C₂₇ to C₃₂ sterols found in Pneumocystis, and opportunistic pathogen of immunocompromised mammals

Abstract

Pneumocystis carinii is the paradigm of opportunistic infections in immunocompromised mammals. Prior to the acquired immunodeficiency syndrome (AIDS) pandemic and the use of immunosuppressive therapy in organ transplant and cancer patients, P. carinii was regarded as a curiosity, rarely observed clinically. Interest in this organism exploded when it was identified as the agent of P. carinii pneumonia (PcP), the direct cause of death among many AIDS patients. Aggressive prophylaxis has decreased the number of acute PcP cases, but it remains among the most prevalent opportunistic infections found within this patient population. The taxonomic assignment of P. carinii has long been argued; molecular genetics data now demonstrate that it is a fungus. Several antimycotic drugs are targeted against ergosterol or its biosynthesis, but these are not as effective against PcP as they are against other fungal infections. This can now be explained in part by the identification of the sterols of P. carinii. The organism lacks ergosterol but contains distinct C28 and C29 delta7 24-alkylsterols. Also, 24-methylenelanost-8-en-3beta-ol (C31) and pneumocysterol, (24Z)-ethylidenelanost-8-en-3beta-ol (C32) were recently identified in organisms infecting humans. Together, the delta7 24-alkylsterols and pneumocysterol are regarded as signature lipids of the pathogen that can be useful for the diagnosis of PcP, since no other lung pathogen is known to contain them. Cholesterol (C27), the dominant sterol component in P. carinii, is probably totally scavenged from the host. De novo synthesis of sterols has been demonstrated by the presence of lovastatin-sensitive 3-hydroxy-3-methylglutaryl-CoA reductase activity, the incorporation of radiolabeled mevalonate and squalene into P. carinii sterols, and the reduction in cellular ATP in cells treated with inhibitors of enzymes in sterol biosynthesis.

Comprehensive and definitive structural identities of Pneumocystis carinii sterols

“…This pooled sample, containing 35 mg total sterols, made it possible to fractionate the components by nondestructive HPLC and to analyze the isolated fractions by NMR. Here we present a comprehensive NMR analysis providing unambiguous structural assignments for 43 P. carinii sterols and correlate their identities with those previously characterized by GLC and GLC-MS (9,11,13). Most of the sterol molecular species were 24-alkylsterols, consistent with the suggestion that the Pneumocystis S -adenosyl-l -methionine:C-24 sterol methyl transferase (SAM: SMT) activity and other sterol biosynthesis reactions in this pathogen are excellent targets for the development of new therapies against PcP (2, 9 Ϫ 22).…”

“…Several of these were identified as 24-alkysterols but ergosterol, the dominant sterol of most fungal pathogens, was not among them (9,11,14,15). The structural assignments of several of these sterol components were made by analyses using TLC, GLC, and GLC-mass spectrometry (MS) in conjunction with chemically synthesized authentic standards.…”

Evidence for Cholesterol Scavenging by Pneumocystis and Potential Modifications of Host‐Synthesized Sterols by the P. carinii SAM:SMT

“…Cholesterol (cholest‐S‐en‐3β‐ol) comprises approximately 75% of the sterol mass in P. carinii [2,3]. We have evidence that the organism in the mammalian lung scavenges most, if not all its cholesterol from its host.…”

“…Pneumocystis carinii and P. jirovecii contain a large number of 24‐alkylsterols [1–5] S‐Adenosy‐L‐methionine: sterol C‐24 methyl transferase (SAM:SMT) is the enzyme that catalyzes the transfer of methyl groups from SAM to the C‐24 position of the sterol side chain. SAM:SMT is an attractive target for chemotherapeutic attack against the pathogen since mammals do not have this enzyme and hence do not synthesize 24‐alkylsterols.…”

“…Similar to the situation in some plants, the P. carinii SAM:SMT can transfer one or two methyl groups to the sterol side chain to produce both C 28 and C 29 sterols. Unlike SAM:SMT from most other fungal pathogens infecting mammals, the enzyme in P. carinii prefers lanosterol instead of zymosterol as substrate [2,4,5] and synthesizes a number of C 31 and C 32 lanosterol derivatives [2,3]. …”