon rank sum tests, p Results: Study cohort: 14 patients with HIV and KSHV-NHL; men (13), woman (1); median (med) age 42 yrs (26, 60); African American (4), African (2), Hispanic (4), white (4). Diagnoses included PEL (12, 5 with extracavitary manifestations) and large cell lymphoma in KSHV-MCD (2). Of the 14, 5 had pathologically confirmed KSHV-MCD, and 10 had cutaneous KS. Laboratory data: med CD4 126 cells/uL (15,1072), HIV viral load Conclusions: In the era of antiretroviral therapy, KSHV-NHL in HIV occurs at a broader range of CD4 counts than previously appreciated, often in the setting of suppressed HIV. Significant overlap exists between KSHV-associated lymphoproliferative disease and KSHV-NHL. Essentially all KSHV-NHL patients presented with inflammatory symptoms, elevated CRP, hypoalbuminemia, and cytopenias, and either had KSHV-MCD or met our working criteria for KICS. Elevated KSHV-infected PBMCs and associated KSHV modulation of host immune response leading to marked elevations in IL10 and IL6 likely contribute to symptoms and KSHV-NHL pathogenesis. KSHV-MCD and KICS Natural History studies are ongoing, and evaluation of curative-intent regimens for KSHV-NHL incorporating strategies to target these unique immunologic and virologic abnormalities is warranted. Disclosures: No relevant conflicts of interest to declare.

Abstract 2631 Background: Epstein-Barr Virus (EBV) is a risk factor for some types of Hodgkin lymphoma (HL). Host response to the virus may play a role in pathogenesis and is in part determined by genetic factors. Therefore, using only traditional unrelated controls may not be appropriate. Methods: We compared EBV DNA copy number in peripheral blood mononuclear cells (PBMCs) collected from 63 young adult HL patients in remission and their unaffected co-twins, and 43 spouse controls. Viral load was measured with TaqMan quantitative PCR assays, tested in triplicate and reported as viral copies per million cells. Samples positive in all 3 replicates were classified as positive, and those positive in 1–2 samples were classified as qualitative positive (QP). EBV copy number was log-transformed for some analyses. A binomial proportion test was used to compare EBV copy number in cases and their unaffected co-twins (n=50 like-sex pairs). Conditional (case vs. unaffected co-twin) and unconditional logistic regression (case and unaffected co-twin vs. spouse controls) were used to estimate the effect of EBV copy number on risk as a categorical (positive, QP, or negative) or continuous variable (SAS 9.1). Unconditional logistic regression analyses were adjusted for sex and age. Results: 26% of case, 18% of unaffected co-twins and 9% of spouse controls had positive EBV copy number measurements. Like-sex young adult HL cases had higher EBV copy numbers compared to their unaffected co-twins (P = 0.04). When EBV copy number was considered as a categorical variable, both cases and monozygotic unaffected co-twins were more likely to be positive compared to controls (cases v. controls: Odds Ratio [OR] = 3.2, 95% confidence intervals [CI] = 0.9, 11.1; unaffected co-twins v. controls: OR= 3.6, 95% CI= 0.9, 14.6). However, there was little difference between dizygotic unaffected co-twins and controls (OR= 0.7, 95% CI = 0.5, 5.5). Conclusions: That young adult HL cases had higher EBV DNA copies than their unaffected co-twins implies that an acquired pre-disease factor or disease affect altered the host response to EBV. However, that both cases and unaffected co-twins had higher EBV DNA copies compared to controls suggests that there is also a heritable component to the host response to EBV. HL cases > co-twins (acquired or disease effect) HL cases and co-twins > controls (heritable effect) Disclosures: No relevant conflicts of interest to declare.

Abstract 1573 Background: Multicentric Castleman disease (MCD) is a polyclonal B-cell lymphoproliferative disorder characterized by flares of severe inflammatory symptoms, including fever and cachexia, with cytopenias and biochemical abnormalities. In the idiopathic form, its pathogenesis is linked to overproduction of interleukin (IL)-6. A distinct form of MCD is caused by Kaposi sarcoma-associated herpesvirus (KSHV, also called human herpesvirus [HHV]-8). KSHV encodes a viral homolog of IL-6, vIL-6, and this has been hypothesized to be central to KSHV-MCD pathogenesis. However, viral and human cytokines have not been examined together in KSHV-MCD, and their contribution to disease activity and symptoms is not known. Methods: Patients with pathologically-proven KSHV-MCD were enrolled on a prospective natural history study incorporating pilot evaluation of novel therapies (NCT00099073). Factors potentially important in pathogenesis were assayed at flare and complete clinical and laboratory remission: KSHV viral load (VL) in peripheral blood mononuclear cells, vIL-6, IL-1β, IL-5, human IL-6 (hIL-6), IL-8, IL-10, IL-12p70, IFN-γ and TNF-α. Paired analyses were performed for each patient comparing initial flare and remission. Associations between cytokines and individual disease manifestations were explored across all flares. Results: 21 patients had at least one flare for analysis (19 [90%] male; med age 44 [range 29–52]). 34 flares were observed (range 1–3 per patient) and followed to remission (20 patients) or death (1). All were HIV infected: CD4 med 252cells/μL (range 24–1319), HIV VL med Factors elevated during initial flares were: KSHV VL (med 14,700/106 PBMCs [range 0–3,913,000] P Conclusions: This prospective analysis shows for the first time that vIL-6 and hIL-6 can independently or together lead to flares of KSHV-MCD. It shows novel associations of KSHV-MCD flares with IL-1β and TNF-α elevation and IL-5 depression while confirming associations with KSHV VL and IL-10. It further suggests that vIL-6 and hIL-6 may jointly contribute to the severity of some symptoms during flares. As vIL-6 can signal independently of the ligand-binding IL-6 receptor-a, these findings have implications for the development of novel KSHV-MCD therapies targeting IL-6 and its downstream signaling. Disclosures: No relevant conflicts of interest to declare.

Human herpes virus 8, also termed KSHV, is an oncogenic gamma herpesvirus associated with Kaposi’s sarcoma (KS), primary effusion lymphoma and MCD. KSHV-associated MCD (KSHV-MCD), a rare B-cell lymphoproliferative disease almost universally found in association with HIV infection, is characterized by recurrent flares of a systemic syndrome of fatigue, fevers, cytopenias, elevated serum C-reactive protein (CRP), and lytic KSHV replication. Prognosis is poor (median survival 14 months). KSHV open reading frames (ORF) 21 and 36 respectively have the ability to phosphorylate zidovudine and ganciclovir, leading to cell death (Cancer Res2007;67(14):7003–10), and this may be exploitable for oncolytic virotherapy. Ten patients (pts) with symptomatic and biopsy confirmed KSHV-associated MCD have been treated with high dose oral zidovudine (HDAZT) 600mg every 6 hours and valganciclovir (VGCV) 900mg every 12 hours. Treatment length of first cycle is dependent on response, and ranges from 7–21 days. Subsequent cycle length is 21 days with 7 treatment days. Pt characteristics: median age 40 (range 33–56); ECOG PS 2 (1–3); median enrollment CD4 count 189 (range 19–1319 UL); median HIV viral load