Ibrutinib in mantle cell lymphoma: a real-world retrospective multi-center analysis of 77 patients treated in the Czech Republic.

Ibrutinib revolutionized therapy for relapsed/refractory (R/R) mantle cell lymphoma (MCL). Real-world data on the outcome of unselected patients are still limited. We analyzed 77 R/R MCL patients receiving ibrutinib with at least one prior systemic anti-lymphoma therapy. After a median follow-up of 14.0 months, 56 patients relapsed/progressed, and 45 died. The overall response rate was 66%, with 31% of complete metabolic remissions on PET/CT. The median progression-free and overall survival (OS) rates were 10.3 and 23.1 months, respectively. The median OS from ibrutinib failure was 3.7 months. High proliferation rate by Ki67 (≥ 30%) and two or more previous therapy lines both negatively correlated with outcome (HR = 2.2, p = 0.04, and HR = 2.06, p = 0.08, respectively). Female gender borderline correlated with better outcome (HR = 0.53, p = 0.08). In multivariate analysis, Ki67 and response to ibrutinib both correlated with OS (p < 0.05). Importantly, ibrutinib appeared to better control nodal and extranodal lymphoma than bone marrow (BM) involvement. From 20 patients with detectable BM infiltration (before ibrutinib initiation) achieving complete (n = 13) or partial (n = 7) metabolic remission, none achieved remission in BM. We confirmed good efficacy of ibrutinib in unselected heavily pre-treated MCL patients. Our findings support the use of a combination of ibrutinib and rituximab in patients with BM involvement.

Complex genetic and histopathological study of 15 patient-derived xenografts of aggressive lymphomas.

Non-Hodgkin lymphomas (NHL) represent the most common hematologic malignancies. Patient-derived xenografts (PDXs) are used for various aspects of translational research including preclinical in vivo validation of experimental treatment approaches. While it was repeatedly demonstrated that PDXs keep majority of somatic mutations with the primary lymphoma samples, from which they were derived, the composition of PDX tumor microenvironment (TME) has not been extensively studied. We carried out a comparative genetic and histopathological study of 15 PDX models derived from patients with various types of NHL including diffuse large B-cell lymphoma (DLBCL; n = 7), Burkitt lymphoma (BL; n = 1), mantle cell lymphoma (MCL; n = 2), and peripheral T-cell lymphomas (PTCL; n = 5). Whole exome sequencing (WES) of the PDXs and primary lymphoma cells was implemented in 13 out of 15 cases with available DNA samples. Standard immunohistochemistry (IHC) was used to analyze the composition of PDX TME. WES data confirmed that PDXs maintained the genetic heterogeneity with the original primary lymphoma cells. In contrast, IHC analysis revealed the following recurrently observed alterations in the composition of PDX tumors: more blastoid lymphoma cell morphology, increased proliferation rate, lack of non-malignant cellular components including T cells and (human or murine) macrophages, and significantly lower intratumoral microvessel density and microvessel area composed of murine vessels. In addition, PDX tumors derived from T-NHL displayed additional differences compared to the primary lymphoma samples including markedly lower desmoplasia (i.e., the extent of both reticular and collagen fibrosis), loss of expression of cytotoxic granules (i.e., perforin, TIA, granzyme B), or loss of expression of T-cell specific antigens (i.e., CD3, CD4, CD8). Our data suggest that despite keeping the same genetic profiles, PDX models of aggressive NHL do not recapitulate the microenvironmental heterogeneity of the original lymphomas. These findings have implications on the relevance of PDX models in the context of preclinical research.

Experimental model of primary intraocular lymphoma based on BALB/CaNn strain and A20 cells is optimal for investigational research.

AIM: The purpose of this project was to compare the characteristics of two experimental murine models of primary intraocular lymphoma (PIOL) and determine which experimental model is most suitable for further investigational research to elucidate the pathophysiology of PIOL and to find new therapeutical strategies. METHODS: In both experimental models PIOL was induced in immunocompetent mice with intravitreal injection of syngeneic B-cell lymphoma cell lines. Murine strain C3H/HeN and cell line 38C13 were used in the first model and BALB/CaNn mice and cell line A20 in the second model. During the experiments, thorough clinical evaluation (using photo documentation, ultrasonography, and MRI) and histological evaluation were performed. RESULTS: In both models, the percentage of PIOL development was high, reaching nearly 80%. Disease progression was faster in C3H/HeN with exophthalmos occurring on average on day 10. Vitreous involvement was a predominant sign in the clinical presentation of this group. In BALB/CaNn mice exophthalmos occurred on average on day 22. The predominant clinical sign in the BALB/CaNn group was tumorous infiltration of the retina, optic disc, and tumorous retinal detachment. CONCLUSION: Slower progression of the disease in BALB/CaNn mice, greater possibility to examine the retina due to mild vitreous involvement, and later occurrence of exophthalmos makes this strain more suitable for further investigational research.

Cyclin dependent kinase 4/6 inhibitor palbociclib synergizes with BCL2 inhibitor venetoclax in experimental models of mantle cell lymphoma without RB1 deletion.

BACKGROUND: Mantle cell lymphoma (MCL) is a chronically relapsing malignancy with deregulated cell cycle progression. We analyzed efficacy, mode of action, and predictive markers of susceptibility to palbociclib, an approved CDK 4/6 inhibitor, and its combination with venetoclax, a BCL2 inhibitor. METHODS: A panel of nine MCL cell lines were used for in vitro experiments. Four patient derived xenografts (PDX) obtained from patients with chemotherapy and ibrutinib-refractory MCL were used for in vivo proof-of-concept studies. Changes of the mitochondrial membrane potential, energy-metabolic pathways, AKT activity, and pro-apoptotic priming of MCL cells were evaluated by JC-1 staining, Seahorse XF analyser, genetically encoded fluorescent AKT reporter, and BH3 profiling, respectively. MCL clones with gene knockout or transgenic (over)expression of CDKN2A, MYC, CDK4, and RB1 were used to estimate impact of these aberrations on sensitivity to palbociclib, and venetoclax. RESULTS: Co-targeting MCL cells with palbociclib and venetoclax induced cytotoxic synergy in vitro and in vivo. Molecular mechanisms responsible for the observed synthetic lethality comprised palbociclib-mediated downregulation of anti-apoptotic MCL1, increased levels of proapoptotic BIM bound on both BCL2, and BCL-XL and increased pro-apoptotic priming of MCL cells mediated by BCL2-independent mechanisms, predominantly palbociclib-triggered metabolic and mitochondrial stress. Loss of RB1 resulted in palbociclib resistance, while deletion of CDKN2A or overexpression of CDK4, and MYC genes did not change sensitivity to palbociclib. CONCLUSIONS: Our data strongly support investigation of the chemotherapy-free palbociclib and venetoclax combination as an innovative treatment strategy for post-ibrutinib MCL patients without RB1 deletion.

Concurrent TP53 and CDKN2A Gene Aberrations in Newly Diagnosed Mantle Cell Lymphoma Correlate with Chemoresistance and Call for Innovative Upfront Therapy.

Mantle cell lymphoma (MCL) is a subtype of B-cell lymphoma with a large number of recurrent cytogenetic/molecular aberrations. Approximately 5-10% of patients do not respond to frontline immunochemotherapy. Despite many useful prognostic indexes, a reliable marker of chemoresistance is not available. We evaluated the prognostic impact of seven recurrent gene aberrations including tumor suppressor protein P53 (TP53) and cyclin dependent kinase inhibitor 2A (CDKN2A) in the cohort of 126 newly diagnosed consecutive MCL patients with bone marrow involvement ≥5% using fluorescent in-situ hybridization (FISH) and next-generation sequencing (NGS). In contrast to TP53, no pathologic mutations of CDKN2A were detected by NGS. CDKN2A deletions were found exclusively in the context of other gene aberrations suggesting it represents a later event (after translocation t(11;14) and aberrations of TP53, or ataxia telangiectasia mutated (ATM)). Concurrent deletion of CDKN2A and aberration of TP53 (deletion and/or mutation) represented the most significant predictor of short EFS (median 3 months) and OS (median 10 months). Concurrent aberration of TP53 and CDKN2A is a new, simple, and relevant index of chemoresistance in MCL. Patients with concurrent aberration of TP53 and CDKN2A should be offered innovative anti-lymphoma therapy and upfront consolidation with allogeneic stem cell transplantation.