Pneumocandin B (PB) is a lipohexapeptide synthesized by and serves as the precursor for the widely used antifungal drug caspofungin acetate (Cancidas®). However, the low titer of PB results in fermentation and purification costs during caspofungin production, limiting its widespread clinical application. Here, we engineered an efficient PB-producing strain of by systems metabolic engineering strategies, including multi-omics analysis and multilevel metabolic engineering. We overexpressed four rate-limiting enzymes: thioesterase GLHYD, two cytochrome P450s GLP450s, and chorismate synthase GLCS; knocked out two competing pathways responsible for producing 6-methylsalicylic acid and pyranidine E; and overexpressed the global transcriptional activator GLHYP. As a result, the PB titer increased by 108.7 % to 2.63 g/L at the shake-flask level through combinatorial strategies. Our study provides valuable insights into achieving high-level production of PB and offers general guidance for developing efficient fungal cell factories to produce polyketide synthase-non-ribosomal peptide synthetase hybrid metabolites.