PurposeThe proteasome inhibitor bortezomib may enhance activity of chemoimmunotherapy in lymphoma. We evaluated dose-escalated bortezomib plus standard cyclophosphamide, doxorubicin, vincristine, and prednisone (CHOP) plus rituximab (R) in patients with diffuse large B-cell lymphoma (DLBCL) and mantle cell lymphoma (MCL).Patients and MethodsSeventy-six subjects with untreated DLBCL (n = 40) and MCL (n = 36) received standard CHOP every 21 days (CHOP-21) with R plus bortezomib at 0.7 mg/m2(n = 4), 1.0 mg/m2(n = 9), or 1.3 mg/m2(n = 63) on days 1 and 4 for six cycles.ResultsMedian age was 63 years (range, 20 to 87), and International Prognostic Index (IPI) scores were generally unfavorable (39% with IPI of 2, and 49% with IPI of 3 to 5), as were Mantle Cell Lymphoma International Prognostic Index scores in patients with MCL (28% intermediate risk and 39% high risk). Toxicity was manageable, including neuropathy in 49 subjects (8% grade 2 and 4% grade 3) and grade 3/4 anemia (13%), neutropenia (41%), and thrombocytopenia (25%). For DLBCL, the evaluable overall response rate (ORR) was 100% with 86% complete response (CR)/CR unconfirmed (CRu; n = 35). Intent-to-treat (ITT, n = 40) ORR was 88% with 75% CR/CRu, 2-year progression-free survival (PFS) of 64% (95% CI, 47% to 77%) and 2-year overall survival (OS) of 70% (95% CI, 53% to 82%). For MCL, the evaluable ORR was 91% with 72% CR/CRu (n = 32). The ITT (n = 36) ORR was 81% with 64% CR/CRu, 2-year PFS 44% (95% CI, 27% to 60%) and 2-year OS 86% (95% CI, 70% to 94%). IPI and MIPI correlated with survival in DLBCL and MCL, respectively. Unlike in DLBCL treated with R-CHOP alone, nongerminal center B cell (non-GCB) and GCB subtypes had similar outcomes.ConclusionBortezomib with R-CHOP-21 can be safely administered and may enhance outcomes, particularly in non-GCB DLBCL, justifying randomized studies.

Apoptosis is a natural process during animal development for the programmed removal of superfluous cells. During apoptosis general protein synthesis is reduced, but the synthesis of cell death proteins is enhanced. Selective translation has been attributed to modification of the protein synthesis machinery to disrupt cap-dependent mRNA translation and induce a cap-independent mechanism. We have previously shown that disruption of the balance between cap-dependent and cap-independent C. elegans eIF4G isoforms (IFG-1 p170 and p130) by RNA interference promotes apoptosis in developing oocytes. Germ cell apoptosis was accompanied by the appearance of the Apaf-1 homolog, CED-4. Here we show that IFG-1 p170 is a native substrate of the worm executioner caspase, CED-3, just as mammalian eIF4GI is cleaved by caspase-3. Loss of Bcl-2 function (ced-9ts) in worms induced p170 cleavage in vivo, coincident with extensive germ cell apoptosis. Truncation of IFG-1 occurred at a single site that separates the cap-binding and ribosome-associated domains. Site-directed mutagenesis indicated that CED-3 processes IFG-1 at a non-canonical motif, TTTD456. Coincidentally, the recognition site was located 65 amino acids downstream of the newly mapped IFG-1 p130 start site suggesting that both forms support cap-independent initiation. Genetic evidence confirmed that apoptosis induced by loss of ifg-1 p170 mRNA was caspase (ced-3) and apoptosome (ced-4/Apaf-1) dependent. These findings support a new paradigm in which modal changes in protein synthesis act as a physiological signal to initiate cell death, rather than occur merely as downstream consequences of the apoptotic event.

Regulated mRNA translation is vital for germ cells to produce new proteins in the spatial and temporal patterns that drive gamete development. Translational control involves the de-repression of stored mRNAs and their recruitment by eukaryotic initiation factors (eIFs) to ribosomes. C. elegans expresses five eIF4Es (IFE-1-IFE-5); several have been shown to selectively recruit unique pools of mRNA. Individual IFE knockouts yield unique phenotypes due to inefficient translation of certain mRNAs. Here, we identified mRNAs preferentially translated through the germline-specific eIF4E isoform IFE-1. Differential polysome microarray analysis identified 77 mRNAs recruited by IFE-1. Among the IFE-1-dependent mRNAs are several required for late germ cell differentiation and maturation. Polysome association of gld-1, vab-1, vpr-1, rab-7 and rnp-3 mRNAs relies on IFE-1. Live animal imaging showed IFE-1-dependent selectivity in spatial and temporal translation of germline mRNAs. Altered MAPK activation in oocytes suggests dual roles for IFE-1, both promoting and suppressing oocyte maturation at different stages. This single eIF4E isoform exerts positive, selective translational control during germ cell differentiation.

Chimeric antigen receptor T cells targeting CD30 (CD30.CAR-T) have shown high response rates, including some durable remissions, in patients with relapsed/refractory (r/r) classical Hodgkin lymphoma (HL) (Ramos et al., JCO 2020). However, some patients are non-responders or eventually relapse after therapy. Because CD30 expression is retained in relapsing tumors, recurrence may be due to the insufficient persistence of CAR-Ts within the highly immunosuppressive tumor microenvironment of HL. We therefore reasoned that with enhanced trafficking to the tumor site, CD30.CAR-Ts would have increased opportunities to eliminate tumors before inhibitory mechanisms become predominant. This is especially relevant for HL, where Hodgkin Reed Sternberg (HRS) cells produce CCL17 (thymus and activation-regulated chemokine) to create a physical and inhibitory barrier to cytotoxic T cells. We have previously shown that CD30.CAR-Ts co-expressing the cognate receptor for CCL17, CCR4 (CCR4.CD30.CAR-Ts), have improved tumor homing and anti-lymphoma activity compared with CD30.CAR-Ts that do not express CCR4 (Di Stasi et al., Blood 2009). CCR4.CD30.CAR-Ts should also be more effective in CD30+ cutaneous T-cell lymphomas (CTCL) due to enhanced trafficking to the skin. We present the preliminary results of a clinical trial assessing the safety (primary objective) of this novel strategy and its efficacy compared to CD30.CAR-Ts lacking CCR4 in patients with r/r HL and CD30+ CTCL (NCT03602157). CCR4.CD30.CAR-Ts are infused in patients in a dose escalation fashion (DL1=2x10 7 CAR-Ts/m 2, DL3=5x10 7 CAR-Ts/m 2, DL5=1x10 8 CAR-Ts/m 2,). To provide definitive conclusions on the role of CAR-T tumor homing, after completion of each dose level, patients receive the dose of CCR4.CD30.CAR-Ts established to be safe in the prior DL, combined with a fixed dose of CD30.CAR-Ts (1x10 8 CAR-Ts/m 2) (DL2, DL4, DL6). All patients receive lymphodepletion with 3 days of bendamustine 70 mg/m 2 and fludarabine 30 mg/m 2. Key inclusion criteria are age ≥ 18 years and r/r HL or CTCL having failed ≥2 prior therapies. At the time of data cut off (8/1/2021), 6 patients were treated on DL1, 3 patients on DL2, and 3 patients on DL3. The median age is 43.5 (range 27-75) with a median of 5.5 prior lines of therapy (range 2-10). Ten patients had HL and 2 patients had CTCL. All patients had received prior brentuximab vedotin. Eleven patients received prior checkpoint inhibitors, 9 had prior autologous stem cell transplant, and 5 had prior allogeneic stem cell transplant. The treatment was well tolerated with no dose limiting toxicities observed. Two patients had grade 2 cytokine release syndrome (CRS) which resolved with tocilizumab, and 1 had self-limiting grade 1 CRS. None of the treated patients developed immune effector cell-associated neurotoxicity syndrome. All of the 8 patients with HL who have had disease assessment have responded with 6 complete responses (CR) (75%) and 2 partial responses (PR). Five patients are in remission to date, with one patient still in CR at 2.5 years post treatment. Two patients with HL have responses pending at time of data cut off. Among the 2 patients with CTCL, 1 patient had stable disease and went on to receive alternative therapy and 1 patient had progressive disease. At a median follow up of 12.7 months, the median progression free survival (PFS) for all 10 evaluable patients is 5.2 months and the median PFS for HL patients has not been reached. The median overall survival for all patients has not been reached. Plasma CCL17, a biomarker of disease response for HL, was reduced by 83±15% by week 2 post infusion in patients treated with CCR4.CD30.CAR-Ts as compared to 52±38% in patients on our previous trial that had received CD30.CAR-Ts lacking CCR4 (p=0.02). In a HL patient on DL1 biopsied 3 weeks post infusion, we found markedly enriched CAR-T signals at the tumor site (14.4 x10 5 copies/ug of DNA) as compared to the signals found at the same time point in the peripheral blood (4.3 x10 5 copies/ug of DNA). Our data confirm the safety of CCR4.CD30.CAR-Ts as well as their promising efficacy in patients with r/r HL. Interestingly, responses are already seen at the lowest dose level, suggesting that early tumor homing driven by CCR4 may allow more fitted cells to better exploit their antitumor potential. Our data serve as a proof of concept for future modifications of CAR-T cells to improve their localization to disease sites. Figure 1 Figure 1. Disclosures Grover: Kite: Other: Advisory Board; Tessa: Consultancy; ADC: Other: Advisory Board; Novartis: Consultancy; Genentech: Research Funding. Morrison: Vesselon: Consultancy. Dittus: BeiGene: Other: Advisory Board; Seattle Genetics: Research Funding; AstraZeneca: Research Funding; Genentech: Research Funding. Dotti: Tessa: Patents & Royalties: Approach for CD30.CAR-T Cells for Hodgkin Lymphoma. Serody: Tessa: Patents & Royalties: Approach for CD30.CAR-T Cells for Hodgkin Lymphoma. Savoldo: Tessa: Patents & Royalties: Approach for CD30.CAR-T Cells for Hodgkin Lymphoma.

During apoptosis, activated caspases cleave the translation initiation factor eIF4G. This cleavage disrupts cap-dependent mRNA translation initiation within the cell. However, a specific subset of mRNAs can still be recruited for protein synthesis in a cap-independent manner by the residual initiation machinery. Many of these mRNAs, including cell death related mRNAs, contain internal ribosome entry sites (IRESes) that promote their enhanced translation during apoptosis. Still other mRNAs have little dependence on the cap recognition mechanism. The expression of the encoded proteins, both anti- and pro-apoptotic, allows for an initial period of attempted cell survival, then commitment to cell death when damage is extensive. In this study we address the translational regulation of the stress and apoptosis-related mRNAs in C. elegans: BiP (hsp-3) (hsp-4), Hif-1 (hif-1), p53 (cep-1), Bcl-2 (ced-9) and Apaf-1 (ced-4). Altered translational efficiency of these messages was observed upon depletion of cap-dependent translation and induction of apoptosis within the C. elegans gonad. Our findings suggest a physiological link between the cap-independent mechanism and the enhanced translation of hsp-3 and ced-9. This increase in the efficiency of translation may be integral to the stress response during the induction of physiological apoptosis.

Our group has recently demonstrated that chimeric antigen receptor T-cell therapy targeting the CD30 antigen (CD30.CAR-T) is highly effective in patients with relapsed and refractory (r/r) classical Hodgkin lymphoma (cHL). Despite high rates of clinical response, relapses and progression were observed in a subset of patients. The objective of this study was to characterize clinical and correlative factors associated with progression-free survival (PFS) after CD30.CAR-T cell therapy. We evaluated correlatives in 27 patients with r/r cHL treated with lymphodepletion and CD30.CAR-T cells. With a median follow-up of 9.5 months, 17 patients (63%) progressed, with a median PFS of 352 days (95% confidence interval: 116-not reached), and 2 patients died (7%) with a median overall survival of not reached. High metabolic tumor volume (MTV, >60 mL) immediately before lymphodepletion and CD30.CAR-T cell infusion was associated with inferior PFS (log rank, P = .02), which persisted after adjusting for lymphodepletion and CAR-T dose (log rank, P = .01 and P = .006, respectively). In contrast, receiving bridging therapy, response to bridging therapy, CD30.CAR-T expansion/persistence, and percentage of CD3+PD-1+ lymphocytes over the first 6 weeks of therapy were not associated with differences in PFS. In summary, this study reports an association between high baseline MTV immediately before lymphodepletion and CD30.CAR-T cell infusion and worse PFS in patients with r/r cHL. This trial was registered at www.clinicaltrials.gov as #NCT02690545.

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Background: The safety and efficacy of treating pts with relapsed/refractory (r/r) CD30+ lymphomas with chimeric receptor modified T cells targeting the CD30 molecule (CD30.CAR-T) has been recently demonstrated (Grover et al., ASH 2018). Expression of PD-1 on CD30.CAR-T cells has been observed post-infusion at the time of disease recurrence. To determine the safety and efficacy of anti-PD-1 mAb therapy (CPI) post CD30.CAR-T cell therapy, we present a retrospective cohort review of pts with r/r Hodgkin lymphoma (HL) who subsequently received CPI after CD30.CAR-T progression. Methods: We followed and analyzed the clinical outcome of 5 pts with r/r HL who progressed after CD30.CAR-T cell therapy at the University of North Carolina at Chapel Hill and subsequently received CPI to treat relapsed disease. Clinical responses in follow up were determined per local radiologist assessment. Results: Prior to CAR-T cell therapy, the average age of the pts in this cohort was 36 years old (range, 26-54). Four of the five had been treated with CPI prior to CD30.CAR-T therapy, with a median duration of response of 224 days (range, 81-280) and best objective response being a partial response (PR) for all 4 pts. The median time from CPI to CD30.CAR-T therapy was 483 days (range, 25-1174 days). A total of 5 pts have been treated with CPI following clinical progression after CD30.CAR-T. All pts were heavily pre-treated with a median of 8 therapies prior to CD30.CAR-T. One pt received a second infusion of CD30.CAR-T after having stable disease (SD) 6 weeks following the first infusion. Following CD30.CAR-T infusion, 1 pt initially had a complete response (CR), 1 pt had a PR, 2 pts had stable disease (SD), and 1 pt had progressive disease (PD). All 5 pts subsequently developed PD. The median time from CD30.CAR-T to CPI was 132 days (range, 67-393 days), (Table 1). Of the 5 pts evaluable for response, 4 pts achieved a CR and 1 pt achieved a PR (Figure 1). At the time of data cutoff, the median duration of response was 212 days (range 139-836) with ongoing response in 3 pts. Two pts developed recurrent disease following CPI and were again treated with CD30.CAR-T cell therapy, achieving a 2nd CR in both cases. CPI use was associated with the expected toxicities. One pt developed neuropathy, leading to therapy discontinuation after 8 months of therapy. Another pt developed grade 3 colitis, resulting in discontinuation of therapy after 25 months of therapy. A third pt developed pneumonitis (prior mediastinal radiation), with discontinuation of therapy after 8 months of therapy. All pts were still alive at last follow up with a median overall survival of 2.44 years (range, 0.93-2.76) from initial CD30.CAR-T cell therapy. Conclusion: To our knowledge, this is first report of pts with r/r HL who received CPI after CD30.CAR-T cell therapy. Though limited by size, this cohort demonstrates clinical efficacy of CPI after CD30.CAR-T even in those pts who had progressed after previous treatment with CPI. All pts achieved an objective response with CPI administered for relapsed disease after CD30.CAR T cell therapy. Four of five of these patients achieved a CR and 3 patients achieved an improved response when compared to CPI prior to CD30.CAR-T. Combination of CPI and CD30.CAR-T may be an attractive study design in the future for r/r HL. Disclosures Grover: Seattle Genetics: Consultancy. Park:BMS: Consultancy, Research Funding; Rafael Pharma: Membership on an entity's Board of Directors or advisory committees; G1 Therapeutics: Consultancy; Teva: Consultancy, Research Funding; Gilead: Speakers Bureau; Seattle Genetics: Research Funding, Speakers Bureau. Savoldo:Baylor College of Medicine: Patents & Royalties: CAR.CD30 patent; Bluebirdbio: Other: research agreement; Cell Medica: Other: Research Agreement; Bellicum: Other: Research Agreement. Serody:Merck: Research Funding; GlaxoSmithKline: Research Funding.