ALBERT

Description: Introduction: Despite significant progress in the treatment of multiple myeloma (MM), the disease remains incurable and typically follows a pattern of multiple responses and relapses. In the relapsed/refractory MM setting (RRMM), outcomes for patients may be particularly discouraging. New treatments that are safe and effective are therefore of urgent need for this population. Since its FDA approval in 1998 for the treatment of rheumatoid arthritis, leflunomide has been used in over 300,000 patients worldwide. Leflunomide is hepatically cleared and has a favorable toxicity profile even when given over long periods. Its primary mechanism of action is inhibition of pyrimidine synthesis by targeting dihydroorotate dehydrogenase, thus achieving anti-proliferative effects on B and T lymphocytes. The anti-neoplastic potential of this agent has been studied in a number of pre-clinical tumor models. Leflunomide's immunoregulatory action may be related to functional inhibition of CD4+ effector T cells, including Th17 cells as well as dysregulation of T regulatory cells (Tregs). We found that leflunomide-treated C57BL/KaLwRij mice engrafted with 5TGM1 cells had more robust expansion of CD8+ cytotoxic T cells and subsequent decrease of CD4+ Tregs compared to untreated mice. We have also noted that leflunomide impairs growth of MM cells at least partly through inhibition of PIM kinases and c-Myc signaling. We present here final results from a phase 1 study of leflunomide in patients with RRMM. Methods: This single center, single agent, phase 1 dose-escalation trial was designed to determine the maximum tolerated dose of leflunomide in patients with RRMM. The trial implemented a modified rolling six phase 1 dose escalation design. The primary objectives were as follows: 1) to determine the maximum tolerated dose and recommended phase 2 dose of leflunomide; 2) to assess the safety and tolerably of leflunomide at each dose level by evaluation of toxicities. Leflunomide was administered at a loading dose of 100 mg daily for the first three days, then daily in 28-day cycles. The starting dose of daily leflunomide was 20 mg daily, with dose escalation in increments of 20 mg/day, up to 60 mg/day. Dose de-escalation in decrements of 10mg/day was planned. Results: A total of 12 patients have been enrolled starting in December 2015 and treated. The median age is 68 (range 48 - 85), and the median number of prior therapies is 5 (range: 3 - 14). Nine patients had prior autologous stem cell transplant. Double refractory (lenalidomide/bortezomib) disease was noted in 9 patients. High-risk cytogenetics were observed in 5 patients including 2 patients with del17p. All 12 subjects were evaluable for toxicity. One subject was not evaluable for response because of non-compliance. Of the eleven patients evaluable for response, the median number of cycles was 3 (range 1- 15). The median follow up was 177 days (range: 42 - 602). Three patients were treated on DL 1 (20 mg) and three on DL3 (40 mg) without incidence of DLT. At DL5 (60 mg), one patient had a DLT with grade III elevation of alanine aminotransferase; an additional three patients were enrolled at this dose level without further DLTs. One out of 12 subjects remains on treatment, 8 patients were removed from study due to disease progression, two due to adverse events (bacteremia at 60 mg, possibly related to study drug and angioedema at 40 mg, not related to study drug) and one from noncompliance. The most common toxicities were hematologic. There were 4 patients with grade 1 or 2 neutropenia on the 20 and 40 mg dose levels and 1 patient with grade 4 lymphopenia on the 40 mg dose. Except for the DLT, all non-hematologic toxicities were ≤ grade 2. Response: Although not all patients were treated at the 60 mg dose, a clinical benefit rate of 90% has been seen, with 9/10 achieving stable disease (SD). The median duration of SD among 9 patients thus far is 56 days (range: 27-401). In the five evaluable patients with high risk cytogenetics, four of them achieved a clinical benefit. Two subjects had SD lasting nearly one year or longer. In this small cohort, no association between dose and benefit was observed. Conclusion: Leflunomide is a safe and well-tolerated oral option for patients with RRMM, with a clinical benefit from single agent dosing. On the basis of our preclinical work showing synergistic inhibition of MM using leflunomide, pomalidomide, and dexamethasone, we plan clinical testing of this drug combination. Disclosures Rosenzweig: Celgene: Speakers Bureau. Krishnan:Janssen: Consultancy, Speakers Bureau; Sutro: Speakers Bureau; Onyx: Speakers Bureau; Takeda: Speakers Bureau; Celgene: Consultancy, Equity Ownership, Speakers Bureau. Forman:Mustang Therapeutics: Other: Licensing Agreement, Patents & Royalties, Research Funding.

Description: Background: Classical Hodgkin lymphoma (HL) expresses surface CD30. Brentuximab vedotin (BV) is an antibody-drug conjugate that selectively delivers a potent cytotoxic agent, monomethyl auristatin E (MMAE), specifically to cells expressing surface CD30. Although BV elicits a high response rate (75%) in HL, patients who do not achieve complete response (CR) will eventually develop resistance to BV. We have developed 2 BV-resistant HL cell models and shown that resistance to BV is due to MDR1 upregulation rather than downregulation of surface CD30. MDR1 upregulation leads to increased drug exporter pump and decreased intracellular MMAE, the cytotoxic agent in BV. Our hypothesis is that by inhibiting MDR1, we can overcome resistance to BV and also achieve synergy with BV in HL cell lines and mouse xenograft model. Methods: HL cell lines (L428 and KMH2) were used for in vitro experiments. The selection of BV resistant cell model (L428R and KMH2) was done using pulsatile treatment with BV. Both BV resistant cell models were confirmed with MTS assay and cell growth assays. MDR1 upreguation was confirmed by qRT-PCR and western blot. Intracellular MMAE concentration was assessed by mass spectrometry. MDR1 inhibitors verapamil (VrP, 10 ug/ml) and cyclosporine (CsA, 5 uM) were chosen. NSG mice were used for mouse xenograft experiments. Results: MTS assay showed that the IC50 to BV increased 12 folds in L428R (32 ug/ml in L428, 391 ug/ml in L428R), and that the IC50 to BV has increased 17 folds in KMH2R (13 ug/ml in KMH2, 219 ug/ml in KMH2R). QRT-PCR shows that L428R and KMH2R had upregulation of MDR1 mRNA, and western blot shows L428R and KMH2R had overexpression of PgP (MDR1) as compared to L428 and KMH2. Mass spectrometry shows that the intracellular MMAE concentration was 7.6 folds lower in L428R as compared to L428 after treatment with BV. We then examined the effect of MDR1 inhibitors (VrP and CsA) on the IC50 of BV in L428R and KMH2R. The addition of VrP to L428R was able to decrease the IC50 from 344 ug/ml to 22.8 ug/ml (15 fold). The addition of VrP to KMH2R was able to decrease the IC50 from 170 ug/ml to 22 ug/ml (7.7 fold). The addition of CsA to L428R was able to decrease the IC50 from 275 ug/ml to 0.0068 ug/ml (40441 fold). The addition of CsA to KMH2 was able to decrease the IC50 from 129 ug/ml to 0.11 ug/ml (1098 fold). Also, the addition of VrP to L428R was able to increase the intracellular MMAE levels of L428R by 6.4 folds. We then examined the effect of MDR1 inhibitors plus BV in BV naïve parental cell lines. The addition of VrP to L428 was able to decrease the IC50 of BV from 69 ug/ml to 0.03 ug/ml (2300 folds). The addition of VrP to KMH2 was able to decrease the IC50 of BV from 6.3 ug/ml to 0.0013 ug/ml (4846 folds). The addition of CsA to L428 was able to decrease the IC50 of BV from 36 ug/ml to 0.0085 ug.ml (4235 folds). The addition of CsA to KMH2 was able to decrease the IC50 of BV from 7.1 ug/ml to 0.0002 ug/ml (32645 folds). Neither the VrP or CsA had individual effects on HL parental or resistant cell lines. Because CsA seems to have the strongest effect in KMH2 cell lines, we examined the effect of CsA plus BV in KMH2 mouse xenograft model. KMH2 was injected into the right flank of the mice to form subcutaneous lymphomas and mice were separated into 3 groups of 5. Group 1 is control without addition of BV or CsA. Group 2 is control plus BV at dose of 0.3 mg/kg intravenously every 4 days. Group 3 is treated with BV at doses of 0.3 mg/kg intravenously every 4 days and CsA intraperitoneally at 15 mg BID. We show that group 2 had a 37% reduction in tumor volume as compared to group 1 and that group 3 had a 70% reduction in tumor volume as compared to group 1 (Figure 1). Conclusion: MDR1 upregulation increases efflux pump activity and decreases intracellular accumulation of MMAE, and leads to resistance to BV in HL. This mechanism of resistance can be overcome with MDR1 inhibitors VrP and CsA. The addition of MDR1 inhibitors to BV is able to increase the intracellular accumulation of MMAE, and restore the sensitive to BV. We are also able to show synergery between MDR1 inhibitors and BV in the parental non-resistant HL cell lines and mouse xenograft model. This is the first study showing synergy between MDR1 inhibition and antibody drug conjugates. A clinical trial has been initiated to examine the safety/efficacy of the combination of MDR1 inhibitors plus BV in patients with relapsed/refractory HL. Figure 1. Figure 1. Disclosures Chen: Seattle Genetics: Consultancy, Honoraria, Research Funding, Speakers Bureau; Merck: Consultancy, Research Funding; Millenium: Consultancy, Research Funding, Speakers Bureau; Genentech: Consultancy, Speakers Bureau. Kwak:Antigenics: Equity Ownership; Celltrion: Consultancy; XEME BioPharma: Consultancy, Equity Ownership; Sella Life Sciences: Consultancy.