Bacteria were first detected in human tumors more than 100 years ago, but the characterization of the tumor microbiome has remained challenging because of its low biomass. We undertook a comprehensive analysis of the tumor microbiome, studying 1526 tumors and their adjacent normal tissues across seven cancer types, including breast, lung, ovary, pancreas, melanoma, bone, and brain tumors. We found that each tumor type has a distinct microbiome composition and that breast cancer has a particularly rich and diverse microbiome. The intratumor bacteria are mostly intracellular and are present in both cancer and immune cells. We also noted correlations between intratumor bacteria or their predicted functions with tumor types and subtypes, patients’ smoking status, and the response to immunotherapy.

Highlights d Human fetuses in 2 nd trimester show T cell diversity with effector-memory phenotype d Fetal organs show diverse bacterial genera that can be cultured and propagated d Bacterial structures with mucin-like threads are visualized in 14-weeks EGA fetal gut d Fetal bacteria induce syngeneic memory T cell activation in fetal mLN T cells

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Neurogenesis, the formation of new neurons in the adult brain, is important for memory formation and extinction. One of the most studied external interventions that affect the rate of adult neurogenesis is physical exercise. Physical exercise promotes adult neurogenesis via several factors including brain-derived neurotrophic factor (BDNF) and vascular endothelial growth factor (VEGF). Here, we identified L -lactate, a physical exercise-induced metabolite, as a factor that promotes adult hippocampal neurogenesis. While prolonged exposure to L -lactate promoted neurogenesis, no beneficial effect was exerted on cognitive learning and memory. Systemic pharmacological blocking of monocarboxylate transporter 2 (MCT2), which transports L -lactate to the brain, prevented lactate-induced neurogenesis, while 3,5-dihydroxybenzoic acid (3,5-DHBA), an agonist for the lactate-receptor hydroxycarboxylic acid receptor 1 (HCAR1), did not affect adult neurogenesis. These data suggest that L -lactate partially mediates the effect of physical exercise on adult neurogenesis, but not cognition, in a MCT2-dependent manner.

Although most lung cancers with EGFR-mutation respond to osimertinib, resistance eventually develops, and there are no approved targeted therapies once resistance to osimertinib occurs. Intra-tumoral bacteria are emerging as a cause of therapy resistance in cancer. In order to determine if intra-tumoral bacteria play a role in the resistance of EGFR-mutant lung cancers to EGFR-TKIs, including osimertinib, we used a bacterial pre-conditioned medium (PCM) screening system which utilized various bacteria identified in lung cancer specimens. We used a cell viability evaluation method in GFP-labeled PC9 cells (EGFR Ex19del) to identify the bacterial PCM that may lead to resistance to EGFR-TKIs. We found that PCM from Chryseobacterium indologenes (C-PCM) markedly rescued PC9 from osimertinib. Furthermore, we found that C-PCM also could increase cell viability in other EGFR-mutated cell lines HCC4006, HCC827, and H1650. We found that proteinase K treatment eliminated the rescue of C-PCM on EGFR-TKI treatment, indicating that proteins within C-PCM are responsible for its effect of EGFR-TKI treatment. Mass spectrometry (MS) analysis showed that C-PCM does not degrade osimertinib, suggesting that C-PCM mediated resistance does not occur through drug degradation. To understand the mechanism by which C-PCM meditates resistance to osimertinib in NSCLC, we performed phosphor-Receptor Kinase Array to detect changes in phosphorylation between C-PCM and non-C-PCM treatment in PC9 cells. This showed no change in phosphorylation levels of EGFR, indicating that rescue from osimertinib is not through re-activation of EGFR pathway. However, phosphorylation levels of MET and IGF1R were significantly increased when C-PCM was applied to PC9 cells regardless of the presence or absence of osimertinib. Knockdown of IGF1R significantly increased the sensitivity of PC9 cell to osimertinib. Similar results were observed in siMET-PC9 cells. Interestingly, we found that C-PCM lost its ability to rescue PC9 cells from osimertinib when IGF1R was knocked down. Also, inhibition of IGF1R activation by the IGF1R inhibitor linsitinib increased PC9 cell sensitivity to osimertinib, and C-PCM rescue ability decreased with increased linsitinib concentration. Even though MET knockdown increased PC9 cell sensitivity to osimertinib, C-PCM still could increase cell viability with MET knockdown. These results together indicate that activation of IGF1R may be the main pathway that mediates Chryseobacterium indologenes resistance to osimertinib. Further mechanistic studies of C-PCM mediated resistance are underway to understand the precise mechanism of bacterial-mediated EGFR-TKI resistance. Additional research on the clinical relevance of these findings may lead to new strategies to overcome EGFR-TKI-resistance. Citation Format: Wendong Li, Keqiang Zhang, Aviva Rotter-Maskowitz, Deborah Nejman, Ravid Straussman, Dan Raz. Role of intra-tumoral bacteria in EGFR-tyrosine kinase inhibitor resistance [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1697.

Mutations of the epidermal growth factor receptor (EGFR) are the most common group of actionable mutations in lung cancer, and they predict response to EGFR-tyrosine kinase inhibitos (EGFR-TKIs). The third-generation EGFR-TKI osimertinib is the standard of care for first-line treatment of patients with advanced EGFR-mutated lung cancer. Although most lung cancers with EGFR-mutation respond to osimertinib, resistance is eventually observed in all patients, and there are no approved targeted therapies once resistance to osimertinib occurs. Mechanisms of resistance to osimertinib are poorly understood and involve both EGFR and non-EGFR related pathways. Intra-tumoral bacteria are emerging as a cause of therapy resistance in cancer.Intra-tumoral bacteria have been reported in many cancers and emerging evidence suggests that the tumor microbiome plays an important role in drug resistance in a variety of cancers. But the link between intra-tumoral bacteria and therapy resistance in lung cancer remains elusive. We previously described the tumor microbiome identified within a cohort of non-small cell lung cancers. In order to determine if intra-tumoral bacteria within lung adenocarcinoma patient with EGFR mutation could play role on the resistance to EGFR-TKIs, such as osimertinib and erlotinib, we used a bacterial pre-conditioned medium (PCM) screening system which utilized various bacteria identified in lung cancer specimens. We used a cell viability evaluation method in GFP-labeled PC9 cells (EGFR Ex19del) to identify the bacterial PCM that may lead to resistance to EGFR-TKIs. We found that PCM from Chryseobacterium indologenes (C-PCM) markedly rescued PC9 cells from osimertinib or erlotinib treatment. We also found that proteinase K treatment eliminated the rescue of C-PCM on EGFR-TKI treatment, suggesting that a proteins within C-PCM are responsible for its effect of EGFR-TKI treatment. Mass spectrometry (MS) analysis showed that C-PCM does not degrade osimertinib or erlotinib, suggesting that C-PCM mediated resistance does not occur through drug degradation. Western blot analysis showed that C-PCM does not affect EGFR, phospho-EGFR, or AKT expression level but C-PCM significantly increased AKT phosphorylation level. Our current studies indicated that intra-tumoral bacteria within lung adenocarcinoma patient may leads to resistance to osimertinib and erlotinib. Further mechanistic studies of C-PCM mediated resistance are under way and may lead to new therapies to overcome osimertinib-resistance. Citation Format: Wendong Li, Aviva Rotter-Maskowitz, Ting Sun, Keqiang Zhang, Ravid Straussman, Dan Raz. Role of intra-tumoral bacteria in EGFR-tyrosine kinase inhibitor resistance [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1327.