EA041924B1 - AN ANALOGUE OF CYCLIC DINUCLEOTIDE, ITS PHARMACEUTICAL COMPOSITION AND APPLICATION - Google Patents

Description

The present application claims priority according to China Patent Application No. CN 201811105973.6 filed September 21, 2018, China Patent Application No. CN 201811276297.9 filed October 30, 2018, China Patent Application No. CN 201910042984.2 filed January 20, 19 and China Patent Application No. CN 201910287528.4, filed April 11, 2019. The content of the above patent applications in

China is incorporated herein by reference in its entirety.

Technical field

The present invention relates to a cyclic dinucleotide analog, a pharmaceutical composition thereof, and its use as an interferon gene stimulator (STING) agonist and for activation of the interferon gene stimulator (STING) pathway.

State of the art

Interferon gene stimulator (STING), also known as T1MEM173, MITA, MPYS, and ERIS, is an important signaling molecule in innate immunity signaling. The protein encoded by this gene contains a five-strand transmembrane structure and plays an important regulatory role in the immune response associated with viral or bacterial infection. The interferon gene stimulator is a pattern recognition receptor that detects exogenous nucleic acids in the cytoplasm and activates signal transduction pathways associated with type I interferon responses. In addition, the interferon gene stimulator has been shown to be involved in the regulation of apoptotic signaling through interaction with major histocompatibility complex type II (MHCII). Studies in human tumors with spontaneous T cell infiltration have shown that CD8+ T cell infiltration is closely related to the transcriptional profile of type I interferons (Harlin et al., Cancer R ^e s, 2009; 69(7): OF1). Mechanistic studies in mice have shown that T-cell activation processes for tumor-associated antigens exhibit abnormalities in experimental animals with defective type I interferon signaling (Diamond et al., J. Exp. Med, 2011; 208(10): 1989; Fuerte et al., J. Exp. Med, 2011; 208(10):2005). Further studies of tumor recognition by the innate immune system in vivo and the signaling pathways involved in this process, such as tumor-triggered antigen presenting cell (APC)-mediated interferon (IFN) expression, showed that the interferon gene stimulator signaling pathway is activated by cytoplasmic DNA and that these exogenous nucleic acids can be recognized by cyclo-GMP-AMP synthetase (cGAS) and then catalyze the formation of cyclic nucleic acids such as cyclic GMP-AMP (cGMP), which can act as endogenous ligands that activate interferon gene stimulator signaling (Sun et al., Science, 2013; 339(15): 786). An activated interferon gene stimulator can subsequently induce TBK1 type kinase autophosphorylation and phosphorylation of type III interferon regulatory factor (IRF-3), and phosphorylated type III interferon regulatory factor can further activate the transcription process of type I interferon genes and regulate the synthesis and secretion of type I interferon, which , in turn, elicits an immune response. Thus, the interferon gene stimulator signaling pathway has been shown to play an extremely important role in the process of tumor recognition by the innate immune system, and activation of this signaling pathway on antigen-presenting cells is directly related to the activation of T cells against tumor-associated antigens. Based on its role in tumor immune recognition, it would be expected that activation of interferon stimulator gene signaling through drugs or other pharmacological approaches could enhance interferon expression and have a positive effect on tumor therapy. Therefore, the development of interferon gene stimulator signaling agonists for the treatment of neoplastic diseases has become a hot topic of research.

In addition, stimulation of interferon gene stimulator signaling pathway activation has also been shown to promote antiviral responses. The loss of a functional response at the cellular or organismal level demonstrates that the viral load cannot be controlled without the stimulator of interferon genes. Activation of the interferon gene stimulator signaling pathway triggers an immune response leading to the production of antivascular and pro-inflammatory cytokines against the virus and mobilizes the innate and adaptive immune systems. Thus, small molecule compounds with an agonistic effect on the interferon gene stimulator signaling pathway have potential for the treatment of chronic viral infections and can be used, for example, in the treatment of hepatitis B virus (HBV).

Several analogues of cyclic dinucleotide with agonistic action on the signaling pathway of the interferon gene stimulator have been described (WO 2014/093936, WO 2014/189805, WO 2014/189806, WO 2016/120305, WO 2016/145102, WO 2017/027645/027. WO 2017/0/093933 WO 2017/123657 WO 2017/123669 WO 2017/161349 WO 2017/186711 WO 2018/009466 WO 2018/009648 060323, WO 82018/0653, WO 2018/0653, WO 2018/065), but currently no interferon gene stimulator agonist has been approved for sale.

- 1 041924

The essence of the present invention

The technical problem to be solved in the present invention is to provide a new analogue of a cyclic dinucleotide, its pharmaceutical composition and its use. The cyclic dinucleotide analog of the present disclosure has a good STING modulating effect and can effectively treat, alleviate and/or prevent various diseases caused by immunosuppression such as tumors, infectious diseases, neurodegenerative diseases, psychiatric disorders or autoimmune diseases.

Provided herein is a type (I) cyclic dinucleotide analogue, isomer, prodrug, isotopically stable derivative or pharmaceutically acceptable salt thereof;

X ³¹

(I) where each of Z1, Z2 independently represents O, S, SO2, CH2, CF2 or Se;

B1 represents ^w ₂ wt 'w ₃ o II z- P ΆΑ ^V 3 q , s 37 11 o=f Ύι A ^V! - JI JN N V< PI V -J Pxq llllg ' B-17 B-18 I B ₂ is Rd AA Rd A N^A-., s^A _x/ 0=( D A ⁰⁼ ^ Yes A n R _c ^ V^R _C ' B-1 ' B-2 ' B-3 a '''"Λ “3^ L % I a A ν _2χ ~N^^WNW Al jvuvw I । ^B ' ¹⁹ or ^B-2 ° ;

- 2 041924

Each L and L1 is independently a bridging bond or CR1R ² ;

each L' and L1' independently represent a connecting bond or CR11R ²¹ ;

L ₂ is O, S or CR3R ⁴ ;

L ₂ ' represents O, S or CR ³¹ R ⁴¹ ;

X ¹ is O, S or CR5R ⁶ ;

X ¹¹ is O, S or CR51R ⁶¹ ;

X ² is O, S or CR7R ⁸ ; And

X ²¹ is O, S or CR71R ⁸¹ ;

each X ³ and X ³¹ are independently OH, SH or BH ₃ ^- ;

each R and R' are independently hydrogen, C _2-6 alkenyl, C _2-6 alkynyl, or C _1-6 alkyl; C _2-6 alkenyl, C _2-6 alkynyl or C _1-6 alkyl is unsubstituted or selectively substituted at any position with 1-3 substituents selected from halo, hydroxyl, amino, azido and cyano;

each R1 and ^R2 are independently hydrogen, halogen, cyano, hydroxyl, thiol, amino, azido, C2-6 alkenyl, C2-6 alkynyl, C1-6 alkyl, halo-C1-6 alkyl, halo-C1-6 alkoxy , halo-C1-6 alkylthio, C1-6 alkylamino, OC(O)R ^a or OR ^a ; C2-6 alkenyl, C _2-6 alkynyl or C _1-6 alkyl is unsubstituted or selectively substituted at any position 1-3 with halogen, hydroxyl, amino, azido and cyano;

each R ¹¹ and R ²¹ are independently hydrogen, halogen, cyano, hydroxyl, thiol, amino, azido, C2-6 alkenyl, C2-6 alkynyl, C1-6 alkyl, halo-C1-6 alkyl, halo-C1-6 alkoxy, halo-C1-6 alkylthio, C1-6 alkylamino, OC(O)R ^a or OR ^a ; C2-6 alkenyl, C _2-6 alkynyl or C _1-6 alkyl is unsubstituted or selectively substituted at any position 1-3 with halogen, hydroxyl, amino, azido and cyano;

alternatively, R ¹ and R ² together form a carbonyl;

alternatively, R ¹¹ and R ²¹ together form a carbonyl;

alternatively, R is co6-CH ₂ -, R ¹ is -O-, R and R ¹ are linked together to form heterocycloalkyl;

alternatively, R' is co6-CH ₂ -, R ¹¹ is -O-, R' and R ¹¹ are linked together to form heterocycloalkyl;

each R ³ and R ⁴ independently represent hydrogen, deuterium, halogen or C _1-6 alkyl;

each R ³¹ and R ⁴¹ independently represent hydrogen, deuterium, halogen or C _1-6 alkyl;

each R ⁵ and R ⁶ independently represent hydrogen, deuterium, halogen or C _1-6 alkyl;

each R ⁵¹ and R ⁶¹ independently represent hydrogen, deuterium, halogen or C _1-6 alkyl;

each R ⁷ and R ⁸ independently represent hydrogen, deuterium, halogen or C _1-6 alkyl;

each R ⁷¹ and R ⁸¹ independently represent hydrogen, deuterium, halogen or C _1-6 alkyl;

- 3 041924 each Y and Y1 independently represent CRE or N;

U is CHRE' or NRD';

U ₁ represents CH or N;

each V, V1, V ₂ and V ₃ are independently CRE or N;

W is O or S;

each W1, W ₂ , W ₃ and W4 are independently N or CRF';

each of RA, RB, R _C , RE, RE', RE, R _F , R _F ' and RG is independently H, halogen, -CN, -NO2, -N ₃ , R ^c , -SR ^c , -OR ^c , -OC(O)R ^c , -OC(O)OR ^c , -OC(O)NR ^b R ^c , -C(O)OR ^c , -C(O)R ^c , -C(O)NR ^b R ^c , -NR ^b R ^c , -NR ^b C(O)R ^c , -N(R ^b )C(O)OR ^c , -N(R ^a )C(O)NR ^b R ^c , -NR ^b S(OhR ^c , -NR ^b C(=NH)R ^c , -NR ^b C(=NR ^c )NH2, -S(O)1-2R ^c , -S(O)2NR ^b R ^c or -NR ^a S(O)2NR ^b R ^c ;

each of R _D and R _D ' is independently H or R ^c ;

each of R ^a and R ^b is independently H, C _2-6 alkenyl, C _2-6 alkynyl, C _1-10 alkyl, halo-C _1-6 alkyl, aryl, heteroaryl, cycloalkyl, heterocycloalkyl, aryl-C _{1 -6} alkyl, heteroaryl-C _1-6 alkyl, cycloalkyl-C _1-6 alkyl or heterocycloαalkyl-C _1-6 alkyl;

each R ^c is independently H, substituted or unsubstituted C _1-10 alkyl, substituted or unsubstituted C _2-8 alkenyl, substituted or unsubstituted C _2-8 alkynyl, substituted or unsubstituted C _3-10 cycloalkyl, substituted or unsubstituted C _{6- 10} aryl, substituted or unsubstituted 3-10 membered heterocycloalkyl, substituted or unsubstituted 5-10 membered heteroaryl, substituted or unsubstituted C _6-10 aryl-C _1-6 alkyl, substituted or unsubstituted C _3-10 cycloalkyl C1-6 alkyl, substituted or unsubstituted 3-10 membered heterocycloalkyl-C _1-6 alkyl, substituted or unsubstituted 5-10 membered heteroaryl-C _1-6 alkyl; C _1-10 alkyl, C _2-8 alkenyl, C _2-8 alkynyl, C _3-10 cycloalkyl, C _6-10 aryl, 3-10 membered heterocycloalkyl, 5-10 membered heteroaryl, C _6-10 aryl- C _1-6 alkyl, C _3-10 cycloalkyl-C _1-6 alkyl, 3-10 membered heterocycloalkyl-C _1-6 alkyl, or 5-10 membered heteroaryl-C _1-6 alkyl is unsubstituted or selectively substituted in any position of one or more Rd;

each Rd is independently halo, halo-C _1-6 alkyl, halo-C _1-6 alkoxy, C _1-6 alkyl, -CN, -N ₃ , -SR ^e , -OR ^e , -C(O)R ^e , -NR ^e R ^e , substituted or unsubstituted C _6-10 aryl, substituted or unsubstituted 5-10 membered heteroaryl, substituted or unsubstituted C _3-10 cycloalkyl, or substituted or unsubstituted 3-10 membered heterocycloalkyl; C _6-10 aryl, 5-10 membered heteroaryl, C _3-10 cycloalkyl, or 3-10 membered heterocycloalkyl is unsubstituted or selectively substituted at any position with one or more substituents selected from halo, hydroxyl, cyano, amino, C1. 4 alkyl, halo-C _1-4 alkyl, C _1-4 alkoxy, C _1-4 alkylamino and halo-C _1-4 alkoxy;

each of R ^e and R ^e is independently C _2-6 alkenyl, C _2-6 alkynyl, C _1-10 alkyl, haloC _1-6 alkyl, aryl, heteroaryl, cycloalkyl, heterocycloalkyl, aryl-C _1-6 alkyl, heteroaryl-C _1-6 alkyl, cycloalkyl-C _1-6 alkyl or heterocycloalkyl-C _1-6 alkyl.

In some embodiments, a type (I) cyclic dinucleotide analog, isomer, prodrug, isotopically stable derivative, or pharmaceutically acceptable salt has the general structural formula

X ³¹

B1 represents

B ₂ is where each Z1, Z ₂ are independently O, S, SO ₂ , CH ₂ , CF2 or Se;

- 4 041924

each L and L1 is independently a bridging bond or CR1R ² ;

each L' and L1' independently represent a connecting bond or CR11R ²¹ ;

L ₂ is O, S or CR3R ⁴ ;

L ₂ ' represents O, S or CR ³¹ R ⁴¹ ;

X ¹ is O, S or CR5R ⁶ ;

X ¹¹ is O, S or CR51R ⁶¹ ;

X ² is O, S or CR7R ⁸ ; And

X ²¹ is O, S or CR71R ⁸¹ ;

each X ³ and X ³¹ are independently OH, SH or BH ₃ ^- ;

each R and R' are independently hydrogen, C _2-6 alkenyl, C _2-6 alkynyl, or C _1-6 alkyl; C _2-6 alkenyl, C _2-6 alkynyl or C _1-6 alkyl is unsubstituted or selectively substituted at any position with 1-3 substituents selected from halogen, hydroxyl, amino, azido and cyano;

each R ¹ and R ² are independently hydrogen, halogen, cyano, hydroxyl, thiol, amino, azido, C2-6 alkenyl, C2-6 alkynyl, C1-6 alkyl, halo-C1-6 alkyl, halo-C1-6 alkoxy, halo-C1-6 alkylthio, C1-6 alkylamino, OC(O)R ^a or OR ^a ; C2-6 alkenyl, C _2-6 alkynyl or C _1-6 alkyl is unsubstituted or selectively substituted at any position 1-3 with halogen, hydroxyl, amino, azido and cyano;

each R ¹¹ and R ²¹ are independently hydrogen, halogen, cyano, hydroxyl, thiol, amino, azido, C2-6 alkenyl, C2-6 alkynyl, C1-6 alkyl, halo-C1-6 alkyl, halo-C1-6 alkoxy, halo-C1-6 alkylthio, C1-6 alkylamino, OC(O)R ^a or OR ^a ; C2-6 alkenyl, C _2-6 alkynyl or C _1-6 alkyl is unsubstituted or selectively substituted at any position with 1-3 substituents selected from halo, hydroxyl, amino, azido and cyano;

alternatively, R ¹ and R ² together form a carbonyl;

alternatively, R ¹¹ and R ²¹ together form a carbonyl;

alternatively, R is co6-CH ₂ -, R ¹ is-O-, R and R ¹ are linked together to form heterocycloalkyl;

alternatively, R' is co6-CH ₂ -, R ¹¹ is-O-, R' and R ¹¹ are linked together to form heterocycloalkyl;

each R ³ and R ⁴ independently represent hydrogen, deuterium, halogen or C _1-6 alkyl;

each R ³¹ and R ⁴¹ independently represent hydrogen, deuterium, halogen or C _1-6 alkyl;

each R ⁵ and R ⁶ independently represent hydrogen, deuterium, halogen or C _1-6 alkyl;

each R ⁵¹ and R ⁶¹ independently represent hydrogen, deuterium, halogen or C _1-6 alkyl;

each R ⁷ and R ⁸ independently represent hydrogen, deuterium, halogen or C _1-6 alkyl;

each R ⁷¹ and R ⁸¹ independently represent hydrogen, deuterium, halogen or C _1-6 alkyl;

each Y and Y1 are independently CRE or N;

- 5 041924

U is CHRE' or NRD';

U ₁ represents CH or N;

each V and V1 are independently CRE or N;

W is O or S;

each of RA, RB, RC, RE, RE', RE, R _F , R _F ' and Rg is independently H, halogen, -CN, -NO ₂ , -N ₃ , R ^c , -SR ^c , -OR ^c , -OC(O)R ^c , -OC(O)OR ^c , -OC(O)NR ^b R ^c , -C(O)OR ^c , -C(O)R ^c , -C(O)NR ^b R ^c , -NR ^b R ^c , -NR ^b C(O)R ^c , -N(R ^b )C(O)OR ^c , -N(R ^a )C(O)NR ^b R ^c , -NR ^b S(O)2R ^c , -NR ^b C(=NH)R ^c , -NR ^b C(=NR ^c )NH2, -S(O)1-2R ^c , -S(O)2NR ^b R ^c or -NR ^a S(O)2NR ^b R ^c ;

each of RD and RD' is independently H or R ^c ;

each of R ^a and R ^b is independently H, C _2-6 alkenyl, C _2-6 alkynyl, C _1-10 alkyl, halo-C _1-6 alkyl, aryl, heteroaryl, cycloalkyl, heterocycloalkyl, aryl-C _{1 -6} alkyl, heteroaryl-C _1-6 alkyl, cycloalkyl-C _1-6 alkyl or heterocycloalkyl-C _1-6 alkyl;

each R ^c is independently H, substituted or unsubstituted C _1-10 alkyl, substituted or unsubstituted C _2-8 alkenyl, substituted or unsubstituted C _2-8 alkynyl, substituted or unsubstituted C _3-10 cycloalkyl, substituted or unsubstituted C _{6- 10} aryl, substituted or unsubstituted 3-10 membered heterocycloalkyl, substituted or unsubstituted 5-10 membered heteroaryl, substituted or unsubstituted C _{6 -10} aryl-C _1-6 alkyl, substituted or unsubstituted C _3-10 cycloalkylC _1-6 alkyl , substituted or unsubstituted 3-10 membered heterocycloalkyl-C _1-6 alkyl, substituted or unsubstituted 5-10 membered heteroaryl-C _1-6 alkyl; C _1-10 alkyl, C _2-8 alkenyl, C _2-8 alkynyl, C _3-10 cycloalkyl, C _6-10 aryl, 3-10 membered heterocycloalkyl, 5-10 membered heteroaryl, C _6-10 aryl- C _1-6 alkyl, C _3-10 cycloαalkyl-C _1-6 alkyl, 3-10 membered heterocycloalkyl-C _1-6 alkyl or 5-10 membered heteroaryl-C _1-6 alkyl is unsubstituted or selectively substituted in any position of one or more R ^d ;

each R ^d is independently halo, halo-C1-6 alkyl, halo-C1-6 alkoxy, C1-6 alkyl, -CN, -N3, -SR ^e , -OR ^e , -C(O)R ^e , - NR ^e R ^e , substituted or unsubstituted C6-10 aryl, substituted or unsubstituted 5-10 membered heteroaryl, substituted or unsubstituted C _3-10 cycloalkyl, or substituted or unsubstituted 3-10 membered heterocycloalkyl; C _6-10 aryl, 5-10 membered heteroaryl, C _3-10 cycloalkyl, or 3-10 membered heterocycloalkyl is unsubstituted or selectively substituted at any position with one or more substituents selected from halogen, hydroxyl, cyano, amino, C _{1 -4} alkyl, halo-C _1-4 alkyl, C _1-4 alkoxy, C _1-4 alkylamino and halo-C _1-4 alkoxy;

each of R ^e and R ^e is independently C _2-6 alkenyl, C _2-6 alkynyl, C _1-10 alkyl, haloC _1-6 alkyl, aryl, heteroaryl, cycloalkyl, heterocycloalkyl, aryl-C _1-6 alkyl, heteroaryl-C _1-6 alkyl, cycloαalkyl-C _1-6 alkyl or heterocycloalkyl-C _1-6 alkyl.

In some embodiments, a type (I) cyclic dinucleotide analog, isomer, prodrug, stable isotope derivative, or pharmaceutically acceptable salt thereof has the general structural formula

B1 represents

B ₂ represents

where each Z1, Z ₂ independently represent O, S, SO ₂ , CH ₂ , CF2 or Se;

- 6 041924

each L and L ₁ independently represent a connecting bond or CR1R ² ;

each L' and L1' independently represent a connecting bond or CR11R ²¹ ;

L ₂ is O, S or CR3R ⁴ ;

L ₂ ' represents O, S or CR ³¹ R ⁴¹ ;

X ¹ is O, S or CR5R ⁶ ;

X ¹¹ is O, S or CR51R ⁶¹ ;

X ² is O, S or CR7R ⁸ ; And

X ²¹ is O, S or CR71R ⁸¹ ;

each X ³ and X ³¹ are independently OH, SH or BH ₃ ^- ;

each R and R' are independently hydrogen, C _2-6 alkenyl, C _2-6 alkynyl, or C _1-6 alkyl; C _2-6 alkenyl, C _2-6 alkynyl or C _1-6 alkyl is unsubstituted or selectively substituted at any position with 1-3 substituents selected from halogen, hydroxyl, amino, azido and cyano;

each R ¹ and R ² are independently hydrogen, halogen, cyano, hydroxyl, thiol, amino, azido, C2-6 alkenyl, C2-6 alkynyl, C1-6 alkyl, halo-C1-6 alkyl, halo-C1-6 alkoxy, halo-C1-6 alkylthio, C1-6 alkylamino, OC(O)R ^a or OR ^a ; C2-6 alkenyl, C _2-6 alkynyl or C _1-6 alkyl is unsubstituted or selectively substituted at any position 1-3 with halogen, hydroxyl, amino, azido and cyano;

each R ¹¹ and R ²¹ are independently hydrogen, halogen, cyano, hydroxyl, thiol, amino, azido, C2-6 alkenyl, C2-6 alkynyl, C1-6 alkyl, halo-C1-6 alkyl, halo-C1-6 alkoxy, halo-C1-6 alkylthio, C1-6 alkylamino, OC(O)R ^a or OR ^a ; C2-6 alkenyl, C _2-6 alkynyl or C _1-6 alkyl is unsubstituted or selectively substituted at any position 1-3 with halogen, hydroxyl, amino, azido and cyano;

alternatively, R ¹ and R ² together form a carbonyl;

alternatively, R ¹¹ and R ²¹ together form a carbonyl;

alternatively, R is co6-CH ₂ -, R ¹ is-O-, R and R ¹ are linked together to form heterocycloalkyl;

alternatively, R' is co6-CH ₂ -, R ¹¹ is-O-, R' and R ¹¹ are linked together to form heterocycloalkyl;

each R ³ and R ⁴ independently represent hydrogen, deuterium, halogen or C _1-6 alkyl;

each R ³¹ and R ⁴¹ independently represent hydrogen, deuterium, halogen or C _1-6 alkyl;

each R ⁵ and R ⁶ independently represent hydrogen, deuterium, halogen or C _1-6 alkyl;

each R ⁵¹ and R ⁶¹ independently represent hydrogen, deuterium, halogen or C _1-6 alkyl;

each R ⁷ and R ⁸ independently represent hydrogen, deuterium, halogen or C _1-6 alkyl;

each R ⁷¹ and R ⁸¹ independently represent hydrogen, deuterium, halogen or C _1-6 alkyl;

each Y and Y1 are independently CRE or N;

- 7 041924

U is CHRE' or NR _D ';

U ₁ represents CH or N;

each V and V ₁ are independently CRE or N;

W is O or S;

Each of RA, RB, R _C , RE, RE', RE, R _F , R _F ' and RG is independently H, halogen, -CN, -NO2, -N3, R ^c , -SR ^c , -OR ^c , -OC(O)R ^c , -OC(O)OR ^c , -OC(O)NR ^b R ^c , -C(O)OR ^c , -C(O)R ^c , -C(O)NR ^b R ^c , -NR ^b R ^c , -NR ^b C(O)R ^c , -N(R ^b )C(O)OR ^c , -N(R ^a )C(O)NR ^b R ^c , -NR ^b S(O)2R ^c , -NR ^b C(=NH)R ^c , -NR ^b C(=NR ^c )NH2, -S(O)i—2R ^c , -S(O)2NR ^b R ^c or - NR ^a S(O)2NR ^b R ^c ;

each of R _d and R _d ' is independently H or R ^c ;

each of R ^a and R ^b independently represents H, C ₂ - ₆ alkenyl, C _2-6 alkynyl, C ₁ . ₁ o alkyl, halo-C _1- b alkyl, aryl, heteroaryl, cycloalkyl, heterocycloalkyl, aryl-C _1-6 alkyl, heteroaryl-C _1-6 alkyl, cycloalkyl-C _1-6 alkyl or heterocycloalkyl-C _1-6 alkyl;

each R ^c is independently H, substituted or unsubstituted C _1-10 alkyl, substituted or unsubstituted C ₂ _ ₈ alkenyl, substituted or unsubstituted C 2-8 alkynyl, substituted or unsubstituted C ₃ _ ₁₀ cycloalkyl, substituted or unsubstituted C ₆ _ ₁₀ aryl, substituted or unsubstituted 3-10 membered heterocycloalkyl, substituted or unsubstituted 5-10 membered heteroaryl, substituted or unsubstituted C ₆ _ ₁₀ aryl-C ₁ . ₆ alkyl, substituted or unsubstituted C ₃ _ ₁₀ cycloalkylC ₁ _ ₆ alkyl, substituted or unsubstituted 3-10-membered heterocycloalkyl-C ₁ . ₆ alkyl, substituted or unsubstituted 5-10 membered heteroaryl-C ₁ . ₆ alkyl; C _1-10 alkyl, C ₂ _ ₈ alkenyl, C ₂ _ ₈ alkynyl, C ₃ _ ₁₀ cycloalkyl, C ₆ _ ₁₀ aryl, 3-10-membered heterocycloalkyl, 5-10-membered heteroaryl, C ₆ _ ₁₀ aryl- _C1 . ₆ alkyl, C ₃ _ ₁₀ cycloalkyl-C ₁ . ₆ alkyl, 3-10 membered heterocycloalkyl-C ₁ . ₆ alkyl or 5-10 membered heteroaryl-C ₁ .6 alkyl is unsubstituted or selectively substituted at any position with one or more R ^d ;

each R ^d is independently halo, halo-C1.6 alkyl, halo-C1.6 alkoxy, C1-6 alkyl, -CN, -N3, -SR ^e , -OR ^e , -C(O)R ^e , - NR ^e R ^e , substituted or unsubstituted C6-10 aryl, substituted or unsubstituted 5-10 membered heteroaryl, substituted or unsubstituted C _3-10 cycloalkyl, or substituted or unsubstituted 3-10 membered heterocycloalkyl; C ₆ _ ₁₀ aryl, 5-10 membered heteroaryl, C ₃ _ ₁₀ cycloalkyl or 3-10 membered heterocycloalkyl is unsubstituted or selectively substituted at any position with one or more substituents selected from halogen, hydroxyl, cyano, amino, Cm alkyl , halogen-C ₁ . ₄ alkyl, C _1-4 alkoxy, Cm alkylamino and halo-C ₁ . ₄ alkoxy;

each of R ^e and R ^e independently represents C ₂ _ ₆ alkenyl, C ₂ _ ₆ alkynyl, C _1-10 alkyl, haloC _1-6 alkyl, aryl, heteroaryl, cycloalkyl, heterocycloalkyl, aryl-C ₁ . ₆ alkyl, heteroaryl-C ₁ . ₆ alkyl, cycloalkyl-C ₁ . ₆ alkyl or heterocycloalkyl-C ₁ . ₆ alkyl.

In some embodiments, each R ^c is independently H, substituted or unsubstituted C _1-10 alkyl, substituted or unsubstituted C _3-10 cycloalkyl, substituted or unsubstituted C _6-10 aryl, substituted or unsubstituted 3-10 membered heterocycloalkyl, substituted or unsubstituted 5-10 membered heteroaryl, substituted or unsubstituted C _6-10 aryl-C ₁ . ₆ alkyl, substituted or unsubstituted C _3-10 cycloalkyl-C ₁ . ₆ alkyl, substituted or unsubstituted 310-membered heterocycloalkyl-C ₁ . ₆ alkyl, substituted or unsubstituted 5-10 membered heteroaryl-C ₁ . ₆ alkyl; C _1-10 alkyl, C ₃ _ ₁₀ cycloalkyl, C ₆ _ ₁₀ aryl, 3-10 membered heterocycloalkyl, 5-10 membered heteroaryl, C ₆ _ ₁₀ aryl-C ₁ . ₆ alkyl, C3-10 cycloalkyl-C ₁ . ₆ alkyl, 3-10 membered heterocycloalkyl-C ₁ . ₆ alkyl or 5-10 membered heteroaryl-C ₁ . ₆ alkyl is unsubstituted or selectively substituted at any position with one or more R ^d ;

R ^d is defined above.

in the definition of R ^c substituted or unsubstituted C _1-10 alkyl is preferably substituted or unsubstituted C _1-6 alkyl such as methyl, ethyl, propyl, isopropyl, tert-butyl, nbutyl; where methyl, ethyl, propyl, isopropyl, tert-butyl or n-butyl is substituted or unsubstituted;

in the definition of R ^c substituted or unsubstituted C _3-10 cycloalkyl is preferably cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, 2,3-dihydro-1H-inden-1-yl, 2,3-dihydro 1H-inden-2-yl; where cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, 2,3-dihydro-1H-inden-1-yl or 2,3-dihydro-1H-inden-2-yl is substituted or unsubstituted;

in the definition of R ^c, substituted or unsubstituted C ₆ _ ₁₀ aryl is preferably substituted or unsubstituted phenyl;

in the definition of R ^c substituted or unsubstituted 3-10-membered heterocycloalkyl is preferably tetrahydrofuran-3-yl, tetrahydro-2H-pyran-3-yl, tetrahydro-2H-pyran-4-yl, pyrrolidin-3-yl, piperidin- 3-yl, piperidin-4-yl; where tetrahydrofuran-3-yl, tetrahydro-2H-pyran-3yl, tetrahydro-2H-pyran-4-yl, pyrrolidin-3-yl, piperidin-3-yl, piperidin-4-yl is substituted or unsubstituted;

in the definition of R ^c, a substituted or unsubstituted 5-10 membered heteroaryl is preferably a substituted or unsubstituted 5-6 membered heteroaryl.

in some embodiments, each R ^c is independently H, C _1-4 alkyl, or halo-C ₁ . ₄ alkyl.

- 8 041924

In some embodiments, each R ^b is independently H or

C _1-4 alkyl.

In some embodiments, Z1 is O.

In some embodiments, Z2 is O.

In some embodiments of the invention, in definition B-4 to B-16, Y is N.

In some embodiments of the invention in the definition of B-4 to B-16, Y is CRE, and RE is preferably H, F, Cl, -CF ₃ , -CH3, -CN, -NH2.

In some embodiments of the invention, in definition B-4 to B-16, Y1 is N.

In some embodiments of the invention in the definition of B-4 to B-16, Y1 is CR _E and R _E is preferably H, F, Cl, -CF ₃ , -CH ₃ , -CN, -NH ₂ .

In some embodiments of the invention in the definition of B-6 to B-9, B-13, B-16 to B-17, U is NRD', and RD' is H or -CH ₃ .

In some embodiments of the invention in the definition of B-1 to B-5, B-10 to B-13, B-15, V is N or CH.

In some embodiments of the invention in the definition of B-1 to B-12, B-17 V1 is N or CH.

In some embodiments, in definition B-1 to B-3, RA is H, halogen, -OR ^c , -NR ^b R ^c ; where R ^b and R ^c are defined above.

In some embodiments of the invention in the definition of B-4, B-5, B-10 to B-12, B-15 RB is H, halogen, -OR ^c , -NR ^b R ^c ; where R ^b and R ^c are defined above.

In some embodiments of the invention in the definition of B-1, B-2, B-4, B-6 to B-8, B-10 to B-13, B-17 RC is H, F, Cl, - OR ^c , -SR ^c , -NR ^b R ^c or R ^c ; where R ^c is defined above.

In some embodiments of the invention in the definition of B-15, B-16 RG is H.

In some embodiments, in the definition of B-15, B-16 RF is H.

In some embodiments of the invention in the definition of B-18 to B-20, V ₃ is N or CH.

In some embodiments of the invention in the definition of B-18 to B-20, V ₂ is CH.

In some embodiments of the invention in the definition of B-18 to B-20, W1 is N or CH.

In some embodiments of the invention in the definition of B-18 to B-20, W ₂ is CRF'; where RF' is H or -NH2.

In some embodiments of the invention in the definition of B-18 to B-20, W ₃ is CH.

In some embodiments of the invention in the definition of B-18, W4 is CH.

In some embodiments of the invention, B1 is any of the following structures:

In some embodiments of the invention, B1 is:

structures:

In some embodiments of the invention, B1 is:

In some embodiments, B1 is any of the following

- 9 041924

In some embodiments, B1 is any of the following

structures:

is any of the following structures:

In some structures:

In some embodiments of the invention B1

implementation of the invention B ₁ embodiments of the following is any is from

In some embodiments, B1 is any of the following

structures:

In some embodiments of the invention, B2 is any structure:

of the following

- 10 041924

In some embodiments of the invention B2 is: _n nh ₂ nh ₂ o oaX ⁿ eL ⁿ w ^C|

PM , OR .

nh ₂ nh ₂ o/yS °=< ⁵ Λ / n^n^ci

In some embodiments of the invention, B2 is: ^{4 or} nh ₂ about

1^20 <'χ\ ^Η

N

In some embodiments, B2 is: ' ^or '

In some embodiments of the invention, when L is a connecting bond, then L1 is CR1R ² ; R ¹ and R ² are defined as described previously.

In some embodiments, when L is CR1R ² , then L1 is a bridging bond; R ¹ and R ² are defined as described previously.

In some embodiments of the invention, when L' is a connecting bond, then L1' is CR11R ²¹ ; R ¹¹ and R ²¹ are defined as previously described.

In some embodiments of the invention, when L' represents CR11R ²¹ then L1' represents a connecting bond; R ¹¹ and R ²¹ are defined as previously described.

- 11 041924

In some embodiments of the invention, L2 is CH2. In some embodiments of the invention L ₂ ' represents CH2. In some embodiments, X ¹ is O. In some embodiments, X ¹¹ is O. In some embodiments, X ² is O. In some embodiments, X ²¹ is O.

In some embodiments, R ^a is C _1-4 alkyl or haloC _1-4 alkyl.

In some embodiments, R ^a is C _1-4 alkyl.

In some embodiments of the invention, each R ¹ and R ² independently represent hydrogen, halogen, hydroxyl, or OR ^a ; R ^a is defined as described previously.

In some embodiments of the invention, each R ¹¹ and R ²¹ independently represent hydrogen, halogen, hydroxyl, or OR ^a ; R ^a is defined as described previously.

In some embodiments of the invention R ¹ represents hydrogen.

In some embodiments of the invention R ² represents hydrogen, fluorine, hydroxyl or methoxy.

In some embodiments of the invention R ¹¹ represents hydrogen.

In some embodiments of the invention R ²¹ represents hydrogen, fluorine, hydroxyl or methoxy.

In some embodiments, R is hydrogen. In some embodiments of the invention R' represents hydrogen.

In some embodiments, certain groups in a compound represented by Formula I, its isomer, prodrug, stable isotope derivative, or pharmaceutically acceptable salt are as defined below, and non-described groups are as defined in any of the above embodiments : Zi is O;

nh ₂ nh ₂ O ^{o \u003d} < ⁵ L ⁿ

V R - ex P R or;

Z2 is O; Bi represents:

nh ₂ o

Ι</ ^ν ύ4χ % 4^^ ^Ν ^ ^ΝΗ 2 .

B ₂ is: ' ^or '' each L and L1 is independently a bridging bond or CR1R ² ;

each L' and L1' independently represent a connecting bond or CR11R ²¹ ; And

L, L ₁ , L' and L ₁ ' are defined in the following combinations:

1) L is a bridging bond, L1 is CR1R ² , L' is CR ¹¹ R ²¹ , L ₁ ' is a bridging bond, or

2) L is CR1R ² , L1 is a bond, L' is a bond, L1' is CR ¹¹ R ²¹ ;

L2 is CH2;

L2' is CH2;

X ¹ is O;

X ¹¹ is O;

X ² is O;

X ²¹ is O;

each X ³ and X ³¹ are independently OH or SH;

each R and R' are independently hydrogen;

each R ¹ and R ² independently represent hydrogen, halogen, hydroxyl or OR ^a ;

each R ¹¹ and R ²¹ independently represent hydrogen, halogen, hydroxyl or OR ^a ;

each R ^a is independently C _1-4 alkyl or halo-C ₁ . ₄ alkyl.

In some embodiments, certain groups in a compound represented by Formula I, its isomer, prodrug, stable isotope derivative, or pharmaceutically acceptable salt are as defined below, and non-described groups are as defined in any of the above embodiments :

Z1 is O;

- 12 041924

Z2 is O; B1 is:

nh ₂ about hr

B2 is: ' ^or '' each L and L1 independently represent a connecting bond or CR1R ² ;

each L' and L1' independently represent a connecting bond or CR11R ²¹ ; And

L, L1, L' and L ₁ ' are defined in the following combinations:

1) L is a bridging bond, L1 is CR1R ² , L' is CR11R ²¹ , L1' is a bridging bond, or

2) L is CR1R ² , L1 is a bond, L' is a bond, L1' is CR11R ²¹ ;

L2 is CH2;

L ₂ ' is CH2;

X ¹ is O;

X ¹¹ is O;

X ² is O;

X ²¹ is O;

each X ³ and X ³¹ are independently OH or SH;

each R and R' are independently hydrogen;

each R ¹ and R ² independently represent hydrogen, halogen, hydroxyl or OR ^a ;

each R ¹¹ and R ²¹ independently represent hydrogen, halogen, hydroxyl or OR ^a ;

each R ^a is independently C _1-4 alkyl or halo-C _1-4 alkyl.

In some embodiments, certain groups in a compound represented by Formula I, its isomer, prodrug, stable isotope derivative, or pharmaceutically acceptable salt are as defined below, and non-described groups are as defined in any of the above embodiments. inventions:

Z ₁ is O;

Z2 is O;

B1 is:

B2 is:

each L and L1 is independently a bridging bond or CR1R ² ;

each L' and L1' independently represent a connecting bond or CR11R ²¹ ; And

L, L ₁ , L' and L ₁ ' are defined in the following combinations:

1) L is a bridging bond, L1 is CR1R ² , L' is CR11R ²¹ , L1' is a bridging bond, or

2) L is CR1R ² , L ¹ is a bridging bond, L' is a bridging bond, L ¹ ' is CR11R ²¹ ;

L2 is CH2;

L2' is CH2;

X ¹ is O;

X ¹¹ is O;

X ² is O;

X ²¹ is O;

each X ³ and X ³¹ independently represent SH;

each R and R' are independently hydrogen;

R ¹ is hydrogen;

- 13 041924

R ² is hydrogen, halogen, hydroxyl or OR ^a ;

R ¹¹ is hydrogen;

R ²¹ is hydrogen, halogen, hydroxyl or OR ^a ;

each R ^a is independently C _1-4 alkyl.

In some embodiments, certain groups in a compound represented by Formula I, its isomer, prodrug, stable isotope derivative, or pharmaceutically acceptable salt are as defined below, and non-described groups are as defined in any of the above embodiments :

Z ₁ is O;

Z ₂ is O;

B represents:

B ₂ is: ' ^or '' each L and L ₁ independently represent a connecting bond or CR1R ² ;

each L' and L1' independently represent a connecting bond or CR11R ²¹ ; And

L, L _b L' and L1' are defined in the following combinations:

1) L is a bridging bond, L ₁ is CR1R ² , L' is CR ¹¹ R ²¹ , L ₁ ' is a bridging bond, or

2) L is CR1R ² , L1 is a bond, L' is a bond, L1' is CR11R ²¹ ;

L ₂ is CH ₂ ; L ₂ ' is CH ₂ ;

X ¹ is O;

X ¹¹ is O; X ² is O;

X ²¹ is O;

X ³ and X ³¹ are SH;

R and R' are hydrogen;

R ¹ is hydrogen;

R ² is hydrogen, halogen, hydroxyl or OR ^a ;

R ¹¹ is hydrogen;

R ²¹ is hydrogen, halogen, hydroxyl or OR ^a ;

each R ^a is independently C _1-4 alkyl.

In some embodiments, a compound of formula I, an isomer, prodrug, stable isotope derivative, or pharmaceutically acceptable salt thereof, is a compound of formula II, III, IV, or V, an isomer, prodrug, stable isotope derivative, or pharmaceutically acceptable salt thereof:

- 14 041924

(iv) (V) where B ₁ , B ₂ , Z ₁ , Z2, R, R', R ¹ , R ² , R ¹¹ , R ²¹ , X ³ , X ³¹ , L2 and L2' are defined as previously described .

Combinations including any of the embodiments of B1, B ₂ , Z1, Z ₂ , R, R', R ¹ , R ² , R ¹¹ , R ²¹ , X ³ , X ³¹ , L2 and L2' as described in Formula I are included within the scope of formula II, III, IV or V in the present invention.

All embodiments of formula II, III, IV or V described below are included within the scope of formula II, III, IV or V in the present invention.

In some preferred embodiments of Formula II, III, IV, or VR ¹ is H; R ² is -OH, F, -N ₃ , -SCF ₃ or -OCH ₃ .

In some preferred embodiments of Formula II, III, IV, or VR ¹¹ is H; R ²¹ is -OH, F, -N ₃ , -SCF ₃ or -OCH ₃ .

In some preferred embodiments of formula II, III, IV, or VR is -CH2-, R ¹ is -O-, R and R ¹ are linked together to form a heterocycloalkyl.

In some preferred embodiments of formula II, III, IV, or VR' is -CH2-, R ¹¹ is -O-, R' and R ¹¹ are linked together to form a heterocycloalkyl.

In some preferred embodiments of Formula II, III, IV, or VZ ₁ is O;

Z ₂ is O.

in some preferred embodiments of Formula II, III, IV, or VL ₂ is -CH ₂ -;

L2' is -CH2-.

In some embodiments, a compound of formula I, an isomer, prodrug, stable isotope derivative, or pharmaceutically acceptable salt thereof, is a compound of formula VI or VII, an isomer, prodrug, stable isotope derivative, or pharmaceutically acceptable salt thereof:

(VI) (VII) where B ₁ , B ₂ , R ² and R ²¹ are defined as described previously.

Combinations comprising any of the embodiments of B ₁ , B ₂ , R ² and R ²¹ as described in formula I are included within the scope of formula VI or VII in the present invention.

All embodiments of Formula VI or VII described below are included within the scope of Formula VI.

- 15 041924 or VII in the present invention.

In some preferred embodiments of Formula VI or VII, R ² is —OH.

In some preferred embodiments of formula VI or VII, R ² is F.

In some preferred embodiments of Formula VI or VII, R ²¹ is —OH.

In some preferred embodiments of Formula VI or VII, ^R21 is F.

In some preferred embodiments of Formula VI or VII, B1 is

hh ₂

In some preferred embodiments of Formula VI or VII, B ₂ is

In some preferred embodiments of Formula VI or VII, the stereoconfiguration is (Sp, Sp), (Sp, Rp), (Rp, Rp), or (Rp, Sp).

In some preferred embodiments of Formula VI, R ² is -OH;

R ²¹ is -OH.

In some preferred embodiments of Formula VII, R ² is -OH or -OCH3;

R ²¹ is -OH or F.

In some embodiments, a Formula I compound, isomer, prodrug, stable isotope derivative, or pharmaceutically acceptable salt has any of the following structures:

- 16 041924

- 17 041924

In some embodiments, a Formula I compound, isomer, prodrug, stable isotope derivative, or pharmaceutically acceptable salt has any of the following structures:

In some embodiments, a Formula I compound, isomer, prodrug, stable isotope derivative, or pharmaceutically acceptable salt has any of the following structures:

- 18 041924

In some embodiments, a Formula I compound, isomer, prodrug, stable isotope derivative, or pharmaceutically acceptable salt most preferably has any of the following structures:

- 19 041924

- 20 041924

In some embodiments, a Formula I compound, isomer, prodrug, stable isotope derivative, or pharmaceutically acceptable salt has any of the following structures:

- 21 041924

- 22 041924

- 23 041924

A compound of formula (I), a pharmaceutically acceptable salt thereof, is synthesized by a general chemical process.

Generally, salt formation is carried out by reacting the free base or acid with an equivalent chemical equivalent or excess of the acid (inorganic or organic acid) or base (inorganic or organic base) in a suitable solvent or mixture of solvents.

The present invention also relates to a pharmaceutical composition containing a therapeutically effective amount of the active ingredient and a pharmaceutically acceptable excipient; the active ingredient comprises one or more analogs of cyclic dinucleotide (I), an isomer thereof, a prodrug, a stable isotope derivative, and a pharmaceutically acceptable salt.

The active ingredient in the pharmaceutical composition may also include other therapeutic agents against viral infections or other infectious diseases (e.g. HIV (HIV), hepatitis B virus (HBV), hepatitis C virus (HCV), etc.), autoimmune diseases (e.g. , rheumatoid arthritis, lupus erythematosus, psoriasis, etc.) or malignant neoplasms.

In a pharmaceutical composition, a pharmaceutically acceptable excipient may include a pharmaceutically acceptable carrier, diluent and/or excipient.

According to the purpose of treatment, the pharmaceutical composition can be formulated into various types of unit dosage forms such as tablets, pills, powders, liquids, suspensions, emulsions, granules, capsules, suppositories and injections (solutions and suspensions) and the like, and preferably liquids, suspensions , emulsions, suppositories and injections (solutions and suspensions), etc.

Any excipient known and commonly used in the art can be used to form the pharmaceutical composition into a tablet. For example, carriers such as lactose, white sugar, sodium chloride, glucose, urea, starch, calcium carbonate, kaolin, crystalline cellulose and silicic acid; binders such as water, ethanol, propanol, total syrup, dextrose solution, starch solution, gelatin solution, carboxymethylcellulose, shellac, methylcellulose and potassium phosphate, polyvinylpyrrolidone, etc.; tablet disintegrating agents such as dry starch, sodium alginate, agar powder, kelp powder, sodium bicarbonate, calcium carbonate, fatty acid esters of polyethylene dehydrated sorbitol, sodium dodecyl sulfate, monoglyceryl stearate, starch and lactose; tablet disintegrating agents such as white sugar, glycerol tristearate, coconut oil and hydrogenated oil; adsorption promoters such as quaternary ammonium bases and sodium dodecyl sulfate; wetting agents such as glycerol, starch, etc.; adsorbents such as starch, lactose, kaolin, bentonite and colloidal silicic acid; and lubricants such as pure talc, stearate, boric acid powder and polyethylene glycol. A conventional coating material can also be used to make a sugar coated tablet, a gelatin film coated tablet, a coated tablet, a film coated tablet, a bilayer film coated tablet, and a multilayer tablet.

Any excipient known and commonly used in the art can be used to form the pharmaceutical composition into a pill, for example, carriers such as lactose, starch, coconut oil, hardened vegetable oil, kaolin, talc, etc.; binders such as gum arabic powder, tragacanth powder, gelatin, ethanol, etc.; disintegrating agents such as agar powder and kelp powder.

Any excipient known and widely used in the art, such as polyethylene glycol, coconut oil, higher alcohols, higher alcohol esters, gelatin, and semi-synthetic glycerides, etc., can be used to form the pharmaceutical composition into a suppository form.

To obtain a pharmaceutical composition in the form of an injection, a solution or suspension can be sterilized (preferably by adding an appropriate amount of sodium chloride, glucose or glycerol, etc.) to obtain an injection that is isotonic with blood. You can also use any of the media commonly used in the art. For example, water, ethanol, propylene glycol, ethoxylated isostearyl alcohol, polyethoxylated isostearyl alcohol, and fatty acid esters of polyethylene dehydrated sorbitol. Additionally, conventional solubilizers, buffers, analgesics can be added.

In the present invention, the content of the composition in the pharmaceutical composition, in particular,

- 24 041924 is not limited and can be selected in a wide range, usually 5-95% by weight, preferably

30-80% by weight.

In the present invention, the method of administering the pharmaceutical composition is not particularly limited. Formulations of various dosage forms can be selected depending on the age, sex and other conditions and symptoms of the patient. For example, tablets, pills, solutions, suspensions, emulsions, granules or capsules are administered orally; injections can be administered alone or in combination with injection solutions (eg glucose solution and amino acid solution); suppositories are injected into the rectum.

The present invention also relates to the use of a cyclic dinucleotide analog (I), an isomer thereof, a prodrug, a stable isotope derivative or a pharmaceutically acceptable salt, or a pharmaceutical composition in the preparation of modulators of STING (interferon gene stimulator) factor stimulating interferon genes. The modulator of STING (interferon gene stimulator) interferon gene stimulating factor is preferably an agonist of STING (interferon gene stimulator) interferon gene stimulating factor. An interferon gene stimulator agonist refers to a compound of formula (I), an isomer, a prodrug, a stable isotope derivative or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition capable of activating interferon gene stimulator signaling.

The present invention also relates to the use of a cyclic dinucleotide (I) analog, isomer, prodrug, stable isotope derivative or pharmaceutically acceptable salt or pharmaceutical composition thereof in the preparation of a medicament for modulating the proliferation of T cells or other immune cells.

The present invention also relates to the use of a cyclic dinucleotide (I) analogue, isomer, prodrug, stable isotope derivative or pharmaceutically acceptable salt or pharmaceutical composition thereof in the preparation of a vaccine adjuvant.

The present invention also relates to the use of a cyclic dinucleotide analog (I), an isomer thereof, a prodrug, a stable isotope derivative or a pharmaceutically acceptable salt, or a pharmaceutical composition in the preparation of a medicament for the treatment and/or alleviation of a neoplastic disease and a non-neoplastic disease mediated by an interferon gene stimulator . Interferon gene stimulator mediated diseases refer to diseases caused by immunosuppression or hyperactivation due to interferon gene stimulator signaling. Types of associated diseases include, but are not limited to: viral infections or other infectious diseases, autoimmune diseases, and malignancies.

The present invention preferably relates to the use of a cyclic dinucleotide (I) analogue, isomer, prodrug, stable isotope derivative or pharmaceutically acceptable salt or pharmaceutical composition thereof in the preparation of a medicament for the treatment and/or amelioration of immunosuppressed cancers.

The present invention further relates to a method for treating viral infections or other infectious diseases, malignancies, autoimmune diseases with a cyclic dinucleotide analog (I), an isomer, a prodrug, a stable isotope derivative or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition comprising: administering to a mammal the required dose of the cyclic dinucleotide analog (I), isomer, prodrug, stable isotope derivative or pharmaceutically acceptable salt thereof, or pharmaceutical composition.

The mammal is preferably a human.

The present invention preferably relates to a compound represented by formula (I), an isomer, a prodrug, a stable isotope derivative or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition for the preparation of a medicament for the treatment and/or alleviation of a disease mediated by an interferon gene stimulator; IFN gene stimulator mediated diseases are diseases caused by IFN gene stimulator mediated immunosuppression, diseases may include: viral infections or other infectious diseases (e.g. HIV (HIV), hepatitis B virus (HBV), hepatitis C virus (HCV) etc.), autoimmune diseases (eg rheumatoid arthritis, lupus erythematosus, psoriasis, etc.) or malignant neoplasms.

The present invention further relates to the use of a compound represented by formula (I), an isomer, a prodrug, a stable isotope derivative, or a pharmaceutically acceptable salt or a pharmaceutical composition thereof, in the preparation of a medicament for the treatment and/or alleviation of cancer.

The present invention further relates to the use of a compound represented by formula (I), an isomer, a prodrug, a stable isotope derivative or a pharmaceutically acceptable salt or a pharmaceutical composition thereof, in the preparation of a medicament for the treatment and/or amelioration of viral or other infections.

The present invention further relates to the use of the compound represented by

- 25 041924 formula (I), its isomer, prodrug, derivative with stable isotopes or pharmaceutically acceptable salt, or pharmaceutical composition in the preparation of a drug for the treatment and/or alleviation of autoimmune diseases.

The present invention further relates to a cyclic dinucleotide (I) analogue, isomer, prodrug, stable isotope derivative or pharmaceutically acceptable salt thereof, or a pharmaceutical composition in combination with one or more other kinds of therapeutic agents and/or therapeutic techniques in the treatment, alleviation and/ or preventing diseases mediated by the interferon gene stimulator. Interferon gene stimulator mediated diseases are caused by IFN gene stimulator mediated immunosuppression and these diseases may include: viral or other infections (eg HIV (HIV), hepatitis B virus (HBV), hepatitis C virus (HCV), etc. .), autoimmune diseases (eg rheumatoid arthritis, lupus erythematosus, psoriasis, etc.) or cancer.

The present invention preferably relates to a cyclic dinucleotide (I) analog, isomer, prodrug, stable isotope derivative or pharmaceutically acceptable salt, or a pharmaceutical composition in combination with one or more other kinds of therapeutic agents and/or therapeutic techniques for treating and/or alleviating cancer.

The present invention preferably relates to a cyclic dinucleotide (I) analog, isomer, prodrug, stable isotope derivative or pharmaceutically acceptable salt, or a pharmaceutical composition in combination with one or more other kinds of therapeutic agents and/or therapeutic techniques for treating and/or alleviating cancer mediated by the interferon gene stimulator.

In the present invention, other types of therapeutic agents (eg, other types of cancer therapeutic agents) can be formulated into a cyclic dinucleotide analog (I) therapeutic dosage form for single administration of the drug or separate therapeutic dosage forms for sequential administration.

The present invention further relates to a combination preparation containing a compound of formula (I), an isomer, a prodrug, a stable isotope derivative or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition and other kinds of therapeutic agents and/or therapeutic methods for treating cancer.

In the present invention, other types of cancer therapeutic agents may include, but are not limited to, one or more of: microtubule protein inhibitors, alkylating agents, type I/II topozyme inhibitors, platinum compounds, antimetabolites, hormones and hormone analogs, signal transduction pathway inhibitors , angiogenesis inhibitors, targeted therapeutic agents (eg, specific kinase inhibitors), immunotherapeutic agents, pro-apoptotic agents, and cell cycle signaling pathway inhibitors.

In the present invention, other types of therapeutic methods for treating cancer may include, but are not limited to, one or more of: tumor immunotherapy and radiation therapy.

In the present invention, other types of cancer therapeutic agents are preferably immunotherapeutic agents.

In the present invention, the microtubulin inhibitor may be selected from, but not limited to, one or more of the vincristine families (e.g., vinblastine, vincristine, vinorelbine, vindesine sulfate), the taxane families (docetaxel, paclitaxel), and eribulin mesylate.

In the present invention, the alkylating agent may be selected from, but not limited to, nitrogen mustard, N-oxo-nitrogen mustard hydrochloride, nitrogen mustard 3-cyclohexyl-3-hydroxybutyrate, uracil mustard, cyclophosphamide, ifosfamide, thiotepa, carboquone, triethyleneiminoquinone, improsulfan tosylate, mannosesufan (mannosesulfan), treosulfan, busulfan, nimustine hydrochloride, dibromomannite, melphalan, dacarbazine, ranimustine, carmustine, lomustine, streptozotocin, temozolomide, procarbazine, ethyleneimine derivatives, methanesulfonates, nitrosourea, triazene.

In the present invention, type I/II topozyme inhibitors can be selected from, but are not limited to, one or more of doxorubicin, daunorubicin, epirubicin, idarubicin, irinotecan, topotecan, rubitecan, belotecan, etoposide, teniposide, adriamycin, dexrazoxane, and camptothecin.

In the present invention, the platinum compound may be selected from, but not limited to, one or more of cisplatin, carboplatin, oxaliplatin, and nedaplatin.

In the present invention, the antimetabolites may be selected from, but are not limited to, one or more of: folic acid antagonists, pyrimidine analogs, purine analogs, adenosine deaminase inhibitors such as: methotrexate, 5-fluorouracil, floxuridine, cytarabine, 6-mercaptopurine, 6-thioguanine, fludarabine phosphate, pentostatin and gemcitabine.

In the present invention, the immunotherapeutic agent may be selected from, but not limited to, one or more of immunomodulators, modulators of the tumor microenvironment, and anti-angiogenic factors. Immunomodulators may include, but are not limited to:

1) protein antagonists (for example, immune checkpoint inhibitors), which inhibit

- 26 041924 T-cell activity: one or more of CTLA4 (for example, one or more of ipilimumab, tremelimumab, abatacept, belatacept, BMS-986249, BMS-986218, AGEN-1884 and KN-046), PD-1 ( for example, one or more of: camrelizumab, toripalimab, sintilimab, cemiplimab, pembrolizumab, nivolumab, tiselizumab, spartalizumab, dostarlimab, genolimzumab, cetrelimab, HLX-10, BCD100, AK-105, MEDI-0680, CS-1003, BAT-1306 , HX-008, sasanlimab, AGEN-2034, BI-754091, GLS-010, MGA-012, AK-104, and AK-103), PD-L1 (eg, one or more of: durvalumab, avelumab, atezolizumab, envafolimab , cocibelimab, CS1001, SHR-1316, lasertinib, bintrafuspa alfa, TQB-2450, CA170, CX-072, BGB-A333, BMS-936559, GEN-1046, KL-A167 and IO-103), LAG3 and TIM3;

2) protein agonists that stimulate T cell activity: one or more of GITR, OX40, OX40L, 4-1BB (CD137), CD27, and CD40;

3) one or more of TLR2 agonists, TLR4 agonists, TLR5 agonists, TLR7 agonists, TLR8 agonists, and TLR9 agonists;

4) IDO inhibitors, CD73 inhibitors.

In the present invention, signal transduction inhibitors (STIs) can be selected from, but are not limited to, one or more of: BCR/ABL type kinase inhibitors, epidermal growth factor receptor inhibitors, her-2/neu type receptor inhibitors, AKT family kinase inhibitors , PI3K-type signaling pathway inhibitors, and cell cycle checkpoint inhibitors.

In the present invention, angiogenesis inhibitors can be selected from, but are not limited to, one or more of VEGF/VEGFR type signaling pathway inhibitors, Src family kinase inhibitors, Src type signaling pathway inhibitors, and c-Fes type kinase inhibitors.

In the present invention, the target therapeutic agents may be selected from, but are not limited to, one or more of: erlotinib, imatinib, apatinib, nilotinib, crizotinib, dasatinib, pazopanib, regorafenib, ruxolitinib, sorafenib, sunitinib, vandetanib, vemurafenib, bosutinib, gefitinib, afatinib, axitinib, dabrafenib, dacomitinib, nintedanib, levatinib, masitinib, midostaurin, neratinib, ponatinib, radotinib, trametinib, brivanib, alaninate, cediranib, cabozantinib malate, ibrutinib, icotinib, lapatinib, cobimetinib, idelicinib, alinitinib, ponatinib, ponatinib , orantinib, rigosertib, tipifarnib, tivozanib, pimacertib, buparlisib, and fedratinib.

In the present invention, tumor immunotherapy can be selected from, but is not limited to, one or more of: antitumor vaccines (eg, synthetic peptides, DNA vaccines, and recombinant viruses), oncolytic viruses, cytokine therapies (eg, IL2 and GM-CSF), and therapies with T-cells with chimeric antigen receptors (CAR-T).

In the present invention, viral and other infections may include: infections caused by viruses such as influenza viruses, hepatitis B virus (HBV), hepatitis C virus (HCV), human papillomavirus (HPV), cytomegalovirus (CMV), Epstein-Barr virus (EBV), poliovirus, varicella-zoster virus, coxsackieviruses, or human immunodeficiency virus (HIV, HIV).

In the present invention, cancers include metastatic and non-metastatic cancers, also include familial and sporadic cancers, and may also include solid and non-solid tumors.

In the present invention, specific examples of solid tumors may include, but are not limited to, tumors of: eye, bone, lung, stomach, pancreas, breast, prostate, brain (including glioblastoma and medulloblastoma), ovary (including these stromal cells, germ cells, and mesenchymal cells derived from epithelial cells), bladder, testis, spinal cord, kidney (including adenocarcinoma, nephroblastoma), mouth, lip, throat, oral cavity (including squamous cell carcinoma), nasal cavity, small intestine, colon, rectum , parathyroid gland, gallbladder, bile duct, cervix, heart, subpharyngeal gland, bronchus, liver, ureter, vagina, anus, larynx, thyroid gland (including thyroid cancer and medullary cancer), esophagus, nasopharyngeal pituitary gland, salivary glands , adrenal gland, intraepithelial neoplasms of the head and neck (including Bowen's disease and Paget's disease), sarcoma (including smooth muscle sarcoma, rhabdomyos rcoma, liposarcoma, fibrosarcoma, osteoblastic sarcoma), skin (including melanoma, Kaposi's sarcoma, basal cell carcinoma and squamous cell carcinoma) and other related tumors.

In the present invention, the solid tumor is preferably one or more of: eye cancer, bone cancer, lung cancer, stomach cancer, pancreatic cancer, breast cancer, prostate cancer, brain cancer (including but not limited to glioblastoma, cell tumor adult neural tube), ovarian cancer, bladder cancer, cervical cancer, testicular cancer, kidney cancer (including but not limited to adenocarcinoma, nephroblastoma), oral cavity cancer (including squamous cell carcinoma), tongue cancer, laryngeal cancer, cancer nasopharynx, head and neck cancer, colon cancer, small intestine cancer, rectal cancer, parathyroid cancer, thyroid cancer, esophageal cancer, gallbladder cancer, bile duct cancer, cervical cancer, liver cancer, lung cancer (including, but not limited to, small cell lung cancer, non-small cell lung cancer), chorionic epithelioma, osteosarcoma,

- 27 041924 Ewing's tumor, soft tissue sarcoma and skin cancer.

In the present invention, specific examples of non-solid tumors (including hematologic tumors) may include, but are not limited to: lymphoid leukemia (including acute lymphoblastic leukemia, lymphoma myeloma, chronic lymphocytic leukemia, Hodgkin's lymphoma, non-Hodgkin's lymphoma, T-cell chronic lymphatic leukemia, B- cellular chronic lymphatic leukemia), myeloid-associated leukemia (including acute myeloid leukemia, chronic myeloid leukemia) and AIDS-associated leukemia.

In the present invention, autoimmune diseases may include, but are not limited to, one or more of: rheumatoid arthritis, systemic lupus erythematosus, mixed connective tissue disease (MCTD), systemic scleroderma (including: CREST syndrome), dermatomyositis, nodular vasculitis, renal failure (including: hemorrhagic pulmonary-renal syndrome, acute glomerulonephritis, primary membranous proliferative glomerulonephritis, etc.), endocrine diseases (including: type I diabetes, gonadal insufficiency, pernicious anemia, hyperthyroidism, etc.), liver diseases (including: primary biliary cirrhosis, autoimmune cholangitis, autoimmune hepatitis, primary sclerosing cholangitis, etc.) and autoimmune reactions caused by infections (eg, AIDS, malaria, etc.).

Unless otherwise indicated, the following terms used in the description and claims of the present invention have the following meanings.

The term alkyl refers to a straight or branched chain saturated hydrocarbon group containing 1 to 20 carbon atoms, preferably 1 to 10 carbon atoms, more preferably 1 to 8, 1 to 6, 1 to 4, 1 to 3 carbon atoms, typical examples of alkyl groups include, but are not limited to: methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, t-butyl, isobutyl, n-pentyl, n-hexyl, n-heptyl, octyl, nonyl, decyl, 1,1-dimethylpropyl, 1,2dimethylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, 1-ethyl-2-methylpropyl, 1,1,2-trimethylpropyl, 1,1-dimethylbutyl, 1,2-dimethylbutyl, 2,2-dimethylbutyl, 1,3-dimethylbutyl, 2,3dimethylbutyl, 2-ethylbutyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, 4,4-dimethylpentyl, 2methylhexyl, 3-methylhexyl, 4-methylhexyl, 5-methylhexyl, 2,3-dimethylpentyl, 2,4-dimethylpentyl, 2,2dimethylpentyl, 3,3-dimethylpentyl, 2-ethylpentyl, 3-ethylpentyl, 2,2,4- trimethylpentyl, undecyl, dodecyl and their various isomers, etc.

The term cycloalkyl refers to a saturated or partially unsaturated (containing 1 or 2 double bonds) monocyclic or polycyclic group containing from 3 to 20 carbon atoms. The term monocyclic cycloalkyl is preferably 3-10 membered monocyclic alkyl, more preferably 3-8 membered monocyclic alkyl such as: cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, cyclodecyl, cyclohexenyl. The term polycyclic cycloalkyl includes bridged cycloalkyl, fused cycloalkyl, and spirocycloalkyl. Monocyclic cycloalkyl or polycyclic cycloalkyl may be linked to the parent molecule via any carbon atom in the ring.

The term heterocycloalkyl refers to a saturated or partially unsaturated (containing 1 or 2 double bonds) non-aromatic cyclic group consisting of carbon atom(s) and heteroatom(s) selected from nitrogen, oxygen or sulfur, which is monocyclic or polycyclic, in the present invention the number of heteroatoms in the heterocycloalkyl group is preferably 1, 2, 3 or 4, the nitrogen, carbon or sulfur atom in the heterocycloalkyl group is optionally oxidized. The nitrogen atom is optionally further substituted with groups to form tertiary amines or quaternary ammonium salts. The monocyclic heterocycloalkyl is preferably a 3-10 membered monocyclic heterocycloalkyl, more preferably a 3-8 membered monocyclic heterocycloalkyl. For example: aziridinyl, tetrahydrofuran-2-yl, morpholin-4-yl, thiomorpholin-4-yl, thiomorpholin-S-oxide-4-yl, piperidin-1-yl, N-alkylpiperidin-4-yl, pyrrolidin-1- yl, N-alkylpyrrolidin-2-yl, piperazin-1-yl, 4-alkylpiperazin-1-yl, etc. Polycyclic heterocycloalkyl includes fused heterocycloalkyl, spiroheterocycloalkyl, and bridged heterocycloalkyl. Monocyclic heterocycloalkyl and polycyclic heterocycloalkyl can be linked to the parent molecule via any ring atom in the ring. The above ring atoms refer in particular to the carbon and/or nitrogen atoms which constitute the ring skeleton.

The term cycloalkylalkyl refers to a cycloalkyl group linked to the parent core structure via an alkyl group. Thus, the term cycloalkylalkyl encompasses the above definitions of alkyl and cycloalkyl.

The term heterocycloalkylalkyl refers to a heterocycloalkyl group linked to the parent core structure via an alkyl group. Thus, the term heterocycloalkylalkyl encompasses the above definitions of alkyl and heterocycloalkyl.

The term alkoxy refers to a cyclic or non-cyclic alkyl group having said carbon atoms attached through an oxygen bridge and includes alkyloxy, cycloalkyloxy, and heterocycloalkyloxy. Thus, alkoxy includes the above definitions of alkoxy

- 28 041924 la, heterocycloalkyl and cycloalkyl.

The term alkenyl refers to a linear, branched or cyclic non-aromatic hydrocarbon group containing at least 1 carbon-carbon double bond. 1 to 3 carbon-carbon double bonds may be present, preferably 1 carbon-carbon double bond. The term C ₂ _ ₄ alkenyl refers to an alkenyl group containing from 2 to 4 carbon atoms, and the term C ₂ . ₆ Alkenyl refers to an alkenyl group containing 2 to 6 carbon atoms, including vinyl, propylene, butenyl, 2-methylbutenyl, and cyclohexenyl. Alkenyl groups may be substituted.

The term alkynyl refers to a linear, branched or cyclic hydrocarbon group containing at least 1 carbon-carbon triple bond. 1 to 3 carbon-carbon triple bonds may be present, preferably 1 carbon-carbon triple bond. The term C ₂ _ ₆ alkynyl refers to an alkynyl group containing 2 to 6 carbon atoms, including ethynyl, propynyl, butynyl, and 3-methylbutynyl.

The term aryl refers to any stable 6-10 monocyclic or bicyclic aromatic group such as: phenyl, naphthyl, tetrahydronaphthyl, 2,3-dihydroindenyl or biphenyl.

The term heteroaryl refers to an aromatic cyclic group formed by replacing at least 1 carbon atom in the ring with a heteroatom selected from nitrogen, oxygen or sulfur, which has a 5-7 membered monocyclic structure or a 7-12 membered bicyclic structure, preferably 5- 6-membered heteroaryl. In the present invention, the number of heteroatoms is preferably 1, 2 or 3 and includes: pyridyl, pyrimidinyl, pyridazin-3(2H)-one group, furanyl, thienyl, thiazolyl, pyrrolyl, imidazolyl, pyrazolyl, oxazolyl, isoxazolyl, 1,2,5 -oxadiazolyl, 1,2,4-oxadiazolyl, 1,2,4-triazolyl, 1,2,3-triazolyl, tetrazolyl, indazolyl, isoindazolyl, indolyl, isoindolyl, benzofuranyl, benzothienyl, benzo[d][1,3] dioxolyl, benzothiazolyl, benzoxazolyl, quinolinyl, isoquinolinyl, quinazolinyl, etc.

The term arylalkyl refers to an aryl group linked to the parent core structure via an alkyl group. Thus, the term arylalkyl encompasses the above definitions of alkyl and aryl.

The term heteroarylalkyl refers to a heterocycloalkyl group linked to the parent core structure via an alkyl group. Thus, the term heteroarylalkyl encompasses the above definitions of alkyl and heteroaryl.

The term halogen means fluorine, chlorine, bromine or iodine.

The term haloalkyl refers to an alkyl group optionally substituted with halogen. Thus, haloalkyl includes the definitions of halogen and alkyl as defined above.

The term haloalkoxy refers to an alkoxy group optionally substituted with halogen. Thus, the term haloalkoxy encompasses the above definitions of halogen and alkoxy.

The term amino refers to -NH ₂ and the term alkylamino refers to that at least one hydrogen atom in the amino group is replaced by an alkyl group, including but not limited to: NHCH3, -N(CH ₃ ) ₂ , -NHCH ₂ CH3, -N(CH ₂ CH ₃ ) ₂ .

The term nitro refers to -NO ₂ .

The term cyano refers to k-CN.

The term azido refers to -N ₃ .

The term room temperature in the present invention refers to 15-30°C.

An isotopically substituted derivative includes an isotopically substituted derivative in which any hydrogen atom of a compound of formula (I) is replaced by 1-5 deuterium atoms, or any carbon atom of a compound of formula (I) is replaced by 1-3 C ¹⁴ atoms, or any oxygen atom compounds of formula I replaced by 1-3 O ¹⁸ atoms.

The term prodrug refers to a compound capable of being converted to its parent active compound after in vivo metabolism. Typically, the prodrug is inactive or less active than the active parent compound, but provides ease of handling, administration, or improved metabolic properties.

The pharmaceutically acceptable salts described in the present invention are discussed in the article Pharmaceutically acceptable salts by Berge, et al., J. Pharm. Sci., 66, 1-19 (1977), and it is apparent to pharmaceutical chemists that salts are essentially non-toxic and provide desirable pharmacokinetic properties, palatability, absorption, distribution, metabolism or excretion, and so on. The compounds referred to herein may have acid groups, basic groups, or amphoteric groups, and typical pharmaceutically acceptable salts include salts obtained by reacting the compounds of the present invention with acids, such as: hydrochloride, hydrobromide, sulfate, pyrosulfate, bisulfate, sulfite, bisulfite , phosphate, hydrogen phosphate, dihydrogen phosphate, metaphosphate, pyrophosphate, nitrate, acetate, propionate, decanoate, octanoate, formate, acrylate, isobutyrate, caproate, heptanoate, oxalate, malonate, succinate, octandioate, benzoate, methyl benzoate, phthalate, maleate, methanesulfonate, p -toluenesulfonate, (DD-tartaric acid, citric acid, maleic acid, (DD-malic acid, fumaric acid, succinate, lactate, trifluoromethanesulfonate, naphthalene-1sulfonate, mandelic acid salt, pyruvate, stearate, ascorbate, salicylate. When the compounds are

- 29 041924 present invention contain acidic groups, their pharmaceutically acceptable salts may also include: alkali metal salts such as lithium, sodium or potassium salts; alkaline earth metal salts such as zinc, calcium or magnesium salts; alkali metal type organic salts such as those formed with ammonia, alkylamines (including, but not limited to: methylamine, triethylamine), hydroxyalkylamines, amino acids (including, but not limited to: lysine, arginine), N-methylglucosamine, etc. d.

The term isomer in the present invention means that the compound of formula (I) of the present invention may have asymmetric centers and racemates, racemic mixtures and individual diastereoisomers, all of which are included in the present invention, including stereoisomers and geometric isomers. In the present invention, individual stereoisomers (enantiomers and diastereoisomers), as well as mixtures thereof, are included within the scope of the present invention when a compound of formula (I) or a salt thereof may be present in stereoisomeric form (for example, the compound contains one or more asymmetric carbon atoms and/or atoms phosphorus). The present invention also includes individual isomers of the compounds represented by formula (I) or salts, as well as mixtures of isomers with one or more reversed chiral centers. The scope of the present invention includes mixtures of stereoisomers as well as purified enantiomeric mixtures or mixtures enriched in enantiomers/diastereoisomers. The present invention includes stereoisomeric mixtures formed by all enantiomers and diastereoisomers in all possible different combinations. The present invention includes all combinations and subsets of stereoisomers of all specific groups as defined above. The compound of formula (I) in the present invention contains chiral P atoms with the conformation Rp or Sp, so that compounds with individual stereoconfiguration (Sp, Sp), (Sp, Rp), (Rp, Rp) or (Rp, Sp) and any of them mixtures are included within the scope of the present invention.

The above preferred conditions of the present invention can be arbitrarily combined without departing from the general knowledge in the art to obtain the preferred embodiments of the present invention.

The reagents and raw materials used in the present invention are commercially available.

Brief description of the drawing

Figure 1 is tumor volume change curves for compounds 6-p3 (1 mg/kg, 2 mg/kg i.t. (intratumoral injection)) and comparative compound 1 (2 mg/kg i.t.) in CT26 colon cancer subcutaneously implanted mouse tumor .

Detailed description of an embodiment of the invention

The following embodiments serve to illustrate the present invention, but the embodiments should not be construed as limiting the scope of the invention. Some of the experimental methods of the following embodiments, which do not specify specific conditions, may be in accordance with widely used reaction conditions and methods, or in accordance with product specifications.

All structures of the compounds in the present invention were confirmed by nuclear magnetic resonance (1H NMR) and/or mass spectra (MS).

Chemical shifts (δ) 1H NMR were recorded in ppm. (10 ^-6 ). NMR spectra were recorded on a Bruker AVANCE-400 spectrometer. Suitable solvents were chloroform-d (CDCl ₃ ), methanol-d ₄ (CD3OD) and dimethylsulfoxide-d ₆ (DMSO-d ₆ ), tetramethylsilane served as an internal standard (TMS).

Low resolution analytical mass spectra (LCMS, Liquid Chromatography-Mass spectrometry, liquid chromatography-mass spectrometry, LCMS) were recorded on an Agilent 1200 HPLC/6120 using XBridge C18, 3.0x50 mm, 3 µm, column temperature: 35°C; or recorded on ThermoUltiMate 3000HPLC/MSQPLUS using XBridge C18, 3.0x50 mm, 3.5 µm, column temperature: 30°C. Agilent Type 1 Gradient Elution Method: 95-5% Solvent A1 and 5-95% Solvent B1 (0-2.0 min) followed by 95% Solvent B1 and 5% Solvent A1 (over 1.1 min). The percentage used in the present description is the volume percentage of the volume of solvent in the total volume of solvent. Solvent A1: 0.01% aqueous trifluoroacetic acid (TFA); Solvent B1: 0.01% trifluoroacetic acid acetonitrile solution. The percentage is the volume of solvent in the total volume of solvent. Type 2 gradient elution method on Thermo: 95-5% Solvent A ₂ and 5-95% Solvent B ₂ (0-2 min) followed by 95% Solvent B ₂ and 5% Solvent A2 (within 1.8 min) . The percentage is the volume of solvent in the total volume of solvent. Solvent A ₂ : 10 mM ammonium bicarbonate aqueous solution; Solvent B ₂ : acetonitrile.

All compounds in the present invention were separated by preparative high performance liquid chromatography or flash column chromatography.

Purification by preparative high performance liquid chromatography (preparative HPLC) was performed on a Shimadzu LC-20 HPLC, chromatographic column: Waters xbridge Pre C18, 10 µm, 19 mm x 250 mm. Separation method type 1 (acidic medium): mobile phase A: 0.05% aqueous solution of three

- 30 041924 fluoroacetic acid, mobile phase B: acetonitrile; elution B was 40%, elution time: 20 min. Separation method type 2 (basic medium): mobile phase A: 10 mmol/l aqueous ammonium bicarbonate solution, mobile phase B: acetonitrile; gradient elution B was 10% to 80%, elution time: 30 min. Separation method type 3 (basic medium): mobile phase A: 10 mmol/l aqueous ammonium bicarbonate solution, mobile phase B: acetonitrile; gradient elution B was 0% to 15%, elution time: 30 min. Separation method type 4 (basic medium): mobile phase A: 10 mmol/l aqueous ammonium bicarbonate solution, mobile phase B: acetonitrile; gradient elution B was 0% to 4%, elution time: 10 min; gradient elution B was 4% to 8%, elution time: 15 min. Separation method type 5 (basic medium): mobile phase A: 10 mmol/l aqueous ammonium bicarbonate solution, mobile phase B: acetonitrile; gradient elution B was 0% to 5%, elution time: 10 min; gradient elution B was 5% to 10%, elution time: 15 min. Separation method type 6 (alkaline): mobile phase A: 10 mmol/l aqueous ammonium bicarbonate solution, mobile phase B: acetonitrile; gradient elution B was 10% to 30%, elution time: 5 min; gradient elution B was 30% to 75%, elution time: 20 min. Separation method type 7 (basic medium): mobile phase A: 10 mmol/l aqueous ammonium bicarbonate solution, mobile phase B: acetonitrile; gradient elution B was 0% to 10%, elution time: 7 min; gradient elution B was 10% to 40%, elution time: 18 min. Detection wavelength: 214 nm and 254 nm; flow rate: 15.0 ml/min

Flash column chromatography (flash/Cheetah™) was performed on an Agela Technologies MP200. The direct phase chromatography column was Flash columm Silica-CS (25 g, 40 g, 80 g, 120 g or 330 g), Agela Technologies, Tianjing. Ethyl acetate/petroleum ether or dichloromethane/methanol was chosen as the elution system. The reverse phase chromatography column was C18 columm (12 g, 20 g or 40 g), Santai Technologies, Changzhou. Acetonitrile/aqueous ammonium bicarbonate (10 mmol/l) was chosen as the elution system.

All compounds in the present description were analyzed using high performance liquid chromatography. High performance liquid chromatography (HPLC) was performed on a Waters e2695, 2498 UV/VIS Detector, chromatography column: Waters Xselect CHS C18 (4.6x150 mm) 5 µm, mobile phase A: acetonitrile, mobile phase B: acetic acid-triethylamine buffer which adjusted to pH 5.0 with acetic acid. The gradient elution of mobile phase B was 95% to 15%, elution time: 30 min. Detection wavelength: 214 nm and 254 nm; column temperature: 35°C.

Implementation Option 1. Synthesis of Intermediate 1-8.

Stage 1. Liquid bromine (60 g, 374 mmol) was added to a suspension of adenosine (50 g, 187 mmol) in a buffer solution of acetic acid/sodium acetate (pH=4.0, 0.5 M, 1 L), the temperature of the system was maintained below 10°C. After the addition, the reaction system was stirred at room temperature for 48 hours. An aqueous saturated sodium bisulfate solution was added to the reaction solution to remove excess bromine, and then adjusted to neutral pH with an aqueous sodium hydroxide solution (1 M), the reaction solution was stirred for 2 h in an ice bath. A precipitate formed which was collected by filtration, dried under vacuum to give intermediate 1-1 (29 g), m/z: [M+H]+ 346.0/348.0.

Step 2 To a suspension of intermediate 1-1 (10 g, 28.9 mmol) in methanol (100 mL) was added sodium methanolate (9.36 g, 173 mmol) and the reaction system was stirred at reflux. within 5 h, the methanol was concentrated under reduced pressure. The residue was dissolved in a mixed solvent of methanol/dichloromethane (1/10). The solution was filtered through a funnel

Buchner, covered with a layer of silica gel. The filtrate was concentrated under reduced pressure to give intermediate 1-2 (3.8 g), m/z: [M+H]+ 298.2.

Step 3 To a solution of intermediate 1-2 (10 g, 336 mmol) in pyridine (40 ml) was added chlorotrimethylsilane (16 ml, 121 mmol) under nitrogen at 0°C. The reaction system was stirred at room temperature for 2 h, benzoyl chloride (9.4 ml, 80.7 mmol) was slowly added to the above reaction system. The resulting solution was stirred at room temperature overnight, and then an ammonium hydroxide solution (25-28%) was added thereto and stirred for 30 minutes. The solvent was concentrated under reduced pressure. The residue was purified by flash column chromatography (0-10% methanol/dichloromethane) to give intermediate 1-3 (7.3 g) as a white solid, m/z: [M+H]+ 402.2.

Step 4 To a solution of intermediate 1-3 (7 g, 17.4 mmol) in anhydrous pyridine (40 ml) was added 4,4'-dimethoxytrityl chloride (DMTrCl, 5.9 g, 17.4 mmol) under nitrogen at 0°C. The reaction system was stirred at room temperature for 3 hours and then quenched by adding water (1 ml). The solvent was concentrated under reduced pressure. The residue was purified by flash column chromatography (0-10% methanol/dichloromethane) to give intermediate (7.4 g) as a yellow solid, m/z: [M+H]+ 704.2.

Step 5 To a solution of intermediate 1-4 (1.2 g, 1.70 mmol) in pyridine (5 mL) was added tert-butyldimethylsilyl chloride (TBSCl, 0.31 g, 2.05 mmol) and imidazole (0.29 g, 4.30 mmol) under nitrogen at 0°C. The reaction system was stirred at room temperature for 16 h and then cooled with ice water, diluted with water (10 ml) and ethyl acetate (50 ml), the organic layer was washed with brine (50 mlx2), the organic layer was separated and dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was purified by flash column chromatography (0-60% ethyl acetate/petroleum ether) to give intermediates 1-5 (less polar, 276 mg, white solid) and 1-6 (more polar, 670 mg, off-white solid) . Intermediate 1-5: m/z: [M+H]+ 818.3; TCXRf=0.42 (DCM/MeOH=15/1); 1H NMR (400 MHz, DMSO^): δ 11.00 (s, 1H), 8.48 (s, 1H), 8.02 (d, J=7.8 Hz, 2H), 7.62 (t , J=7.4 Hz, 1H), 7.53 (t, J=7.6 Hz, 2H), 7.39 (d, J=7.8 Hz, 2H), 7.29-7.16 (m, 7H), 6.83 (d, J=8.6 Hz, 4H), 5.90 (d, J=5.8 Hz, 1H), 5.13 (d, J=6.0 Hz , 1H), 5.08 (t, J=5.5 Hz, 1H), 4.27 (dd, J=9.6, 5.2 Hz, 1H), 4.07 (d, J=4, 0 Hz, 1H), 4.00 (s, 3H), 3.71 (s, 6H), 3.27-2.23 (m, 1H), 3.14-3.10 (m, 1H), 0.74 (s, 9H), 0.10 (d, J=6.6 Hz, 6H). Intermediate 1-6: m/z: [M+H]+ 818.3, TLC Rf=0.23 (DCM/MeOH=10/1); 1H NMR (400 MHz, DMSO-d6): δ 11.00 (s, 1H), 8.50 (s, 1H), 8.01 (d, J=7.6 Hz, 2H), 7.62 ( m, 1H), 7.52 (m, 2H), 7.32 (d, J=7.5 Hz, 2H), 7.26-7.16 (m, 7H), 6.82 (dd, J = 8.8, 2.3 Hz, 4H), 5.83 (d, J=5.2 Hz, 1H), 5.40 (d, J=6.1 Hz, 1H), 5.06 (m , 1H), 4.59 (t, J=4.5 Hz, 1H), 4.06 (s, 3H), 3.71 (s, 6H), 3.35-3.33 (m, 1H) , 3.31-3.26 (m, 1H), 3.09-3.01 (m, 1H), 0.85 (s, 9H), 0.08 (d, J=7.8 Hz, 6H ).

Step 6 To a solution of intermediate 1-6 (400 mg, 0.48 mmol) in pyridine (4 ml) was added diphenyl phosphite (460 mg, 1.92 mmol) followed by stirring at room temperature for 30 minutes. Triethylamine (0.4 ml) and water (0.4 ml) were added successively to the above reaction system and stirred for 30 minutes. Dichloromethane (5 ml) and aqueous sodium bicarbonate solution (5 ml, 5%) were added successively to the above reaction system. The organic layer was washed with water, and then the separated organic layer was dried over anhydrous sodium sulfate, filtered and concentrated. The residue was purified by flash column chromatography (0-10% methanol/dichloromethane) to give intermediate 1-7 (triethylamine salt, 600 mg) as a white solid, m/z: [M+H]+ 882.3.

Step 7 To a solution of intermediate 1-7 (2.8 g, 2.85 mmol) in a mixed solvent of dichloromethane (20 ml) and water (0.3 ml) was added a dichloromethane solution of dichloroacetic acid (DCA) (0.6 M, 23.7 ml) and then stirred at room temperature for 1 hour. Pyridine (20 ml) was added to the reaction solution, and the resulting mixture was stirred at room temperature for 10 minutes. The solvent was concentrated under reduced pressure to give intermediate 1-8 (pyridinium salt, crude product), m/z: [M+H]+580.1.

Implementation Option 2. Synthesis of compounds 1-p1, 1-p2, 1-p3 and 1-p4.

- 32 041924

Step 1. Intermediate 1-8 (2.85 mmol) was dissolved in anhydrous acetonitrile (15 ml), and then the solvent was concentrated under reduced pressure, repeated twice and the last time left 10 ml of acetonitrile, to which was added a molecular sieve type 4A ( 0.8 g). Compound 1-9 (CAS number: 104992-55-4, 3.3 g, 3.42 mmol) was dissolved in anhydrous acetonitrile (15 ml), and then the solvent was concentrated under reduced pressure, repeated twice and the last time left 5 ml acetonitrile. A solution of compound 19 in acetonitrile was slowly added to a solution of 1-8 in acetonitrile at 0°C, the reaction system was stirred at room temperature for 0.5 h, and ((dimethylaminomethylidene)amino)-3H-1,2,4- dithiazolin-3-thione (DDTT, 697 mg, 3.42 mmol) and stirred for an additional 40 minutes. The molecular sieve was removed by filtration, and the filtrate was concentrated under reduced pressure to give compound 1-10 (7.5 g), m/z: [M+H]+ 1499.3.

Step 2 Compound 1-10 (3.7 g, 2.35 mmol) was dissolved in dichloromethane (35 ml) and water (0.7 ml); a dichloromethane solution of dichloroacetic acid (DCA, 0.6 M, 31 ml, 18.8 mmol) was added dropwise thereto at room temperature. The reaction system was stirred at room temperature for 2 hours. Triethylsilane (20 ml) was added dropwise thereto, and the reaction solution was stirred at room temperature for an additional 1 hour. Pyridine (10 ml) was added dropwise thereto, and the reaction solution was concentrated under reduced pressure. The residue was purified by preparative HPLC to give compound 1-11 (400 mg) as a white solid, m/z: [M+H] ⁺ 1196.2.

Steps 3 and 4 Diphenyl chlorophosphate (DPCP, 1 g, 3.8 mmol) was slowly added dropwise to pyridine (5 ml) at -40°C, anhydrous dichloromethane solution (5 ml) of compound 1- was slowly added dropwise to the above solution. 11 (230 mg, 0.19 mmol) at -40°C, and then stirred at this temperature for 30 min, and a reaction solution of compound 1-12 was obtained. 3H-1,2-benzodithiol-3-one (64 mg, 0.38 mmol) was directly added to a solution of compound 1-12 and stirred for 1 h. Water (68 mg, 0.38 mmol) was added to it and stirred for an additional 1 h. The reaction solution was diluted with ethyl acetate and washed with aqueous sodium bicarbonate solution (2.7%, 30 ml), the organic layer was separated and concentrated to obtain compound 1-13. Compound 1-13 was separated by preparative HPLC (type 2 separation method) to give 3 isomers: 1-13-p1 (60 mg) as a yellow solid, 1-13-p2 (30 mg) as a white solid and 1-13-p3 (40 mg) as a white solid.

Step 5 To a solution of compound 1-13-p1 (80 mg, 0.066 mmol) in acetonitrile (2 ml) was added tert-butylamine (2 ml). The reaction system was stirred at room temperature for 0.5 h. The solvent was concentrated under reduced pressure. The residue was dissolved in methanol (4 ml), methanolic hydrochloric acid (2M, 4 ml) was added thereto, the reaction solution was stirred at 45° C. for 1 hour, and the solvent was concentrated under reduced pressure to obtain compound 1-14 -p1 (100 mg) as yellow solid, m/z: [M+H]+ 1143.1.

Step 6 To a solution of compound 1-14-p1 (100 mg) in methanol (6 ml) was added ammonium hydroxide solution (6 ml). The reaction system was stirred at 45° C. overnight. The solvent was concentrated under reduced pressure. The residual liquid was lyophilized to obtain the compound

1-15-p1 (100 mg, crude compound) as yellow solid, m/z: [M+H]+ 935.2.

Step 7 Compound 1-15-p1 (45 mg) was azeotropically dehydrated three times with anhydrous pyridine (10 ml), then dissolved in pyridine (2 ml), and then triethylamine (0.66 ml) was added thereto under nitrogen atmosphere and triethylamine trihydrofluoride (387 mg). The resulting solution was stirred at 45° C. for 3 hours. The solvent was concentrated under reduced pressure. The residue was purified by preparative HPLC (type 3 separation method) to give compound 1-p1 (0.34 mg, m/z: [M+H]+ 706.8, HPLC retention time: 8.584 min) and 1-p2 (0 .30 mg, m/z: [M+H]+ 706.8, HPLC retention time: 8.662 min) as white solids.

Synthesis of compound 1-p3.

Similarly, to a solution of compound 1-13-p2 (50 mg, 0.041 mmol) in acetonitrile (2.0 ml) was added tert-butylamine (2 ml) and stirred at room temperature for 0.5 h, the solvent was concentrated under reduced pressure . The residue was dissolved in methanol (1 ml), a methanolic hydrochloric acid solution (2 ml, 2 M) was added thereto, the reaction solution was stirred at 40° C. for 4 hours, and then the solvent was concentrated under reduced pressure. The residue was dissolved in methanol (1 ml), ammonium hydroxide solution (1 ml) was added thereto and stirred at 50° C. for 16 hours, and then the reaction solution was purged with nitrogen to remove most of the ammonia. The solvent was concentrated under reduced pressure. The residual liquid was lyophilized. The crude compound was azeotropically dehydrated three times with anhydrous pyridine (10 ml) and then dissolved in pyridine (2 ml) and triethylamine (0.66 ml) and triethylamine trihydrofluoride (0.36 ml) were added under nitrogen. The resulting solution was stirred at 50° C. for 2 hours. The solvent was concentrated under reduced pressure. The residue was purified by preparative HPLC (type 3 separation method) to give compound 1-p3 (0.40 mg, m/z: [M+H]+ 706.8, HPLC retention time: 9.789 min) as an off-white solid substances.

Synthesis of compound 1-p4.

Compound 1-p4 (2.4 mg, m/z: [M+H]+ 706.8, HPLC retention time: 9.960 min) was obtained as a white solid in a similar manner to compound 1-p3 using compound 1-13-p3 (40 mg, 0.033 mmol) as starting material. ¹ H NMR (400 MHz, DMSO-db+D2O): δ 8.37 (s, 1H), 8.16 (s, 1H), 7.97 (s, 1H), 5.93 (d, J =7.8Hz, 1H), 5.74(d, J=8.0, 1H), 5.35(m, 1H), 5.14(m, 1H), 4.70(m, 1H) , 4.59 (s, 1H), 4.18 (s, 1H), 4.15 (m, 1H), 3.85 (m, 2H), 3.53 (m, 1H), 3.06 ( m, 1H); ³¹ P NMR (161 MHz, DMSO-d6+D2O): δ 58.47, 46.58.

Implementation Option 3. Synthesis of intermediates 2-8 and 2-9.

Ac0 'OAc

ΝΗΒς chlorotrimethylsilane U J ^{WATER Nb|I} solution to lithium hydroxide benzoyl chloride AcO ^- x q ν Ν pyridine \__/ acetonitrile

AcO' OAc

DMTrCI pyridine

DMTrO

H0 He

2-4

NHBz

T8SCI imidazole pyridine

dichloromethane/ I d^d pyridine 1

Step 1 To a solution of 7-aminothiazolo[4,5-d]pyrimidin-2(3H)-one (see J. Med. Chem. 1990, 33, 407415, compound 28) (6.5 g, 38.7 mmol) and tetraacetylribose (11 g, 46.4 mmol) in acetonitrile (120 ml)

- 34 041924 K,O-bis(trimethylsilyl)acetamide (BSA, 13.6 g, 116 mmol) was added and then stirred at reflux for 1 hour. Trimethylsilyl trifluoromethylsulfonate (TMSOTf, 17.2 g, 77.4 mmol) was added to the above reaction solution after cooling to room temperature, and stirred at reflux for an additional 48 hours. The reaction solution was cooled to room temperature, and saturated aqueous sodium bicarbonate solution, the mixture was extracted with ethyl acetate (150 mlx3), the combined organic layers were washed with water, the separated organic layer was dried over anhydrous sodium sulfate, filtered and concentrated. The residue was purified by flash column chromatography (ethyl acetate/petroleum ether=3/4) to give intermediate 2-1 (4.3 g) as a light yellow solid, m/z: [M+H]+ 427.0.

Step 2 To a solution of intermediate 2-1 (0.5 g, 1.17 mmol) in pyridine (5 mL) was added chlorotrimethylsilane (0.07 mL, 0.58 mmol) at 0° C. under nitrogen and stirred under nitrogen. over 5 minutes, benzoyl chloride (0.32 ml, 2.81 mmol) was added to the above reaction solution, the reaction system was stirred at room temperature overnight and then quenched by adding water (50 ml). The aqueous layer was extracted with ethyl acetate (50 mlx2), the combined organic layers were washed with water, and the separated organic layer was dried over anhydrous sodium sulfate, filtered and concentrated. The residue was purified by flash column chromatography (ethyl acetate/petroleum ether=9/10) to give intermediate 2-2 (0.45 g) as a light yellow solid, m/z: [M+H]+ 634.8.

Step 3 To a solution of intermediate 2-2 (3 g, 4.7 mmol) in acetonitrile (150 ml) was added an aqueous solution of lithium hydroxide (47 ml, 1 M), the reaction system was stirred at room temperature for 15 min, and then neutralized to pH 6 with hydrochloric acid (2 M). The reaction solution was concentrated to 1/3 of the total volume, the solid was precipitated, filtered off, the filter cake was washed 3 times with water, and then dried under vacuum to obtain intermediate compound 2-3 (1 g) as a yellow solid, m/ z: [M+H]+ 405.0.

Step 4 To a solution of intermediate 2-3 (7.6 g, 18.8 mmol) in pyridine (95 ml) was added 4,4'-dimethoxytrityl chloride (DMTrCl, 9.5 g, 28.5 mmol) under nitrogen atmosphere . The reaction system was stirred at room temperature overnight. The solvent was concentrated under reduced pressure. The residue was purified by flash column chromatography (methanol/dichloromethane=1/25) to give intermediate 2-4 (8.3 g) as an off-white solid, m/z: [M+H]+ 706.8 .

Step 5 To a solution of intermediate 2-4 (8.3 g, 11.7 mmol) and imidazole (2 g, 29.3 mmol) in pyridine (60 mL) was added tert-butyldimethylsilyl chloride (TBSCl, 2.1 g, 14 mmol) at 0°C under nitrogen atmosphere. The reaction system was stirred at room temperature overnight and then quenched by adding water (1 ml) and saturated aqueous sodium bicarbonate (100 ml), the aqueous layer was extracted with ethyl acetate (200 mlx2), the combined organic layers were washed with water, the organic layer was dried over anhydrous sodium sulfate, filtered and concentrated, the residue was purified by flash column chromatography (methanol/dichloromethane=1/50~1/20) to obtain intermediate compound 2-5 (4 g, off-white solid, retention time LCMS (Thermo ): 2.813 min) and 2-6 (2 g, off-white solid, retention time LCMS (Thermo): 2.763 min Intermediate 2-5: m/z: [M+H]+ 821.1; 1H NMR (400 MHz, DMSO-d6): δ 11.71 (s, 1H), 8.66 (s, 1H), 8.05-8.03 (m, 2H), 7.68-7.64 (m , 1H), 7.56-7.52 (m, 2H), 7.43-7.32 (m, 2H), 7.27-7.17 (m, 7H), 6.84-6.82 (m, 4H), 6.05-6.04 (m, 1H), 5.29-5.28 (m, 1H), 4.88-4.84 (m, 1H), 4.53-4 .50 (m, 1H), 4.05-3.98 (m, 1H), 3.71 (s, 6 H), 3.35-3.27 (m, 1H), 3.08-3.04 (m, 1H), 0.82 (s, 9H), 0.05 (s, 3H), 0.01 (s, 3H); Intermediate 2-6: m/z: [M+H]+ 821.1; 1H NMR (400 MHz, DMSOd6): δ 11.73 (s, 1H), 8.62 (s, 1H), 8.06-8.04 (m, 2H), 7.68-7.65 (m , 1H), 7.57-7.53 (m, 2H), 7.41-7.36 (m, 2H), 7.27-7.17 (m, 7H), 6.85-6.82 (m, 4H), 6.09-6.08 (m, 1H), 5.00-4.95 (m, 2H), 4.31-4.27 (m, 1H), 4.05-4 .03 (m, 1H), 3.72 (s, 6H), 3.23-3.16 (m, 2H), 0.95 (s, 9H), 0.05 (s, 6H).

Step 6 To a solution of intermediate 2-5 (0.5 g, 0.61 mmol) in pyridine (5 ml) was added diphenyl phosphite (0.57 g, 1.43 mmol) at 0°C under nitrogen atmosphere, the reaction system was stirred for 1 h, and then triethylamine (0.6 ml) and water (0.6 ml) were added thereto. The resulting mixture was stirred at room temperature for 5 minutes and then diluted with water (50 ml), the aqueous layer was extracted with dichloromethane (30 ml x 2), the combined organic layers were washed with water, and the separated organic layer was dried over anhydrous sodium sulfate, filtered and concentrated. The residue was purified by flash column chromatography (methanol/dichloromethane=1/10) to give intermediate 2-7 (triethylamine salt, 0.6 g) as an off-white solid, m/z: [M+H]+ 884.5.

Step 7 To a solution of intermediate 2-7 (0.6 g, 0.61 mmol) in a mixed solvent of dichloromethane (10 ml) and water (1 ml) was added a dichloromethane solution of dichloroacetic acid (DCA, 0.6 M, 9 .1 ml). The reaction mixture was stirred for 0.5 h, and then pyridine (20 ml) was added thereto, the resulting mixture was concentrated under reduced pressure to obtain intermediate compound 2-8 (pyridinium salt, crude product), m/z: [M+H]+582.9.

Synthesis of Intermediate 2-9: Intermediate 2-9 (pyridinium salt) was prepared in the same manner as Intermediate 2-8 using Intermediate 2-6 as starting material, m/z: [M+H]+ 583.0.

Implementation 4. Synthesis of compounds 2-p1, 2-p2, 2-p3 and 2-p4.

Step 1 Intermediates 2-8 (0.68 mmol) and 1-9 (0.81 g, 0.82 mmol) were azeotropically dehydrated twice with anhydrous acetonitrile (10 ml), respectively, and then dissolved in acetonitrile (5 ml ) respectively to use. To a solution of intermediate 2-8 in acetonitrile, which contained a type 4A molecular sieve, was slowly added a solution of intermediate 1-9 in acetonitrile at 0°C under nitrogen atmosphere, the resulting mixture was stirred for 1 hour. To the above reaction system was added (( dimethylaminomethylidene)amino)-3H1,2,4-dithiazolin-3-thione (DDTT, 0.16 g, 0.79 mmol) and stirred for an additional 1 h. The molecular sieve was removed by filtration and the filtrate was concentrated under reduced pressure to obtain compounds 2-9 (1 g), m/z: [M+H]+ 1501.5.

Step 2 To a solution of compound 2-9 (0.33 g, 0.22 mmol) in a mixed solvent of dichloromethane (3 ml) and water (0.3 ml) was added a dichloromethane solution of dichloroacetic acid (DCA, 0.6 M, 2.93 ml) under a nitrogen atmosphere. The reaction system was stirred for 0.5 h, then pyridine (1 ml) was added thereto and concentrated under reduced pressure, and the residue was purified by flash column chromatography (acetonitrile/ammonium bicarbonate aqueous solution (10 mmol/l)=60%) to obtain compound 2-10 (0.25 g, pyridinium salt) as a white solid, m/z: [M+H]+ 1199.6.

Step 3 and 4 Compound 2-10 (250 mg, 0.21 mmol) was azeotropically dehydrated three times with pyridine (1 ml) and then dissolved in a mixed solvent of pyridine (2 ml) and dichloromethane (2 ml). 2-Chloro-5,5-dimethyl-1,3,2-dioxaphosphorinan-2-oxide (DMOCP, 775 mg, 4.2 mmol) was added to the mixture, and the mixture was stirred at room temperature for 10 min to obtain a reaction solution of the compound 2-11. Water (756 mg, 42 mmol) and 3H-1,2-benzodithiol-3-one (71 mg, 0.42 mmol) were added directly to the above reaction solution of compound 2-11, and stirred at room temperature for 20 min, to an aqueous solution of sodium bicarbonate (2.7%, 50 ml) was added thereto. The aqueous layer was extracted with ethyl acetate and the separated organic layer was concentrated. The residue was purified by flash column chromatography (acetonitrile/ammonium bicarbonate aqueous solution (10 mmol/l) = 10-80%) to give compound 2-12-p1 (30 mg, LCMS retention time (Thermo): 2.147 min), 2 -12-p2 (35 mg, LCMS retention time (Thermo): 2.247 minutes) and 2-12-p3 (50 mg, LCMS retention time (Thermo): 2.327 minutes) as white solids.

Step 5. To a solution of compound 2-12-p1 (30 mg, 0.01 mmol) in acetonitrile (2 ml) was added tert-butylamine (2 ml), the reaction system was stirred at room temperature for 0.5 h, and then directly concentrated to obtain compound 2-13-p1 (30 mg, crude product), m/z: [M+H]+ 1159.9.

Step 6. To a solution of compound 2-13-p1 (30 mg, crude) in methanol (2 ml) was added ammonium hydroxide solution (2 ml), the reaction system was stirred at 45°C overnight in a sealed tube, and then the reaction solution concentrated under reduced pressure. The residue was azeotropically dehydrated three times with pyridine (2 ml) and then dissolved in pyridine (2 ml). Triethylamine (0.35 g, 3.5 mmol) and triethylamine trihydrofluoride (0.28 g, 1.75 mmol) were added to the above reaction system under nitrogen atmosphere. The resulting mixture was stirred at 45° C. for 6 hours. The solvent was concentrated under reduced pressure. The residue was adjusted to pH 8 with aqueous ammonium bicarbonate (1 M) and then purified by preparative HPLC (type 3 separation method) to give compound 2-p1 (diammonium salt, 2 mg, m/z: [M+H] + 723.8, HPLC retention time: 8.116 min) and 2-p2 (diammonium salt, 2 mg, m/z: [M+H]+ 723.8, HPLC retention time: 10.121 min) as white solids.

Synthesis of compound 2-p3.

To a solution of compound 2-12-p2 (30 mg, 0.01 mmol) in acetonitrile (2 ml) was added tert-butylamine (2 ml). The mixture was stirred at room temperature for 0.5 h and then concentrated under reduced pressure to give compound 2-13-p2. Ammonium hydroxide solution (2 ml) was added to a solution of compound 2-13-p2 in methanol (2 ml), the reaction system was stirred in a sealed tube at 45° C. overnight, and then the reaction solution was directly concentrated under reduced pressure. The residue was azeotropically dehydrated three times with pyridine (2 ml) and then dissolved in pyridine (2 ml). Triethylamine (0.35 g, 3.5 mmol) and triethylamine trihydrofluoride (0.28 g, 1.75 mmol) were added to the above reaction system under nitrogen atmosphere. The resulting mixture was stirred at 45° C. for 6 hours. The solvent was concentrated under reduced pressure. The residue was adjusted to pH 8 with aqueous ammonium bicarbonate (1 M) and then purified by preparative HPLC (type 3 separation method) to give compound 2-p3 (diammonium salt, 0.3 mg, m/z: [M+ H]+ 723.7, HPLC retention time: 10.121 min) as a white solid.

Synthesis of compound 2-p4.

Compound 2-p4 (diammonium salt, 0.4 mg, m/z: [M+H]+ 723.7, HPLC retention time: 9.632 min) was obtained as a white solid in a similar manner to compound 2-p3. using compound 2-12-p3 (17 mg, 0.01 mmol) as starting material.

Implementation variant 5. Synthesis of compounds 3-p1, 3-p1/3-p2 and 3-p3.

Step 1 Compound 3-1 (CAS number: 129451-95-8) (2.36 g, 2.4 mmol) and Intermediate 2-9 (1.05 g, 2 mmol) were azeotropically dehydrated twice with anhydrous acetonitrile (10 ml) respectively, and then dissolved in acetonitrile (5 ml) respectively for use. To a solution of intermediate 2-9 in acetonitrile, which contained a molecular sieve type 4A, was slowly added a solution of compound 3-1 in acetonitrile at 0°C under nitrogen atmosphere, the resulting mixture was stirred for 1 hour. To the above reaction system was added ((dimethylamino - 37 041924 methylidene)amino)-3H-1,2,4-dithiazolin-3-thione (DDTT, 0.49 g, 2.4 mmol) and stirred for an additional 1 hour. The molecular sieve was removed by filtration and the filtrate was concentrated under reduced pressure to obtain compound 3-3 (1.3 g), m/z: [M+H]+ 1501.5.

Step 2 To a solution of compound 3-3 (1 g, 0.66 mmol) in a mixed solvent of dichloromethane (3 ml) and water (0.3 ml) was added a dichloromethane solution of dichloroacetic acid (DCA, 0.6 M, 10 ml ). The reaction system was stirred for 0.5 h. Pyridine (3 ml) was added thereto, and the mixture was concentrated under reduced pressure. The residue was purified by flash column chromatography (acetonitrile/ammonium bicarbonate aqueous solution (10 mmol/l)=60%) to give compound 3-4 (390 mg, pyridinium salt) as a white solid, m/z: [M+ H]+ 1199.6.

Step 3 and 4 Compound 3-4 (250 mg, 0.21 mmol) was azeotropically dehydrated three times with pyridine (1 ml) and then dissolved in a mixed solvent of pyridine (2 ml) and dichloromethane (2 ml). 2-Chloro-5,5-dimethyl-1,3,2-dioxaphosphorinan 2-oxide (DMOCP, 775 mg, 4.2 mmol) was added to the above reaction system, the mixture was stirred at room temperature for 10 min to obtain a reaction solution of the compound 3-5. Water (756 mg, 42 mmol) and 3H-1,2-benzodithiol-3-one (71 mg, 0.42 mmol) were added to the reaction solution of compound 3-5 and stirred at room temperature for 20 min, and then to an aqueous sodium bicarbonate solution (2.7%, 50 ml) was added thereto, the aqueous layer was extracted with ethyl acetate, and the separated organic layer was concentrated. The residue was purified by flash column chromatography (acetonitrile/ammonium bicarbonate aqueous solution (10 mmol/l) = 10-80%) to give compound 3-6-p1 (50 mg, LCMS retention time (Thermo): 2.143 min), 3 -6-p2 (15 mg, LCMS retention time (Thermo): 2.230 min) and 3-6-p3 (18 mg, LCMS retention time (Thermo): 2.320) as white solids.

Step 5. To a solution of compound 3-6-p1 (20 mg, 0.02 mmol) in acetonitrile (2 ml) was added tert-butylamine (2 ml), the reaction system was stirred at room temperature for 0.5 h, and then directly concentrated to obtain compound 3-7-p1 (25 mg, crude product), m/z: [M+H]+ 1159.9.

Step 6 To a solution of compound 3-7-p1 (25 mg, crude) in methanol (2 ml) was added ammonium hydroxide solution (2 ml), the reaction system was stirred in a sealed tube at 45° C. overnight, and then the reaction solution was concentrated under reduced pressure. The residue was azeotropically dehydrated three times with pyridine (2 ml) and then dissolved in pyridine (2 ml). Triethylamine (0.35 g, 3.5 mmol) and triethylamine trihydrofluoride (0.28 g, 1.75 mmol) were added to the above reaction system under nitrogen atmosphere. The resulting mixture was stirred at 50° C. for 6 hours. The solvent was concentrated under reduced pressure. The residue was adjusted to pH 8 with aqueous ammonium bicarbonate (1 M) and then purified by preparative HPLC (type 3 separation method) to give compound 3-p1 (diammonium salt, 0.7 mg, m/z: [M+ H]+ 723.8, HPLC retention time: 9.726 min) and 3-p1/3-p2 mixture (diammonium salt, 1.1 mg, m/z: [M+H]+ 723.8, HPLC retention time: 9.726 min and 11.161 min).

Synthesis of compound 3-p3.

To a solution of compound 3-6-p3 (50 mg, 0.04 mmol) in acetonitrile (2 ml) was added tert-butylamine (2 ml). The mixture was stirred at room temperature for 0.5 h and then concentrated under reduced pressure to give compound 3-7-p3. Ammonium hydroxide solution (2 ml) was added to a methanol (2 ml) solution of compound 3-7-p3, the reaction system was stirred in a sealed tube at 45° C. overnight, and then the reaction solution was directly concentrated under reduced pressure. The residue was azeotropically dehydrated three times with pyridine (2 ml) and then dissolved in pyridine (2 ml). Triethylamine (0.35 g, 3.5 mmol) and triethylamine trihydrofluoride (0.28 g, 1.75 mmol) were added to the above reaction system under nitrogen atmosphere. The resulting mixture was stirred at 45° C. for 6 hours. The solvent was concentrated under reduced pressure. The residue was adjusted to pH 8 with aqueous ammonium bicarbonate (1 M) and then purified by preparative HPLC (type 3 separation method) to give compound 3-p3 (diammonium salt, 2.2 mg, m/z: [M+ H]+ 723.7, HPLC retention time: 11.76 min) as a white solid.

Embodiment 6. Synthesis of Intermediate 4-3.

Step 1 To a solution of 2'-fluoro-2'-deoxyadenosine (CAS number: 64183-27-3) (9.7 g, 36.0 mmol) in pyridine (110 ml) was added chlorotrimethylsilane (23.5 g, 216 mmol) at 0°C in a nitrogen atmosphere. The mixture was stirred at room temperature for 2 hours, and then benzoyl chloride (7.6 g, 54 mmol) was added thereto. The resulting mixture was stirred at room temperature overnight. Water (40 ml) was added to the reaction solution and stirred for 1 hour, and then an ammonium hydroxide solution (40 ml) was added thereto and stirred for an additional 2 hours. Additional water (40 ml) was added thereto, the mixture was extracted with ethyl acetate (500 ml x 2) and the combined organic layers were dried over anhydrous sodium sulfate, and then concentrated to 1/5 of the total volume, filtered, the filter cake was dried under vacuum to obtain intermediate compound 4-1 (10 g) as a white solid substances, m/z: [M+H]+ 374.0.

Step 2 To a solution of intermediate 4-1 (1.2 g, 3.2 mmol) in pyridine (15 ml) was added 4,4'-dimethoxytrityl chloride (DMTrCl, 1.6 g, 4.8 mmol) under nitrogen atmosphere . The reaction system was stirred at room temperature overnight. Water (50 ml) was added to the reaction solution, the mixture was extracted with ethyl acetate (40 ml×3), the combined organic layers were dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was purified by flash column chromatography (2% methanol/dichloromethane) to give intermediate 4-2 (1.9 g) as a light yellow solid, m/z: [M+H]+ 676.0.

Step 3 To a solution of intermediate 4-2 (1.35 g, 2 mmol) in dichloromethane (10 mL) was added X,L-diisopropylethylamine (1.1 mL, 6 mmol) and 2-cyanoethyl-X,L-diisopropylchlorophosphoramidite (947 mg, 4 mmol) at 0°C under nitrogen. The resulting mixture was stirred at room temperature for 2 hours. Water (50 ml) and a saturated sodium bicarbonate aqueous solution (20 ml) were added to the reaction solution. The resulting mixture was extracted with ethyl acetate (50 mlx3) and the combined organic layers were dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was purified by flash column chromatography (3% methanol/dichloromethane) to give intermediate 4-3 (1.2 g) as a yellow solid, m/z: [M+H]+ 876.1.

Embodiment 7. Synthesis of compounds 4-p1, 4-p2 and 4-p3.

Step 1: To a solution of Intermediate 4-4 (4-4 was prepared as a mixture of stereoisomers in a similar manner as Steps 1-4 of Embodiment 4, using Intermediates 4-3 and 2-8 as starting materials) (50 mg, 0.05 mmol) in acetonitrile (2 ml) was added tert-butylamine (2 ml). The resulting mixture was stirred at room temperature for 0.5 h and then concentrated under reduced pressure. The residue was purified by flash column chromatography (acetonitrile/ammonium bicarbonate aqueous solution (10 mmol/l)=10-80%) to give compound 4-5 (15 mg) as a white solid, m/z: [M+H]+ 1047.8.

Step 2. To a solution of compound 4-5 (30 mg, 0.03 mmol) in methanol (2 ml) was added an ammonium hydroxide solution (2 ml), the reaction solution was stirred in a sealed tube at 45° C. overnight, and then the solvent concentrated under reduced pressure. The residue was azeotropically dehydrated three times with anhydrous pyridine (2 ml) and then dissolved in pyridine (1 ml), and then triethylamine (0.91 g, 9 mmol) and triethylamine trihydrofluoride (0.58 g, 3 .6 mmol). The resulting mixture was stirred at 50° C. for 6 hours. The solvent was concentrated under reduced pressure. The residue was adjusted to pH 8 with ammonium hydroxide solution and then purified directly by preparative HPLC (type 3 separation method) to give compound 4-p1 (diammonium salt, 0.76 mg, m/z: [M+H]+ 725.8, HPLC retention time: 9.04 min), 4-p2 (diammonium salt, 0.82 mg, m/z: [M+H]+ 725.8, HPLC retention time: 10.45 min), and 4-p3 (diammonium salt, 0.97 mg, m/z: [M+H]+ 725.8, HPLC retention time: 10.35 min) as white solids.

Embodiment 8. Synthesis of compounds 5-p1, 5-p2, 5-p3 and 5-p4.

- 39 041924

Step 1 To a solution of compound 5-1-p1 (5-1-p1, 5-2-p2 and 5-3-p3 were prepared in the same manner as steps 1-4 of Embodiment 4 using intermediate 3-2 and 3'-tert-butyldimethylsilylisobutyrylguanosinephosphoramidite (3'-TBDMS-IBU-RGphosphoramidite) (CAS number: 1445905-51-0) as starting materials, LCMS retention times (Thermo) for 5-1-p1, 5-2-p2 and 5 -3-p3 was 2.11 min, 2.14 min and 2.31 min respectively) (30 mg, 0.03 mmol) in acetonitrile (2 ml) t-butylamine (2 ml) was added. The resulting mixture was stirred at room temperature for 0.5 h and then concentrated to give compound 5-2-p1 (30 mg, crude product), m/z: [M+H]+ 1141.6.

Step 2 To a solution of compound 5-2-p1 (30 mg, crude) in methanol (2 ml) was added ammonium hydroxide solution (2 ml), the reaction solution was stirred in a sealed tube at 45° C. overnight, and then the solvent was concentrated at reduced pressure. The residue was azeotropically dehydrated three times with anhydrous pyridine (2 ml) and then dissolved in pyridine (1 ml), and then triethylamine (0.35 g, 3.5 mmol) and triethylamine trihydrofluoride (0.28 g , 1.75 mmol). The resulting mixture was stirred at 50° C. for 6 hours. The solvent was concentrated under reduced pressure. The residue was adjusted to pH 8 with ammonium hydroxide solution and then purified directly by preparative HPLC (type 3 separation method) to give compound 5-p1 (diammonium salt, 0.89 mg, m/z: [M+H]+ 739 8, HPLC retention time: 6.495 min) as a white solid.

Synthesis of compounds 5-p2 and 5-p3.

Compound 5-p2 (diammonium salt, 13 mg, m/z: [M+H]+ 739.8, HPLC retention time: 10.666 min, ^1H NMR (400 MHz, DMSO-d6+D2O): δ 8.22 (s, 1H), 8.10 (s, 1H), 5.99 (d, J=7.7 Hz, 1H), 5.85 (d, J=8.5 Hz, 1H), 5.34 -5.28(m, 1H), 5.21-5.25(m, 1H), 5.12-5.16(m, 1H), 4.43-4.31(m, 1H), 4 .16-4.12 (m, 1H), 4.084.06 (m, 1H), 4.01-3.97 (m, 2H), 3.66 (d, J=11.8 Hz, 1H), 3.59-3.55 (m, 1H), ³¹ P NMR (162 MHz, DMSO^): δ 59.36, 57.52) and 5-p3 (diammonium salt, 0.7 mg, m/z: [M+H]+ 739.9, HPLC retention time: 10.663 min) were obtained as white solids in a similar manner to compound 5-p1 using compound 5-1-p2 (76.5 mg, crude) in as source material.

Synthesis of compound 5-p4.

Compound 5-p4 (diammonium salt, 1.41 mg, m/z: [M+H]+ 739.9, HPLC retention time: 11.973 min) was obtained as a white solid in a similar manner to compound 5-p1, using compound 5-1-p3 (100 mg, crude) as starting material.

Embodiment 9. Synthesis of Intermediate 6-14.

carbonate swept ^Ac0 ^V°x/ ^C iodine l "alium DMTrCI l sodium hydride

Υ Ύ _____AcO D__ / V- _____- BUT \ / , DMTrO D__ / V- _________y _v acetone < methanol - pyridine '-q 'χ DMF

AcO OAc ^AcO = . NO ^and NO υ

6-1 6-2 6-3

6-4

DCA

benzoyl chloride

6-5

6-6

OH-o' Yh

6-7

ΒζΟ

DMTrO dichloromethane tri and fluoroacetic acid ΒζΟ

Step 1: Tetraacetylribose (150 g, 472 mmol) was dissolved in acetone (1 L) and iodine (11.9 g, 47.2 mmol) was added thereto at 0°C. After the addition, the reaction system was stirred at room temperature for 12 hours. Saturated aqueous sodium bisulfate was added to the reaction solution to remove excess iodine, followed by extraction with ethyl acetate (500 mL×3), and the combined organic layers were washed with brine, the separated organic layer was dried over anhydrous sodium sulfate, the filtrate was concentrated under reduced pressure. The residue was purified by flash column chromatography (0-60% ethyl acetate/petroleum ether) to give Intermediate 6-1 (100 g) as a yellow oil, m/z: [M+H]+ 275.0.

Step 2 To a suspension of intermediate 6-1 (100 g, 365 mmol) in methanol (100 ml) was added potassium carbonate (150 g, 1.09 mmol). The reaction system was stirred at room temperature for 12 hours and then filtered, the filtrate was concentrated under reduced pressure. The residue was purified by flash column chromatography (0-10% methanol/dichloromethane) to give Intermediate 6-2 (60.3 g) as a white solid, m/z: [M+H]+ 191 ,0.

Step 3 To a solution of intermediate 6-2 (60 g, 316 mmol) in pyridine (300 ml) was added 4,4'-dimethoxytrityl chloride (DMTrCl, 128 g, 379 mmol) in small portions at 0°C under nitrogen atmosphere. The reaction system was stirred at room temperature for 12 h and then quenched by adding water (100 ml). The solvent was concentrated under reduced pressure. The residue was purified by flash column chromatography (0-50% petroleum ether/ethyl acetate) to give Intermediate 6-3 (150.3 g) as a white solid.

Step 4 To a solution of intermediate 6-3 (150 g, 305 mmol) in DMF (500 ml) was added sodium hydride (18.3 g, 458 mmol, 60%) under nitrogen at 0°C. The reaction system was stirred at 0°C for 0.5 h. Iodomethane (56.3 g, 396 mmol) was added to the reaction system and stirred at room temperature for 3 h. The mixture was diluted with water (100 ml) and ethyl acetate (300 ml x 3 ). The organic layer was washed with brine and then separated and dried over anhydrous sodium sulfate, the filtrate was concentrated under reduced pressure to give intermediate 6-4 (150 g) as a white solid, m/z: [M+H]+ 507, 1.

Step 5 To a solution of intermediate 6-4 (150 g, 296 mmol) in dichloromethane (500 ml) was added dichloroacetic acid (DCA, 344 g, 2.67 mol) at 0°C. The reaction system was stirred at 0°C for 3 h and then quenched by adding saturated aqueous sodium carbonate solution, the aqueous layer was extracted with dichloromethane (300 ml x3), the combined organic layers were washed with brine (150 ml x2), the separated organic layer was dried over anhydrous sodium sulfate, and then concentrated under reduced pressure. The residue was purified by flash column chromatography (0-10% methanol/dichloromethane) to give intermediate 6-5 (50.2 g) as a white solid, m/z: [M+H]+205.0.

Step 6 To a solution of intermediate 6-5 (50 g, 245 mmol) in dichloromethane (500 ml) was added benzoyl chloride (41.2 g, 294 mmol) at 0°C. The reaction system was stirred at room temperature for 12 h, and then the reaction was quenched by adding water, the aqueous layer was extracted with dichloromethane (150 ml x 3), the combined organic layers were washed with brine (150 ml x 2), the separated organic layer was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was purified by flash column chromatography (0-70% petroleum ether/ethyl acetate) to give intermediate 6-6 (65.3 g) as a white solid, m/z: [M+H]+309.0.

Step 7 Intermediate 6-6 (65 g, 211 mmol) was dissolved in an aqueous solution of trifluoroacetic acid (150 ml, 80%), the reaction system was stirred at room temperature for 5 hours. Most of the solvent was removed under reduced pressure, and then the reaction solution was washed with saturated sodium bicarbonate aqueous solution, the aqueous layer was extracted with dichloromethane (100 ml x 3), the organic layer was washed with brine (150 ml x 2), the separated organic layer was dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain intermediate 6-7 (50.2 g) as an off-white solid.

Step 8 To a solution of intermediate 6-7 (50 g, 187 mmol) in pyridine (150 ml) was slowly added acetic anhydride (114 g, 1.11 mol), the reaction system was stirred at room temperature for 12 hours. the solvent was removed under reduced pressure, and then the reaction solution was washed with saturated sodium bicarbonate aqueous solution, the aqueous layer was extracted with ethyl acetate (100 ml x 3), the combined organic layers were washed with brine (100 ml x 2), the separated organic layer was dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure. The residue was purified by flash column chromatography (0-70% petroleum ether/ethyl acetate) to give intermediate 6-8 (60.2 g) as an off-white solid.

Step 9 To a solution of 7-aminothiazolo[4,5-d]pyrimidin-2(3H)-one (6.5 g, 38.7 mmol) and intermediate 6-8 (16.3 g, 46.4 mmol ) in acetonitrile (120 mL) was added N,Obis(trimethylsilyl)acetamide (BSA, 13.6 g, 116 mmol), the reaction system was stirred at reflux for 1 hour. trimethylsilyl trifluoromethylsulfonate (TMSOTf, 17.2 g, 77.4 mmol) was added and stirred at reflux for an additional 72 h, and then after the reaction solution was cooled to room temperature, a saturated sodium bicarbonate aqueous solution was slowly added to the reaction solution, and after aqueous the layer was extracted with ethyl acetate (150 ml x3), the combined organic layers were washed with water, the separated organic layer was dried over anhydrous sodium sulfate, filtered and concentrated. The residue was purified by flash column chromatography (ethyl acetate/petroleum ether=3/4) to give intermediate 6-9 (5 g) as a light yellow solid, m/z: [M+H]+ 461.0.

Step 10 To a solution of intermediate 6-9 (5 g, 10.9 mmol) in pyridine (5 mL) was added benzoyl chloride (5.3 g, 38 mmol) at 0°C. The reaction system was stirred at room temperature overnight and then quenched by adding water (50 ml), the aqueous layer was extracted with dichloromethane (150 mlx3), the combined organic layers were washed with water, the separated organic layer was dried over anhydrous sodium sulfate, filtered and concentrated. The residue was purified by flash column chromatography (petroleum ether/ethyl acetate=1/1) to give intermediate 6-10 (6.8 g) as a white solid, m/z: [M+H]+ 669.1.

Step 11 To a solution of intermediate 6-10 (6.8 g, 10.1 mmol) in a mixed solvent of tetrahydrofuran and methanol (60/20 ml) was added an aqueous solution of lithium hydroxide (36 ml, 1 M) at 0°C , the reaction system was stirred at 0°C for 2 h, and then neutralized to pH 6 with acetic acid. The reaction solution was concentrated to 1/3 of the total volume, the solid was precipitated, filtered off, the filter cake was washed 3 times with water, and then dried under vacuum to obtain intermediate compound 6-11 (3.8 g) as a yellow solid, m/z: [M+H]+ 419.0.

Step 12 To a solution of intermediate 6-11 (3.8 g, 9.09 mmol) in pyridine (60 ml) was added 4,4'-dimethoxytrityl chloride (DMTrCl, 3.6 g, 10.9 mmol) under nitrogen atmosphere . The reaction system was stirred at room temperature overnight. The solvent was concentrated under reduced pressure. The residue was purified by flash column chromatography (petroleum ether/ethyl acetate=1/1) to give Intermediate 6-12 (6 g) as a light yellow solid.

Step 13 To a solution of intermediate 6-12 (5 g, 6.94 mmol) in pyridine (5 ml) was added diphenyl phosphite (4.9 g, 20.8 mmol), the reaction solution was stirred at room temperature for 1 h. Triethylamine (2 ml) and water (1 ml) were successively added thereto. The reaction solution was stirred at room temperature for 5 min and then diluted with water (50 ml) and extracted with dichloromethane (60 mlx3), the combined organic layers were washed with aqueous sodium bicarbonate solution (5%), the separated organic layer was dried over anhydrous sodium sulfate, filtered and concentrated. The residue was purified by flash column chromatography (methanol/dichloromethane=1/10) to give intermediate 6-13 (5.1 g, triethylamine salt) as an off-white solid, m/z: [M+H]+ 886.3.

Step 14 To a solution of intermediate 6-13 (3 g, 3.39 mmol) in dichloromethane (20 ml) was added a dichloromethane solution of dichloroacetic acid (DCA, 0.6 M, 50 ml) at 0°C. The reaction mixture was stirred at room temperature for 1 hour, and then triethylsilane (1 ml) and pyridine (3 ml) were added thereto, the resulting mixture was stirred at room temperature for an additional 10 minutes, the solvent was concentrated under reduced pressure to obtain intermediate 6 -14. (2.5 g, pyridinium salt).

Implementation 10. Synthesis of compounds 6-p1, 6-p2, 6-p3 and 6-p4.

Step 1 To a solution of Compound 6-15 (6-15 was prepared as a mixture of stereoisomers in a similar manner as Steps 1-4 of Embodiment 4 using intermediates 6-14 and 4-3 as starting materials) (50 mg , 0.05 mmol) in acetonitrile (2 ml) was added tert-butylamine (2 ml). The resulting mixture was stirred at room temperature for 0.5 h and then concentrated under reduced pressure to obtain compound 6-16 (40 mg). m/z: [M+H] + 948.0.

Step 2. To a solution of compound 6-16 (40 mg, crude) in methanol (1 ml) was added ammonium hydroxide solution (1 ml), the mixture was stirred in a sealed tube at 50°C for 5 h, and then cooled to room temperature and quenched by adding acetic acid, the mixture was lyophilized. The residue was purified by preparative HPLC (type 4 separation method) to give compound 6-p1 (diammonium salt, 3.6 mg, m/z: [M+H]+ 740.0, HPLC retention time: 10.814 min), 6- p2 (diammonium salt, 3.2 mg, m/z: [M+H]+ 740.0, HPLC retention time: 11.380 min), 6-p3 (diammonium salt, 3.9 mg, m/z: [M +H]+ 740.0, HPLC retention time: 10.370 min, 1H NMR (400 MHz, D2O): δ 8.36 (s, 2H), 8.12 (s, 1H), 6.61 (s, 1H ), 6.11 (s, 1H), 5.79 (s, 1H), 5.39 (d, J=51.6 Hz, 1H), 4.93-5.02 (m, 1H), 4 .36-4.48 (m, 3H), 3.84-4.03 (m, 4H), 3.51 (s, 3H), ³¹ P NMR (162 MHz, D ₂ O): δ 56.49 , 51.13; ¹⁹ F NMR (162 MHz, D2O): δ-202.92) and 6-p4 (diammonium salt, 2.3 mg, m/z: [M+H]+ 740.0, retention time HPLC: 11.650 min) as white solids.

Embodiment 11: Synthesis of Intermediate 7-1.

- 42 041924

ABOUT

IN %

HO—y _o N^N NHBz

4 ' h θ ⁺ Ru TBSO O-PH and O 7-1

Intermediate 7-1 was obtained as a yellow solid in a similar manner as Intermediate 2-8 of Embodiment 3 using tetraacetylribose and 5-aminothiazolo[4,5-d]pyrimidine-2,7(3H,6H)- dione (see J. Med. Chem. 1990, 33, 407-415, compound 4) as starting materials, m/z: [M+H]+ 599.1.

Embodiment 12. Synthesis of compounds 7-p1, 7-p2, 7-p3 and 7-p4.

Step 1 To a solution of Compound 7-2 (7-2 was prepared as a mixture of stereoisomers in a similar manner as Steps 1-4 of Embodiment 4, using intermediates 7-1 and 4-3 as starting materials) (330 mg , 0.3 mmol) in acetonitrile (2 ml) was added tert-butylamine (2 ml). The resulting mixture was stirred at room temperature for 1 hour and then concentrated under reduced pressure. The residue was purified by flash column chromatography (acetonitrile/ammonium bicarbonate aqueous solution (10 mmol/l)=0~60%) to obtain compound 7-3 (100 mg) as a white solid, m/z: [M+H ]+ 1063.8.

Step 2. To a solution of compound 7-3 (100 mg, 94 μmol) in methanol (2 ml) was added an ammonium hydroxide solution (2 ml), the reaction solution was stirred in a sealed tube at 55°C for 5 h, and then the solvent was concentrated under reduced pressure. The residue was subjected to azeotropic dehydration three times with anhydrous pyridine (2 ml) and then dissolved in pyridine (1 ml), and then triethylamine (1 ml) and triethylamine trihydrofluoride (0.5 ml) were added thereto under nitrogen atmosphere. The resulting mixture was stirred at 50° C. for 3 hours. The solvent was concentrated under reduced pressure. The residue was neutralized with ammonium hydroxide solution and then directly purified by preparative HPLC (type 5 separation method) to give compound 7-p1 (diammonium salt, 1.29 mg, m/z: [M+H]+ 741.3, HPLC retention time: 9.058 min), 7-p2 (diammonium salt, 4.41 mg, m/z: [M+H]+ 741.3, HPLC retention time: 9.590 min), 7-p3 (diammonium salt, 2 .3 mg, m/z: [M+H]+ 741.3, HPLC retention time: 10.438 min) and 7-p4 (diammonium salt, 24 mg, m/z: [M+H]+ 741.7, HPLC retention time: 10.929 min), as white solids.

Embodiment 13. Synthesis of compounds 8-p1, 8-p2, 8-p3 and 8-p4.

Step 1 To a solution of Compound 8-1 (8-1 was prepared as a mixture of stereoisomers in a similar manner as Steps 1-4 of Embodiment 4, using intermediates 2-9 and 4-3 as starting materials) (618 mg , 0.61 mmol) in acetonitrile (3 ml) was added tert-butylamine (3 ml). The resulting mixture was stirred at room temperature for 1 hour and then concentrated under reduced pressure. The residue was purified by flash column chromatography (acetonitrile/ammonium bicarbonate aqueous solution (10 mmol/L)=10~80%) to obtain compounds 82-p1 (67 mg), 8-2-p2 (40 mg), 8-2 -p3 (40 mg) and 8-2-p4 (87 mg) as white solids, m/z: [M+H]+ 1047.7.

Step 2. Ammonium hydroxide solution (1 ml) was added to a solution of compound 8-2-p1 (50 mg, 52 μmol) in methanol (1 ml), the reaction solution was stirred in a sealed test tube at 50°C in

- 43 041924 for 4 hours, and then the solvent was concentrated under reduced pressure. The residue was subjected to azeotropic dehydration three times with anhydrous pyridine (1 ml) and then dissolved in pyridine (1 ml), and then triethylamine (1 ml) and triethylamine trihydrofluoride (0.5 ml) were added thereto under nitrogen atmosphere. The resulting mixture was stirred at 55° C. for 4 hours. The solvent was concentrated under reduced pressure. The residue was neutralized with ammonium hydroxide solution and then directly purified by preparative HPLC (type 5 separation method) to give compound 8-p1 (diammonium salt, 7 mg, m/z: [M+H]+ 725.5, retention time HPLC: 11.702 min) as a white solid.

Synthesis of compound 8-p2.

Compound 8-p2 (diammonium salt, 1.98 mg, m/z: [M+H]+ 725.6, HPLC retention time: 10.602 min) was obtained as a white solid in a similar manner to compound 8-p1. using compound 8-2-p2 (40 mg, 0.04 mmol) as starting material.

Synthesis of compound 8-p3.

Compound 8-p3 (diammonium salt, 1.1 mg, m/z: [M+H]+ 726.1, HPLC retention time: 10.556 min) was obtained as a white solid in a similar manner to compound 8-p1. using compound 8-2-p3 (40 mg, 0.04 mmol) as starting material.

Synthesis of compound 8-p4.

Compound 8-p4 (diammonium salt, 7.5 mg, m/z: [[M+H] ⁺ 725.6, HPLC retention time: 12.102 min) was obtained as a white solid in a similar manner to compound 8-p1 . using compound 8-2-p4 (80 mg, 0.08 mmol) as starting material.

Embodiment 14: Synthesis of intermediates 9-4 and 9-5.

Step 1: A solution of isocarbostyril (25 g, 172 mmol), tetraacetylribose (137 g, 431 mmol) and N,Obis(trimethylsilyl)acetamide (BSA, 105 g, 517 mmol) in anhydrous acetonitrile was stirred at reflux for 1 h, and then after the reaction solution was cooled to room temperature, trimethylsilyl trifluoromethylsulfonate (TMSOTf, 62 mL, 344 mmol) was added thereto, the mixture was stirred at reflux for an additional 5 h, after the reaction solution was cooled to room temperature, saturated an aqueous sodium bicarbonate solution to adjust the pH to about 7, the aqueous layer was extracted with ethyl acetate (150 ml x3), the combined organic layers were washed with water, the separated organic layer was dried over anhydrous sodium sulfate, filtered and concentrated. The residue was purified by flash column chromatography (petroleum ether/ethyl acetate=2/1) to give intermediate 9-1 (41.1 g) as an orange oil, m/z: [M+H]+404.0.

Step 2 To a solution of intermediate 9-1 (41.1 g, 102 mmol) in acetonitrile (300 ml) was added an aqueous solution of lithium hydroxide (510 ml, 1 M), and then the reaction system was stirred at room temperature for 1. 5 h. Precipitated some white solid, filtered, the filter cake was washed 3 times with water, and then dried under vacuum to obtain intermediate compound 9-2 (18.5 g) as a white solid, m/z: [M +H]+278.0.

Step 3 Intermediate 9-2 (17.5 g, 63.1 mmol) was azeotropically dehydrated three times with anhydrous pyridine and then dissolved in pyridine (100 ml) under nitrogen. A pyridine solution of 4,4'-dimethoxytrityl chloride (DMTrCl, 22.5 g, 66.3 mmol, 50 ml) was added to the above reaction solution at 0°C. The resulting mixture was stirred at this temperature for 3 hours. The reaction was quenched by adding water (50 ml), the aqueous layer was extracted with ethyl acetate (100 ml x2) and the combined organic layers were concentrated under reduced pressure. The residue was purified by flash column chromatography (petroleum ether/ethyl acetate=1/1) to give intermediate 9-3 (32.6 g) as an off-white solid, m/z: [M+Na]+ 602, 0.

Step 4 Intermediate 9-3 (12 g, 20.7 mmol) and imidazole (4.9 g, 72.5 mmol) 3 times

- 44 041924 was subjected to azeotropic dehydration with anhydrous pyridine, and then dissolved in pyridine (50 ml) under nitrogen atmosphere, tert-butyldimethylsilyl chloride (TBSCl, 3.4 g, 22.8 mmol) was added to the mixture at 0°C. The reaction system was stirred at room temperature overnight and water (50 ml) was added thereto, the aqueous layer was extracted with ethyl acetate (100 ml x 2), the combined organic layers were washed with water, dried over anhydrous sodium sulfate, filtered and concentrated, the residue was purified by flash column -chromatography (petroleum ether/ethyl acetate=10/1~6/1) to give intermediates 9-4 (4.3 g, off-white solid, less polar) 9-5 (3.5 g, off-white solid, more polar). 9-4: m/z: [M+Na]+ 716.0; 1 H NMR (400 MHz, DMSO-d ₆ ): δ 8.28 (d, J=8.4 Hz, 1H), 7.71 (t, J=8.0 Hz, 2H), 7.65 (d , J=8.0 Hz, 1H), 7.53-7.45 (m, 3H), 7.36-7.32 (m, 6H), 7.26 (t, J=8.0 Hz, 1H), 6.93 (d, J=8.4 Hz, 4H), 6.46 (d, J=8.0 Hz, 1H), 6.29 (d, J=4.0 Hz, 1H) , 5.14 (d, J=6.0 Hz, 1H), 4.28-4.26 (m, 1H), 4.20-4.16 (m, 1H), 4.14-4.20 (m, 1H), 4.05-4.00 (m, 1H), 3.74 (s, 6H), 3.37-3.35 (m, 1H), 0.82 (m, 9H), 0.03 (s, 3H), 0.01 (s, 3H); 9-5: m/z: [M+Na]+ 716.0; ¹ H NMR (400 MHz, DMSO-de): δ 8.27 (d, J=8.0 Hz, 1H), 7.75-7.70 (m, 2H), 7.66-7.64 ( m, 1H), 7.52 (t, J=8.0 Hz, 1H), 7.45-7.43 (m, 2H), 7.35-7.25 (m, 7H), 6.92 (d, J=8.4 Hz, 4H), 6.50 (d, J=7.6 Hz, 1H), 6.27 (d, J=3.6 Hz, 1H), 5.39 (d , J=6.0 Hz, 1H), 4.28-4.26 (m, 1H), 4.05-4.00 (m, 2H), 3.74 (s, 6H), 3.46- 3.44 (m, 1H), 3.22-3.18 (m, 1H), 0.78 (m, 9H), 0.03 (s, 3H), 0.01 (s, 3H).

Embodiment 15. Synthesis of compounds 9-p1, 9-p2 and 9-p3.

Step 1 To a solution of intermediate 9-5 (3.1 g, 4.47 mmol) in pyridine (20 ml) was added diphenyl phosphite (3.1 g, 13.4 mmol) at 0° C. under nitrogen, the reaction system was stirred for 0.5 h, and then triethylamine (3 ml) and water (3 ml) were added thereto. The resulting mixture was stirred at room temperature for 5 min and then diluted with water (50 ml), extracted with propan-2-ol/chloroform (30 mlx2), the combined organic layers were washed with water, the separated organic layer was dried over anhydrous sodium sulfate, filtered and concentrated. The residue was purified by flash column chromatography (dichloromethane/methanol/triethylamine=100/5/1) to give compound 9-6 (4.5 g, triethylamine salt) as an off-white solid, m/z: [M- H] ^- 756.0.

Step 2 To a solution of compound 9-6 (3 g, 3.49 mmol) in a mixed solvent of dichloromethane (28 ml) and water (1 ml) was added a dichloromethane solution of dichloroacetic acid (DCA, 0.6 M, 46.5 ml ). The mixture was stirred at room temperature for 30 minutes, triethylsilane (28 ml) was added to the reaction solution, and stirred for an additional 1 hour. When the color of the reaction solution changed from brown to colorless, pyridine (28 ml) was added thereto, the solvent was concentrated under reduced pressure to obtain compound 9-7 (3 g, pyridinium salt, crude product). m/z: [M+H]+ 445.0.

Step 3 and 4 Compound 9-7 (3 g, crude) was dissolved in anhydrous acetonitrile (15 ml) and then concentrated under reduced pressure, repeating three times. The residue was dissolved in acetonitrile (50 ml) and a 4A type molecular sieve (1 g) was added thereto. 3'-tert-butyldimethylsilylisobutyrylguanosinephosphoramidite (CAS number: 1445905-51-0, 3.4 g, 3.49 mmol) was dissolved in anhydrous acetonitrile (15 mL) and concentrated under reduced pressure by repeating three

- 45 041924 times. The residue was dissolved in acetonitrile (20 ml) and a 4A type molecular sieve (2 g) was added thereto. A solution of 3'-tert-butyldimethylsilylisobutyrylguanosinephosphoramidite in acetonitrile was slowly added to a solution of 9-7 in acetonitrile at 0°C, the resulting mixture was stirred at room temperature for 0.5 h, and then ((dimethylaminomethylidene)amino)-3H1, 2,4-dithiazolin-3-thione (DDTT, 697 mg, 3.42 mmol) and stirred at room temperature for an additional 3 hours. The molecular sieve was removed by filtration. Water (1 ml) was added to the mixture, and then dichloroacetic acid (DCA, 3.6 g, 27.9 mmol) was slowly added dropwise, the resulting mixture was stirred at room temperature for 2 hours, triethylsilane (28 ml) was added thereto and stirred for an additional 1 hour, pyridine (28 ml) was added to the mixture, and the reaction solution was concentrated under reduced pressure. The residue was purified by flash column chromatography (acetonitrile/ammonium bicarbonate aqueous solution (10 mmol/l)=50%) to give compound 9-10 (680 mg) as a white solid, m/z: [M+H]+ 1053, 9.

Step 5 To a solution of compound 9-10 (200 mg, 0.19 mmol) in anhydrous pyridine (8 ml) was added 2-chloro-5,5-dimethyl-1,3,2-dioxaphosphorinan-2-oxide (DMOCP, 0.7 g, 3.8 mmol) per batch, the mixture was stirred at room temperature for 0.5 h, 3H-1,2-benzodithiol-3-one (38 mg, 0.23 mmol ) and then stirred at room temperature for 0.5 h, the reaction was quenched by adding an aqueous solution of sodium bicarbonate (2.7%). The mixture was extracted with ethyl acetate (50 ml x 2) and the combined organic layers were washed with brine, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was purified by preparative HPLC (type 5 separation method) to give compound 9-11-p1 (13.5 mg, LCMS retention time (Thermo): 2.023 min), 9-11-p2 (30.2 mg, LCMS retention time (Thermo): 2.157 min) and 9-11-p3 (38.0 mg, LCMS retention time (Thermo): 2.300 min) as white solids, m/z: [M+H]+ 1067.9.

Step 6 Compound 9-11-p1 (10 mg, 0.094 mmol) was dissolved in an ethanol solution of methylamine (1 ml, 30%) and stirred at room temperature for 1 h and then concentrated under reduced pressure, the residue was dissolved in pyridine ( 0.5 ml), triethylamine (0.5 ml) and triethylamine trihydrofluoride (0.25 ml) were added thereto, the reaction system was stirred at 50° C. for 5 hours and then concentrated under reduced pressure. The residue was diluted with methanol, and then an ammonium hydroxide solution was slowly added dropwise to reach a pH of 8~9. The resulting mixture was purified by preparative HPLC (type 5 separation method) to give compound 9-p1 (4.07 mg, HPLC retention time: 11.311 min; m/z: [M+H]+ 716.8; 1H NMR (400 MHz , DMSO-d ₆ ): δ 8.26 (d, J=6.0 Hz, 1H), 8.19 (s, 1H), 7.73 (t, J=7.6 Hz, 1H), 7 .68 (d, J=8.0 Hz, 1H), 7.53 (t, J=8.0 Hz, 1H), 7.42 (d, J=7.6 Hz, 1H), 6.69 (d, J=7.6 Hz, 1H), 6.51-6.47 (m, 2H), 6.22 (d, J=4.0 Hz, 1H), 5.90-5.82 ( m, 2H), 5.52-5.51 (m, 1H), 5.08-5.06 (m, 1H), 4.95-4.90 (m, 1H), 4.88-4, 86 (m, 1H), 4.65-4.61 (m, 1H), 4.57-4.55 (m, 1H), 4.40-4.36 (m, 2H), 4.25- 4.19 (m, 2H), 4.10-3.99 (m, 2H), 3.94-3.87 (m, 2H), ³¹ P NMR (162 MHz, DMSO-d ₆ ): δ 56 ,49, 54,10) as a white solid.

Synthesis of compounds 9-p2.

Compound 9-p2 (4.45 mg, HPLC retention time: 11.389 min; m/z: [M+H]+ 706.8; 1H NMR (400 MHz, DMSO-d ₆ ): δ 8.26 (d, J=8.0Hz, 1H), 8.02(s, 1H), 7.74(t, J=7.2Hz, 1H), 7.67(d, J=7.6Hz, 1H) , 7.53 (t, J=7.6 Hz, 1H), 7.37 (d, J=7.6 Hz, 1H), 6.91-6.34 (m, 4H), 5.87- 5.85 (m, 1H), 5.37-5.19 (m, 2H), 4.38-4.37 (m, 1H), 4.21-4.19 (m, 1H), 4. 11-4.05 (m, 2H), 3.74-3.70 (m, 1H), 3.01-2.99 (m, 8H), ³¹ P NMR (162 MHz, DMSOd ₆ ): δ 59 ,18, 56.65, 54.32, 47.74) was obtained as a white solid in a similar manner to compound 9-p1 using compound 9-11-p2 as starting material and purified by preparative HPLC ( separation method type 5).

Synthesis of compounds 9-p3.

Compound 9-p3 (7.0 mg, HPLC retention time: 10.912 min; m/z: [M+H]+ 716.8; 1H NMR (400 MHz, DMSO-d6): δ 8.26 (d, J =8.0 Hz, 1H), 7.97 (s, 1H), 7.73 (t, J=7.6 Hz, 1H), 7.65 (d, J=7.6 Hz, 1H), 7.53 (t, J=7.6 Hz, 1H), 7.44 (d, J=7.6 Hz, 1H), 6.62 (d, J=8.0 Hz, 1H), 6, 56-6.52 (m, 1H), 6.33 (d, J=6.8 Hz, 1H), 5.86 (d, J=8.4 Hz, 1H), 5.27-5.23 (m, 1H), 5.06-5.04 (m, 2H), 4.46-4.40 (m, 2H), 4.22-4.18 (m, 1H), 4.12-4 .08 (m, 2H), 3.96-3.92 (m, 2H), 3.78-3.75 (m, 1H), 3.01-2.95 (m, 4H), ³¹ P NMR (162 MHz, DMSO^): δ 57.77, 50.27) was obtained as a white solid in a similar manner to compound 9-p1 using compound 9-11-p3 as starting material and purified by preparative HPLC (Type 5 Separation Method).

Embodiment 16. Synthesis of compounds 10-p1, 10-p2 and 10-p3.

Synthesis of intermediates 10-4 and 10-5.

- 46 041924

Intermediates 10-4 (2.0 g, more polar) and 10-5 (3.4 g, less polar) were obtained as white solids in a similar manner to compounds 9-4 and 9-5 using quinazoline -4(3H)-one as starting material, m/z: [M+H]+ 695.0; 10-4: 1H NMR (400 MHz, DMSO-d ₆ ):

δ 8.50 (s, 1H), 8.16 (d, J=8.0 Hz, 2H), 7.86-7.82 (m, 1H), 7.67 (d, J=8.0 Hz, 2H), 7.58-7.54 (m, 1H), 7.40 (d, J=8.0 Hz, 2H), 7.31-7.22 (m, 7H), 6.89 -6.87 (m, 4H), 6.09 (d, J=4.0 Hz, 1H), 5.10 (d, J=8.0 Hz, 1H), 4.38-4.36 ( m, 1H), 4.13-4.09 (m, 2H), 3.72 (s, 6H), 0.81 (s, 9H) 0.01-0.00 (s, 6H); 10-5: 1H NMR (400 MHz, DMSO-d ₆ ): δ 8.53 (s, 1H), 8.15 (d, J=8.0 Hz, 1H), 7.86-7.82 ( m, 1H), 7.67 (d, J=8.0 Hz, 1H), 7.58-7.54 (m, 1H), 7.41-7.39 (m, 2H), 7.30 -7.19 (m, 7H), 6.85 (d, J=8.0 Hz, 4H), 6.06-6.05 (m, 1H), 5.36 (d, J=8.0 Hz, 1H), 4.31-4.26 (m, 2H), 4.02-4.00 (m, 1H), 3.71 (s, 6H), 0.79 (s, 9H), 0 .02-0.00 (s, 6H).

Synthesis of compounds 10-11-p1, 10-11-p2 and 10-11-p3: compounds 10-11-p1 (7.0 mg, retention time LCMS (Thermo): 2.020 min), 10-11-p2 (26 mg, LCMS retention time (Thermo): 2.170 min) and 10-11p3 (19 mg, LCMS retention time (Thermo): 2.350 min) were obtained as white solids in a similar manner to steps 1-5 of Embodiment 15, with using Compound 10-5 as starting material, and purified by preparative HPLC (Type 6 separation method). m/z: [M+H]+ 1068.9.

Synthesis of compound 10-p1.

An ethanol solution of methylamine (1 ml, 30%) was added to compound 10-11-p1 (7.0 mg, 0.006 mmol), the reaction system was stirred at room temperature for 1 h and then concentrated under reduced pressure, the residue was dissolved in anhydrous pyridine (0.5 ml), triethylamine (0.5 ml) and triethylamine trihydrofluoride (0.25 ml) were added thereto under nitrogen atmosphere, the reaction solution was stirred at 50° C. for 1 hour. The solvent was concentrated under reduced pressure, the residue diluted with methanol, and then slowly added dropwise ammonium hydroxide solution to achieve a pH of 8~9. The mixture was purified by preparative HPLC (type 4 separation method) to give compound 10-p1 (0.73 mg, HPLC retention time: 10.671 min, m/z: [M+H]+ 717.8) as a white solid.

Synthesis of compound 10-p2.

Compound 10-p2 (7.10 mg, HPLC retention time: 10.826 min; m/z: [M+H]+ 717.8; 1H NMR (400 MHz, DMSO-de): δ 8.41 (s, 1H ), 8.25-8.20 (m, 1H), 8.03 (s, 1H), 7.89-7.85 (m, 1H), 7.72-6.69 (m, 1H), 7.61-7.57(m, 1H), 7.61-7.57(m, 1H), 6.52-6.48(m, 2H), 6.07-6.00(m, 1H ), 5.91-5.79 (m, 1H), 0.56 (s, 1H), 5.34-5.30 (m, 1H), 5.20-5.19 (m, 1H), 5.09-5.01 (m, 1H), 4.70-4.64 (m, 1H), 4.36-4.33 (m, 1H), 4.27-4.18 (m, 1H ), 4.14-4.05 (m, 2H), 3.95-3.70 (m, 3H), 1.24 (s, 2H)) was obtained as a white solid in a similar manner to compound 10 -p1 using compound 10-11-p2 as starting material and purified by preparative HPLC (type 4 separation method).

Synthesis of compound 10-p3.

Compound 10-p3 (5.46 mg, HPLC retention time: 10.925 min; m/z: [M+H]+ 717.8; 1H NMR (400 MHz, DMSO-d ₆ ): δ 8.44 (s, 1H), 8.22-8.20(m, 1H), 7.99(s, 1H), 7.89-7.85(m, 1H), 7.68(d, J=8.0 Hz , 1H), 7.61-7.57 (m, 1H), 6.56 (s, 2H), 6.09 (d, J=4.0 Hz, 2H), 5.83 (d, J= 8.0 Hz, 1H), 5.34-5.32(m, 1H), 5.26(s, 1H), 5.08(m, 1H), 4.71-4.68(m, 1H), 4.46-4.45(m, 1H), 4.25(m, 1H), 4.14-3.95(m, 5H), 3.79-3.76(m, 1H) , 1.24 (s, 2H)) was obtained as a white solid in a similar manner to compound 10-p1 using compound 10-11-p3 as starting material and purified by preparative HPLC (type 4 separation method).

- 47 041924

Embodiment 17. Synthesis of compounds 11-p1, 11-p2, 11-p3 and 11-p4.

Synthesis of intermediates 11-4 and 11-5.

Synthesis of compounds 11-4 and 11-5: compounds 11-4 (650 mg, more polar) and 11-5 (610 mg, less polar) were obtained as white solids in a similar manner as compounds 9-4 and 9- 5 using pyrido[2,3-d]pyrimidin-4-one as starting material, m/z: [M+H]+ 696.0; 114: ¹ H NMR (400 MHz, DMSO-d ₆ ): δ 8.97 (d, J=4.0 Hz, 1H), 8.74 (s, 1H), 8.56 (d, J=8 .0 Hz, 1H), 7.60-7.57. (m, 1H), 7.42 (d, J=8.0 Hz, 2H), 7.31-7.20 (m, 1H), 6.88 (d, J=12.0 Hz, 4H) , 6.02 (s, 1H), 5.40 (d, J=4.0 Hz, 1H), 4.30-4.29 (m, 2H), 4.01-3.98 (m, 1H ), 3.71 (s, 6H), 3.40-3.37 (m, 1H), 3.21-3.18 (m, 1H) 0.74 (s, 9H), 0.00 (s , 6H); 11-5: 1H NMR (400 MHz, DMSO-de): δ 8.94 (d, J=4.0 Hz, 1H), 8.67 (s, 1H), 8.55 (d, J=8 .0 Hz, 1H), 7.567.53. (m, 1H), 7.41 (d, J=8.0 Hz, 2H), 7.29-7.25 (m, 1H), 6.86 (d, J=8.0 Hz, 4H) , 6.00 (s, 1H), 5.13 (d, J=4.0 Hz, 1H), 4.13-4.08 (m, 2H), 4.00-3.98 (m, 1H ), 3.39 (s, 6H), 3.36-3.32 (m, 1H), 3.27-3.25 (m, 1H) 0.79 (s, 9H), 0.00 (s , 6H).

Synthesis of compounds 11-11-p1, 11-11-p2, 11-11-p3 and 11-11-p4: compounds 11-11-p1 (27 mg), 11-11-p2 (15 mg), 11-11 -p3 (19 mg) and 11-11-p4 (36 mg) were obtained as white solids in a similar manner to steps 1-5 of Embodiment 15 using compound 11-5 as starting material and purified by preparative HPLC (Type 6 Separation Method).

Synthesis of compound 11-p1.

Ethanol solution of methylamine (2 ml, 30%) was added to compound 11-11-p1 (20 mg, 0.019 mmol), the reaction system was stirred at room temperature for 10 hours and then concentrated under reduced pressure, the residue was dissolved in anhydrous pyridine ( 0.5 ml), triethylamine (0.5 ml) and triethylamine trihydrofluoride (0.25 ml) were added thereto under nitrogen atmosphere, the reaction solution was stirred at 50° C. for 1 hour. The solvent was concentrated under reduced pressure. The residue was diluted with methanol, and then ammonium hydroxide solution was slowly added to lower the pH to 8~9. The mixture was purified by preparative HPLC (type 7 separation method) to give compound 11-p1 (2.45 mg, HPLC retention time: 8.963 min, m/z: [M+H] ⁺ 718.8) as a white solid.

Synthesis of compound 11-p2.

Compound 11-p2 (3.07 mg, HPLC retention time: 8.527 min, m/z: [M+H] ⁺ 718.7) was obtained as a white solid in a similar manner to compound 11-p1 using compound 11-11-p2 as starting material and purified by preparative HPLC (type 5 separation method).

Synthesis of compound 11-p3.

Compound 11-p3 (1.28 mg, HPLC retention time: 9.103 min, m/z: [M+H] ⁺ 719.1) was obtained as a white solid in a similar manner to compound 11-p1 using compound 11-11-p3 as starting material and purified by preparative HPLC (type 5 separation method).

Synthesis of compound 11-p4.

Compound 11-p4 (3.03 mg, HPLC retention time: 9.403 min, m/z: [M+H] ⁺ 718.6) was obtained as a white solid in a similar manner to compound 11-p1 using compound 11-11-p4 as starting material and purified by preparative HPLC (type 5 separation method).

- 48 041924

Embodiment 18: Synthesis of Intermediate 12-1.

Step 1 2,6-dichloronicotinonitrile (25 g, 145 mmol) and ammonium hydroxide solution (250 ml) were charged into a sealed tube. The reaction system was stirred at 120° C. for 48 h and then cooled to 10° C., the solid was filtered off and washed with cold water, the filter cake was dried under vacuum to give 2,6-diaminonicotinonitrile (12.5 g) as light yellow solids, m/z: [M+H]+ 135.0.

Step 2 To a solution of 2,6-diaminonicotinonitrile (12.5 g, 93.2 mmol) in acetic acid (120 ml) was added slowly concentrated sulfuric acid (3 ml), the reaction system was stirred at reflux for 9 hours and then cooled to room temperature and concentrated under reduced pressure. The residue was triturated with ammonium hydroxide solution, the solid was filtered off, the filter cake was washed with cold water and dried under vacuum to give 7-aminopyrido[2,3-d]pyrimidin-4(3H)-one (15.1 g) as light -yellow solid, m/z: [M+H]+ 163.0.

Step 3. 7-aminopyrido[2,3-d]pyrimidin-4(3H)-one (15.1 g, 93.1 mmol) was dissolved in pyridine (1.5 L) and isobutyryl chloride (29. 7 g, 279 mmol), the reaction system was stirred at room temperature for 18 h, and then water (1.5 L) was added thereto, the reaction solution was extracted with chloroform/propan-2-ol (1.5 L), the organic layer concentrated under reduced pressure. The residue was purified by flash column chromatography (0-6.2% methanol/dichloromethane) to give compound 12-1 (11.9 g) as a white solid, m/z: [M+H]+ 233.1.

Embodiment 19. Synthesis of compounds 12-p1, 12-p2, 12-p3 and 12-p4.

Synthesis of intermediates 12-4 and 12-5.

Intermediates 12-4 (1.73 g, more polar) and 12-5 (1.1 g, less polar) were obtained as white solids in a similar manner to compounds 9-4 and 9-5 using the compound 12-1 as starting material, m/z: [M+H]+ 781.0; 12-4: 1H NMR (400 MHz, DMSO-d ₆ ): δ 11.15 (s, 1H), 8.70 (s, 1H), 8.52 (d, J=8.8 Hz, 1H) , 8.33 (d, J=8.8 Hz, 1H), 7.43 (d, J=7.6 Hz, 2H), 7.32 (d, J=7.6 Hz, 2H), 7 .29 (d, J=8.4 Hz, 4H), 6.89 (d, J=8.4 Hz, 4H), 6.03 (d, J=2.8 Hz, 1H), 5.40 (d, J=6.0 Hz, 1H), 4.30 (t, J=3.2 Hz, 1H), 4.05-4.02 (m, 2H), 3.73 (s, 6H) , 3.40-3.36(m, 1H), 3.25-3.22(m, 1H), 2.83-2.79(m, 1H), 1.12(s, 3H), 1 .11 (s, 3H), 0.77 (s, 9H), 0.03 (s, 3H), -0.04 (s, 3H); 12-5: Ή NMR (400 MHz, DMSO^): δ 11.13 (s, 1H), 8.67 (s, 1H), 8.54 (d, J=8.8 Hz, 1H), 8 .31 (d, J=8.8 Hz, 1H), 7.45 (d, J=7.6 Hz, 2H), 7.33 (d, J=7.6 Hz, 2H), 7, 31 (d, J=8.4 Hz, 4H), 6.90 (d, J=8.4 Hz, 4H), 6.04 (d, J=2.8 Hz, 1H), 5.16 ( d, J=6.0 Hz, 1H), 4.40 (t, J=3.2 Hz, 1H), 4.17-4.12 (m, 2H), 3.74 (s, 6H), 3.43-3.40 (m, 1H), 3.30-3.27 (m, 1H), 2.82-2.79 (m, 1H), 1.13 (s, 3H), 1, 11 (s, 3H), 0.85 (s, 9H), 0.05 (s, 6H).

Synthesis of compounds 12-11-p1, 12-11-p2, 12-11-p3 and 12-11-p4: compounds 12-11-p1 (25 mg), 12-11-p2 (7 mg), 12-11 -p3 (40 mg) and 12-11-p4 (32 mg) were obtained as white solids in a similar manner to steps 1-5 of Embodiment 15 using compound 12-5 as starting material, and purified by preparative HPLC (type 6 separation method).

Synthesis of compound 12-p1.

An ethanol solution of methylamine (2 ml, 30%) was added to compound 12-11-p1 (25 mg, 19 μmol), the reaction system was stirred at room temperature for 3 h and then concentrated under reduced pressure, the residue was dissolved in anhydrous pyridine ( 0.5 ml), triethylamine (0.5 ml) and triethylamine trihydrofluoride (0.25 ml) were added thereto under nitrogen atmosphere, the reaction solution was stirred at 50° C. for 1 hour. The solvent was concentrated under reduced pressure. The residue was diluted with methanol, and then an ammonium hydroxide solution was slowly added dropwise to reach a pH of 8~9. The mixture was purified by preparative HPLC (type 4 separation method) to give compound 12-p1 (0.9 mg, HPLC retention time: 8.891 min, m/z: [M+H]+ 733.7) as a white solid.

Synthesis of compound 12-p2.

Compound 12-p2 (0.5 mg, HPLC retention time: 8.571 min, m/z: [M+H]+ 733.7) was obtained as a white solid in a similar manner to compound 12-p1 using compound 12-11-p2 as starting material and purified by preparative HPLC (Type 4 separation method).

Synthesis of compound 12-p3.

Compound 12-p3 (1.5 mg, HPLC retention time: 10.020 min, m/z: [M+H]+ 733.8) was obtained as a white solid in a similar manner to compound 12-p1 using compound 12-11-p3 as starting material and purified by preparative HPLC (Type 4 separation method).

Synthesis of compound 12-p4.

Compound 12-p4 (3.07 mg, HPLC retention time: 9.468 min, m/z: [M+H]+ 733.8) was obtained as a white solid in a similar manner to compound 12-p1 using compound 12-11-p4 as starting material and purified by preparative HPLC (Type 4 separation method)

Embodiment 20: Synthesis of intermediates 13-4 and 13-5.

Synthesis of imidazo[1,2-c]pyrimidin-5(6H)-one: suspension of cytosine (18 g, 0.16 mol), aqueous solution of chloroacetaldehyde (63.6 g, 0.32 mol, 40%) and sodium acetate (32.8 g, 0.40 mol) in water (180 ml) was stirred at 80°C for 3 h, and then the reaction system was cooled to 10°C, the solid was filtered off and washed with cold water, the filter cake was dried under vacuum to obtain imidazo[1,2-c]pyrimidin-5(6H)-one (17.8 g) as a brown solid, m/z: [M+H]+ 136.0.

Step 1. To a suspension of imidazo[1,2-c]pyrimidin-5(6H)-one (17.8 g, 0.13 mmol), tetraacetylribose (46.1 g, 0.14 mmol) and N,O- bis(trimethylsilyl)acetamide (BSA, 39.7 g, 0.19 mmol) in acetonitrile (200 ml) was added dropwise trimethylsilyl trifluoromethylsulfonate (TMSOTf, 43.3 g, 0.19 mol) at 0°C, the reaction system was stirred at 60°C for 3 h, and then concentrated under reduced pressure to obtain compound 13-1 (100 g, crude product) as a brown oil, m/z: [M+H]+ 394.0.

Step 2 Compound 13-1 (45 g, crude) was dissolved in methanol (100 ml), ammonia in methanol (81.7 ml, 7 M) was added slowly, the reaction system was stirred at room temperature overnight, solid filtered and washed with methanol, the filter cake was dried under vacuum to obtain compound 13-2 (8 g) as a brown solid, m/z: [M+H]+ 268.0.

Step 3 Compound 13-2 (12.5 g, 5.76 mmol) was azeotropically dehydrated three times with anhydrous pyridine (20 ml) and then dissolved in pyridine (100 ml). To the above reaction solution, 4,4'-dimethoxytrityl chloride (DMTrCl, 19.52 g, 5.76 mmol) was added in small portions at 0~5° C. under a nitrogen atmosphere. The reaction system was stirred at room temperature for

- 50 041924 nights, the solvent was concentrated under reduced pressure and the residue was diluted with chloroform (100 ml). The organic layer was washed with saturated aqueous sodium bicarbonate (30 ml x2), the organic layer was separated and dried over anhydrous sodium sulfate, filtered and concentrated. The residue was purified by flash column chromatography (methanol/dichloromethane=0-3%) to give compound 13-3 (16.65 g) as a brown solid, m/z: [M+Na]+ 570.0.

Step 4 Compound 13-3 (5.38 g, 9.44 mmol) was azeotropically dehydrated three times with anhydrous pyridine (20 ml) and then dissolved in pyridine (60 ml). To the mixture were added tert-butyldimethylsilyl chloride (TBSCl, 2.13 g, 14.2 mmol) and imidazole (2.25 g, 33.1 mmol) at 0° C. under a nitrogen atmosphere. The reaction system was stirred at room temperature overnight, the solvent was concentrated under reduced pressure, and the residue was diluted with chloroform (100 ml). The organic layer was washed with saturated aqueous sodium bicarbonate (30 mlx2), the separated organic layer was dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by flash column chromatography (0-3% methanol/dichloromethane) to give compounds 13-4 (1.32 g, less polar) and 13-5 (3.68 g, more polar) as white foamy solids. , m/z: [M+H]+ 684.0; 13-4: ¹ H NMR (400 MHz, DMSOd6): δ 7.88 (d, J=5.6 Hz, 1H), 7.82 (s, 1H), 7.76 (s, 1H), 7 .40-7.26 (m, 9H), 6.92-6.90 (m, 4H), 6.68 (d, J=6.0 Hz, 1H), 5.90 (d, J=9 .2 Hz, 1H), 5.33 (d, J=4.8 Hz, 1H), 4.50 (t, J=4.4 Hz, 1H), 4.15-4.14 (m, 2H ), 3.74 (s, 6H), 3.28-3.26 (m, 2H), 0.77 (s, 9H), 0.02 (s, 3H), -0.12 (s, 3H ). 13-5: 1H NMR (400 MHz, DMSO-d ₆ ): δ 7.92 (d, J=6.0 Hz, 1H), 7.78 (s, 2H), 7.36-7.21 ( m, 9H), 6.88-6.86 (m, 4H), 6.68 (d, J=8.8 Hz, 1H), 5.81 (d, J=4.8 Hz, 1H), 5.63 (d, J=6.0 Hz, 1H), 4.40-4.35 (m, 2H), 4.27 (t, J=4.8 Hz, 1H), 4.06-4 .04 (m, 1H), 3.74 (s, 6H), 3.20-3.16 (m, 1H), 0.80 (s, 9H), 0.06 (s, 3H), 0. 01 (s, 3H)

Embodiment 21. Synthesis of compounds 13-p1, 13-p2, 13-p3 and 13-p4.

Synthesis of compounds 13-11-p1, 13-11-p2, 13-11-p3 and 13-11-p4: compounds 13-11-p1 (55 mg, retention time LCMS (Thermo): 1.874, 1.944 min), 13-11-p2 (25 mg, LCMS retention time (Thermo): 2.030 minutes) and 13-11-p3 (11 mg, LCMS retention time (Thermo): 2.170 minutes) were obtained as white solids in a similar manner as Steps 1-5 of Embodiment 15 using Compound 13-5 as starting material and purified by flash column chromatography (acetonitrile/ammonium bicarbonate aqueous solution (10 mmol/L)=40%). m/z: [M+H]+ 1057.9.

Synthesis of compounds 13-p1 and 13-p2.

Compound 13-11-p1 (43 mg, 0.04 mmol) was dissolved in an ethanol solution of methylamine (3 ml, 30%) and stirred at room temperature for 3 h, and then concentrated under reduced pressure, the residue was dissolved in anhydrous pyridine ( 0.5 ml), triethylamine (0.5 ml) and triethylamine trihydrofluoride (0.25 ml) were added thereto under nitrogen atmosphere, the reaction solution was stirred at 50° C. for 3 hours and then concentrated under reduced pressure. The residue was diluted with methanol and ammonium hydroxide solution was slowly added dropwise to reach pH 8~9. The mixture was purified by preparative HPLC (type 5 separation method) to give compounds 13-p1 (2.28 mg, HPLC retention time: 6.048 min, m/z: [M+H]+ 706.8) and 13-p2 (0 .68 mg, HPLC retention time: 7.781 min, m/z: [M+H]+ 706.8) as white solids.

Synthesis of compound 13-p3.

Compound 13-p3 (3.82 mg, HPLC retention time: 6.445 min, m/z: [M+H]+ 706.8) was obtained as a white solid in a similar manner to compounds 13-p1/13-p2 , using compound 13-11-p2 as starting material, and purified by preparative HPLC (type 4 separation method).

Synthesis of compound 13-p4.

Compound 13-p4 (4.03 mg, HPLC retention time: 10.395 min, m/z: [M+H]+ 706.8) was obtained as a white solid in a similar manner to compounds 13-p1/13-p2 , using compound 13-11-p3 as starting material, and purified by preparative HPLC (type 5 separation method).

- 51 041924

Embodiment 22: Synthesis of compounds 14-p1, 14-p2, 14-p3 and 14-p4.

Synthesis of intermediates 14-4 and 14-5.

Intermediates 14-4 (1.3 g, more polar) and 14-5 (0.56 g, less polar) were obtained as white solids in a similar manner to compounds 13-4 and 13-5 using imidazo [5,1-f][1,2,4]triazin-4(3H)-one (CAS No.: 865444-76-4) as starting material, m/z: [M+H]+ 685.0 .

Synthesis of compounds 14-11-p1, 14-11-p2, 14-11-p3 and 14-11-p4: compounds 14-11-p1 (5 mg, retention time LCMS (Thermo): 1.850 min), 14-11 -p2 (17 mg, LCMS retention time (Thermo): 1.880 min), 14-11-p3 (22 mg, LCMS retention time (Thermo): 2.020 min) and 14-11-p4 (18 mg, LCMS retention time ( Thermo): 2.140 min) was obtained as white solids in a similar manner to steps 1-5 of Embodiment 15 using compound 14-5 as starting material and purified by preparative HPLC (type 7 separation method). m/z: [M+H]+ 1059.0.

Synthesis of compound 14-p1.

Compound 14-11-p1 (5.0 mg, 0.05 mmol) was dissolved in an ethanol solution of methylamine (3.0 ml, 30%), the mixture was stirred at room temperature for 3 h, and then concentrated under reduced pressure, the residue was dissolved in anhydrous pyridine (0.5 ml), triethylamine (0.5 ml) and triethylamine trihydrofluoride (0.3 ml) were added thereto under nitrogen atmosphere, the reaction solution was stirred at 50° C. for 2 hours. The solvent was concentrated under reduced pressure. The residue was diluted with methanol, and then an ammonium hydroxide solution was slowly added dropwise to reach a pH of 8~9. The mixture was purified by preparative HPLC (type 5 separation method) to give compound 14-p1 (3.65 mg, HPLC retention time: 1.914 min, m/z: [M+H]+ 707.5) as a white solid .

Synthesis of compound 14-p2.

Compound 14-p2 (17.5 mg, HPLC retention time: 3.123 min, m/z: [M+H]+ 707.5) was obtained as a white solid in a similar manner to compound 14-p1 using compound 14-11-p2 as starting material and purified by preparative HPLC (Type 4 separation method).

Synthesis of compound 14-p3.

Compound 14-p3 (1.04 mg, HPLC retention time: 3.193 min, m/z: [M+H]+ 707.5) was obtained as a white solid in a similar manner to compound 14-p1 using compound 14-11-p3 as starting material and purified by preparative HPLC (Type 4 separation method).

Synthesis of compound 14-p4.

Compound 14-p4 (1.04 mg, HPLC retention time: 36.036 min, m/z: [M+H]+ 707.5) was obtained as a white solid in a similar manner to compound 14-p1 using compound 14-11-p4 as starting material and purified by preparative HPLC (Type 4 separation method).

Options for the implementation of biological methods of analysis

Embodiment 1. Type I interferon activation assay.

THP-1 Dual cells (Invivogen) were seeded at 100,000 cells/well in a 96-well plate, then phorbol-12-myristate-13-acetate (PMA) was added, and the final concentration of phorbol-12-myristate-13-acetate was 30 ng /ml After 24 hours of incubation, the cells were washed twice with fresh medium and compounds were added to the appropriate wells for a threefold dilution. Compounds were diluted with buffer PB (50 mM HEPES buffer, 100 mM KCl, 3 mM MgCl ₂ , 0.1 mM DTT (dithiothreipol, DTT), 85 mM sucrose, 1 mM ATP, 0.1 mM GTP (glutamyltransferase, GTP), 0 .2% bovine serum albumin and 5 μg/ml digitonin) and the maximum dose was 10 μM, the minimum dose was 0.0015 μM. Cells were incubated for 30 min and then washed twice with fresh medium again. Fresh medium was added and cells were incubated for an additional 24 hours. After incubation, 10 µl of supernatant and 50 µl of QUANTI-Luc (Invivogen) were added to a 96-well plate, and fluorescence was read using TECAN. The level of interferon expression was proportional to the fluorescence intensity. A curve was built using Graphpad Prism and the EC50 compounds were analyzed.

Connection number EC50 (µM) Connection number EC50 (µM) 1-p1 >10 8-rZ 99.53 1-r2 >10 8-r4 38.80 1-rZ >10 9-p1 >100 1-r4 25.615 9-r2 >100 2-p1 45.755 9-rZ 4.865 2-r2 >100 10-p1 70.69 2-rZ 97.411 10-r2 118.8 2-p4 2.077 10-rZ 0.494 4-p1 6.834 11-r1 60.81 4-r2 0.625 I -p2 9.805 4-rZ 0.417 I -p3 17.50 5-p1 >10 I -p4 0.421 5-r2 51.981 12-p1 >100 5-rZ 58.238 12-r2 19.01 5-r4 3.223 12-rZ 8.628 6-r1 2.273 12-r4 0.565 6-p2 1.063 13-r1 188.1 6-rZ 0.429 13 -r2 351.6 6-r4 0.311 13-rZ 176.0 7-r1 3.464 13 -p4 4.991 7-r2 40.62 14-r1 >100 7-rZ 0.275 14-r2 >100 7-r4 2.307 14-rZ 40.35 8-p1 >100 14-r4 21.52 8-r2 >100 Reference Compound 1 2.942

Embodiment 2. Interferon-P gene stimulator secretion assay.

40 μl of THP-1 type cells (ATCC) were seeded at 16,000 cells/well in a 96-well plate (Corning, 3596). Three-fold dilution compounds were added to the appropriate wells. Compounds were diluted with buffer (RPMI1640+2mM L-glutamine+1x non-essential amino acids+1mM sodium pyruvate+0.5% fetal bovine serum) and the maximum compound dose was 100 μM, the minimum dose was 1.23 μM. After 5 hours of incubation, 2 μl of supernatant was added to a 384-well plate (Greiner, Cat: 784075) and interferon-β secretion was determined using the AllphaLISA IFN-β kit (PerkinElmer, Cat: AL577C). A 1x receptor and donor dilution buffer was prepared, and then 4 μl of 20 μg/ml Anti-pIFNP AlphaLISA type receptor was added to each well. After 30 min incubation at room temperature, 4 μl of 2 nM biotinylated Anti-pIFNe type antibody was added to a 384-well plate and incubated overnight at 4°C. 10 µl of 40 µg/ml Streptavidin type donor (SA) was added, which was diluted 1x with buffer and then incubated at room temperature for 30 min. Fluorescence was read using TECAN. The expression level of interferon-β was proportional to the fluorescence intensity. Created a curve with

Graphpad Prism and analyzed EC ₅₀ compounds.

Connection number EC so (µM) 2-p4 47.64 4-p2 36.22 4-rZ 94.39 6-p2 58.09 6-rZ 9.84 6-r4 9.40

Option 3. Efficacy study in a CT-26 colon tumor xenograft model in mice.

Cell culture.

Colon tumor cells type CT26 (ATCC) were maintained in vitro as a single layer culture in RPMI1640 medium supplemented with 10% fetal bovine serum at 37° C. in an atmosphere of 5% CO2 in air. Tumor cells were typically passaged twice a week with trypsin-EDTA treatment. Cells growing in the exponential growth phase were harvested and counted for tumor inoculation.

Animals: BALB/c nude mice, 6-8 weeks old, 19-22 g, animal supplier: Beijing Vital River Laboratory Animal Technology Co., Ltd.

Created 4 groups as follows. ___________________________________

Group Number Remedy for treatment Dose Path dosing Current scheme 1 8 Drug carrier medium - i.t. Q3Dx3 doses 2 8 Reference Compound 1 2 mg/kg i.t. Q3Dx3 doses 3 8 Compound 6-p3 1 mg/kg i.t. Q3Dx3 doses 4 8 Compound 6-p3 2 mg/kg i.t. Q3Dx3 doses

Note: i.t.: intratumoral injection, Q3D: once every 3 days.

Experimental Method: Each mouse was subcutaneously inoculated with tumor cells of the CT-26 type (1x10 ⁵ ) in 0.1 ml on the right flank. Tumor growth was observed regularly when tumor volume reached 150 mm ³ , mice were randomized based on tumor volume and body weight and treated according to the schedule. Weight and tumor size of mice were measured 2-3 times a week throughout the experiment.

Tumor size formula:

tumor volumes (mm ³ )=0.5 x(tumor size on the long axis χ tumor size on the short axis ² ).

Curves of tumor growth for different 4 groups are shown in the drawing. The result showed that compared with the positive control of Comparative Compound 1, the compound of the present invention can show better efficacy in the CT26 type subcutaneous colon tumor model in mice.

Note: Comparative compound 1 used in biological assay embodiments 1 and 3 was MLRR-CDA (ammonium salt), CAS No.: 1638750-96-5, the synthesis method may refer to compound 22 of PCT patent application WO 2014/ 189805 A1.

Claims (12)

CLAIM 1. An analogue of a cyclic dinucleotide, which is a compound of general structural formula (VI) or (VII), its stereoisomer or a pharmaceutically acceptable salt; -54041924 where w< *W ₃ RaO Η ³ T °0 AA ⁰⁼ ^ AA ^'n^w 0 ^{Vl Rc} 0 ^V 1 R _c X o .. _r „ ' B-2 ' V-17 , _lp B-18 B1 represents , ^or , R _B W Yi^0 _x/ Yl^X|i /jyy/T y 400¾400¾ B-4B-6 The ₂ represents ^or ' R ² is independently hydrogen, halogen, cyano, hydroxyl, thiol, amino, azido, C2-6 alkenyl, C2-6 alkynyl, C1-6 alkyl, halo-C1-6 alkyl, halo-C1-6 alkoxy, halo- C1-6 alkylthio, C1-6 alkylamino, OC(O)R ^a or OR ^a ; C2-6 alkenyl, C _2-6 alkynyl or C _1-6 alkyl is unsubstituted or selectively substituted at any position with 1-3 substituents selected from halo, hydroxyl, amino, azido and cyano; R ²¹ is independently hydrogen, halogen, cyano, hydroxyl, thiol, amino, azido, C2-6 alkenyl, C2-6 alkynyl, C1-6 alkyl, halo-C1-6 alkyl, halo-C1-6 alkoxy, halo- C1-6 alkylthio, C1-6 alkylamino, OC(O)R ^a or OR ^a ; C2-6 alkenyl, C _2-6 alkynyl or C _1-6 alkyl is unsubstituted or selectively substituted at any position 1-3 with halogen, hydroxyl, amino, azido and cyano; each Y and Y1 are independently CRE or N; U is CHRE' or NRD'; each V and V1 is independently CRE or N, each V2 and V ₃ is independently N or CH; W is O or S; each W1 is N or CH, each W2 is CRF', W ₃ and W4 are independently CH; each of R _a , R _b , Rc, RE, RE', RE and RF' is independently H, halogen, -CN, -NO2, -N ₃ , R ^c , -SR ^c , -OR ^c , -OC( O)R ^c , -OC(O)OR ^c , -OC(O)NR ^b R ^c , -C(O)OR ^c , -C(O)R ^c , -C(O)NR ^b R ^c , - NR ^b R ^c , -NR ^b C(O)R ^c , -N(R ^b )C(O)OR ^c , -N(R ^a )C(O)NR ^b R ^c , -NR ^b S(OhR ^c , -NR ^b C(=NH)R ^c , -NR ^b C(=NR ^c )NH2, -S(O)1-2R ^c , -S(O)2NR ^b R ^c , or -NR ^a S(O) 2NR ^b R ^c ; R _D ' is independently H or R ^c ; each of R ^a and R ^b independently represents H, C _2-6 alkenyl, C _2-6 alkynyl, C _1-10 alkyl, halo-C _1-6 alkyl; each R ^c is independently H, substituted or unsubstituted C _1-10 alkyl, substituted or unsubstituted C _2-8 alkenyl, substituted or unsubstituted C _2-8 alkynyl, substituted or unsubstituted C _3-10 cycloalkyl, substituted or unsubstituted C _{6- 10} aryl, substituted or unsubstituted 3-10 membered heterocycloalkyl, substituted or unsubstituted 5-10 membered heteroaryl, substituted or unsubstituted C _6-10 aryl-C _1-6 alkyl, substituted or unsubstituted C _3-10 cycloalkylC _1-6 alkyl , substituted or unsubstituted 3-10 membered heterocycloalkyl-C _1-6 alkyl, substituted or unsubstituted 5-10 membered heteroaryl-C _1-6 alkyl; C _1-10 alkyl, C _2-8 alkenyl, C _2-8 alkynyl, C _3-10 cycloalkyl, C _6-10 aryl, 3-10-membered heterocycloalkyl, 5-10-membered heteroaryl, C _6-10 aryl- C _1-6 alkyl, C _3-10 cycloalkyl-C _1-6 alkyl, 3-10 membered heterocycloalkyl-C _1-6 alkyl, or 5-10 membered heteroaryl-C _1-6 alkyl is unsubstituted or selectively substituted in any position of one or more R ^d ; each R ^d is independently halo, halo-C1-6 alkyl, halo-C1-6 alkoxy, C1-6 alkyl, -CN, -N3, -SR ^c , -OR ^c , -C(O)R ^c , - NR ^c R ^c , substituted or unsubstituted C6-10 aryl, substituted or unsubstituted 5-10 membered heteroaryl, substituted or unsubstituted _C3-10 cycloalkyl, or substituted or unsubstituted 3-10 membered heterocycloalkyl; C _6-10 aryl, 5-10 membered heteroaryl, C _3-10 cycloalkyl or 3-10 membered heterocycloalkyl is unsubstituted or selectively substituted at any position with one or more substituents selected from halogen, hydroxyl, cyano, amino, Cm alkyl, halo-Cl _-4 alkyl, Cm alkoxy, Cm alkylamino and halo-CM alkoxy; each of R ^e and R ^e is independently C _2-6 alkenyl, C _2-6 alkynyl, C 1-10 alkyl, haloCM alkyl; where each heteroaryl independently includes 1, 2 or 3 heteroatoms selected from nitrogen, oxygen or sulfur; and each heterocycloalkyl independently comprises 1, 2, 3, or 4 heteroatoms selected from nitrogen, oxygen, or sulfur. 2. A cyclic dinucleotide analog which is a compound of general structural formula (VI) or (VII), its stereoisomer or pharmaceutically acceptable salt according to claim 1, where B ₁ is any of the following structures: and/or B ₂ is any of the following structures: 3. An analogue of a cyclic dinucleotide, which is a compound of general structural formula (VI) or (VII), its stereoisomer or pharmaceutically acceptable salt according to claim 1, where R ² independently represents hydrogen, halogen, hydroxyl or OR ^a ; and/or R ²¹ independently represents hydrogen, halogen, hydroxyl or OR ^a ; and/or each R ^a is independently Cm alkyl or halo-CM alkyl. 4. Analog of a cyclic dinucleotide, which is a compound of general structural formula (VI) or (VII), its stereoisomer or pharmaceutically acceptable salt according to claim 1, where - 56 041924 Bi AT 2 R ² is is independently hydrogen, halogen, hydroxyl or OR ^a ; R ²¹ independently represents hydrogen, halogen, hydroxyl or OR ^a ; each R ^a is independently C _1-4 alkyl or halo-C _1-4 alkyl. 5. Analog of a cyclic dinucleotide, which is a compound of general structural formula (VI) or (VII), its stereoisomer or pharmaceutically acceptable salt according to claim 1, where Bi AT 2 R ² is is independently hydrogen, halogen, hydroxyl or OR ^a ; R ²¹ independently represents hydrogen, halogen, hydroxyl or OR ^a ; each R ^a is independently C _1-4 alkyl or halo-C _1-4 alkyl. 6. Analog of a cyclic dinucleotide, which is a compound of general structural formula (VI) or (VII), its stereoisomer or pharmaceutically acceptable salt according to claim 1, where the stereoconfiguration P is (Sp, Sp), (Sp, Rp), ( Rp, Rp) or (Rp, Sp). 7. A cyclic dinucleotide analogue which is a compound of general structural formula (VI) or (VII), its stereoisomer or pharmaceutically acceptable salt according to claim 6, wherein B ₁ is and/or B ₂ is in formula VI, R ² is —OH; R ²¹ is -OH; in formula VII R ² is -OH or -OCH ₃ ; R ²¹ is -OH or F. 8. A cyclic dinucleotide analogue which has any of the following structures, a stereoisomer or a pharmaceutically acceptable salt thereof: - 57 041924 - 58 041924 - 59 041924 - 60 041924 - 61 041924 9. Pharmaceutical composition as an agonist of STING (interferon gene stimulator), containing a therapeutically effective amount of the compound, its stereoisomer or pharmaceutically acceptable salt according to at least one of claims 1 to 8, and a pharmaceutically acceptable excipient. 10. The use of a compound, a stereoisomer thereof or a pharmaceutically acceptable salt thereof according to at least one of claims 1 to 8 or a pharmaceutical composition according to claim 9 for the preparation of a STING agonist, vaccine adjuvant or medicament for the treatment, alleviation and/or prevention of a disease mediated by STING, or to regulate the proliferation of T cells or other immune cells, or to treat and/or alleviate cancer. 11. Use according to claim 10, wherein the STING-mediated disease is a viral infection or other infectious diseases, autoimmune diseases, or malignancies. 12. Combination composition containing a compound, its stereoisomer or pharmaceutically acceptable salt according to at least one of claims 1 to 8 or a pharmaceutical composition according to claim 9 and other types of therapeutic agents for the treatment of cancer.