CK1δ and CK1ε Signaling Sustains Mitochondrial Metabolism and Cell Survival in Multiple Myeloma.

Multiple myeloma remains an incurable malignancy due to acquisition of intrinsic programs that drive therapy resistance. Here we report that casein kinase-1δ (CK1δ) and CK1ε are therapeutic targets in multiple myeloma that are necessary to sustain mitochondrial metabolism. Specifically, the dual CK1δ/CK1ε inhibitor SR-3029 had potent in vivo and ex vivo anti-multiple myeloma activity, including against primary multiple myeloma patient specimens. RNA sequencing (RNA-seq) and metabolic analyses revealed inhibiting CK1δ/CK1ε disables multiple myeloma metabolism by suppressing genes involved in oxidative phosphorylation (OxPhos), reducing citric acid cycle intermediates, and suppressing complexes I and IV of the electron transport chain. Finally, sensitivity of multiple myeloma patient specimens to SR-3029 correlated with elevated expression of mitochondrial genes, and RNA-seq from 687 multiple myeloma patient samples revealed that increased CSNK1D, CSNK1E, and OxPhos genes correlate with disease progression and inferior outcomes. Thus, increases in mitochondrial metabolism are a hallmark of multiple myeloma progression that can be disabled by targeting CK1δ/CK1ε. SIGNIFICANCE: CK1δ and CK1ε are attractive therapeutic targets in multiple myeloma whose expression increases with disease progression and connote poor outcomes, and that are necessary to sustain expression of genes directing OxPhos.

Whole-Exome sequencing analysis identified TMSB10/TRABD2A locus to be associated with carfilzomib-related cardiotoxicity among patients with multiple myeloma.

Background: Proteasome inhibitor Carfilzomib (CFZ) is effective in treating patients with refractory or relapsed multiple myeloma (MM) but has been associated with cardiovascular adverse events (CVAE) such as hypertension, cardiomyopathy, and heart failure. This study aimed to investigate the contribution of germline genetic variants in protein-coding genes in CFZ-CVAE among MM patients using whole-exome sequencing (WES) analysis. Methods: Exome-wide single-variant association analysis, gene-based analysis, and rare variant analyses were performed on 603,920 variants in 247 patients with MM who have been treated with CFZ and enrolled in the Oncology Research Information Exchange Network (ORIEN) at the Moffitt Cancer Center. Separate analyses were performed in European Americans and African Americans followed by a trans-ethnic meta-analysis. Results: The most significant variant in the exome-wide single variant analysis was a missense variant rs7148 in the thymosin beta-10/TraB Domain Containing 2A (TMSB10/TRABD2A) locus. The effect allele of rs7148 was associated with a higher risk of CVAE [odds ratio (OR) = 9.3 with a 95% confidence interval of 3.9-22.3, p = 5.42*10-7]. MM patients with rs7148 AG or AA genotype had a higher risk of CVAE (50%) than those with GG genotype (10%). rs7148 is an expression quantitative trait locus (eQTL) for TRABD2A and TMSB10. The gene-based analysis also showed TRABD2A as the most significant gene associated with CFZ-CVAE (p = 1.06*10-6). Conclusions: We identified a missense SNP rs7148 in the TMSB10/TRABD2A as associated with CFZ-CVAE in MM patients. More investigation is needed to understand the underlying mechanisms of these associations.

Replicative Instability Drives Cancer Progression.

In the past decade, defective DNA repair has been increasingly linked with cancer progression. Human tumors with markers of defective DNA repair and increased replication stress exhibit genomic instability and poor survival rates across tumor types. Seminal studies have demonstrated that genomic instability develops following inactivation of BRCA1, BRCA2, or BRCA-related genes. However, it is recognized that many tumors exhibit genomic instability but lack BRCA inactivation. We sought to identify a pan-cancer mechanism that underpins genomic instability and cancer progression in BRCA-wildtype tumors. Methods: Using multi-omics data from two independent consortia, we analyzed data from dozens of tumor types to identify patient cohorts characterized by poor outcomes, genomic instability, and wildtype BRCA genes. We developed several novel metrics to identify the genetic underpinnings of genomic instability in tumors with wildtype BRCA. Associated clinical data was mined to analyze patient responses to standard of care therapies and potential differences in metastatic dissemination. Results: Systematic analysis of the DNA repair landscape revealed that defective single-strand break repair, translesion synthesis, and non-homologous end-joining effectors drive genomic instability in tumors with wildtype BRCA and BRCA-related genes. Importantly, we find that loss of these effectors promotes replication stress, therapy resistance, and increased primary carcinoma to brain metastasis. Conclusions: Our results have defined a new pan-cancer class of tumors characterized by replicative instability (RIN). RIN is defined by the accumulation of intra-chromosomal, gene-level gain and loss events at replication stress sensitive (RSS) genome sites. We find that RIN accelerates cancer progression by driving copy number alterations and transcriptional program rewiring that promote tumor evolution. Clinically, we find that RIN drives therapy resistance and distant metastases across multiple tumor types.

CancerCellTracker: a brightfield time-lapse microscopy framework for cancer drug sensitivity estimation.

MOTIVATION: Time-lapse microscopy is a powerful technique that relies on images of live cells cultured ex vivo that are captured at regular intervals of time to describe and quantify their behavior under certain experimental conditions. This imaging method has great potential in advancing the field of precision oncology by quantifying the response of cancer cells to various therapies and identifying the most efficacious treatment for a given patient. Digital image processing algorithms developed so far require high-resolution images involving very few cells originating from homogeneous cell line populations. We propose a novel framework that tracks cancer cells to capture their behavior and quantify cell viability to inform clinical decisions in a high-throughput manner. RESULTS: The brightfield microscopy images a large number of patient-derived cells in an ex vivo reconstruction of the tumor microenvironment treated with 31 drugs for up to 6 days. We developed a robust and user-friendly pipeline CancerCellTracker that detects cells in co-culture, tracks these cells across time and identifies cell death events using changes in cell attributes. We validated our computational pipeline by comparing the timing of cell death estimates by CancerCellTracker from brightfield images and a fluorescent channel featuring ethidium homodimer. We benchmarked our results using a state-of-the-art algorithm implemented in ImageJ and previously published in the literature. We highlighted CancerCellTracker's efficiency in estimating the percentage of live cells in the presence of bone marrow stromal cells. AVAILABILITY AND IMPLEMENTATION: https://github.com/compbiolabucf/CancerCellTracker. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Racial and ethnic differences in clonal hematopoiesis, tumor markers, and outcomes of patients with multiple myeloma.

Multiple myeloma (MM) incidence, mortality, and survival vary by race and ethnicity, but the causes of differences remain unclear. We investigated demographic, clinical, and molecular features of diverse MM patients to elucidate mechanisms driving clinical disparities. This study included 495 MM patients (self-reported Hispanic, n = 45; non-Hispanic Black, n = 52; non-Hispanic White, n = 398). Hispanic and non-Hispanic Black individuals had an earlier age of onset than non-Hispanic White individuals (53 and 57 vs 63 years, respectively, P < .001). There were no differences in treatment by race and ethnicity groups, but non-Hispanic Black patients had a longer time to hematopoietic cell transplant than non-Hispanic White patients (376 days vs 248 days; P = .01). Overall survival (OS) was improved for non-Hispanic Black compared with non-Hispanic White patients (HR, 0.50; 95% CI, 0.31-0.81; P = .005), although this association was attenuated after adjusting for clinical features (HR, 0.62; 95% CI, 0.37-1.03; P = .06). Tumor mutations in IRF4 were most common in Hispanic patients, and mutations in SP140, AUTS2, and SETD2 were most common in non-Hispanic Black patients. Differences in tumor expression of BCL7A, SPEF2, and ANKRD26 by race and ethnicity were observed. Clonal hematopoiesis was detected in 12% of patients and associated with inferior OS in non-Hispanic Black patients compared with patients without clonal hematopoiesis (HR, 4.36; 95% CI, 1.36-14.00). This study provides insight into differences in molecular features that may drive clinical disparities in MM patients receiving comparable treatment, with the novel inclusion of Hispanic individuals.

Plasma cell dependence on histone/protein deacetylase 11 reveals a therapeutic target in multiple myeloma.

The clinical utility of histone/protein deacetylase (HDAC) inhibitors in combinatorial regimens with proteasome inhibitors for patients with relapsed and refractory multiple myeloma (MM) is often limited by excessive toxicity due to HDAC inhibitor promiscuity with multiple HDACs. Therefore, more selective inhibition minimizing off-target toxicity may increase the clinical effectiveness of HDAC inhibitors. We demonstrated that plasma cell development and survival are dependent upon HDAC11, suggesting this enzyme is a promising therapeutic target in MM. Mice lacking HDAC11 exhibited markedly decreased plasma cell numbers. Accordingly, in vitro plasma cell differentiation was arrested in B cells lacking functional HDAC11. Mechanistically, we showed that HDAC11 is involved in the deacetylation of IRF4 at lysine103. Further, targeting HDAC11 led to IRF4 hyperacetylation, resulting in impaired IRF4 nuclear localization and target promoter binding. Importantly, transient HDAC11 knockdown or treatment with elevenostat, an HDAC11-selective inhibitor, induced cell death in MM cell lines. Elevenostat produced similar anti-MM activity in vivo, improving survival among mice inoculated with 5TGM1 MM cells. Elevenostat demonstrated nanomolar ex vivo activity in 34 MM patient specimens and synergistic activity when combined with bortezomib. Collectively, our data indicated that HDAC11 regulates an essential pathway in plasma cell biology establishing its potential as an emerging theraputic vulnerability in MM.

IAP and HDAC inhibitors interact synergistically in myeloma cells through noncanonical NF-κB- and caspase-8-dependent mechanisms.

Interactions between the inhibitor of apoptosis protein antagonist LCL161 and the histone deacetylase inhibitor panobinostat (LBH589) were examined in human multiple myeloma (MM) cells. LCL161 and panobinostat interacted synergistically to induce apoptosis in diverse MM cell lines, including those resistant to bortezomib (PS-R). Similar interactions were observed with other histone deacetylase inhibitors (MS-275) or inhibitors of apoptosis protein antagonists (birinapant). These events were associated with downregulation of the noncanonical (but not the canonical) NF-κB pathway and activation of the extrinsic, caspase-8-related apoptotic cascade. Coexposure of MM cells to LCL161/LBH589 induced TRAF3 upregulation and led to TRAF2 and NIK downregulation, diminished expression of BCL-XL, and induction of γH2A.X. Ectopic expression of TRAF2, NIK, or BCL-XL, or short hairpin RNA TRAF3 knock-down, significantly reduced LCL161/LBH589 lethality, as did ectopic expression of dominant-negative FADD. Stromal/microenvironmental factors failed to diminish LCL161/LBH589-induced cell death. The LCL161/LBH589 regimen significantly increased cell killing in primary CD138+ cells (N = 31) and was particularly effective in diminishing the primitive progenitor cell-enriched CD138-/19+/20+/27+ population (N = 23) but was nontoxic to normal CD34+ cells. Finally, combined LCL161/LBH589 treatment significantly increased survival compared with single-agent treatment in an immunocompetent 5TGM1 murine MM model. Together, these findings argue that LCL161 interacts synergistically with LBH589 in MM cells through a process involving inactivation of the noncanonical NF-κB pathway and activation of the extrinsic apoptotic pathway, upregulation of TRAF3, and downregulation of TRAF2/BCL-XL. Notably, this regimen overcomes various forms of resistance, is active against primary MM cells, and displays significant in vivo activity. This strategy warrants further consideration in MM.

Therapeutic Targeting of Protein Disulfide Isomerase PDIA1 in Multiple Myeloma.

Multiple myeloma is a genetically complex hematologic neoplasia in which malignant plasma cells constantly operate at the maximum limit of their unfolded protein response (UPR) due to a high secretory burden of immunoglobulins and cytokines. The endoplasmic reticulum (ER) resident protein disulfide isomerase, PDIA1 is indispensable for maintaining structural integrity of cysteine-rich antibodies and cytokines that require accurate intramolecular disulfide bond arrangement. PDIA1 expression analysis from RNA-seq of multiple myeloma patients demonstrated an inverse relationship with survival in relapsed or refractory disease, supporting its critical role in myeloma persistence. Using a structure-guided medicinal chemistry approach, we developed a potent, orally bioavailable small molecule PDIA1 inhibitor CCF642-34. The inhibition of PDIA1 overwhelms the UPR in myeloma cells, resulting in their apoptotic cell death at doses that do not affect the normal CD34+ hematopoietic stem and progenitor cells. Bortezomib resistance leads to increased PDIA1 expression and thus CCF642-34 sensitivity, suggesting that proteasome inhibitor resistance leads to PDIA1 dependence for proteostasis and survival. CCF642-34 induces acute unresolvable UPR in myeloma cells, and oral treatment increased survival of mice in the syngeneic 5TGM1 model of myeloma. Results support development of CCF642-34 to selectively target the plasma cell program and overcome the treatment-refractory state in myeloma.

Metabolic Changes Are Associated with Melphalan Resistance in Multiple Myeloma.

Multiple myeloma is an incurable hematological malignancy that impacts tens of thousands of people every year in the United States. Treatment for eligible patients involves induction, consolidation with stem cell rescue, and maintenance. High-dose therapy with a DNA alkylating agent, melphalan, remains the primary drug for consolidation therapy in conjunction with autologous stem-cell transplantation; as such, melphalan resistance remains a relevant clinical challenge. Here, we describe a proteometabolomic approach to examine mechanisms of acquired melphalan resistance in two cell line models. Drug metabolism, steady-state metabolomics, activity-based protein profiling (ABPP, data available at PRIDE: PXD019725), acute-treatment metabolomics, and western blot analyses have allowed us to further elucidate metabolic processes associated with melphalan resistance. Proteometabolomic data indicate that drug-resistant cells have higher levels of pentose phosphate pathway metabolites. Purine, pyrimidine, and glutathione metabolisms were commonly altered, and cell-line-specific changes in metabolite levels were observed, which could be linked to the differences in steady-state metabolism of naïve cells. Inhibition of selected enzymes in purine synthesis and pentose phosphate pathways was evaluated to determine their potential to improve melphalan's efficacy. The clinical relevance of these proteometabolomic leads was confirmed by comparison of tumor cell transcriptomes from newly diagnosed MM patients and patients with relapsed disease after treatment with high-dose melphalan and autologous stem-cell transplantation. The observation of common and cell-line-specific changes in metabolite levels suggests that omic approaches will be needed to fully examine melphalan resistance in patient specimens and define personalized strategies to optimize the use of high-dose melphalan.

Transcriptional programming drives Ibrutinib-resistance evolution in mantle cell lymphoma.

Ibrutinib, a bruton's tyrosine kinase (BTK) inhibitor, provokes robust clinical responses in aggressive mantle cell lymphoma (MCL), yet many patients relapse with lethal Ibrutinib-resistant (IR) disease. Here, using genomic, chemical proteomic, and drug screen profiling, we report that enhancer remodeling-mediated transcriptional activation and adaptive signaling changes drive the aggressive phenotypes of IR. Accordingly, IR MCL cells are vulnerable to inhibitors of the transcriptional machinery and especially so to inhibitors of cyclin-dependent kinase 9 (CDK9), the catalytic subunit of the positive transcription elongation factor b (P-TEFb) of RNA polymerase II (RNAPII). Further, CDK9 inhibition disables reprogrammed signaling circuits and prevents the emergence of IR in MCL. Finally, and importantly, we find that a robust and facile ex vivo image-based functional drug screening platform can predict clinical therapeutic responses of IR MCL and identify vulnerabilities that can be targeted to disable the evolution of IR.

A pharmacodynamic model of clinical synergy in multiple myeloma.

BACKGROUND: Multiagent therapies, due to their ability to delay or overcome resistance, are a hallmark of treatment in multiple myeloma (MM). The growing number of therapeutic options in MM requires high-throughput combination screening tools to better allocate treatment, and facilitate personalized therapy. METHODS: A second-order drug response model was employed to fit patient-specific ex vivo responses of 203 MM patients to single-agent models. A novel pharmacodynamic model, developed to account for two-way combination effects, was tested with 130 two-drug combinations. We have demonstrated that this model is sufficiently parameterized by single-agent and fixed-ratio combination responses, by validating model estimates with ex vivo combination responses for different concentration ratios, using a checkerboard assay. This new model reconciles ex vivo observations from both Loewe and BLISS synergy models, by accounting for the dimension of time, as opposed to focusing on arbitrary time-points or drug effect. Clinical outcomes of patients were simulated by coupling patient-specific drug combination models with pharmacokinetic data. FINDINGS: Combination screening showed 1 in 5 combinations (21.43% by LD50, 18.42% by AUC) were synergistic ex vivo with statistical significance (P < 0.05), but clinical synergy was predicted for only 1 in 10 combinations (8.69%), which was attributed to the role of pharmacokinetics and dosing schedules. INTERPRETATION: The proposed framework can inform clinical decisions from ex vivo observations, thus providing a path toward personalized therapy using combination regimens. FUNDING: This research was funded by the H. Lee Moffitt Cancer Center Physical Sciences in Oncology (PSOC) Grant (1U54CA193489-01A1) and by H. Lee Moffitt Cancer Center's Team Science Grant. This work has been supported in part by the PSOC Pilot Project Award (5U54CA193489-04), the Translational Research Core Facility at the H. Lee Moffitt Cancer Center & Research Institute, an NCI-designated Comprehensive Cancer Center (P30-CA076292), the Pentecost Family Foundation, and Miles for Moffitt Foundation.

BCL2 Amplicon Loss and Transcriptional Remodeling Drives ABT-199 Resistance in B Cell Lymphoma Models.

Drug-tolerant "persister" tumor cells underlie emergence of drug-resistant clones and contribute to relapse and disease progression. Here we report that resistance to the BCL-2 targeting drug ABT-199 in models of mantle cell lymphoma and double-hit lymphoma evolves from outgrowth of persister clones displaying loss of 18q21 amplicons that harbor BCL2. Further, persister status is generated via adaptive super-enhancer remodeling that reprograms transcription and offers opportunities for overcoming ABT-199 resistance. Notably, pharmacoproteomic and pharmacogenomic screens revealed that persisters are vulnerable to inhibition of the transcriptional machinery and especially to inhibition of cyclin-dependent kinase 7 (CDK7), which is essential for the transcriptional reprogramming that drives and sustains ABT-199 resistance. Thus, transcription-targeting agents offer new approaches to disable drug resistance in B-cell lymphomas.

PLK1 stabilizes a MYC-dependent kinase network in aggressive B cell lymphomas.

Concordant activation of MYC and BCL-2 oncoproteins in double-hit lymphoma (DHL) results in aggressive disease that is refractory to treatment. By integrating activity-based proteomic profiling and drug screens, polo-like kinase-1 (PLK1) was identified as an essential regulator of the MYC-dependent kinome in DHL. Notably, PLK1 was expressed at high levels in DHL, correlated with MYC expression, and connoted poor outcome. Further, PLK1 signaling augmented MYC protein stability, and in turn, MYC directly induced PLK1 transcription, establishing a feed-forward MYC-PLK1 circuit in DHL. Finally, inhibition of PLK1 triggered degradation of MYC and of the antiapoptotic protein MCL-1, and PLK1 inhibitors showed synergy with BCL-2 antagonists in blocking DHL cell growth, survival, and tumorigenicity, supporting clinical targeting of PLK1 in DHL.

Resistance to Ibrutinib in B Cell Malignancies: One Size Does Not Fit All.

Ibrutinib resistance, as a result of coordinated rewiring of signaling networks and enforced tumor microenvironment (TME)-lymphoma interactions, drives unrestrained proliferation and disease progression. To combat resistance mechanisms, we must identify the compensatory resistance pathways and the central modulators of reprogramming events. Targeting the transcriptome and kinome reprogramming of lymphoma cells represents a rational approach to mitigate ibrutinib resistance in B cell malignancies. However, with the apparent heterogeneity and plasticity of tumors shown in therapy response, a one size fits all approach may be unattainable. To this end, a reliable and real-time drug screening platform to tailor effective individualized therapies in patients with B cell malignancies is warranted. Here, we describe the complexity of ibrutinib resistance in B cell lymphomas and the current approaches, including a drug screening assay, which has the potential to further explore the mechanisms of ibrutinib resistance and to design effective individualized combination therapies to overcome resistance and disable aggressive lymphomas (see Outstanding Questions).

Classifying the evolutionary and ecological features of neoplasms.

Neoplasms change over time through a process of cell-level evolution, driven by genetic and epigenetic alterations. However, the ecology of the microenvironment of a neoplastic cell determines which changes provide adaptive benefits. There is widespread recognition of the importance of these evolutionary and ecological processes in cancer, but to date, no system has been proposed for drawing clinically relevant distinctions between how different tumours are evolving. On the basis of a consensus conference of experts in the fields of cancer evolution and cancer ecology, we propose a framework for classifying tumours that is based on four relevant components. These are the diversity of neoplastic cells (intratumoural heterogeneity) and changes over time in that diversity, which make up an evolutionary index (Evo-index), as well as the hazards to neoplastic cell survival and the resources available to neoplastic cells, which make up an ecological index (Eco-index). We review evidence demonstrating the importance of each of these factors and describe multiple methods that can be used to measure them. Development of this classification system holds promise for enabling clinicians to personalize optimal interventions based on the evolvability of the patient's tumour. The Evo- and Eco-indices provide a common lexicon for communicating about how neoplasms change in response to interventions, with potential implications for clinical trials, personalized medicine and basic cancer research.

A preclinical assay for chemosensitivity in multiple myeloma.

Accurate preclinical predictions of the clinical efficacy of experimental cancer drugs are highly desired but often haphazard. Such predictions might be improved by incorporating elements of the tumor microenvironment in preclinical models by providing a more physiological setting. In generating improved xenograft models, it is generally accepted that the use of primary tumors from patients are preferable to clonal tumor cell lines. Here we describe an interdisciplinary platform to study drug response in multiple myeloma, an incurable cancer of the bone marrow. This platform uses microfluidic technology to minimize the number of cells per experiment, while incorporating three-dimensional extracellular matrix and mesenchymal cells derived from the tumor microenvironment. We used sequential imaging and a novel digital imaging analysis algorithm to quantify changes in cell viability. Computational models were used to convert experimental data into dose-exposure-response "surfaces," which offered predictive utility. Using this platform, we predicted chemosensitivity to bortezomib and melphalan, two clinical multiple myeloma treatments, in three multiple myeloma cell lines and seven patient-derived primary multiple myeloma cell populations. We also demonstrated how this system could be used to investigate environment-mediated drug resistance and drug combinations that target it. This interdisciplinary preclinical assay is capable of generating quantitative data that can be used in computational models of clinical response, demonstrating its utility as a tool to contribute to personalized oncology.

Evolutionary approaches to prolong progression-free survival in breast cancer.

Many cancers adapt to chemotherapeutic agents by upregulating membrane efflux pumps that export drugs from the cytoplasm, but this response comes at an energetic cost. In breast cancer patients, expression of these pumps is low in tumors before therapy but increases after treatment. While the evolution of therapeutic resistance is virtually inevitable, proliferation of resistant clones is not, suggesting strategies of adaptive therapy. Chemoresistant cells must consume excess resources to maintain resistance mechanisms, so adaptive therapy strategies explicitly aim to maintain a stable population of therapy-sensitive cells to suppress growth of resistant phenotypes through intratumoral competition. We used computational models parameterized by in vitro experiments to illustrate the efficacy of such approaches. Here, we show that low doses of verapamil and 2-deoxyglucose, to accentuate the cost of resistance and to decrease energy production, respectively, could suppress the proliferation of drug-resistant clones in vivo. Compared with standard high-dose-density treatment, the novel treatment we developed achieved a 2-fold to 10-fold increase in time to progression in tumor models. Our findings challenge the existing flawed paradigm of maximum dose treatment, a strategy that inevitably produces drug resistance that can be avoided by the adaptive therapy strategies we describe.

Buffer Therapy for Cancer.

Oral administration of pH buffers can reduce the development of spontaneous and experimental metastases in mice, and has been proposed in clinical trials. Effectiveness of buffer therapy is likely to be affected by diet, which could contribute or interfere with the therapeutic alkalinizing effect. Little data on food pH buffering capacity was available. This study evaluated the pH and buffering capacity of different foods to guide prospective trials and test the effect of the same buffer (lysine) at two different ionization states. Food groups were derived from the Harvard Food Frequency Questionnaire. Foods were blended and pH titrated with acid from initial pH values until 4.0 to determine "buffering score", in mmol H+/pH unit. A "buffering score" was derived as the mEq H+ consumed per serving size to lower from initial to a pH 4.0, the postprandial pH of the distal duodenum. To differentiate buffering effect from any metabolic byproduct effects, we compared the effects of oral lysine buffers prepared at either pH 10.0 or 8.4, which contain 2 and 1 free base amines, respectively. The effect of these on experimental metastases formation in mice following tail vein injection of PC-3M prostate cancer cells were monitored with in vivo bioluminescence. Carbohydrates and dairy products' buffering score varied between 0.5 and 19. Fruits and vegetables showed a low to zero buffering score. The score of meats varied between 6 and 22. Wine and juices had negative scores. Among supplements, sodium bicarbonate and Tums® had the highest buffering capacities, with scores of 11 and 20 per serving size, respectively. The "de-buffered" lysine had a less pronounced effect of prevention of metastases compared to lysine at pH 10. This study has demonstrated the anti-cancer effects of buffer therapy and suggests foods that can contribute to or compete with this approach to manage cancer.

Adaptation to survival in germinal center is the initial step in onset of indolent stage of multiple myeloma.

Aberrant mutations of centrocytes in germinal centers (GC) can generate two completely different diseases: B-cell lymphomas and monoclonal gammopathy of undetermined significance (MGUS). In this article we use computational models to examine the evolutionary dynamics by which initial adaptation to survival in the GC allows naive MGUS cells to proliferate in the bone marrow and initiate the evolutionary process that will lead to aggressive multiple myeloma (MM). Our simulations show that MGUS cells may generate bone marrow tumors ranging from indolent to aggressive, depending on the original adaptation in the GC. All these tumors, however, are limited to approximately 15% of the marrow cellularity due to hypoxia-induced quiescence (this correlates with the cellularity that separates MGUS and MM, ∼10%). Resistance to hypoxia-induced quiescence and cell death was one of the two major bone marrow adaptations that allowed continued tumor growth and establishment of paracrine cytokine loops, known to increase MM cell replication and de novo multidrug resistance. The second major adaptation was an increase in IL-6-independent growth rate, which correlates with the mutations observed in advanced stage patients. Even though there was an increase in the microvessel density in all simulations, the "angiogenic switch" was not due to a MM angiogenic phenotype, but rather the response of MM cells to the regional hypoxia caused by the increased tumor burden. These results indicate that treatments targeting the adaptation to survival and proliferation in hypoxia, in conjunction with currently available therapies, may have synergistic effects, by delaying tumor growth and reducing cytokine paracrine loops mediated by angiogenic factors.

Evolution of tumor invasiveness: the adaptive tumor microenvironment landscape model.

Interactions between cancer cells and their microenvironment are crucial for promoting tumor growth and invasiveness. In the tumor adaptive landscape model, hypoxic and acidic microenvironmental conditions reduce the fitness of cancer cells and significantly restrict their proliferation. This selects for enhanced motility as cancer cells may evolve an invasive phenotype if the consequent cell movement is rewarded by proliferation. Here, we used an integrative approach combining a mathematical tumor adaptive landscape model with experimental studies to examine the evolutionary dynamics that promote an invasive cancer phenotype. Computer simulation results hypothesized an explicit coupling of motility and proliferation in cancer cells. The mathematical modeling results were also experimentally examined by selecting Panc-1 cells with enhanced motility on a fibroblast-derived 3-dimensional matrix for cells that move away from the unfavorable metabolic constraints. After multiple rounds of selection, the cells that adapted through increased motility were characterized for their phenotypic properties compared with stationary cells. Microarray and gene depletion studies showed the role of Rho-GDI2 in regulating both cell movement and proliferation. Together, this work illustrates the partnership between evolutionary mathematical modeling and experimental validation as a potentially useful approach to study the complex dynamics of the tumor microenvironment.