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26S proteasome non-ATPase subunits 1 (PSMD1) and 3 (PSMD3) were recently identified as prognostic biomarkers and potential therapeutic targets in chronic myeloid leukemia (CML) and multiple solid tumors. In the present study, we analyzed the expression of 19S proteasome subunits in acute myeloid leukemia (AML) patients with mutations in the FMS-like tyrosine kinase 3 (FLT3) gene and assessed their impact on overall survival (OS). High levels of PSMD3 but not PSMD1 expression correlated with a worse OS in FLT3-mutated AML. Consistent with an oncogenic role for PSMD3 in AML, shRNA-mediated PSMD3 knockdown impaired colony formation of FLT3+ AML cell lines, which correlated with increased OS in xenograft models. While PSMD3 regulated nuclear factor-kappa B (NF-κB) transcriptional activity in CML, we did not observe similar effects in FLT3+ AML cells. Rather, proteomics analyses suggested a role for PSMD3 in neutrophil degranulation and energy metabolism. Finally, we identified additional PSMD subunits that are upregulated in AML patients with mutated versus wild-type FLT3, which correlated with worse outcomes. These findings suggest that different components of the 19S regulatory complex of the 26S proteasome can have indications for OS and may serve as prognostic biomarkers in AML and other types of cancers.

Tyrosine kinase inhibitors (TKIs) targeting BCR-ABL1 have revolutionized therapy for chronic myeloid leukemia (CML), paving the way for clinical development in other diseases. Despite success, targeting leukemic stem cells and overcoming drug resistance remain challenges for curative cancer therapy. To identify drivers of kinase-independent TKI resistance in CML, we performed genome-wide expression analyses on TKI-resistant versus sensitive CML cell lines, revealing a nuclear factor-kappa B (NF-κB) expression signature. Nucleocytoplasmic fractionation and luciferase reporter assays confirmed increased NF-κB activity in the nucleus of TKI-resistant versus sensitive CML cell lines and CD34+ patient samples. Two genes that were upregulated in TKI-resistant CML cells were proteasome 26S subunit, non-ATPases 1 (PSMD1) and 3 (PSMD3), both members of the 19S regulatory complex in the 26S proteasome. PSMD1 and PSMD3 were also identified as survival-critical genes in a published small hairpin RNA library screen of TKI resistance. We observed markedly higher levels of PSMD1 and PSMD3 mRNA in CML patients who had progressed to the blast phase compared with the chronic phase of the disease. Knockdown of PSMD1 or PSMD3 protein correlated with reduced survival and increased apoptosis in CML cells, but not in normal cord blood CD34+ progenitors. Luciferase reporter assays and immunoblot analyses demonstrated that PSMD1 and PSMD3 promote NF-κB protein expression in CML, and that signal transducer and activator of transcription 3 (STAT3) further activates NF-κB in scenarios of TKI resistance. Our data identify NF-κB as a transcriptional driver in TKI resistance, and implicate PSMD1 and PSMD3 as plausible therapeutic targets worthy of future investigation in CML and possibly other malignancies.

Monocytes and macrophages are early sentinels of infection. The peritoneum contains two resident populations: large and small peritoneal macrophages (LPMs and SPMs). While LPMs self-renew, circulating monocytes enter the peritoneum and differentiate into SPMs. We lack information on the dynamics of monocyte–macrophage trafficking during abdominal sepsis, reflecting an important knowledge gap. In this study, we characterize the presence of LPMs, SPMs, and monocytes in the peritoneum of mice following cecal ligation and puncture (CLP)–induced sepsis and sham surgery. LPMs rapidly disappeared from the peritoneum and were scarce at 18–66 h after CLP or sham surgery. By 14 d, LPMs returned for sham mice, but they remained scarce in CLP mice. Depletion of LPMs from the peritoneum of CD11b-DTR mice greatly increased animal mortality. These data imply that LPMs are critical for sepsis survival. Monocytes rapidly infiltrated the peritoneum and were abundant at 18–66 h after CLP or sham surgery. Surprisingly, SPMs only increased at 14 d post-CLP. Therefore, monocytes may defend hosts from acute sepsis mortality without generating SPMs. More monocytes were present in mice predicted to survive sepsis versus mice predicted to die. However, altering monocyte numbers via CCR2 deficiency or adoptive transfer did not significantly affect animal survival. We reasoned that animals destined to survive sepsis may exhibit a different monocyte phenotype, rather than merely enhanced numbers. Indeed, mice predicted to survive possessed more CD31+, CXCR4hi transitional premonocytes in their abdomen. Inhibition of CXCL12–CXCR4 signaling via AMD3100 exacerbated sepsis. These data imply that recruitment of transitional premonocytes to the abdomen promotes sepsis survival.

Despite significant advances in the treatment of myeloid malignancies, many patients become resistant to therapy and ultimately succumb to their disease. Accumulating evidence over the past several years has suggested that the inadequacy of many leukaemia therapies results from their failure to target the leukaemic stem cell (LSC). For this reason, the LSC population currently represents the most critical target in the treatment of myeloid malignancies. However, while LSCs are ideal targets in the treatment of these diseases, they are also the most difficult population to target. This is due to both their heterogeneity within the LSC population, and also their phenotypic similarities with normal haematopoietic stem cells. This review will highlight the current landscape surrounding LSC biology in myeloid malignancies, with a focus on altered energy metabolism, and how that knowledge is being translated into clinical advances for the treatment of chronic and acute myeloid leukaemia and myelodysplastic syndromes.

BACKGROUND The Texas/Chihuahua (US/Mexico) border is a medically underserved region with many reported barriers for health care access. Although Hispanic ethnicity is associated with health disparities for many different diseases, the population-based estimates of incidence and survival for patients with blood cancer along the border are unknown. The authors hypothesized that Hispanic ethnicity and border proximity is associated with poor blood cancer outcomes. METHODS Data from the Texas Cancer Registry (1995-2016) were used to investigate the primary exposures of patient ethnicity (Hispanic vs non-Hispanic) and geographic location (border vs non-border). Other confounders and covariates included sex, age, year of diagnosis, rurality, insurance status, poverty indicators, and comorbidities. The Mantel-Haenszel method and Cox regression analyses were used to determine adjusted effects of ethnicity and border proximity on the relative risk (RR) and survival of patients with different blood cancer types. RESULTS Hispanic patients were diagnosed at a younger age than non-Hispanic patients and presented with increased comorbidities. Whereas non-Hispanics had a higher incidence of developing blood cancer compared with Hispanics overall, Hispanics demonstrated a higher incidence of acute lymphoblastic leukemia (RR, 1.92; 95% CI, 1.79-2.08; P < .001) with worse outcomes. Hispanics from the Texas/Chihuahua border demonstrated a higher incidence of chronic myeloid leukemia (RR, 1.28; 95% CI, 1.07-1.51; P = .02) and acute myeloid leukemia (RR, 1.17; 95% CI, 1.04-1.33; P = .0009) compared with Hispanics living elsewhere in Texas. CONCLUSIONS Hispanic ethnicity and border proximity were associated with a poor presentation and an adverse prognosis despite the younger age of diagnosis. Future studies should explore differences in disease biology and treatment strategies that could drive these regional disparities.

Ever since the ubiquitin proteasome system was characterized, efforts have been made to manipulate its function to abrogate the progression of cancer. As a result, the anticancer drugs bortezomib, carfilzomib, and ixazomib targeting the 26S proteasome were developed to treat multiple myeloma, mantle cell lymphoma, diffuse large B-cell lymphoma, colorectal cancer, and thyroid carcinoma. Despite the success demonstrated by these treatments, adverse side effects and drug resistance are prominent, raising the need for alternative therapeutic options. The 26S proteasome is composed of two 19S regulatory subunits and a 20S catalytic core, leaving ample opportunity for new targets. We recently demonstrated that knockdown of the 19S regulatory subunits, proteasome 26S subunit, non-ATPases 1 (PSMD1) and 3 (PSMD3), resulted in increased apoptosis of tyrosine kinase inhibitor (TKI)-resistant chronic myeloid leukemia (CML) cells, but had no effect on normal cord blood controls, suggesting they may be good targets for therapy. Therefore, we hypothesized that PSMD1 and PSMD3 are potential targets for anticancer therapeutics and that their relevance stretches beyond CML to other types of cancers. In the present study, we analyzed PSMD1 and PSMD3 expression in other types of cancers versus normal controls using data from The Cancer Genome Atlas (TCGA) and the Clinical Proteomic Tumor Analysis Consortium (CPTAC), comparing expression with overall survival (OS). Data were analyzed using UALCAN (http://ualcan.path.uab.edu) and GEPIA2 (http://gepia2.cancer-pku.cn/). At the mRNA level, PSMD1 was found to be overexpressed in 14/24 (58%) TCGA cancers compared with normal controls. This was confirmed at the protein level for breast (p=4.2e-4), colon (p=7.1e-20), clear cell renal carcinoma (p=1.8e-4), and endometrial cancer (p=2.0e-22). PSMD3 was found upregulated at the mRNA level in 18/24 (75%) TCGA cancers, which was confirmed at the protein level in breast (p=8.3e-4), colon (p=1.0e-27), ovarian (p=4.2e-6), clear cell renal carcinoma (p=4.0e-19), and endometrial cancer (p=1.5e-19). Upon correlation of PSMD1 expression with OS, we saw that when overexpressed, individuals did significantly worse with adrenocortical carcinoma, lower grade glioma, lung adenocarcinoma, mesothelioma, and uveal melanoma. In contrast, diffuse large B cell lymphoma (DLBCL) and stomach adenocarcinoma patients with lower PSMD1 expression had a worse OS. Similarly, patients with high levels of PSMD3 mRNA expression had a significantly worse OS in kidney chromophobe, skin cutaneous melanoma, uveal melanoma, and mesothelioma. Our analysis revealed that differential expression of PSMD1 and PSMD3 is correlated with survival in several different cancer types. Future directions will identify PSMD1 and PSMD3 post-translational modifications that may be novel targets for anticancer therapeutics. In conclusion, we highlight PSMD1 and PSMD3 as potential therapeutic targets for the development of novel proteasome inhibitors to treat cancer patients with less toxicity. Citation Format: Andres J. Rubio, Alfonso E. Bencomo-Alvarez, James E. Young, Vanessa V. Velazquez, Anna M. Eiring. PSMD1 and PSMD3 as putative targets for cancer therapy [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2021 Oct 7-10. Philadelphia (PA): AACR; Mol Cancer Ther 2021;20(12 Suppl):Abstract nr P150.

Ever since the ubiquitin proteasome system was characterized, efforts have been made to manipulate its function to abrogate the progression of cancer. As a result, the anti-cancer drugs bortezomib, carfilzomib, and ixazomib targeting the 26S proteasome were developed to treat multiple myeloma, mantle cell lymphoma, and diffuse large B-cell lymphoma, among others. Despite success, adverse side effects and drug resistance are prominent, raising the need for alternative therapeutic options. We recently demonstrated that knockdown of the 19S regulatory components, 26S proteasome non-ATPase subunits 1 (PSMD1) and 3 (PSMD3), resulted in increased apoptosis of chronic myeloid leukemia (CML) cells, but had no effect on normal controls, suggesting they may be good targets for therapy. Therefore, we hypothesized that PSMD1 and PSMD3 are potential targets for anti-cancer therapeutics and that their relevance stretches beyond CML to other types of cancers. In the present study, we analyzed PSMD1 and PSMD3 mRNA and protein expression in cancerous tissue versus normal controls using data from The Cancer Genome Atlas (TCGA) and the Clinical Proteomic Tumor Analysis Consortium (CPTAC), comparing expression with overall survival. Altogether, our data suggest that PSMD1 and PSMD3 may be novel putative targets for cancer prognosis and therapy that are worthy of future investigation.

Cancer is a challenging, multifaceted disease that involves a combination of biological and nonbiological factors. Aside from COVID-19, cancer is the second leading cause of death in the United States and the first among Hispanic Americans. The Hispanic population is the largest minority group in the United States, which is rapidly growing in size. Unfortunately, U.S. Hispanics and other minority groups experience many different health disparities, resulting in poor survival outcomes and a reduced quality of life. Factors such as genomic mutations, lower socioeconomic status, lack of education, reduced access to health care, comorbidities, and environmental factors all contribute to these health-care inequalities. In the context of blood cancer health disparities, Hispanic patients are often diagnosed at a younger age and have worse outcomes compared with non-Hispanic individuals. In this commentary, we highlight the existing knowledge about cancer health disparities in the Hispanic population, with a focus on chronic and acute leukemia. In our experience at the U.S./Mexican border, analysis of several different blood cancers demonstrated that younger Hispanic patients with acute lymphoid or myeloid leukemia have higher incidence rates and worse prognoses. A combined approach, involving improved health-care access and better knowledge of the underlying factors, will allow for more timely diagnoses and the development of intervention strategies aimed at reducing or eliminating the disparities.

Tyrosine kinase inhibitors (TKIs) targeting BCR-ABL1 have turned chronic myeloid leukemia (CML) from a fatal to a chronic disease. However, resistance is a clinical problem, and TKIs do not target CML leukemic stem cells (LSC), which are independent from BCR-ABL1 kinase activity. To understand mechanisms driving BCR-ABL1-independent resistance, we analyzed the transcriptional signature of TKI-naïve CD34+ cells from patients with or without a response to imatinib after 12 months of therapy (McWeeney et al. Blood 2010). Among the genes most profoundly downregulated in patients destined to fail imatinib was G0/G1 switch gene 2 (G0S2) (>3-fold, n=59, p50%) correlated with a longer overall survival (n=30 responders, n=16 non-responders, p=0.036). We next analyzed G0S2 mRNA expression in CD34+ cells from normal cord blood and from primary CD34+ cells lacking BCR-ABL1 kinase domain mutations. G0S2 was 4-fold downregulated in newly diagnosed CML (n=6) compared to normal cord blood (n=5, p3-fold in TKI-resistant (n=2) and BP-CML samples (n=5, p Citation Format: Mayra A. Gonzalez, Alfonso E. Bencomo, Christian Barreto-Vargas, Andres J. Rubio, Idaly M. Olivas, Joshua J. Lara, Anna Senina, Jonathan Ahmann, Katherine T. Varley, Luis F. Jave-Suarez, O'Hare Thomas, Michael W. Deininger, Anna M. Eiring. Role of G0S2 in chronic myeloid leukemia and TKI resistance [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 648.

Introduction: Hispanics represent the largest minority group in the United States (U.S.), with 57.5 million individuals. The majority of Hispanics in the U.S. reside in the Southwest region, and >11 million live in the state of Texas. Health disparities for Hispanic cancer patients have previously been linked to disproportionate poverty and other barriers to optimal healthcare. In the case of acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL), Hispanics were found to be diagnosed at a younger age and have worse overall survival (OS) compared with non-Hispanic whites (NHWs) (ACS Cancer Facts & Figures for Hispanics/Latinos 2018-2020). However, little is known about incidence and survival for Hispanic blood cancer patients residing at the U.S./Mexico border. Methods: We retrospectively reviewed data from the Texas Cancer Registry for hematologic malignancies diagnosed in the state of Texas between 1995 and 2016, focusing our analysis on chronic and acute leukemia's (both myeloid and lymphoid), myelodysplastic syndrome (MDS), and myeloproliferative neoplasms (MPNs). Survival for Hispanic and NHW groups was compared using the log-rank test, and Cox regression analyses adjusting for age and diagnosis. Differences in age at diagnosis were evaluated using t-tests and generalized linear models. Research was conducted according to a local IRB-approved protocol in accordance with the Declaration of Helsinki. Results: Of the 69,941 cases of hematologic malignancies with available information throughout the state of Texas, 18.29% identified as Hispanic. We found that Hispanic patients were diagnosed at a significantly younger age in all diseases analyzed (p Conclusions: Hispanics are diagnosed at a significantly younger age than NHWs in all blood malignancies analyzed. Hispanic patients with AML, MDS, and CML had significantly better OS compared to NHWs in unadjusted analyses, which could be explained by the reduced age of diagnosis. Hispanics with ALL, AML, or CML diagnosed near the U.S./Mexico border demonstrate worse OS compared with Hispanics diagnosed in other areas of Texas. In age-adjusted analyses, Hispanic patients with ALL or APL have a worse OS compared with NHWs. There appears to be evidence that disparities in outcome by ethnicity may be different in El Paso compared with the rest of Texas. Future Directions: Our data demonstrates blood cancer disparities present in our region. Further study is required to identify factors responsible for the disparity in OS, and to identify ways to address it. Citation Format: Alfonso E. Bencomo, Andres J. Rubio, Mayra A. Gonzalez, Idaly M. Olivas, Joshua J. Lara, Kiran Ghimire, Osvaldo Padilla, Angelica Padilla, Nawar Hakim, Attilio Orazi, Javier Corral, Alexander Philipovskiy, Sumit Gaur, Anna M. Eiring. Retrospective study of incidence and survival for patients with hematological malignancies residing at the U.S./Mexico border [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 4343.

Tyrosine kinase inhibitors (TKIs) targeting BCR-ABL1 are effective at eliminating most BCR-ABL1+ cells in chronic myeloid leukemia (CML), but do not target CML leukemic stem cells (LSCs), which are independent of BCR-ABL1 kinase activity. Our previous work showed that BCR-ABL1-independent resistance is largely driven by STAT3 (Eiring et al. Leukemia 2015). To understand how STAT3 contributes to TKI resistance, we performed RNA sequencing on TKI-sensitive K562-S cells versus TKI-resistant K562-R cells, which demonstrate kinase-independent resistance. Surprisingly, gene set enrichment analysis did not reveal STAT3-mediated transcription (p=1.0), but was reminiscent of TNFα signaling via NF-κB (p=0.024). Nucleocytoplasmic fractionation revealed higher levels of phospho-NF-κB in the nucleus of K562-R vs. K562-S controls, and in CD34+ progenitors from TKI-resistant CML patients (n=3) compared to TKI responders (n=2) and normal individuals (n=2). These data suggest NF-κB may be driving the transcriptional signature of TKI resistance, and implicate non-canonical functions for STAT3. To confirm increased NF-κB transcriptional activity in TKI resistance, K562-S and K562-R cells were transduced with an NF-κB luciferase reporter construct or a scrambled control. K562-R cells demonstrated increased NF-κB reporter activity compared to K562-S cells. To assess whether STAT3 activates NF-κB, reporter cells were co-infected with an inducible lentiviral shRNA targeting STAT3 (shSTAT3). STAT3 knockdown decreased NF-κB reporter activity in TKI-resistant cells, but increased reporter activity in TKI-sensitive controls. These data suggest that STAT3 may cooperate with NF-κB in the setting of TKI resistance. NF-κB and STAT3 have been shown to cooperatively bind to gene promoters of cytokines, specifically IL-6. ELISA assays demonstrated that K562-R cells produce autonomous IL-6, but not TNFα. Since IL-6 is a potent activator of STAT3, we treated TKI-resistant cells with the IL-6 receptor inhibitor, tocilizumab (100 and 1000 ng/ml). As expected, tocilizumab treatment reduced STAT3 Y705 phosphorylation as assessed by immunoblot analysis, but surprisingly had no effect on survival or NF-κB reporter activity in K562-R cells. These data suggest that STAT3 phosphorylation is not required for survival or NF-κB activation in TKI resistance. The role of unphosphorylated STAT3 in TKI resistance is currently being explored. We correlated our RNA sequencing data in TKI-resistant cell lines with microarray data in TKI-resistant patient samples (McWeeney et al. Blood 2010), identifying upregulation of the proteasome components, PSMD1 and PSMD3. qRT-PCR confirmed upregulation of PSMD3 but not PSMD1 in TKI resistance, and only the PSMD3 promoter was bound by NF-κB as assessed by ChIP. Lentiviral shRNAs targeting PSMD3 (shPSMD3) reduced colony formation of K562-S and K562-R cells, which correlated with induction of apoptosis, an increase of global protein ubiquitination, and decreased NF-κB activation, with little effects observed in normal cord blood. Altogether, our data implicate a positive feedback loop involving NF-κB and PSMD3 in TKI resistance and suggest that PSMD3 may be a novel target for treatment of TKI-resistant CML. Citation Format: Andres J. Rubio, Idaly M. Olivas, Alfonso E. Bencomo, Mayra A. Gonzalez, Joshua J. Lara, Rebecca Ellwood, Carme Ripoll-Fiol, Georgios Nteliopoulos, Alistair Reid, Dragana Milojkovic, Jane Apperley, Jamshid Sorouri-Khorashad, Anna M. Eiring. NF-kB associates with tyrosine kinase inhibitor resistance in chronic myeloid leukemia [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 647.

Introduction: Hispanics represent the largest minority group in the United States (U.S.), with 57.5 million individuals (18% of the population). Most U.S. Hispanics are of Mexican origin (63.2%), followed by Puerto Rican (9.5%), Cuban (3.9%), Salvadoran (3.8%), and Dominican (3.3%), but distribution varies by state. The majority of Hispanics in the U.S. reside in the Southwest region, and >11 million live in the state of Texas. Cancer is the leading cause of death in the Hispanic population, accounting for 21% of deaths in people of all ages. Health disparities for Hispanic cancer patients have previously been linked to disproportionate poverty and other barriers to optimal healthcare, and in the case of acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL), Hispanics were found to be diagnosed younger and to have worse overall survival (OS) than Non-Hispanic whites (NHWs) (ACS. Cancer Facts & Figures for Hispanics/Latinos 2018-2020). However, little is known about incidence and survival for Hispanic blood cancer patients residing at the U.S./Mexico border. To understand the impact of Hispanic ethnicity on outcomes for blood cancer patients diagnosed in this area, we examined OS in adult patients with hematologic malignancies throughout the state of Texas compared to Texas Health Service Region (HSR) 10, encompassing El Paso County. Methods: We retrospectively reviewed data available from the Texas Cancer Registry for hematologic malignancies diagnosed in the state of Texas between 1995 and 2016, focusing our analysis on chronic and acute leukemias (both myeloid and lymphoid), myelodysplastic syndrome (MDS), and myeloproliferative neoplasms (MPNs). Survival for Hispanic and NHW groups was compared using the log-rank test, and Cox regression analyses adjusting for age and diagnosis. Differences in age at diagnosis were evaluated using t-tests and generalized linear models. Similar analyses compared Hispanic patients from HSR 10 versus Hispanic patients from the rest of Texas. Research was conducted according to a local Institutional Review Board-approved protocol in accordance with the Declaration of Helsinki. Results: Of the 69,941 cases of hematologic malignancies with available information throughout the state of Texas, 18.29% self-identified as Hispanic. Surprisingly, in unadjusted analyses, Hispanic patients had significantly better OS than NHWs diagnosed with AML (p Conclusions: While Hispanic patients with AML, MDS, and CML had significantly better OS compared to NHWs in Texas as a whole, this could be explained by a significant reduction in the age of diagnosis for Hispanics. However, when comparing across Texas, El Paso Hispanics with ALL, AML, and CML have a worse prognosis than in the rest of the state. There appears to be evidence that disparities in outcome by ethnicity may be different in El Paso compared with the rest of Texas. Further study is required to identify factors responsible for the disparity in OS. Disclosures No relevant conflicts of interest to declare.